S. Kiss, M. Simihăian (Auth.) - Improving Efficiency of Urea Fertilizers by Inhibition of Soil Urease Activity-Springer Netherlands (2002) PDF

IMPROVING EFFICIENCY OF UREA FERTILIZERS
BY INHIBITION OF SOIL UREASE ACTIVITY

Improving Efficiency
of Urea Fertilizers
by Inhibition
of Soil Urease Activity
by
S. Kiss
Babe§-Bolyai University,
Department of Plant Physiology,
Laboratory for Environmental Enzymology and Microbiology, Romania
and
M. Simih3ian
Environmental Protection Agency,
Department of Environmental Management, Romania
SPRINGER-SCIENCE+BUSINESS MEDIA, B.V.

Library of Congress Cataloging-in-Publication Data
ISBN 978-90-481-5966-6 ISBN 978-94-017-1843-1 (eBook)

DOI 10.1007/978-94-017-1843-1
Printed on acid-free paper
AII Rights Reserved

© 2002 Springer Science+Business Media Dardrecht
Originally published by Kluwer Academic Publishers in 2002
Softcover reprint ofthe hardcover Ist edition 2002
No part of the material protected by this copyright notice may be reproduced or
utilized in any form ar by any means, electronic or mechanical,
including photocopying, recording or by any information storage and
retrieval system, without written permission from the copyright owner.
I
Preface
The purpose of our present work is to review the fundamental studies on inhibition of
soil urease activity and the applied studies on improving efficiency of urea fertilizers by
inhibition of soil urease activity. The general literature on these topics covers 65 years,
and the patent literature comprises a period of nearly 40 years.
Studies related to inhibition of soil urease activity were performed in a great number
of countries' well representing all the continents. Full texts of the papers describing
these studies were published in one of 18 languages·'.
The literature data reviewed are structured into 10 chapters, 81 subchapters, and 224
sections. The bibliographical list consists of 830 papers cited.
·In alphabetical order: Argentina, Armenia, Australia, Austria, Belgium, Belorussia, Brazil. Bulgaria, Canada,
China, Costa Rica, Cuba. Czech RepUblic, Egypt, Estonia, France, Georgia (Gruzia), Germany, Hungary,
India, Iraq, Ireland, Israel, Italy. Japan, Kazakhstan, Lithuania, Malaysia, Moldova, Netherlands, New
Zealand, Pakistan, Philippines, Poland, Romania, Russia, Saudi Arabia, Slovakia, South Africa, South
Korea, Spain, Sri Lanka. Sudan, Sweden, Thailand, Turkey, Ukraine, United Kingdom, United States of
America. Uzbekistan .
•• In alphabetical order: Afrikaans, Bulgarian, Chinese, Czech, English, French, German, Hungarian, Italian,
Japanese, Korean, Polish, Portuguese, Romanian, Russian, Spanish, Swedish, Ukrainian.
III
Acknowledgements
The authors wish to express their gratitude to Mr.Arno Flier, Publishing Editor, Biosciences
Unit, Kluwer Academic Publishers, and to his collaborator, Ms. Claire van Heukelom, for
their kind co-operation in publishing this book. We are also grateful to Professors Colleen
Sanders, Rita Moore, and Debra Taevs for their precious collegial help with revision of the
English language of our manuscript, and to Informatician Alina Veza, of the Editorial Board
of the journal Studia Universitatis Babe~-Bolyai. for her precious collegial advice in giving
the manuscript a camera-ready form. We also thank Dr. Engineer Marian Proorocu,
Director. and Economist Florica pacurar. Head of the Economics Department of the
Environmt:-'I1tal Protection Agt:-'llCY of Cluj County, for the valuable moral and material
support we received from them during the whole period of the elaboration of our
manuscri pt.
v
CONTENTS
Preface III
Acknowledgements IV
INTRODUCTION
Chapter 1. Inorganic Compounds Tested for Evaluation of Their
Inhibiting Effect on Soil Urease Activity, Urea Hydrolysis,
Ammonia Volatilization,and Nitrous Oxide Emission 5
1.1. HEAVY METAL COMPOUNDS 5
1.2. LIGHT METAL COMPOUNDS 19
1.3. SALTS OF ALKALI METALS AND ALKALINE EARTH METALS 20
1.3 .1. Effect ofAlkali Metal and Alkaline Earth Metal Salts
on Urease Activity and Urea Hydrolysis 20
1.3.2. Effect ofAlkali Metal and Alkaline Earth Metal Salts
on Ammonia Volatilization 26
1.4. BORON COMPOUNDS 30
1.5. FLUORIDES 33
1.6. ARSENIC COMPOUNDS 34
1.7. SULFUR COMPOUNDS 35
1.8. OTHER INORGANIC COMPOUNDS 42
Chapter 2. Organic Compounds Tested for Evaluation of Their

Inhibiting Effect on Soil Urease Activity,Urea Hydrolysis,
Ammonia Volatilization, and Nitrous Oxide Emission 43
2.1. ORGANIC MERCURY COMPOUNDS 43
2.2. ORGANO BORON ACID COMPOUNDS 45
2.3. FORMALDEHYDE 46
2.4. HEXAMETHYLENETETRAMINE 47
2.5. UREA DERIVATIVES 47
2.6. DITHIOCARBAMATES 52
2.7. THIURAM DISULFIDES AND SULFIDES 56
2.8. XANTHATES 58
2.9. HYDROXAMIC ACIDS 60
2.9.1. Monohydroxamic Acids 60
2.9.2. Dihydroxamic Acids 65
2.10. MALEIMIDES 66
2.11. MALEIC HYDRAZIDE 69
2.12. MUCOCHLORIC ACID 69
2.13. BROMO-NITRO COMPOUNDS 70
2.14. HETEROCYCLIC SULFUR COMPOUNDS 71
2.14.1. Tetrahydro-l. 3.5 -thiadiazine-2-thiones 71
2.14.2. 1. 3. 4-Thiadiazoline-2-thiones 72
VI
2.14.3. Rhodanine-5-acetic Acid and Its Derivatives 75

2.14.4.2-Mercaptobenzothiazole 77
2.14.5.2-Thiocarboxamidothiazoles 77
2.15. MONOHYDRIC PHENOLS 78
2.16. POL YHYDRIC PHENOLS AND QUINONES 82
2.17. THIOPYRIDINE-N-OXIDES, THIOPYRIDINES,
AND THIOPYRIMIDINES 101
2.18. N,N' -DIHALO-2-IMIDAZOLIDINONES AND
N-HALO-2-0XAZOLIDINONES 103
2.19. y-L-GLUTAMYL NITROANILIDES 104
2.20. PHOSPHORO(MONO)AMIDATES, PHOSPHORODIAMIDATES,
AND THIOPHOSPHORODIAMIDATES 105
2.20.1. The Patented Compounds and the First Studies on
Their Inhibitory Effect on Soil Urease Activity 106
2.20.2. Further Studies on Phenylphosphorodiamidate (PPDA)
and Some Other Phosphoroamides 113
2.20.3. Effect of Phenylphosphorodiamidate (PPDA) on
Immobilization of Urea-N in Soils 129
2.20.4. Stability of Phenylphosphorodiamidate (PPDA) 130
2.20.4.1. Stability ofPPDA in Solutions (Induding Urea Melt)
and in Solid State 130
2.20.4.2. Stability ofpPDA in Soils 134
2.21. 2-AMINE-2-0XIDE-l,3,2-BENZODIOXAPHOSPHOLE AND
ITS PHOSPHORODIAMIDIC ACID ESTERS 138
2.22. POLYPHOSPHORODIAMIDES 139
2.23. OXIMATED O-DIAMINOPHOSPHINYL DERIVATIVES 140
2.24. OXIDIZED DIAMINOPHOSPHINYL SULFUR DERIVATIVES 140
2.25. DIAMIDOPHOSPHOROTHIOLATE AND
DIAMIDOTHIOPHOSPHOROTHIOLATE COMPOUNDS 141
2.26. PHOSPHORIC TRIAMIDES AND THIOPHOSPHORIC TRIAMIDES 142
2.26.1. The Patented Compounds and the First Studies on
Their Inhibitory Effect on Soil Urease Activity 142
2.26.2. Further Studies on Phosphoric Triamides and
Thiophosphoric Triamides 146
2.27. CYCLOTRIPHOSPHAZATRIENE DERIVATIVES 155
2.28. PHOSPHORYLATED 2-0XIMINOPHENYLACETONITRILE
COMPOUNDS 160
2.29. 2-THIONO-5,5-DIMETHYL-l ,3,2-DIOXAPHOSPHORINANE
COMPOUNDS 161
2.30. ANTIMETABILITES 162
2.30.l. Su({anilamide 162
2.30.2. Other Antimetabolites 163
2.31. NATURAL PRODUCTS 165
2.31.1. Coal and Peat 165
2.31.2. Humic Substances. Lignins. and Tannins 166
2.31.3. Plant Residues and Extracts 169
VII
2.31.3.1. Residues and Extracts Containing Polyphenols 169

2.31.3.2. Extracts Containing Saponins 171
2.31.3.3. Neem Cake, Oil, and Extracts 171
2.31.3.4. Karanja Cake 175
2.31.3.5. Mahua Cake 176
2.31.3.6. Other Plant Materials 177
2.31.4. Microbial Products 177
2.32. A MISCELLANEOUS GROUP OF ORGANIC COMPOUNDS 178
Chapter 3. Combined Use of Inhibitors of Soil Urease Activity 179

3.1. COMBINED USE OF HEAVY METAL COMPOUNDS
WITH OTHER INHIBITORS 179
3.2. COMBINED USE OF FORMALDEHYDE OR ANOTHER
ALDEHYDE WITH OTHER INHIBITORS 179
3.3. COMBINED USE OF HEXAMETHYLENETETRAMINE (HMT A)
3.4. COMBINED USE OF DITHIOCARBAMATES WITH
OTHER INHIBITORS 180
3.5. COMBINED USE OF HYDROXAMIC ACIDS WITH
3.6. COMBINED USE OF POLYHYDRIC PHENOLS AND QUINONES
3.7. COMBINED USE OF PHENYLPHOSPHORODIAMIDATE (PPDA)
3.8. COMBINATIONS CONTAINING HUMIC SUBSTANCES 183
3.9. COMBINATIONS CONTAINING LIGNOSULFONATES 184
Chapter 4. Comparative Studies on the Efficiency of Different

Inhibitors on Soil Urease Activity 187
4.1. COMPARISON OF HEAVY METAL COMPOUNDS WITH
4.2. COMPARISON OF ALKALINE EARTH METAL SALTS
4.3. COMPARISON OF INORGANIC BORON COMPOUNDS
4.4. COMPARISON OF INORGANIC SULFUR COMPOUNDS
4.5. COMPARISON OF ORGANIC MERCURY COMPOUNDS
4.6. COMPARISON OF UREA DERIVATIVES WITH OTHER
INHIBITORS 191
4.7. COMPARISON OF POLYHYDRIC PHENOLS AND QUINONES
4.8. COMPARISON OF PHENYLPHOSPHORODIAMIDATE (PPDA)
VIII
4.9. COMPARISON OF PHOSPHORIC TRIAMIDE (PTA) AND

THIOPHOSPHORIC TRIAMIDE (TPT A) COMPOUNDS WITH
4.9.1. Comparative Studies on the Effect ofPTA and TPTA
Compounds and Other Inhibitors on Soil Urease Activity,
Urea Hydrolysis, and Ammonia Volatilization 198
4.9.1.1. Comparison ofnBTPTA with Ammonium Thiosulfate (ATS) 198
4.9.1.2. Comparison ofnBTPTA with Pyrite, Phosphogypsum, and KCI 198
4.9.1.3. Comparison of nBTPTA and/or TPTA, PTAs, and
Alkyl-PDAs with PPDA 199
4.9.1.4. Comparison ofnBTPTA, PTAs, and Alkyl-PDAs
with PPDA and Hydroquinone (HQ) 201
4.9.1.5. Comparison ofnBTPTA with PPDA and
Ammonium Thiosulfate (ATS) 204
4.9.1.6. ComparisonofnBTPTAwithPPDA 205
4.9.2. Comparative Studies on the Effect ofnBTPTA and Other
Inhibitors on the Immobilization ofUrea-N in Soils 211
4.9.3. Comparative Studies on the Stability ofPTA and TPTA
Compounds and Other Inhibitors 213
4.9.3.1. Comparative Studies on the Stability of Phosphoryl
Triamide (PTA), Thiophosphoryl Triamide (TPT A),
nBTPTA. and Other Inhibitors in Solutions
(Including Urea Melt) and in Solid State 213
4.9.3.2. Comparative Studies on the Stability of PTA, TPTA,
nBTPT A, and Other Inhibitors in Soils 216
4.10. COMPARISON OF CYCLOTRIPHOSPHAZATRIENE (CTPAT)
DERIVATIVES WITH OTHER INHIBITORS 220
Chapter 5. Compounds Tested for Evaluation of Their Inhibiting

Effect on Both Soil Urease Activity and Nitrification 221
5.1. EFFECT OF NITRIFICATION INHIBITORS ON SOIL
UREASE ACTIVITY 221
5.1.1. Effect ofNitrapyrin and Other Nitrification Inhibitors.
Except Azide and Dicyandiamide, on Soil Urease Activity 221
5.1.2. Effect of Sodium and Potassium Azide on Soil Urease Activity 228
5.1.3. Effect ofDicyandiamide on Soil Urease Activity 229
5.2. UREASE INHIBITORS ALSO POSSESSING NITRIFICATION-
INHIBITING CAPACITY 233
5.2.1. Inorganic Compounds 233
5.2.1.1.Heavy Metal Compounds 233
5.2.1.2. Salts of Alkali Metals and Alkaline Earth Metals 234
5.2.1.3. Fluorides 234
5.2.1.4. Sulfur Compounds 235
5.2.2. Organic Compounds 235
5.2.2.1. Organic Mercury Compounds 235
5.2.2.2. Urea Derivatives 235
IX
5.2.2.3. Thiuram Disulfides 236

5.2.2.4. Xanthates 236
5.2.2.5.Heterocyclic Sulfur Compounds 236
5.2.2.6. Monohydric Phenols 236
5.2.2.7. Polyhydric Phenols and Quinones 237
5.2.2.8. N-Halamine Compounds 238
5.2.2.9. Phosphorodiamides 239
5.2.2.10. Phosphoric Triamides 239
5.2.2.11. Humic Substances and Lignosulfonates 241
5.2.2.12. Plant Residues 241
5.2.3. Mixtures of Inorganic and Organic Compounds 242
Chapter 6. Soil Urease Inhibitors Used in Combination with

Nitrification and/or Algal Inhibitors 243
6.1. COMBINED USE OF UREASE AND NITRIFICATION INHIBITORS 243
6.2. COMBINED USE OF UREASE AND ALGAL INHIBITORS 247
6.3. COMBINED USE OF UREASE, NITRIFICATION, AND ALGAL
INHIBITORS 249
Chapter 7. Effect of Soil Urease Inhibitors on Germination,

Growth, and Yield of Plants 251
7.1. EFFECT OF UREASE INHIBITORS ON MAIZE (Zea mays) 251
7.1.1. Effect ofAlkali Metal and Alkaline Earth Metal Salts 251
7.1.2. Effect ofFluorides 251
7.1.3. Effect ofInorganic Sulfur Compounds 252
7.1.4. Effect of Organic Mercury Compounds 253
7.1.5. Effect of Polyhydric Phenols and Quinones 253
7.1.6. Effect ofPh osphorodiamides 254
7.1.7. Effect of Phosphoric Triamide (PTA) and Thiophosphoric
Triamide (TPTA) Compounds 256
7.1.8. Effect ofCyclotriphosphazatriene (CTPAT) Derivatives 260
7.1.9. Effect ofAntimetabolites 261
7.1.10. Effect ofLignosulfonates 261
7.1.11. Effect of Plant Materials 262
7.1.12. Effect of Combined Urease and Nitrification Inhibitors 262
7.2. EFFECT OF UREASE INHIBITORS ON WHEAT (Triticum aestivum) 263
7.2.4. Effect of Dithiocarbamates 264
7.2.5. Effect ofHydroxamic Acids 265
7.2.6. Effect ofBromo-nit,.o Compounds 265
7.2.8. Effect of Ph osphorodiamides 269
x
7.2.10. Effect of Lignosulfonates 279

7.3. EFFECT OF UREASE INHIBITORS ON RICE (Oryza sativa) 280
7.3.1. Effect of Inorganic Boron Compounds 280
7.3.3. Effect of Poly hydric Phenols and Quinones 280
7.304. Effect ofPh osphorodiamides 281
7.3 .6. Effect ofPlant Materials 287
7.3.7. Effect of Combined Urease, Nitrification and/or Algal Inhibitors 287
704. EFFECT OF UREASE INHIBITORS ON BARLEY (Hordeum vulgare) 288
704.1. Effect of Heavy Metal Compounds 288
704.2. Effect of Organic Mercury Compounds 288
704.3. Effect of Urea Derivatives 289
70404. Effect of Polyhydric Phenols and Quinones 289
704.5. Effect of Phosphorodiamides 289
704.6. Effect ofPhosphoric Triamide (PTA) and Thiophosphoric
704.7. Effect ofCyclotriphosphazatriene (CTPAT) Derivatives 291
704.8. Effect o.fCombined Urease and Nitrification Inhibitors 291
7.5. EFFECT OF UREASE INHIBITORS ON OATS (Avena sativa) 292
7.5.1. Effect of Heavy Metal Compounds 292
7.5.2. Effect ofInorganic Boron Compounds 292
7.504. Effect of Urea Derivatives 293
7.5.5. Effect of Phosphorodiamides 293
7.6. EFFECT OF UREASE INHIBITORS ON RYE (Secale cereale) 294
7.7. EFFECT OF UREASE INHIBITORS ON SORGHUM (Sorghum bicolor) 295
7.7.1. Effect o.f Inorganic Sulfur Compounds 295
7.704. Effect ofPh osphorodi amides 296
7.7.5. Effect ofPhosphoric Triamide (PTA) and Thiophosphoric
7.7.6. Effect o.fCyclotriphosphazatriene (CTPAT) Derivatives 297
7.8. EFFECT OF UREASE INHIBITORS ON GRASSES 297
7.8.2. Effect ofInorganic Boron Compounds 298
XI
7.8.4. Effect of Urea Derivatives 299

7.8.5. Effect ofDithiocarbamates 300
7.8.6. Effect ofXanthates 300
7.8.7. Effect ofBromo-nitro Compounds 300
7.8.9. Effect ofPh osphorodiamides 303
7.8.10. Effect ofPhosphoric Triamide (PTA) and Thiophosphoric
7.8.11. Effect ofLignin 311
7.8.12. Effect ofPlant Materials 311
7.8.13. Effect of Combined Urease Inhibitors 312
7.8.14. E.ffect of Combined Urease and Nitrification Inhibitors 312
7.9. EFFECT OF UREASE INHIBITORS ON LEGUMINOUS PLANTS 313
7.9.1. Effect ofHeavy Metal Compounds 313
7.9.4. Effect ofThiuram Disulfides 314
7.9.5. Effect ofHydroxamic Acids 314
7.9.7. Effect ofPh 0 sphorodiamides 314
7.9.8. E.ffect of Phosphoric Triamide (PTA) and Thiophosphoric
7.9.9. Effect ofCyclotriphosphazatriene (CTPAT) Derivatives 315
7.10. EFFECT OF UREASE INHIBITORS ON OTHER PLANTS 315
7.10.6. Effect ofHeterocyclic Sulfur Compounds 316
7.10.10. Effect ofHumic Substances 318
7.10.12. Effect of Combined Urease Inhibitors 318
Chapter 8. Effect of Urease Inhibitors on Other Enzyme Activities,

Microbial Counts, and Biomass as well as on
Respiration and Other Microbial Processes in Soils 321
8.1. EFFECT OF UREASE INHIBITORS ON OTHER ENZYME
ACTIVITIES IN SOILS 321
8.1.2. Effect ofAlkali and Alkaline Earth Metal Salts 322
XII
8.1.3. Effect of Fluorides 323

8.1.7. Effect ofThiuram Disulfides 324
8.1.8. Effect ofHeterocyclic Sulfur Compounds 324
8.1.9. Effect ofMonohydric Phenols 324
8.1.10. Effect ofPolyhydric Phenols and Quinones 324
8.1.11. Effect ofPhosphorodiamides 328
8.l.l2. Effect ofPhosphoric Triamide (PTA) and Thiophosphoric
8.1.13. Effect ofHumic Substances 328
8.1.14. Effect ofLignosu(fonates 328
8.1.15. Effect ofPlant Materials 329
8.2. EFFECT OF UREASE INHIBITORS ON MICROBIAL COUNTS
AND BIOMASS AS WELL AS ON RESPIRAnON AND OTHER
MICROBIAL PROCESSES IN SOILS 329
8.2.1. Effect on Microbial Counts and Biomass 329
8.2.1.1. Effect of Heavy Metal Compounds 329
8.2.1.2. Effect of Fluorides 330
8.2.1.3. Effect of Urea Derivatives 330
8.2.1.4. Effect ofDithiocarbamatcs 330
8.2.1.5. Effect of Thiuram Disulfides 331
8.2.1.6. Effect of Heterocyclic Sulfur Compounds 331
8.2.1.7. Effect of Monohydric Phenols 331
8.2.1.8. Effect of Phosphorodiamides 331
8.2.1.9. Effect of Phosphoric Triamide (PTA) and Thiophosphoric
8.2.1.10. Effect ofCyclotriphosphazatriene (CTPAT) Derivatives 332
8.2.l.l1. Effect of Humic Substances 332
8.2.1.12. Effect of Lignosulfonates 332
8.2.2. Effect on Respiration 334
8.2.2.2. Effect of Alkali Metal Salts 334
8.2.2.3. Effect ofInorganic Sulfur Compounds 334
8.2.2.4. Effect ofDithiocarbamates 334
8.2.2.5. Effect of Polyhydric Phenols and Quinones 334
8.2.2.6. Effect of Phosphoric Triamide (PTA) and Thiophosphoric
Triarnide (TPTA) Compounds 335
8.2.2.7. Effect ofCyclotriphosphazatriene (CTPAT) Derivatives 335
8.2.3. Effect on Cellulose Decomposition 335
8.2.3.1. Effect of Alkali Metal and Alkaline Earth Metal Salts 335
8.2.3.2. Effect of Phosphorodiami des 335
8.2.4. Effect on Methane Emission and Oxidation 336
8.2.4.1. Effect of Combined Urease and Nitrification Inhibitors 336
XIII
8.2.4.2. Effect of Phosphoroamides 336

8.2.5. Effect on Nitrogen Mineralization 336
8.2.5.2. Effect of Phospho roamides 336
8.2.6. Effect on Nitrification 337
8.2.7. Effect on Denitrification 337
8.2.7.2. Effect of Organic Mercury Compounds 339
8.2.7.3. Effect of Polyhydric Phenols and Quinones 339
8.2.7.4. Effect of Phospho roam ides 339
8.2.7.5. Effect of Combined Urease and Nitrification Inhibitors 340
8.2.8. Effect on N2 Fixation 340
8.2.8.2. Effect of Fluorides 340
8.2.8.3. Effect of Dithiocarbamates 340
8.2.8.4. Effect of Thiuram Disulfides 341
8.2.8.5. Effect of Humic Substances 341
8.2.9. Effect on Sulfur Oxidation 341
8.2.10. Effect on Adenosine Triphosphate (ATP) Content 341
8.2.10.2. Effect of Dithiocarbamates 342
8.2.10.3. Effect ofThiuram Disulfides 342
Chapter 9. Use of Urease Inhibitors in the Analysis of Urea

and/or Ammonium from Urea-treated Soils 343
Chapter 10. Urease Inhibitors Used with Another Purpose

than Inhibition of Soil Urease Activity 347
10.1. UREASE INHIBITORS AS FERTILIZERS 347
10.1.1. Inorganic Boron Compounds 347
10.1.2. Inorganic Sulfur Compounds 347
10.1.3. Hexamethylenetetramine 347
10.1.4. Urea Derivatives 347
10.1.5. Phosphoroamides 348
10.1.6. Cyclophosphazene Compounds 350
10.1.7. Plant Materials 351
10.2. UREASE INHIBITORS USED FOR CONTROLLING AMMONIA
(AND ODOR) EMISSION FROM LIVESTOCK WASTES 352
10.3. UREASE INHIBITORS USED FOR PREVENTING TOXICITY
OF UREA-AMENDED FODDERS 353
10.4. UREASE INHIBITORS USED FOR TREATMENT AND
PREVENTION OF SOME HUMAN DISEASES 356
10.5. UREASE INHIBITORS USED FOR PREVENTING
ACID MINE DRAINAGE 358
XIV
CONCLUSIONS 359
REFERENCES 361
SUBJECT INDEX 393
INTRODUCTION
The increase of food production to meet the growing needs related to demographic
explosion is largely conditioned by the efficiency of agricultural fertilizers.
The use of urea as a nitrogen fertilizer has increased tremendously in the last 3--4
decades in both developed and developing countries. According to all predictions, the
increasing trend in fertilizer urea usage will continue. TIlliS, urea becomes gradually the
most important fertilizer in world agriculture.
The growing importance of fertilizer urea should be attributed to its advantages over
other nitrogen fertilizers. TI1ese advantages include: high nitrogen content (46%); low
cost of manufacture, transportation, storage, and distribution; ease of handling (without
fire and explosion hazard); high solubility in water; reduced COITosivity. In addition,
urea is suitable for production of compound fertilizers; it can be applied to soil in solid
state or in solution; its solution can be used as a filliar spray for some crops and also in
association with many pesticides.
In soil, urea is transformed into ammonium carbonate under the action of a
hydrolytic enzyme, the urease. Thus, urea, a neutral compound, gives rise to an alkaline
product, the ammonium carbonate. The an1I11onium cation may be retained by the
adsorptive complex of the soil, but ammonium carbonate, being an unstable compound,
decomposes producing two gases (ammonia and carbon dioxide) and water:
urease
2H~
• • 2 NH3 + c~ + H:P .
The urease in soil is of microbial origin, but a part of it may originate from plants
and animals.
In soil, urease is present as an accumulated enzyme, adsorbed on organic and
mineral soil particles and/or complexed with humic substances. Activity of the
accumulated urease is much higher than that of the urease mzymes produced by the
momentarily proliferating microorganisms.
In most soils, activity of the accumulated urease is too high, resulting in a rapid
hydrolysis of urea with concomitant rise in pH at the site of hydrolysis and liberation of
ammonia. The free ammonia may dan1age the germinating seeds and young plants and
may be lost through volatilization. The highest losses occur in calcareous soils and in
light-textured ones (low cation-exchange capacity) as well as in soils under pasture. The
high ammonium concentration and pH hinder the bacterial oxidation of nitrites into
nitrates. Consequently, the nitrites accumulate in toxic concentrations. Volatilization of
ammonia may also constitute a problem of air pollution. Moreover, ammonia volatilized
from soil may entL'f lakes and streams in the vicinity and may promote eutrophication.
It is estimated that - due to excessi ve urease activity in different soils - a significant
part (up to 70%) of the applied urea-N is lost through volatilization of an1I11onia; the
average loss is considered to be about 20 or 33%. In other words, the output of one of
five or three fertilizer urea factories is lost and, at the same time, pollutes the
environment.
2
Urea-N may be lost into the atmosphere also as nitrous oxide which contributes to
the greenhouse effect and the destruction of the stratospheric ozone layer.
Prevention of the undesirable effects of excessive urea hydrolysis in soil aims at
increasing the efficiency of urea fertilizers. Any increase in this efficiency will increase
the agronomic and economic value of the fertilizers as a means of increasing crop
production, will conserve energy and raw materials needed to manufacture the fertilizers
and will minimize the adverse effects on the environment that may result from
inefficient fertilizer use.
To prevent the excessive urea hydrolysis in soils, investigations were carried out
along four lines.
I. Urea was transforml-'d with aldehydes into compounds that are sparingly soluble
in water (ureaforms, isobutylidene diurea, crotonylidene diurea, etc.). These aldehyde
condensation products slowly decompose in soil. Urea bydrolysis is slow because of the
limited amounts of free urea. Urease activity is not inhibited, and urea remains the
substrate on which the urease will act.
2. Fertilizer urea granules were coated with hydrophobic (water-resistant) materials
(asphalt, waxes, oils, plastics, etc.) or with powders (kaolin, clay, sulfur, Si0 2 , A1 2 0 3 ,
etc.) to limit dissolving of urea. As in the first case, urea hydrolysis is slow because of
the limited amounts of dissolved urea. Urease activity is not inhibited, and urea remains
the substrate of enzyme.
3. Urea was used in association with inhibitor(s) of soil urease activity.
4. Urea was replaced: by adducts of urea with mineral or organic acids (nitric,
phosphoric, boric, oxalic or succinic acid) or with mineral salts or hydroxides of heavy
and ligllt metals (ferric sulfate, aluminium sulfate, felTic hydroxide); by complexes of
urea with stearic acid or with other unbranched, unsubstituted aliphatic compounds
containing at least 6 carbon atoms. Urea in adducts with acids or with metal-containing
compounds hydrolyzes more slowly because of the acidity and metals, respectively,
whereas hydrolysis of urea in complexes with aliphatic compounds is slower because of
the limited amounts of free urea.
To summarize: in the investigations along the first two lines, urea remains the
substrate, and urease is not inhibited; in those along the third line, urea remains the
substrate, but urease is inhibited; in those along the fourth line, the substrate is
modified, whereas the enzyme is inhibited only weakly or is not inhibited at all.
In this review work, we will deal only with the investigations aiming at inhibiting
soil urease activity (the third line of investigations).
In the first investigations this aim was exclusively theoretical: obtaining of
supplementary evidence concerning the enzymatic nature of urea hydrolysis in soil. The
first data on the effects of chemicals on soil urease activity were published by Rotini
(University of Pisa, Italy) in 1935, and by Conrad (California State University, Davis)
in 1940.
Studying hydrolysis of urea in absence and presence of antiseptics, Rotini prepared
reaction mixtures from 50 g of soil + 30 ml of 0.1 'Vo urea solution + 1 m1 of water or 5%
phenol solution. After incubation (42°C/4 hours) , the residual urea was determined. Its
amount was found to be nearly equal in the untreated and phenol-treated soil. The
conclusion was drawn that phenol stopped the growth of microorganisms but did not
lyze them and, thus, did not lead to release of urease from the microbial cells; urease
3
activity measured in the presence of phenol was due to those urease molecules that
existed in soil in a free state even before addition of phenol.
In Conrad's experiment, urea dissolved in 50% ethanol was hydrolyzed in soil at a
lower rate than the urea dissolved in distilled water; the inactivating effect of ethanol on
soil urease was only partial. Treatment of the soil with HgCh, hydroquinone or catechol
led to strong or nearly complete inhibition of soil urease activity. The inhibitions
observed prove that urea hydrolysis in soil is, really, an enzymatically catalyzed
process, thus, confimting the other evidence (e.g.. heat lability of the catalyst, i.e.. of the
urease).
The literature appeared in the 1941-1959 period does not offer information on the
inhibition of soil urease activity, excepting a finding by Kuprevich (1951), according to
which the antibiotic preparation "BIN No.?", containing usnic acid, did not influence
urease activity in soil.
Beginning with 1960 and up to now, the aim of the investigations (ID inhibition of
soil urease activity is not (mly theoretical, i.e.. to obtain supplementary evidence
concerning the enzymatic nature of urea hydrolysis in soil. Contrarily, this aim is rather
practical, Le.. to identify inorganic and organic compounds which, in agricultural
practice, may be used in form of additives to urea fertilizers as inhibitors of soil urease
activity. Accentuation of the practical aim of these investigations was determined by the
world-wide increase in use of urea fertilizers, by evidentiation of the undesirable effects
of the excessivc hydrolysis of urca in soil and, consequently, by the need to increase the
efficiency of urea fertilizers.
The number of chemical compounds tested for evaluation of their effect on soil
urease activity is impressive: more than 14,000 compounds and mixtures of compounds
were tested with this aim. A great number of compounds and mixtures of compounds,
having inhibitory effect on soil urease activity, were patented as inventions in the
U.S.A, Great Britain, France, Germany, in the former U.S.S.R., in Romania and P.R. of
China. Application of some of these patents was also examined by the European Patent
Office. The first patent in this domain was obtained by Hyson (1963) in the U.S.A.
Most patents describing inhibitors of soil urease activity were elaborated by German
and North American investigators.
Of course, all investigations aiming at identification of soil urease inhibitors should
begin with the laboratory phase. This is continued by the phase in which the
investigations arc carried out in vegetation pots and is ended by the phase of
investigations under field conditions.
Effect of the inhibitor should manifest itself not only in a significant reduction of the
urea hydrolysis rate but, consequently, also in limitation of the N losses via al11l11onia
volatilization and nitrous oxide errtission. The inhibitor should be unpolluting, free of
injurious side effects, i.e .. it should have no negative effects on processes related to soil
fertility and should not be toxic to plants, animals, and man. At the same time, the
inl1ibitor should be a stable compound that does not decompose during manufacture,
transportation, and storage of the urea fertilizers. Of course, the inl1ibitor should have a
relatively low cost, and the benefit obtained from its use in agriculture should exceed its
price at purchase and other expenses related to its use. In other words, it should be
4
agronomically, environmentally, and economically better to use than not to use the
inhibitor (see Appendix to References)".
* *
*
Due to space limitation:
a) only the pesticides patented as inhibitors of soil urease activity will be dealt
with;
h) the studies on the inhibition of soil urease activity by high urea concentrations
(substrate excess) (e.g.. Rachhpal-Singh and Nyc, 1984a; Cabrera and Kissel,
1984; Monreal et af.. 1986; MergeI' et af.. 1987; Savant et af.. 1987b; Cabrera
et al.. 1991;Thormiihler and du Preez, 1992; Zhang el af.. 1994) will not be
reviewed; and
c) the studies on the inhibition of ammonia volatilization from urea by activation
of nitrification (e.g.. Fleisher and Hagin, 1981; Praveen-Kumar and Aggarwal,
1988; Goos and de Padua Cruz, 1999) will not be considered .
• The use of soil urease inhibitors is contraindicated only on those flooded rice fields on which the water
control is poor; therefore. urea is lost in the runolrwaters from these fields more easily than the ammonium
ion that can be retained more strongly than urea by the adsorptive complex of the soil (Craswell and Vlek.
1982; Keeney and Sahrawat. 1986).
5
Chapter 1. Inorganic Compounds Tested for Evaluation of Their Inhibiting Effect

on Soil Urease Activity, Urea Hydrolysis, Ammonia Volatilization, and
Nitrous Oxide Emission
1.1. HEAVY METAL COMPOUNDS
Comad (I940) studied the effect of HgCh (mercuric chloride, sublimate) on urease
activity in two California soils. Treating the samples of the first soil (fine sandy loam)
with HgCh· caused a significant reduction in urease activity. At the same time, heating
the samples of the moistened soil to 85°C for 48 hours led to disappearance of urease
activity. This means that in this soil the effect of heat was stronger than that of HgC12'
The other soil (loam) exhibited higher urease activity than the first soil. In samples of
the loam soil, treated with HgCh (1 <X" 4 glkg soil), urease activity was reduced almost
to Zt:ro, the reduction being more pronounced than that caused by heating the moistened
samples at 85°C. In other words, in the loam soil the effect of HgCh was stronger than
that of the heat.
Mitsui et al. (1960) added 5-g air-dry samples of a Japanese paddy soil of volcanic
ash origin to 50 ml of 1 or 0.5 M urea solution containing 0.1 % HgCh (on air-dry soil
weight basis). The mixtures were shaken for 1 or 12 hours, then analyzed for NH/.
Mixtures without HgCh were the controls. It was established that HgCh strongly
inhibited the urease activity; the degree of inhibition was 92 and 85-86% after 1 and 12
hours of shaking, respectively.
Ba'led on the finding by Yolk and Sweat (1955) that urea hydrolysis is retarded in
those Florida soils which contain copper spray residues, Yolk (1961) carried out a
laboratory experiment, using urea pellets (of 1-2-mm diameter) coated with cupric
sulfate (CUS04) dust (0.25% Cu by weight of urea). The pellets were applied on the
surface of a bare moist soil (fine sand, pH 5.6), at a rate of 112 kg N/ha. Pellets without
CUS04 served for comparison. The N losses as ammonia volatilized during 7 days were
determined. The average N losses from the CUS04-coated and uncoated urea pellets
were 32.8 and 36.2%, respectively; CUS04 did not significantly reduce NH.l
volatilization, having only a weak inhibitory effect on soil urease activity. Copper
sulfate was ineffective even when its rate was increased to 0.4% Cu by weight of urea.
Briggs and Segal (1963) obtained, from approximately 25 kg of surface forest soil
(Upper Hutt Valley, New Zealand), a crystalline urease preparation (about 12 mg),
consisting of a mixture of proteins possessing urease activity. The authors mention that
catalytic activity of the preparation was markedly inhibited by Ag+, Hg2+, and Cu 2+, but
they do not give any details on the conditions under which the inhibitory effect was
tested.
Sor et al. (1966) and Sor (1968, 1969), of the Esso Research and Engineering
Company (Delaware), described in their patented inventions that they prepared pellets
from urea, cupric sulfate and lead acetate (0.45% Cu or Pb by weight of urea, the limits
of the Cu or Pb amount being 0.01 and 10%), and a hydrophobic material (solid
hydrocarbon binder). The preferred hydrophobic material was a blend of asphalt (90%)
and microcrystalline wax (10%), representing 10% by weight of urea (the limits being 3
"The rate at which HgCb was added to this soil is not specified in the paper.
6
and 251Vo). nle urea crystals were heated up to 60-80°C, then the inhibitor was added to
the wann urea and mixed thoroughly. This warm mixture received the asphalt-
microcrystalline wax blend previously softened by heating up to 104.5°C. All the
substances were thoroughly blended and this mixture was pelletized by extruding it
from a pellet mill. Tn the pellets obtained, urea, inhibitor, and hydrophobic material
were uniformly distributed. Urea prills without inhibitor and hydrophobic material and
urea pellets without inhibitor, but with hydrophobic material were used as controls.
For evaluation of the effectiveness of copper sulfate and lead acetate, the pellets and
priUs (at an amount equivalent to rates of urea generally applied under field conditions)
were placed on the surface of soil samples (sandy loam, pH 5.5) containing 13 and 5%
moisture, respectively. During incubation at 25°C for 16 and 31 days, respectively, the
volatilized ammonia was determined cumulatively. It was found that less NH3 was lost
from the urea peliets with Cu or Pb than from the control prills and peliets. In the soil
with 13% moisture, Cu and Pb had a similar effect. For example, the cumulative NH3
loss after 16 days of incubation was about 2%, from the Cu- or Pb-containing urea
pellets and 3.5-9SVo from the control prills and pellets. In the soil with 5% moisture, Cu
inhibited more strongly than did Pb. For example, during 28 days of incubation, the Cu-
containing pellets lost via NH3 volatilization about 1.8%, the Pb-containing pellets 8%
and the control prills and pellets about 11-13% of their initial urea-N content.
Pugh and Waid (l969a,b) determined the ammonia losses from lOO-g samples of
three English soils (sandy loam, loamy sand, and clay loam) which were treated with
CUS04 (640 ppm on moist soil weigllt basis) and with urea (4,000 ppm). Samples not
treated with CUS04 served as controls. All the samples of the three soils were incubated
at 20°C for 30, 77, and 148 days, respectively. Copper sulfate caused slight delays in
NH3 loss: the half-loss time (the number of days that elapsed until the NH1loss was half
of the total loss observed in the treatment where urea had been applied alone) was
prolonged under the influence of CUS04 from 4 to 6.5 days (in sandy loam), from 11.5
to 17 days (in loamy sand), and from 34 to 40.5 days (in clay loam). But the total NH3
loss from the three soils during their whole incubation period (30, 77, and 148 days,
respectively) was reduced by CUS04 only to a slight extent (the reductions were of 2.6,
4.6, and 8.4%, respectively).
For determination of urease activity in 21 virgin and 21 cultivated volcanic ash soils,
in 16 virgin and 16 cultivated nonvolcanic ash soils, and in 30 paddy soils from Japan,
Tanabe and Ishi711wa (1969) used 10 ml of a llVo HgCb solution to inactivate the
enzyme in reaction mixtures containing 10 g of soil and 70 ml of aqueous phase.
According to Bhavanandan and Fernando (1970), copper (l,OO() ppm on soil weight
basis), added in the form of CUS04 or CU20 to samples of a tea soil from Sri Lanka,
inhibited urease activity by 30.9 and 43.7%, respectively. The inhibitory effect
increased with the rate of copper. For example, at an 8-fold rate the two forms of copper
brought about 56.2 and 87.5% inhibitions, respectively, in the urease activity.
Surprisingly, the insoluble form of copper (CU20) caused a greater inhibition than did
the soluble copper sulfate at equivalent amounts.
Bremner and Douglas (l971) evaluated, by means of the 5-hour incubation test
described by Douglas and Brenmer (1971), the effect of numerous heavy metal
compounds on the urease activity in two Iowa soils (silty clay loam and clay loam).
This test involves determination of the effect of the test compound on the anlOunt of
urea hydrolyzed by incubation of IO-g samples of soils with 1 ml of toluene, 5 ml of
7
urea solution containing 10 mg of urea-N (1,000 ppm N on soil basis), and 5 m1 of

distilled water or 5 ml of the solution of test compound, at 37°C for 5 hours. After
incubation, the unhydrolyzed urea is extracted with 100 m1 of 2 M KCl solution
containing 0.5 mg of phenylmercuric acetate (urease inhibitor) and estimated
colorimetrically. Concentration of cations in aqueous solutions of the tested compounds
was equivalent to 50 ~g1g soil (50 ppm). Table 1 shows that the cations of heavy metals
inhibited urease activity of the two soils studied, in the following decreasing order:
TABLE I. Effect of heavy metal compounds on soil urease activity"
Inhibition of urease activity (%)

Compound Heavy-metallic cation
Silty clay loam Clay loam
Silver nitrate Ag+ 65 60
Silver sulfute Ag' 63 61
c
Cuprous chloride Cu 16 14
Lead nitrate Ph+ 3 2
Mercuric chloride Hg2+ 42 38

Mercuric sultille Hg2c 40 36
Cupric chloride eu'+ 16 13
Cupric sultllte Cu'+ 14 15
Lead chloride Ph2+ 4 4
Cobaltous chloride Co2 + 4 6
Nickelous chloride Ni 2+ 2
Gold chloride (HAue!.,) Au·l + 18 20

Chromium chloride Cr'+ 3 2
Zinc chloride Z,n 2~ () ()

Manganous chloride Mn'+ () ()
Ferric chloride FeH () ()
"From Bremner and Douglas (1971). by permission of Pergamon Press PLC.
The inhibitory effect was folubstantial only in the case of silver, mercury, gold, and
copper. Comparison of the inllibitions caused by Cu2+ and Cu+, and Pb2+ and Pb+,
respectively, showed that the valency state of these cations did not influence their
inhibitory capacity. A similar comparison of the nitrate or chloride and sulfates of Ag+
and Hg2+ indicated that the anions in these compounds did not affect soil urease activity.
In the case of HgCIz, the influence of concentration (50, 100, 200, and 300 ppm Hg)
on the urease activity in clay loam was also studied, registering the following
inhibitions: 39, 45, 56, and 61 %, respectively, i.e.. the inhibition was not complete at
the highest HgCIz concentration, either.
The effect of the preincubation of soil samples with HgCh and CUS04 (50 ppm) fi}r
3, 7, and 14 days at 30"C, before applying the 5-hour incubation test, was also dealt
with, and it wafol found that the inhibitory effect of both metallic salts decreased, to some
extent, with prolongation of the preincubation time.
8
The effect of heavy metal salts used as micronutrients on the urease activity in
Siberian soils was studied by Gamzikova and Gamzikov (1971). Samples taken from
the 0-20-cm layer of a chernozem and a grey forest soil were treated with one of five
metal salts at rates of 1 mg Mn, 0.5 mg Cu and Zn, 0.3 mg Co and Mo per 100 g soil.
The control samples received no heavy metal salt. The samples moistened to 60% of
water-holding capacity (WHC) were incubated at 2X~C for 21 days, then submitted to
urease activity determination. In the chemozem, urease activity was reduced by Cu,
stimulated by Mn and Mo, and not affected by Zn and Co. In the forest soil, Zn and Co
reduced, Mo stimulated, and Mn and Cu did not affect urease activity.
Pel'tser (1972) studied the effect of Cu(N0 3h, HgCh, and AgNO] on urea
hydrolysis in a slightly acid (pH 6.7) soddy-podzolic soil (Timiryazev Agricultural
Academy, Moscow), by using 14C-labeled urea and assaying the released 14C0 2 • The
reaction mixtures were prepared from 10 mg of urea and 15 rnl of 0.1 % solution of
metallic salt for 100 g of soil. During incubation (at 22QC), moisture content of the soil
was maintained at 60% of WHC. The results showed that complete hydrolysis of urea
took place in about 1.5 days in the control (untreated) soil, in 2 days in the Cu(N0 3 h
treatment, and in nearly 3 days in the HgCl 2 and AgN0 3 treatments.
Kozlovskaya el af. (1972) and Runkov el af. (1974) found that CUS04 used in a high
concentration (X4 mM in the reaction mixtures) strongly inhibited urease activity in peat
bog soils.
In two Sudan soils (clay and sandy loam), 1 rnl of 1% HgCl 2 solution added to 109
of soil stopped urease activity (Said, 1972).
Studying the factors affecting urea hydrolysis in three Alberta soils (a cultivated silt
loam and a virgin and a cultivated loam), Gould et af. (1973) added 5 ml of 1% HgCl 2
solution to 25-g soil samples to stop urease activity.
Lloyd and Sheaffe (1973) used HgCh at 0.1 % final concentration in reaction
mixtures for complete inhibition of urease activity in different samples of an Australian
red-yellow podzolic soil.
Andersson and Bengtsson (1975) studied samples of spruce needle mor collected in
Dalby Kronopark (located in the area of Lund, Sweden). Fresh samples, each equivalent
to 1 g dry matter, were treated with copper or/and zinc acetate solution corresponding to
0,200, 1,000,4,000, 10,000, and 25,000 ppm Cu, Zn, and Cu+Zn (on dry matter basis).
The samples were incubated at 22 QC for 1 and 5 weeks, then analyzed for determination
of their urease activity. It was found that the urease activity decreased significantly at
added metal concentrations ~ 1,000 ppm (after 1 week) and ~ 4,000 ppm (after 5
weeks). The activity could not be detected at ~ 10,000 ppm Cu+Zn and Cu and at
25,000 ppm Zn after both incubation times.
Tabatabai (1977) selected six Iowa soils with a wide range in pH, texture, organic
matter content, and urease activity for studying the effect of numerous metallic
compounds on soil urease activity, assayed by the method of Tabatabai and Brenmer
(1972). The air-dry soil samples (5 g) were treated with 1.5 rnl solution containing
either 2.5 or 25 ~moles of test compound (0.5 or 2.5 11TI10les/g soil), then 0.2 m! of
toluene, 7.5 ml of 0.05 M Tris buffer (pH 9.0), and 1 rnl of 0.2 M urea solution were
added. TIle reaction mixtures were incubated at 37<!C for 2 hours, after which time iliey
were brought to 50 ml with 2.5 M KCI solution containing 100 ppm of Ag2 S04 (as
urease inhibitor) for extracting the NH4 + that had formed. This was then analyzed by
9
steam distillation with MgO. The control reacti<m mixtures contained 1.5 ml of distilled
water instead of the test compound solution.
The results indicated that at the higher rate the heavy-metallic cations inhibited
urease activity in the six soils studied, in the following order:
Ag+ > Hg2+> Cu2+ ~ Cd2+ > Zn2 + > Sn2+ > Mn 2+.
Generally, Fe3+ and Cu 2+ had a str<mger effect than Fe2 + and Cu +, respectively. Also
at the higher rate, Ni 2+, Co2+, Pb2+, Cr3+, V4+, and Mo6+ inhibited urease activity only in
some soils. Lead nitrate was more effective than lead acetate in three of the soils, had a
similar effect as lead acetate in two soils, and was less effective than lead acetate in <me
soil. At the lower rate, the inhibitory effect of all metallic cations was weaker.
By applying the 5-hour incubation test of Douglas and Bremner (1971), Osminkina
and Mir7.aev (1979) studied the kinetics of urea hydrolysis in a calcareous grey soil
from Uzbekistan. The soil samples were treated with CUS04 at rates ranging from 0.01
to 0.1 mg Cu/g soil. The conclusion was drawn that the inhibitory effect of Cu 2+ on
urease activity of this soil corresponded to a reversible competitive inhibition,
characterized by an inhibition constant (ki) equal to 4.9 mg Cu 2+/l 00 g soil.
The English inventors Lewis and Slater (1979a,b) used ferric salts (nitrate, sulfate,
chloride), oxide and hydroxide, and ir<m ore (89.2% Fe203.H20) for obtaining fertilizer
compositions in which urea is complexed or physically mixed with a Fe compound.
Preferred are the compositions containing complexed urea together with different
amounts of free (uncomplexed) urea. The ratio between the number of Fe atoms and the
number of urea molecules should be at least 1:40. The preferred ratios range from 1:5 to
1:20. Laboratory experiments showed that these compositions applied 00 various soils
lost less arnm(mia through volatilization than treatments with urea alone. They were
applied at rates equivalent to 60-330 kglha. pH of the soils varied between 5.0 and 8.1,
and their moisture content ranged from 1.5 to 15.0%.
In a laboratory experiment Zukowska-Wieszczek (1979) added an aqueous PbO
suspension to samples taken from the 5-10-cm layer of four park soils and three street
lawn soils in Warsaw, Poland. Rates of addition were 0,10,50,100,200, and 400 ppm
Pb. Urease activity in the samples was measured after their 6-week incubation at
ambient temperature. It was found that PbO addition at each rate led to inhibition of
urease activity in each soil and the inhibition degree increased with the increasing rate
of PbO. The initially more urease-active park soils were less sL'I1sitive to inhibition than
the initially less urease-active street lawn soils. For example, 50 ppm Pb caused 20 and
81 % inhibitions in urease activity of the park soils and street lawn soils, respectively.
The inhibition became total at 200 ppm Pb in the park soils and at 100 ppm in the street
lawn soils. .
In a short report, the Russian investigators Zyrin el af. (1980) described pot
experiments in which soddy-pod7£llic, chernozemic, and peat bog soils were treated
with Cd or Pb nitrate at different rates, in the spring of 1978. The urease activity was
determined in March and June 1979. At the rate of 100 mg Cd/kg soil, the soddy-
pod7£llic soils exhibited about 50%, whereas the chemozemic and peat bog soils showed
80-90% of the urease activity measured in the untreated controls. Lead was less
inhibitory. Thus, at the rate of 2,000 mg Pb/kg soil, inhibition of urease activity was less
than 20% in the soddy-podwlic soils and 0% in the chernoz$!mic and peat bog soils.
Schinner el af. (19KO) installed experimental plots in a forest nursery on alluvial soil
in Austria. Cembra pine (Pinus cembra) seeds were sown in the plots. In May 1978,
10
when the seeds began to germinate, the plots were treated with mineral fertilizers (6 g
N, 6 g P2 0 S, and 9 g Klm2 ) with or without CUS04 (25 g/m2 ). Untreated plots served for
comparison. After 6 weeks, the soil urease activity was assayed and it was found that
the activity represented 77% in the soil treated with NPK and 48% in the soil treated
with NPK + CUS04 in comparison with that of the untreated soil (100%). This means
that CUS04, used at a high rate, intensified the depressive effect of mineral fertilizers on
the soil urease activity.
Daif and van Beusichem (1981) selected eight soils in the province of Badajoz,
Spain to obtain a wide range of properties. Samples taken from the 0-30-cm soil layer
wt.'fe allowed to air-dry, then crushed to pass a 2-mm screen and used for determination
of urease activity in reaction mixtures, to which Fe, Cu or Zn was added as the sulfate
of the bivalent cation. Rates of addition were: 0, 5, 10 or 20 Ilg Fe, Cu or Zn/g soil. The
results showed that the inhibitory effect of the metallic ions was obvious only when
they had been applied at the rate of 20 Ilg/g soil. The degree of inhibition varied from 7
to 20% depending on the soil and metallic ion. Thus, the light-textured soils were more
sensitive to inhibition than the heavier ones. Cu2+ was more inhibitory than Fe2+ or Zn 2+.
The effect of the three metallic ions (applied only at the rate of 20 Ilg/g soil) on the
microbial production of urease was also studied, but only in two soils. It was found that
these ions, although applied at a relatively high rate, did not inhibit the synthesis of
urease by soil microorganisms.
Marusina (1981) and Krasnova' (1985) described a field experiment, carried out on
1_m2 microplots installed on a soddy-podzolic soil in Russia. The experiment comprised
four variants:l. NPK-fertilized; 2. NPK-fertilized and treated with NiCb; 3. NPK-
fertilized and treated with ZnCb; 4. NPK-fertilized and treated with MnCh. The
fertilizers and heavy metal chlorides were introduced into the O-S-cm layer of the plots
at the following rates: 150 or 60 kg Nlha as ammonium nitrate, 100 or 60 kg Plha as
superphosphate, 100 or 60 kg Klha as potassium chloride; 120 mg Ni, 150 mg Zn or
750 mg Mn per kg soil. Within each variant there were plots left unsown and plots sown
with barley. During the vegetation period, urease activity was determined in the 0-5-
and S-10-cm layers. In both sown and unsown plots, urease activity decreased under the
influence of heavy metals. Ni and Mn were more inhibitory than was Zn. The inhibition
was more pronounced in the O-S-cm layer than in the 5-1 O-cm one.
According to the data published by Tu (1981a), treatment of the samples of a
Canadian clay loam soil (pH 7.2) with HgCh (70 Ilg/g soil) brought about a significant
inhibition of urease activity; degree of inhibition was 58.3% after 2 days and 29.4%
after 7 days of incubation. But in samples of a Canadian organic soil (PH 7.2), the same
amount of HgCb stimulated urease activity; the stimulation was insignificant after 7
days and significant after 14 days of incubation (Tu, 1981b). In another study
(Tu, I 990), urease activity was determined in HgClrtreated samples of a sandy loam
(pH 7.6) and an organic soil (pH 6.8) from Canada. Incubation lasted 7 and 14 days (at
2S"C). HgCh significantly inhibited urease activity of the sandy loam (67 and 81%
inl1ibitions after 7 and 14 days of incubation, respectively). In the organic soil, urease
activity was not affected by HgCl2 (after 7 days) or was significantly enhanced by it
(after 14 days). In other studies, HgCh, applied at a rate of 80 Ilg/g soil, inhibited
'N.M. Krasnova is N.M. M!ll1Isina's new name; she got it, probably. after her marriage.
11
signifil:antiy or did not affect urease activity of the sandy loam incubated for 7 days (Tu,
1992a,b).
By increasing the rate of Cu2+, Zn 2 +, and Mn2+ from 500 to 1,000 ppm in the acid
(pH 5.9) soil studied by the Japanese investigators Komai et 01. (1981), the degree of
the inhibition of urease al:tivity changed only to a small extent, nan1cly from 83 to 87%,
from 64 to 71 (Yo, and from 41 to 40%, respecti vel y.
NOT (1982) assayed urease activity in real:tion mixtures prepared from 109 of soil
(on dry weight basis), 1,200 Ilg of Ag+ or Cu2+, 5 mg of urea and distilled water, and
incubated at 37QC. Reaction mixtures without inhibitors were the controls. After
incubation- the residual urea was determined. Ag+ wmpletely inhibited urease activity
in each of the four l\1alaysian soils studied. eu 2 + had a similar effect in two of the soils
(clay loam, pH 3.8; sandy clay loam, pH 5.1) and a partial (77.5 and 91.7%) inhibitory
effect in the other two soils (sandy clay loam, pH 4.8; loamy sand, pH 5.1).
Grigoryan (1982) added 1-10 mg of Cu, Pb, Ni, Mn (in form of nitrates) or Mo (as
ammonium molybdate) to 200-g samples of flood plain meadow soils, brown forest
soils, and leached chernozems from Armenia. The heavy metalfol, especially Mn at the
lowest rate stimulated urease activity in all soilfol. But folignificant negative correlations
were registered between the higher rates of each heavy metal and urease activity in all
soils.
Krasnova (1983) prepared mixtures from samples of a s(xtdy-podzolic soil and
solution of Mn, Zn or Ni sulfate (at rates of 250-2,000 mg Mn, 25-200 mg Zn or 20-
160 mg Ni/kg foloil). The mixtures moistened to 60% ofWHC were incubated at 30!!C
for 2 weeks, then analyzed for determination of their urease activity. The activity was
inhibited in parallel with tbe rate of eal:h heavy metal. For comparing the inhibitory
effect of the three heavy metals, some data referring to four samples are specified
below: urease activity (expressed in mg of NH.1 produced by 1 g of soil in 24 hours) was
0.10 (untreated control), 0.60 (250 mg Mnlkg soi!), 0.58 (200 mg Zn/kg soil), and 0.40
(160 mg Ni/kg soi\). Thus, the metallic ions inhibited urease activity in the order: Ni >
Zn>Mn.
For studying the effect of Pb and Cd on soil urease activity, Brunner and Schim1er
(1984) used samples taken from the Ah-horizon of an alluvial soil in Tyrol, Austria. The
samples were treated with 0, 50, 200, and 1,000 Ilg ofPb (as lead acetate)/g soil or with
(), 0.5, 5, and 50 Ilg of Cd (as cadmium sulfate)/g soil. The samples were kept at 22°C
for 16 weeks, during which the water content of samples was maintained at 15-20% and
the urease activity was measured at 3-4-day intervals during the first weeks, then at 4-
week intervals. The activity oscillated in both Pb- and Cd-treated samples at each Pb
and Cd concentration: the activity increased during tbe first week, then decreased and
lat er increased again and, at week 16, approached the values found in the untreated
samples.
The Russian investigators Skvortsova et al. (l984) stated that in a soddy-podzolic
soil, treated with mercuric nitrate solution at rates of 0, t, 2.5, 10, 20, 50, 200, and 500
mg Hglkg soil, urease activity suffered a nearly lO-fold decrease under the influence of
50 mg Hg/kg soil. However, numerical data on the activity are not presented in their
paper.
'Ouration of incubation is not specilied in the paper.

12
Badr EI-Din et al. (1985) applied the 5-hour test of Douglas and Bremner (1971) to
study the effect of 15 heavy metal salts on urease activity in three Egyptian soils. The
salts were used at a rate of 50 ppm (soil basis). One can see from Table 2 that the
TABLE 2. Inhibition of soil urease activity by heavy metal salts"

Heavy metal salt
Silty day Sandy clay loam Loamy sand
Silver nitrate 50.0 61.4 64.1
Silver sultate 49.1 62.0 66.9
MercUlic chloride 36.3 43.2 48.2
Mercuric sultate ~R.2 44.0 4R.8
Cuprous sultate 34.9 3R.I 39.S
Cupric sulfate 35.1 38.5 40.8
Cupric chloride 35.0 17.9 41.5
Ferric sultate 20.0 21.1 25.5
Ferric chloride 20.2 20.4 23.9
Ferrous sultate 20.2 22.9 24.3
Cobaltous sulfate 21.4 25.1 27.6
Lead acetate 1.4 3.2 6.8
Zinc chloride 12.5 15.5 19.4
Bismuth nitrate 10.0 15.5 16.0
Stannous chloride 7.4 8.6 IJ.I
"From Badr EI-Din et al. (1985). by permission ofVCH Verlagsgesellschatl.
cations, in respect of their inhibitory effect, can be arranged in the following order:
Ag+ > Hg 2+> Cu+ =Cu 2+ > Co2+ > Fe2 += Fe3+ > Zn2+ > Be+ > Sn2+ > Pb2+.
Table 2 also shows that the valency state of Cu and Fe and the nature of anions had
almost no influence on the inhibitory action. Urease activity in the loamy sand was
more sensitive to inhibition than that in the silty clay.
In the case of the silty clay, longer incubations were also carried out to study the
effect of Cu(ll), Fe(III), and Co(ll) sulfates on urease activity and volatilization of
ammonia from urea. In the absence of inhibitors, urea was hydrolyzed in a single day,
but in their presence complete hydrolysis of urea required 7 days. Cu 2 + proved to be
more effective than Fe3+ or C0 2+ in retarding urea hydrolysis and reducing volatile NHJ
losses.
Dalton et al. (11.)85) studied nine soils from western Oregon and one from eastern
North Carolina. In an experiment, 25-g (dry weight basis) soil samples, after the
addition of 12.5 ml of a solution containing glucose and urea (2 mg and 114 llg/g dry
soil, respectively) with or without NiCh (0.25 llmoles/g dry soil), were preincubated at
2 7"C. Urease assays were performed at both zero time and after 3 days of preincubation.
Urease activity was detem1ined by measurement of 14C02 evolved from 14C-urea in
reaction mixtures prepared from 1 g of soil (dry weigl1t basis), 0.3 ml of 1 M 14C_urea
(3.4 llCi/nID10l) in 5 ml Hepes buffer (pH 7.1) and sufficient Hepes to bring the total
soil water volume to I ml, and then incubated at 37~C for I hour.
At zero time of preincubation, urease activity of the Ni-treated sample versus the
untreated sample did not significantly change in any of the I () soils studied. After 3 days
of preincubation, urease activity showed increased values in all soils, in both Ni-treated
and untreated samples. A comparison of the samples revealed that NiCl 2 at the rate
13
applied did not have significant effect on urease activity in the nine Oregon soils,
whereas in the North Carolina soil (a low-nickel loamy sand; total Ni content = 13
ppm), Ni at the same 0.25 ~oles/g dry soil rate caused a significant (2.5-fold)
stimulation of urease activity.
With this low-nickel soil other preincubation experiments were also carried out, all
followed by urease assays. The results showed that urease activity was highest in
samples treated with 5 ~oles Ni/g dry soil. At the concentration of 5 I!moles metal/g
dry soil, Cu2+ and C02+ (in form of chlorides) significantly inhibited urease activity, but
Mn2+ and Fe 2+ (also in form of chlorides) had no such effects. Nickel became inhibitory
at a concentration of 50 I!moles (2,940 I!g)/g dry soil.
Ushikubo et al. (1985) studied the effect of Cd, Hg, and Cr on urease activity in a
sandy loam soil (PH 6.4) and a loamy sand soil (pH 6.9) collected from a woodlot and
an uncultivated old field, respectively, near the Inland Lake Research and Study Center
of Michigan State University. Fifty-g air-dried soil samples were treated with 100 ml of
Cd, Hg or Cr salt solution containing 0.1, 0.5 or 1 mg Cd or HgIl or 10, 25 or 50 mg
Cr/l. The untreated (control) samples received 100 ml of water. The mixtures were
preincubated at room temperature for 1,2,5 or 10 days, then filtered; the residue (soil)
was air-dried, then submitted to enzymological analyses.
The urease-inhibiting effect of heavy metals showed a tendency to increase with
their concentration and with the preincubation time. The percent inhibitions of urease
activity in the two extreme cases (lowest heavy metal concentration and shortest, i.e.. 1-
day preincubation, and highest heavy metal concentration and longest, i.e.. lO-day
preincubation, respectively) were the following: 6.92 and 40.48 (Cd), 29.73 and 40.48
(Hg), and 2.78 and 19.05 (Cr), respectively, in the sandy loam; 20,00 and 46.67 (Cd and
Hg), and 20.00 and 47.62 (Cr), respectively, in the loamy sand.
In sandy loam, the untreated samples were 14 times more urease-active than
untreated samples of loamy sand, and less sensitive to inhibition than the loamy sand. In
the sandy loam, Cd and Hg, although low in concentration, were more inhibitory than
Cr, in much higher concentrations. In the loamy sand, the inhibiting effect of Cd, Hg,
and Cr was similar.
Studying an Indian clay loam soil, Yadav et al. (1986) treated soil samples with salts
(sulfates and chlorides) of heavy metals, at three rates relative to the cations (100, 500,
and 1,000 ppm on soil weight basis). The reaction mixtures, consisting of 20 g of soil, 1
ml of toluene, 5 ml of urea solution containing 200 ppm N and aqueous salt solution,
were incubated at 28!'C for 12 hours. The residual urea was then extracted and
determined. At the rate of 100 ppm, Ag+ and Hi+ caused more than 60%, whereas
Cu2 +, Zn2 +, C02 +, and Cd2+ exhibited 10-30% inhibitions of urease activity. The
inhibitions given by Pbz+, Cr.l+, Mnz+, and Fe z+ were lower than 10%. At higher
concentrations, the metallic cations had a stronger inhibitory effect. The highest (96%)
inhibition was brought about by 1,000 ppm Ag+ and Hg2+, followed by Cu2 + and Cd2+
(60% inhibition); Pb2+ showed the least effect (25% inhibition). The inhibitory effect of
cations at 1,000 ppm presented the following order:
~=H~>~>~>~>~>~>~>~>Mn2+>~~
The time necessary for complete hydrolysis of urea was also estimated in 20-g soil
samples treated with Cr3 +, Cu2+ or Hg2+ at 100,500, and 1,000 ppm and urea at 200 ppm
N, and incubated at 28!'C. In the no inhibitor control sample, urea hydrolysis was
complete in 3 days. Cr 3+ at 100 ppm did not prolong this duration. In samples treated
14
with Cr3+ at 500 and 1,000 ppm and in samples treated with Cu 2+ at the three rates,
complete hydrolysis of urea took place in 7 days. In the presence of Hg2+, complete
hydrolysis of urea required 14-2X days.
Skujins et at. (19X6) used surface (0-20 cm) samples collected from a deciduous
forest soil of sandy clay loam texture (pH 7.0), in the area of Ultuna, Sweden. After air-
drying, the samplefl were sprayed separately with CuCh and CrCh solutions at
concentrations of 50, 200, 500, and 1,000 ~g Cu or Cr/g soil. Additional water was
given up to XO% of WHC, which was maintained constant during the incubation of
samples at 20"C for 20 days before determination of their urease activity. The activity
was found to decreased logaritlunically with increasing amounts of Cu 2+ over the entire
range from 0 to 1,000 ~g Cu/g soil. The initial decrease of urease activity was more
pronounced in the Cr-treated samples. The logarithmic part of the response to Cr,
however, wafl refltricted to concentrations s 200 ~g Cr/g soil. At Cr concentrations>
200 ~g/g soil, the reflponfle to Cr was retarded toward'l a limit value of 50%.
Doelman and Haanstra (19X6) studied samples of five different soils (sand, sandy
loam, silty loam, clay, and sandy peat) collected from various areas in the Netherlands.
The field-moist samples of the five soils (55-70% of WHC) were amended with 0, 55,
150, 400, 1,000, 3,000, and X,OOO mg/kg Cd, Cr, Cu, Ni, Pb, and Zn (in form of
chlorides), then incubated at 20 QC. Urease activity was measured after 6 weeks and 18
months of incubation. The results showed that the ecological dose-50% (ED50) -
dcfincd as the heavy metal concentration (in mgikg soil), at which urease activity is half
of the initial (uninhibited) level- tended to decrease (i.e .. the inhibiting effect of heavy
metal tended to increase) from week 6 to month 1X for Cd, Cr, Cu, and Zn. For Ni and
Pb, ED50 generally did not change significantly (i.e.. the inhibiting effect stabilized) in
sandy loam, silty loam, and sandy peat, increased in sand and decreased in clay in the
period between week 6 and month 1X. The average ED50 values of Zn were, after 18
months, the lowest (varying between 100 and 3(0), which means that the inhibiting
t-'ffect of Zn on urease activity was the strongest.
Tn an experiment carried out in vegetation pots (Badura et at., 19X6), surface soil
samples taken from a Polish beech forest were treated with Zn orland Cd sulfate at rates
of 5,000 ppm Zn or Cd and 2,500 ppm Zn + 2,500 ppm Cd, respectively, and incubated
for 42 days. Urease activity, determined several times during the incubation period, did
not show any considerable differences between the Zn-, Cd-, and Zn+Cd-treated
samples and between these samples and the untreated ones.
The Chinese investigators Xue and Li (1987) compared the inhibitory effect of three
Cu(II) salts and of ZnS04 on the urease activity in a periodically water-logged paddy
soil. The reaction mixtures were prepared from 5 g of dry soil, 10 rnl of 10% urea
solution also containing the inhibitor at 20-100 ppm (on soil weight basis), and
incubated at 37"C for 48 hours. The results in Table 3 show that CUS04 was the most
effective and ZnS04 the least effective inhibitor. The degree of inhibition increased with
increasing inhibitor concentration, but did not become 100% at the highest inhibitor
concentration either. It was also established that the inhibitory effect of CUS04 was
evident even after 12 days of incubation and this effect of CUS04 showed a tendency to
decrease with increasing urea concentration (1, 2, 4,6, X, and 10%) and to increase with
increasing temperature (22, 37, and 45"C).
15
TABLE 3. Effect of inorganic salts on urease activity in a periodically water-logged

paddy soil"
Concentration of salt (ppm of soil)
Salt 20 40 60 80 100
CuS04.5H2 0 41.91 48.70 51.69 58.48 58.88
CuCh.2H20 43.71 45.71 48.30 51.69 55.08
Cu(NO,),.3H2 0 38.72 43.91 45.90 47.30 49.50
ZnS04.7HzO 30.13 33.33 35.31 39.32 43.31
aAdapted from Xue and Li (1987).
In the laboratory experiment described by Todorov et al. (1987), surface samples of

an alluvial meadow soil (pH 7.24) collected in the area of Kostinbrod, Bulgaria, were
treated with 0,50,500, and 1,000 mg Pb/kg soil in form oflead acetate, then moistened
to 60-65% of WHC and incubated at 26-28~C for 50 days. Urease activity determined
after 5, 20, 35, and 50 days of incubation was surprisingly higher in Pb-treated rather
than in the untreated samples. The stimulating effect of Pb increased with its rate. The
maximum value of urease activity was registered in all samples after 15 days of
incubation. The activity increase was attributed to enhanced production of urease by the
soil microorganisms utilizing the acetate anion as carbon and energy source.
Carrying out a laboratory experiment, Aliev (1988) added heavy metal salts: COS04,
Cr2(S04)], TiCh, and (NH4)6M07024 at rates of 0,5, 15, and 45 kglha to a Siberian dark-
chestnut soil. Urease activity in soil increased under the influence of each rate of each
heavy metal salt; the increase was inversely proportionate to the rate of the addition of
these salts.
Samples of an Italian sandy soil were treated, by Benedetti et al. (1990), with Cr203
(at a rate of 100 mg Cr/kg soil), then moistened and incubated at 30°C for 116 days.
Samples not treated with Cr were the controls. Urease activity was determined after 0,
21, 70, and 116 days of incubation. At day 0, urease activity was lower in the Cr-treated
than in the control samples. During incubation, urease activity continuously decreased,
but surprisingly the decrease was less pronounced in the Cr-treated samples than in the
control ones.
Kandeler et at. (1990) measured urease activity, in May 1990, in samples of two
Austrian brown soils (a sandy loam and a clay loan1) which were submitted, in 1987, in
a pot experiment, to a combined treatment with Zn (300 ppm), Cu (100 ppm), Ni and V
(50 ppm), and Cd (3 ppm). The results obtained indicated that inhibition of urease
activity by the heavy metals was 45% in the sandy loam and only 15% in the clay loam.
Winiarski (1990) added 328 mg urea and 0, 0.5, 1 or 2% Cu (on urea weight basis)
in the form of CuC0 3.Cu(OH)2 to 2-kg samples of a light- and a heavy-textured Polish
soil. The mixtures moistened to 50% of WHC were incubated at 22 QC. The amounts of
ammonia volatilized during 5 and 10 days were determined. Cu reduced NH]
volatilization from both urea-treated soils. The reduction was most pronounced under
the influence of 1% Cu: it was 27.1 % in the light soil during the 10-day incubation and
35.5% in the heavy soil during the 5-day incubation.
The experiments describt,'(] by Chernykh (1991) were carried out in vegetation pots
during the 1985-1988 period. Several soddy-pod:l.olic soils from the Moscow region and
a common chernozem from the Kursk region were used. The soils were treated with 2.5,
5, 10, 20 or 50 mg Cd (as nitrate)/kg soil, 125, 250, 500, 1,000 or 2,000 mg Pb (as
16
nitrate) or Zn (as oxide)/kg soil. No heavy metal was added to the control soils. The test
plant was barley. At three growth phases (germination, tillering, and ripening) of barley,
soil urease activity was determined. The results led to the conclusions that the inhibitory
effect of heavy metals increased in the order: Cd > Pb ~ Zn; the soddy-podzolic soils
were more sensitive to inhibition than was the common chernozem. For example, urease
activity in a soddy-podzolic soil decreased to 7S% under the effect of 100 mg Cd/kg soil
and to 70% when the treatments were done with 2,000 mg Pb or Zn/kg soil. The
minimum heavy metal addition for a statistically significant inhibition of urease activity
was 10 mg Cd or 500 mg Pb or 500 mg Zn/kg soil in a soddy-podzolic soil and 20 mg
Cd or 2,000 mg Pb/kg soil in the common chernozem (in which Zn even at the highest
rate applied - 2,000 mg/kg soil- had no inhibitory effect).
Kucharski and Niklewska (1992) performed a pot experiment using a brown soil of
heavy loamy sand texture (pH 7.1) from Poland. Zn (as sulfate) was added to the soil at
rates of 0, 10, 100, and 1,000 ppm. The test plant was broadbean. Soil urease activity
wa" assessed at the cutting of broadbean (at time of its flowering). The activity showed
10 and 4% increases in the treatments with 10 and 100 ppm Zn, respectively, and a
slight (3.S%) decrease at the 1,000 ppm Zn rate.
Gupta and Chaudhry (1994) studied the effect of four heavy metals on urea
hydrolysis in surface (0-15 cm) samples of a sandy loam soil collected from the farm of
the Agricultural University in Hisar, Haryana, India. The samples were amended with
urea (OJ g N/kg soil) and with Ni, Zn, Pb or Hg at a rate of 0, 0.2 or 0.4 g/kg soil, then
moistened to 60% ofWHC and incubated at 15 or 30QC. The heavy metals retarded urea
hydrolysis in the order: Hg> Zn> Ni > Pb at both temperatures. However, irrespective
of the metal added, urea was completely hydrolyzed within 7 and 14 days at 30Q C and
15~C, respectively.
As a part of a complex study on the effects of Cr(VI) on soil biological properties,
Speir et at. (1995) determined urease activity in three New Zealand soils (Egmont black
loam, Kaitoke silt loam, and Foxton loamy sand). TIle topsoil samples, each equivalent
to 100 g dry weight, were moistened then amended with 10 ml of K2 Cr 2 0 7 solutions at a
concentration range of 0-50 Ilmoles Cr(VI)/g soil or with water in the controls. Urease
activity was measured 3 days after amendment and 'again on the same soil samples 60
days after amendment. The samples were stored at room temperature (l5-22°C) over the
intervening period. In the three soils studied, the ecological dose-50% (EDSO), i. e.. the
Cr(VI) concentration or "dose" (Ilmoles/g soil), resulting in 50% inhibition of urease
activity, was 41.5,22.2, and 20.3, respectively, after 3 days and only 5.61, 9.56, and
4.30, respectively, after 60 days, indicating that the inhibitory effect of Cr(VI) increased
with time.
The effects of Cd, Zn, and Pb additions on urease activity in soddy-podzolic soils (a
clay loam, pH 4.5 and a loamy sand, pH 4.05) from the Moscow region were studied in
pot experiments and under field conditions (in microplots) by Lebedeva et at. (199S).
Rates of additions per kg soil were 0,10,15 or 20 mg Cd (as nitrate), 0, SO, 100,300,
400 or 500 mg Zn (as sulfate) or Pb (as nitrate). Before the addition of heavy metal
saits, the soils were NPK-fertilized and limed or not limed. Urease activity was assessed
in 1991 and 1992, during the growth period of test plants (carrot, beet, dill or rye). Cd
addition even at the highest rate led to no inhibition of urease activity in 1991 and to a
slight inhibition in 1992. Contrarily, Zn and Pb caused inhibition of urease activity in
both years. According to calculations, the minimum rate for a significant inhibition of
17
urease activity averaged 125 mg Zn and 90 mg Pb/kg soil. Lime (20-40 t/ha) addition
brought about a considerable increase in urease activity of both untreated and Cd-, Zn-
or Pb-treated soils. All results were similar in the two soils studied and in the pot and
microplot experiments.
Kozdr6j (1995) used a sandy loam soil (pH 4.5) from the O-lO-cm layer in a beech
forest of the sanctuary "Zloty Potok" (Poland). The 1OO-g soil samples moistened to
40% ofWHC were treated with 0 or 1 or 2 mg of Cu or Cd as chlorides. Urease activity,
determined after I, 3, 5, and 7 weeks of incubation, was inhibited to a larger extent by 2
rather than by 1 mg of heavy metal and by Cu more than Cd during the whole
incubation period, excepting the treatment with 1 mg Cd, in which urease activity was
significantly higher than in the other treatments and at weeks 3, 5, and 7 reached the
level recorded in the control samples.
Richards (1995) patented a urea fertilizer containing urease-inhibiting ferric nitrate
and urea. The fertilizer may be in the form of a liquid urea-ammonium nitrate (solution
or slurry). The molar ratio offerric nitrate-urea complex/urea in the fertilizer is 1: 1-15.
Studying the effects of heavy metals on biological properties, including urease
activity of Egyptian soils, Hernida et al. (1997) added Cu or Zn (as sulfates) at rates of
0, 0.2 or 2 ~lg/g soil to 500-g samples of a clay soil (pH 7.5) and a sandy soil (pH 7.4)
from the area of Assiut. The samples moistened to 28% ofWHC were incubated at 28 QC
and analyzed after 1, 4, and 12 weeks of incubation. It was found after each incubation
time that both Cu and Zn inhibited urease activity in both soils. At the lower rate Cu
was more inhibitory (significantly in the clay soil and insignificantly in the sandy soil)
than Zn. But at the higher rate, both Cu and Zn caused complete inhibition of urease
activity in both soils.
Kucharski (1997) summarized the results of pot experiments in which the effects of
Zn, Pb, and Cd on soil enzymes and yield of the test plant, yellow lupine, were studied.
Some of the results are reproduced in Table 4.
TABLE 4. EtTects ofZn, Ph, and Cd on soil urease and dehydrogenese activities and yield of yellow lupine"
Rate of addition Urease activity Dehydrogenase activity Lupine yield
Heavy metal
(mglkg soil) (%) (%) (%)
2 102.1 1.11.0 79.1
Zn 4 86.5 149.6 SUI
40 84.4 113.8 43.2
50 89.9 99.0 96.4
Ph 500 77.5 45.5 73.9
1000 60.9 29.6 49.7
0.5 91.0 83.8 57.7
Cd 5 68.3 51.8 0
15 71.6 34.2 0
"Adapted Irom Kucharski (1997).
lbe activities and yield are expressed as percentages of the values registered in the untreated, control soil.
It is evident from this table that at the higher rates each heavy metal greatly reduced
urease activity and, excepting Zn, dehydrogenase activity too. Phytotoxicity of Cd
exceeded that of Zn and Pb.
18
Leir6s et al. (1999) treated 400-g samples, taken from the Ah horizon (0-5 cm) of a
loamy soil developed under Atlantic oakwood climax vegetation, with CuCI 2 at rates of
0, 1,2,5, and 10 mg Cu/g soil. The samples were then moistened, and analyzed before
and after incubation (28 days at 25°C) for determination of several biochemical
parameters, including urease activity. This activity (expressed in fllUol NH3/g soil/hour)
decreased with increasing Cu rate both before incubation (19.10, 13.84,6.13,2.09, and
1.01) and after incubation (23.74, 15.77,9.55,2.32, and 0.48).
Blaise et al. (1996, 1997) described two laboratory experiments in which a loess
brown earth (pH 6.8) from cultivated fields at Dumast, Freising, Germany was used for
studying the effects of agricultural grade pyrite (consisting of pyrite, FeS2, and other
sulfides and containing 20% Fe and 22% S) on the volatilization of ammonia and
emission of nitrous oxide from urea-treated samples.
Tn the first experiment, 200-g air-dried soil samples were flooded to a depth of 2 cm,
amended with glucose (500 mg C/kg soil) for enhancing reduced conditions and then
pre incubated at 20QC for 2 weeks. After preincubation, two series of samples were
amended with urea (100 mg N/kg soil) with and without ground pyrite (equivalent to
100 mg S/kg soil) and incubated for 7 days, during which the amounts of NH3
volatilized from one series of samples and the amounts of NzO emitted from the other
series of samples were determined. The cumulative losses of N from the initially added
20 mg urca-NI200-g soil sample were thc following: 1.49 mg N as NH3 and 2.00 mg N
as N20 from the samples amended with urea only, and 0.94 mg N as NH3 and 0.91 mg
N as NzO from the samples amended with urea + pyrite, i.e., the total gaseous N loss of
3.49 mg N/sample was reduced, in the presence of pyrite, to 1.85 mg N/sample, thus the
reduction was 47.3%.
In the second experiment, 100-g air-dried soil samples were moistened to 60% of
WHC, then preincubated at 20Q C for 2 weeks. Following preincubation, the samples
were treated with urea (100 mg N/kg soil) and with 0,0.1,0.5, 1,5, and 10 g of ground
pyrite/kg soil, then submitted to incubation. For estimation of NH3 volatilization the
incubation lasted 15 days. The cumulative NH3-N loss from the initially added 10 mg
urea-N/IOO-g soil sample was 0.72 mg in the urea-alone treatment and significantly
less in the urea + pyrite treatments. The reduction in NH3 volatilization was 7-22% at
the lower rates (0.1-1 glkg soil) of pyrite, and 53 and 86% at the 5 and 109 pyrite/kg
soil, respectively.
Reduction of NH3 volatilization from urea was attributed to the acidic nature of
pyrite (to sulfuric acid formed upon oxidation of pyrite), but the role of pyrite as a
urease inhibitor is not excluded, as it is known (Bayan and Eivazi, 1999) that the iron
oxides (goethite, hematite) inhibit urease activity. To explain the reduction of N 20
emission from urca, Blaise et al. consider it most plausible that pyrite, due to the
sulfides it contains is directly involved in inhibiting at least one of the reductase
enzymes catalyzing the processes NO,' --> NO z' --> NO --> N 20, because pyrite reduced
the N 20 emission from KN0 3 also.
In a previous experiment, Blaise and Prasad (1995) used urea at a much higher rate:
100-g samples of an Indian sandy clay loam soil (pH 8.1) were amended with 1 part
urea (1,000 mg N/kg soil) blended with two parts pyrite. Soil samples amended only
with urea were the controls. All samples were then incubated under aerobic and
anaerobic conditions for 8 days, during which the volatile ammonia was determined.
19
The cumulative NH3 losses from the applied urea-N were significantly reduced by
pyrite from 27.5% (control) to 8.9% under aerobic conditions, and from 19.3% (control)
to 16.9% under anaerobic conditions.
In a greenhouse experiment, Wyszkowska et al. (2001) treated samples (3 kg/pot) of
a loamy sand soil (pH in KCI 6.6) with K2Cr207 at rates of 0, 40, 80, and 120 mg Cr/kg
soil. Nutrients were also applied (glkg soil): 0.15 N as urea, 0.1 Pas K2HP04, 0.15 K as
K2HP04 + KCl, and 0.05 Mg as MgS04.7H20. Some pots were amended with fmely
ground barley straw (4 g/kg soil) and some pots were sown with oats (25 plants/pot).
There were four variants: 1. not amended with straw and not sown; 2. amended with
straw but not sown; 3. not amended with straw but sown; and 4. amended with straw
and sown. The plants were harvested at panicle emergence stage (at day 51 of growth).
Dry matter yield of oats was determined and the soil was submitted to enzymological
and microbiological analyses.
Urease activity in soil not treated with Cr increased in the four variants in the
order: 1 < 2 < 3 < 4. Chromium inhibited urease activity in all variants. Within the same
variant the degree of inhibition increased with the increasing rate of Cr, but the residual
urease activity at the same Cr rate presented in the four variants the same order as
urease activity in the untreated soil. In other words, the amendment with straw and/or
growth of plants led to diminution of the urease-inhibiting effect of Cr.
Moreno et al. (2001) treated 100-g samples of two Italian soils (a sandy loam, pH
8.1 and a sand, pH 4.8) with 5 ml of CdS04 solutions to give a Cd concentration ranging
from 3 to 4,000 mg/kg soil. Untreated samples served as controls. The soil moisture was
adjusted to 55% of WHC, then all samples were incubated at 25°C and submitted to
several analyses, including determination of urease activity, after 3 hours, 7 and 28 days
of incubation. The analytical data were used for calculation of the ecological dose-50%
(EDso).
The EDso of urease activity, i.e., the Cd concentration (in mg/kg soil) that inhibited
urease activity by 50%, had the following values after the three incubation times:
1,538.5 (3 hours), 1,162.8 (7 days), and 1,190.5 (28 days) in the sandy loam, and
4,166.7 (3 hours), 909.1 (7 days), and no inhibition (28 days) in the sand. Thus, EDso
was lowest (and sensitivity of urease activity to Cd was highest) in both soils after 7
days of incubation. Urease activity in the sandy loam compared to urease activity in the
sand was more sensitive to Cd after both the 3-hour and 28-day incubations.
1.2. LIGHT METAL COMPOUNDS
Mutatkar and Pritchett (J967) prepared mixtures from samples of two Florida soils: a
fine sandy soil (containing about 2% organic matter) and a muck (containing 62%
organic matter), by mixing the sandy soil with 8% (by weight) of muck and by adding
0, 90, 180, 360, and 720 ppm of Al in form of AI2(S04)3. The pH of the mixtures was
adjusted to 4.0, 4.8, 5.5 or 6.5 by adding either dilute HCl or NaOH. Urea was applied
at the rate of 200 ppm N (on soil mixture basis). Moisture content of mixtures was
brought to 15%. Incubation took place at 28°C; the NH4+ formed in the mixtures was
analyzed at 14-day intervals. The NH/ content increased as the pH increased. The
highest amount of NH4 + was produced in mixtures initially adjusted to pH 4.0,
regardless of the amounts of AI added. It seemed that urea was readily converted to
20
NH4 +, indicating that concentrations of AI3+ had little effect on urease activity and
ammonification processes.
Under the conditions of the 5-hour test, Bremner and Douglas (1971) established
that aluminium chloride (AICb), applied at a rate of 50 ppm Al (on soil weight basis)
did not exhibit any inhibitory effect on urease activity in two Iowa soils (silty clay loam
and clay loam) studied. At the same time, Tabatabai (1977) showed an inhibitory action
of AICh on urease activity in six soils, when the rate of AICh was 5 Ilmoles/g soil (the
inhibition varied between 12 and 50%). At the 0.5 flIDoles/g soil rate, which was applied
only to two of the soils, the degree of inhibition was 3 and 5%, respectively.
Lewis and Slater (1979 a,b) also used - besides mineral iron compounds (see page
9) - AI(N03h.9H 20 and Ah(S04)3.16H20 to obtain fertilizer compositions containing
complexed and uncomplexed urea which when applied on soils led to diminution of
ammonia volatilization from urea. The ratio between the number of Al atoms and the
number of urea molecules should be equal to at least 1:40. Compositions were also
prepared from three components: urea complexed with AI(N03)3 + urea complexed with
Fe(N03)3 + uncomplexed urea.
Badr EI-Din et al. (1985) established that in the three Egyptian soils studied (see
page 12), aluminium sulfate (50 ppm) caused 22.2, 27.2, and 25.8% inhibitions in the
urease activity assayed with the 5-hour test. Ae+ proved to be a weaker inhibitor than
Cu 2+, but a little stronger one than Co2+. AI3+ inhibited urease activity and volatilization
of ammonia from urea even during longer incubations, of at least 7 days. These effects
of Ae+ were weaker than those of Cu 2+, but more marked than those of Fe3+. It should
be mentioned that for the experiments with longer incubations only one soil (silty clay)
was used.
According to findings by Yadav et at. (1986), Ae+ added (in from of AlCb), at rates
of 100, 500, and 1,000 ppm, to samples of a clay loam soil had at each rate a weaker
inhibitory effect on urease activity than any of the heavy-metallic cations tested under
identical conditions (see page 13).
The urease-inhibited urea fertilizer patented by Richards (1995) (see also page 17)
may contain aluminium nitrate instead of ferric nitrate or both aluminium and ferric
nitrates. The molar ratio of aluminium nitrate-urea complex/urea is 1: 1-18.
1.3. SALTS OF ALKALI METALSANDALKALINEEARTHMETALS
In the relation between these salts and urea added to soil one can delineate two aspects:
- their effect on urease activity and urea hydrolysis in soil;
- their effect on volatilization of ammonia resulted from urea hydrolysis.
1.3.1. Effect ofAlkali Metal and Alkaline Earth Metal Salts on Urease Activity and Urea
Hydrolysis
Tomlinson (1964) mixed urea (100 ppm N) and a chemically equivalent amount ofKCl,
K2S04, KH 2 P04, CaCh, MgCl z or KF with samples of an English soil (noncalcareous
sandy loam, pH 6.5). The control was treated with urea alone. After 2 and 7 days of
incubation at lOoC, at a moisture level a little below field capacity, the NH/ released
from urea was detennined. Similar ~ + amounts were found in the salt-treated samples
as in the control. The only significant exception was the KH 2 P04 treatment, in which the
amount of N~ + after 2-day incubation (but not after 7-day incubation) was greater than
21
in the other treatments. This means that the neutral salts studied did not affect urea
hydrolysis.
Under the conditions of the 5-hour test, Bremner and Douglas (1971) established
that NaC!, Na2S04, KCl, CaC!2, and BaCh at a rate of 50 ppm cation (on soil weight
basis) did not inhibit urease activity in the two soils studied.
Tabatabai (1977) studied the influence of BaCh on urease activity in six soils,
through 2-hour incubations at 37°C. When the rate of BaCh was 5 Ilmoles/g soil, the
inhibitions were 2-3% (in three soils), 7% (in two soils), and 12% (in one soil). At a rate
of 0.5 J..l11loles/g soil, which was applied only to two of the soils, BaCh did not have any
effect on their urease activity.
Studying a leached chernozem (pH 6.6) from Armenia and a soddy-podzolic soil
(pH 4.5) from the Moscow region, Abramyan and Galstyan (1981) treated soil samples
(each 1 g) with Ca, Mg, K or Na chloride (1-12 mg cation) + 5 ml of 3% urea solution
in phosphate buffer (pH 6.7) + 0.2 ml of toluene. After incubation (30°C/24 hours), the
NH4 + released from urea was analyzed. In chemozem, CaCl2 and MgCh in low
concentrations (1-2 mg cation) stimulated and in higher concentrations (4-12 mg cation)
inhibited urease activity; KCl was stimulating and NaCI inhibitory in all the
concentrations. In podzol, the low concentrations of Ca, Mg, and Na chlorides had a
stimulating effect, whereas in higher concentrations manifested a tendency to inhibit
urease activity. KCl in all concentrations had, in this soil, too, a stimulating effect on
urease activity.
In continuation of these investigations, Abramyan (1982) worked with samples of
three Armenian soils: leached chernozem (pH 6.6), meliorated solonetz-solonchak (PH
7.6), and irrigated brown meadow soil (pH 8.1).
In an experiment, Abramyan studied the influence of the nature of anion in six
sodium salts (NaC!, Na2S04, Na2C03, Na2Si03, Na2B407, and CHrCOONa) on soil
urease activity. Each salt was added at a rate of 5 milliequivalents of Nail 00 g soil. In
each soil, the weakest inhibitory effect was produced by NaC! and Na2S04 .The most
inhibitory salts were Na2B407 (in chernozern and meadow soil) and Na2CO), Na2Si03,
and Na2B407 (in solonetz-solonchak). The other salts brought about inhibitions of
intermediary extent.
In another experiment, soil samples (100 g) were treated with the increasing
amounts of Na2C03 (1-10 milliequivalents of Na). Urease activity decreased with the
increasing rate of Na2CO], less markedly in the chernozern than in the other two soils.
For example, at 10 milliequivalents of Na, the chernozem retained 50% of its initial
urease activity, whereas urease was completely inactivated in the other two soils.
In a laboratory experiment performed by Fenn et af. (1981b), urea (at a rate
equivalent to 1,100 kg N/ha) with or without CaCh was applied on the surface of wetted
samples of a calcareous silty clay loam soil from Texas. Previously, the soil was
adjusted to 15% CaC0 3 by weight and received 1% (by wei gilt) fresh organic matter
(bluegrass clippings). CaCh was applied at chemically equivalent Ca:urea-N ratios of
0.25 and 0.50. After 3, 6, and 9 days of incubation at 32°C, the urea was extracted and
determined. The results proved that under the influence of CaCh the rate of urea
hydrolysis diminished. In samples treated with urea alone, urea was not detectable after
3 days of incubation, but in the CaCh-treated samples a significant part of the added
urea remained unhydrolyzed even after 9 days of incubation, namely 45-46% at
Ca:urea-N = 0.25 and 24-55% at Ca:urea-N = 0.50, respectively. As diminution of the
22
rate of urea hydrolysis was associated with diminution of the extractable Ca2+ content,
the authors assume the formation of some type of Ca-urea complex which is less
hydrolyzable than the uncomplexed urea.
EI-Shilmawi and EI-Shimi (198Ia) treated 5-g air-dried samples of two Egyptian
soils (alluvial clay soil, pH 7.85 and calcareous sandy loam, pH 8.43) with 15
milliequivalentsll 00 g soil of Na, K, Ca, and Mg sulfate, carbonate, and chloride, then
the samples were moistened up to 60% of WHC and incubated at 30°C for 30 days.
During incubation, urease activity was determined periodically.
The results indicated stimulating or inhibiting actions in dependence of the nature of
anions and cations from the added salts and of the nature of soils. Thus, the sulfates
stimulated urease activity of the alluvial soil in the order: Mg > Ca > K > Na. In the
calcareous soil, Mg and Ca sulfates stimulated urease activity, whereas Na and K
sulfates inhibited it. Ca and Mg carbonates highly accelerated the hydrolysis of urea in
the alluvial soil, but Na and K carbonates decreased it. In the calcareous soil, the effect
ofNa and K carbonates was stimulating, that of the MgC0 3 was inconsiderable and that
of CaC0 3 was inhibiting. The chlorides had a stimulating effect in the order K > Ca >
Mg > Na in the alluvial soil and manifested an inhibitory effect in the order Ca > Mg >
Na "" K in the calcareous soil. TIle most stimulating salts were CaCO.l in the alluvial soil
and K2CO) in the calcareous soil, whereas the most inhibiting salts were Na2CO.l in the
alluvial soil and CaCl 2 in the calcareous soil.
In other incubation experiments (30°C/30 days), EI-Shil1l1awi and EI-Shimi (1981b)
found that NaCi (at 15 milliequivalentsll 00 g soil) had a weak stimulating effect on
urease activity of the alluvial soil, and caused only inconsiderable changes in urease
activity of the calcareous soil. Depending on the soil moisture content, expressed as a
percentage of WHC, urease activity increased in the order 60 > 80 > 100 > 40% (in
samples of both soils untreated with NaCl), 60 > 80 > 40 > 100% (in the NaCI-treated
alluvial soil), and 80> 60 > 40 > 100% (in the NaCl-treated calcareous soil). At 60% of
WHC, the mixtures ofNaCI + CaCl2 (3,000 ppm), in which the Na:Ca ratio was 1:1,2:1
or 3:1, were also studied. It was established that urease activity increased in the alluvial
soil and decreased in the calcareous soil under the influence of the NaCI + CaCb
mixtures in the following order of the Na:Ca ratios: 1:1> 2:1> 3:1.
Samples of the same two soils were treated by Shehata et a1. (1982) with different
amounts of either NaCi or Na2C03 (0, 10, 20, and 25 milliequivalents/l 00 g soil). To
some samples starch (2% on soil weight basis) was also added. Moisture content in the
soil was kept at 60% ofWHC. In continuation, the experimental procedure already used
by EI-Shilmawi and EI-Shimi (l98Ia,b) was applied. NaCI at a rate of 10
milliequivalentsll 00 g soil stimulated, while at higher rates inhibited urease activity in
both soils. Na2CO.l behaved like NaCi in the alluvial soil, but in the calcareous soil it
had a stimulating d'fect inversely proportionate to its rates. In samples incubated with
added starch, the depressive effect of thc high rates of salts was attenuated, which can
be attributed to the synthesis of new urease molecules by the microorganism~ using the
starch as carbon and energy source during the incubation. This action of the starch was
more marked in the calcareous soil than in the alluvial one.
Based on the results of a laboratory experiment, in which samples of a typical
chemozem from Russia were treated with neutral salts in different amounts, Yarovenko
et a1. (1982) concluded that the salts increased soil urease activity in the order: NaCI >
23
MgCl z "" MgCl 2 + NaCI in samples without crop residues, and MgCI2 > MgCh + NaCI >
NaCI in san1ples containing residues of vetch-oats.
Frankenberger and Bingham (1982) treated samples of a California sandy clay loam
soil (pH 6.82) with four rates ofNaCI, NaZS04, and CaCl2 solutions applied to produce
electrical conductivity (Ee) readings of saturation extracts (Ee,) ranging from 2.2 to
22.4 dS/m (NaCI), from 3.8 to 20.0 dS/m (Na 2 S04), and from 2.6 to 21.6 dS/m (CaCh).
The range of Ec,. values includes threshold salinity levels associated with reduced
yields of agronomic crops. To 5-g samples (oven-dry basis) I ml of the appropiate salt
solution was added to give a moisture content of approximately 60% of WHC. The
mixtures were allowed to equilibrate at 25°C for 7 days, then were assayed for urease
activity. This activity decreased with increasing Ec,., in the following order when
compared at the same Ec,. levels: NaCI > CaCh > NaZS04. Thus, the highest Ec,. (salt
concentration) elicited an inhibition of about 21% (NaCl) , 13% (CaCh), and 4%
(Na ZS04).
In a similar study performed by McClung and Frankenberger (1985), three
California soils (clay loam, pH 8.0; sandy clay loam, pH 5.8, and sandy loam, pH 7.0)
were used. The NaCl, Na ZS04, and CaCh solutions applied at four rates produced Ec,.
values of 5, 10, 15, and 20 dS/m in each soil. The reaction mixtures, consisting of 10-g
samples (on a dry weight equivalent basis of field-moist soils) and 1 ml of the
appropiate salt solution, were allowed to equilibrate at 25°C for 7 days, then treated
with urea (200 J..lg N/g soil) in 0.5 or 1 ml solution, and incubated at 30°C for 14 days.
During incubation, the an1ffionia volatilized was determined, and after incubation, the
NH/, N0 3-, and NO z- contents in the reaction mixtures were analyzed. The results
indicated that the added salts, regardless of their type and amount, did not affect
hydrolysis of urea in any of the three soils studied. Their effect on volatilization of
ammonia from urea will be dealt with on page 30.
Of 18 Utah and California soils studied, Kumar and Wagenet (1984) selected three
(differing from each other by their urease activity and physicochemical properties, e.g.,
pH 7.2, 7.8, and 8.0, respectively) for assessing the effect of CaC0 3 addition on soil
urease activity. Ten-g soil samples were treated with 10 mg of urea-N + finely divided
amorphous CaC03 (0, 2, 4, and 8% on soil weight basis) + water up to field capacity,
then incubated at 37°C for 5 hours and, thereafter, the residual urea was determined. It
was found that addition of CaC03 decreased urease activity in each soil. The decrease
was low in the mixtures treated with 2-4% CaC0 3 , but at the 8% CaC0 3 level a
considerable decrease (68, 49, and 29%, respectively) occurred in urease activity of the
three soils studied. The authors hypothesize that the decrease in urease activity was due
to inactivation of urease by amorphous CaC03 and/or to effect of CaCO) on soil pH
and/or to direct influence of CaC0 3 on the reaction of formation of ~)ZC03 from
urea.
In another laboratory experiment, Kumar and Wagenet (1985) studied the effect of
CaCh on urea hydrolysis in two Utah soils (silty loam, pH 7.7 and fine sandy loam, pH
8.0). Soil columns were constructed of acrylic plastic tubing (7.6 em in diameter and 30
em long) fitted at both ends with porous fritted glass plates. The columns were filled
with soil to a uniform bulk density, wetted from the bottom with CaCl z carrier solution
of 1, 5 or 10 dS/m until saturated, and then reversed to a downward direction of flow.
Addition of carrier solution was maintained until a steady downward flow was achieved,
24
then a 100-ml pulse of the carrier solution containing 500 mg Nil as urea was added to
the column, followed once again by the carrier until all added N was leached. Effluent
samples were collected and analyzed for urea and NH4 +. Based on the analytical data it
was calculated from first-order kinetics that the half-life of urea increased (with
increasing CaCh concentration in carrier solution) from 9 to 47 hours in the silty loam
and from 47 to 77 hours in the fine sandy loam. This means that under the influence of
CaCh urea hydrolysis was reduced in both soils.
In an Indian clay loam studied by Yadav el al. (1986), Sr2+ and Ba2+, used in the
form of chlorides and applied at rates of 100,500, and 1,000 ppm cation (on soil basis),
caused, under the conditions specified on page 13, the following inhibitions of urease
activity: 5 and 13% (at 100 ppm), 19 and 25% (at 500 ppm), and 29 and 50% (at 1,000
ppm), respectively.
Working with a nonsaline sandy loan1 soil (pH 8.1) from Punjab, Singh and Bajwa
(1986) treated 3-kg air-dried soil samples with solutions containing 100
milliequivalents/l ofNaCl, Na 2S04, NaHC0 3 or NaCl + CaCb.2H20 (10.3:1) to produce
EC, values of approximately 10 dS/m. The salt-treated soil samples were submitted to
three wetting (with distilled water) and drying cycles and then assayed for the rate of
urea hydrolysis. The reaction mixtures were prepared from 10 g of soil + 10 ml of urea
solution (200 Ilg N/g soil) and incubated at 11°C for 1-28 days. After incubation, the
residual urea, NH/, and N0 3- contents were determined. The results obtained indicated
that the salts inhibited urea hydrolysis in the order: NaCl + CaCh > NaHC0 3 > NaCl >
Na2S04. We should note that complcte hydrolysis of urea took place in 2 days in the
control soil (not treated with salts) and in 14 days in the NaCI+CaClrtreated soil.
Urease activity determinations also showed that the least inhibitory salt was Na2S04.
The experiment conducted on three American soils by Kumar and Wagenet (1984)
(see page 23) was repeated by Kumar et al. (1988) with a sandy loam soil (PH 8.3) from
India. The reaction mixtures had the following composition: 109 of soil + 0, 2, 4 or 8%
of CaC0 3 (by weight of soil) + urea (200 ppm N by soil weight) + water up to 50% of
WHC. After different durations of incubation at 25°C, the residual urea was determined.
It was found that CaC03 markedly inhibited hydrolysis of urea. Thus, after a 3-day
incubation, the residual urea-N was 2,44,69, and 72 ppm in the samples treated with 0,
2, 4, and 8% of CaC0 3 , respectively. All the added urea was hydrolyzed in 6 days at 0
and 2% CaC03 , whereas at 4 and 8% CaC03 hydrolysis of urea was not complete until
day 12.
Ten-g san1ples of another sandy loam soil (pH 7.8; EC 0.33 dS/m) from India were
treatcd with different quantities of NaCl, CaCI 2, and MgS04 (maintaining the ratio of
Na:Ca+Mg at 1: 1) to produce EC values of 4.33 and 16.64 dS/m. After addition of 200
ppm urea-N and water up to 50% ofWHC and incubation at 25°C, the residual urea was
analyzed. The results showed that the added salts reduced the rate of urea hydrolysis. For
example, after 3 days of incubation, 5, 20, and 43 ppm of urea remained unhydrolyzed in
the samples having EC values of 0.33, 4.33, and 16.64 dS/m, respectively. Complete
hydrolysis of urea needed 6 days at EC = 0.33 and 4.33 dS/m and more than 6 but less
than 12 days at EC = 16.64 dS/m.
In comparison with urea, the fertilizer Cardonite, consisting of urea + 5.5% of Mg in
the form of dolomite, caused a significant (33%) increase in urease activity of a
chemozemic brown forest soil (pH 6.0) from Hungary. The reaction mixtures, containing
25
o or 243.5 mg of urea or 400 mg of Cardonitell 00 g soil, were incubated at 37°C for 2

hours (Domb6vari and Kiss, 1988).
Aliev (1988) found that urease activity (expressed in mg NH3 produced by 1 g of soil
in 24 hours) in samples of a dark-chestnut soil treated with Li 2 S04 at rates equivalent to
0,5, 15, and 45 kg/ha had the following values: 2.04, 3.06,2.72, and 2.38, respectively.
In other words, the increase in urease activity was inversely proportionate to the rate of
Li 2 S04 addition.
loppolo et al. (1989) compared the effect of the NaCI on urease activity of an Italian
soil (silty loam, pH 7.94) and on that of a purified jackbean (Canavalia enstformis)
urease. The reaction mixtures consisted of fresh soil equivalent to 2.5 g of oven-dry
matter + 0.2 ml of toluene + 10 ml of 50 mM urea solution with NaCl (at 0-4 M
concentrations). After incubation (20°C/5 hours), the NH/ released from urea was
TABLE 5. Effect of sodium chloride on activity of soil urease and of purified

jackbean urease"
Concentration ofNaCI Urease activity (%/'
(moles/I) Soil lackbean
0.05 100.S±1.27 97.9±1.05
0.10 98.8±1.48 105.I±O.85
0.15 92.6±3.51 IOS.4±2.35*
0.20 88.3±0.1O I 13.3±1.62*
0.25 87.2±O.S2* IIS.2±3.94*
0.50 79.3±O.21 * 162.1±1.1O***
1.00 69.4±0.87*** 96.3±2.33
2.00 48.8±2.80*** S4.6±3'()9**
3.00 43.3±1.19*** 13.0±0.19***
4.00 38.4±O.96*** 7.6±O.28***
"From Ioppolo et al. (1989).
"Soil urease activity in the control rea~1ion mixture: 1.04±0.0 16 ftmoles of urea
hydrolyzed/g soillhour (100%). lackbean urease activity in the control reaction
*.
mixture: 0.826±O.OI69 Ilmoles of urea hydrolyzedlE.lT.lminute (100%).
'Significance level (p): * 0.05: (UII: ••• 0.001.
determined. Table 5 shows that at concentrations of 0.1-4 M, NaCl inhibited urease

activity of soil. In contrast, the activity of jackbean urease was stimulated by low NaCl
concentrations and inhibited only by the high concentrations.
Gomah et al. (1990) mixed salt solutions containing equal equivalent weights of
NaCl and CaCh with 200-g air-dried samples of a calcareous silt loam soil (from Saudi
Arabia), either unamended or amended with 8 g of air-dried digested sewage sludge
(equivalent to 80 tlha) to give salt concentrations of 7.5, 15, 30, and 60 mglg soil. The
moist mixtures were incubated at 30°C and after 0, 1,2,4, 8, and 12 weeks their urease
activity was determined. It was found that urease activity significantly (p=0.05)
decreased with increasing salt concentration. TIle inhibitions caused by the salts at their
four concentrations were, on an average, 8.0, 18.1, 50.2, and 58.2%, respectively. When
sewage sludge was added to soil, the salt effect on urease activity was mitigated.
In contrast, in samples of two calcareous salinized Iraqi soils, having the same
salinity level, urea hydrolysis was not inhibited, but was stimulated by salts in the order:
NaCI > CaCh > Na 2S04 (AI-Rashidi and AI-Jabri, 1990).
26
Laura and Parshad (1991) studied the effect of NaHCO.l addition (and pH increase)
on the hydrolysis of urea in samples (20 g each on oven-dry basis) of an Indian sandy
loam soil. The samples received 0, 0.544, 0.771, 1.027, and 1.284% NaHCO.l (for
increasing pH from 7.53 up to 9.92) and O. 100. 200. and 400 J.lg N as urea/g soil. The
mixtures were moistened to 70% of WHC and incubated at laboratory temperature (20-
30°C). The periodic determination of the amounts of unhydrolyzed urea revealed that
the rate of urea hydrolysis had a perfect negative correlation (r;::: -1.00) with the amount
of NaHC0 3 added (and, thus, with pH).
In a pot experiment, Garcia and Hemandez (1996) used I-kg samples of a calcareous
soil (pH 7.98) from southeastem Spain. The samples were treated with 240 ml solutions
(60% ofWHC) of increasing NaCI and Na2S04 concentrations (0.1, 0.3, 0.6, 0.8, 1.0, and
1.3 M). The control samples received 240 ml of distilled water. At 0.1 to 0.8 M salt
concentrations, NaCI was less inhibitory on urease activity than Na2S04, but the reverse
was true at 1.0 and 1.3 M salt concentrations. However, the inhibition of urease activity
never reached 20(Vo (see also Garcia et al., 2000).
Analyzing the results of the investigations described above one can deduce that, at
unpolluting concentrations, the salts of alkali metals and alkaline earth metals are
ineffective inhibitors of urease activity in many soils or are even stimulators of this
activity in some soils.
1.3.2. Etf(xt of Alkali Metal and Alkaline Earth Metal Salts on Ammonia Volatilization
Reduction of the volatilization of ammonia from urea under the action of CaCb, KCl,
and other neutral salts was described in a series of papers (e.g., Anderson, 1962;
Tomlinson, 1964; Fenn et aI., 1981a,b, 1982; Prusinkiewicz and J6zetkowicz-Kotlarz,
1982; Rappaport and Axley, 1984; Fenn and Hossner, 1985; Gascho, 1986; Fenn, 1988)
and in two patents (Fenn, 1982, 1985). This action of CaCh is attributed to the reaction:
Thus, CaCl 2 reacts with (NH4hC0 3 formed during urea hydrolysis: the carbonate
ion from (NH4hCO.l precipitates as CaC0 3 • The resulting NH 4Cl is a weakly acidic
compound less conducive to ammonia than (NH4hC0 3 ; around the urea granules the
pH decreases reducing the volatility of ammonia.
The action of KCl is explained as follows: K+ replaces Ca2+ on the exchange sites in
the adsorptive complex of soil, then the released Ca2+ and the cr from KCl react with
(NH4hC0 3 producing, according to the reaction equation cited above, NH4Cl and
CaCO).
This mechanism of the action of CaCb and KCl is in close agreement with the
observation by Watkins et al. (1972), according to which NH3 losses from forest floor
were less from mixtures of NH4Cl and urea crystals than from urea crystals and pellets
alone.
CaCIz was more effective (to a 30-40% extent) than was KCl in reducing NH3 loss.
CaCIz was also found to be more effective than MgCb, which can be explained by the
observation that precipitation of CaC03 takes place at a less alkaline pH than that of
MgC0 3 .Mg(OH)2, forming during the reaction between MgCb and CNH4hC0 3 • The
chlorides are more effective than the sulfates of the same metals.
27
In a field experiment on a clay loam at Rothamsted, Rodgers et al. (1984) applied urea
priUs (375 kg Nlha), without or with CaCI 2, as a single dressing for fertilization of a
perelmial ryegrass ley. Ammonia volatili7A1tion losses were measured during 4 weeks after
fertilizer application. It was found that CaCh slightly reduced the NH, loss.
Fenn et at. (1987) studied the influence of plant residues and urea rate illl the
effectiveness of CaCh to inhibit anID10nia volatilization from urea. In an experiment,
samples of a calcareous silty clay loam were unamended or amended with grass clippings
(l % (m soil weight basis)', then treated on surface with urea (11, 55, and 110 g N/m2) with
or without CaCl 2 , at a Ca:urea molar ratio of 0.50, and submitted to incubation at 22°C for
12 days, during which time the volatilized NH3 was determined. In the case of samples not
amended with grass clippings, the cumulative NH3 losses from the three urea amounts were
28, 48, and 48(Yo, respectively, in treatments without CaCh addition, and 19, 5, and 1%,
respectively, in treatments with CaCl 2 addition. In the case of samples amended with grass
clippings, the cumulative NH3 loss from each urea amount was approximately 65%, and was
reduced, under the influence of CaCI 2 , to 35, 30, and 10%, respectively. Thus, incubation of
soil with grass clippings, which enhanced proliferation of microorganisms and synthesis of
new urease molecules, diminished the inhibitory effectiveness of CaCh on volatilization of
NH, from urea. At the same time, the cumulative NH3 losses, expressed as percentages of
the applied urea-N, decreased with increasing rate of urea application, when CaCl 2 had also
been added. TIlese findings were confil111ed with samples of three other soils.
In another experiment, samples of a urease-free sand were treated with Caco,
(15% on
sand weight basis) and amended with increasing amounts of grass clippings (0.01-10%).
Then, urea was applied at the same three rates as in the first experiment; the Ca:urea molar
ratio also was the same (0.50). Based on the determination of NH, volatilized during the
incubation (22°C/12-17 days), the conclusion was drawn that the inhibitory effectiveness of
CaCI 2 on NH3 volatilization progressively decreased with increasing amounts of grass
clippings added to the sand samples.
For studying volatilization of ammonia from urea and urea-MgS04 .1H 2 0 mixture
(UMM) under laboratory conditions, von Rheinbaben (1987) used three sandy and three
loess soils from Germany. The soils were first brought to 30% of their WHC, then filled into
1,250-ml pots; the weight of moist soils depending on their texture was 1,300-1,600 g/pot.
Urea or UMM was added to the soil surface at a uniform rate of 500 mg N/pot. N:Mg ratio
ofUMM varied from I :0.07 to I :0.50. The pots were kept at 25°C for 14 days, during which
the volatilized NH, was measured.
The cumulative NH3 losses from urea and UMM were compared in the six soils at a
single N:Mg ratio (1:0.21). The losses were significantly lower from UMM than from urea
in an acid loess and a sandy soil and insignificantly lower in the other soils. The influence of
N:Mg ratio on NH3 volatilization was studied in two sandy soils. Reduction of NH3
volatilization from UMM was insignificant at the N:Mg ratio of 1:0.07 and very significant
at the ratio of I :0.50 in both soils. The influence of the form (solid or liquid) of urea and
'Before addition of grass clippings, the soil was adjusted to 15% caeo" by weight.
28
UMM on NH3 volatilization was also studied (in a sandy soil). Cumulative NH3 losses were
lower when urea and UMM (N:Mg=I:0.21) were applied as solutions, not as solids on soil
surface, but the loss was insignificantly lower from the UMM solution than from the urea
solution.
Bundy and Eberle (1988) conducted field studies to determine the amount of ammonia
volatilized from several N fertilizers surface-applied on silty loam soils cultivated with
maize in Wisconsin, namely at Arlington and Lancaster. In some experiments, the N
fertilizers were urea-CaCh and urea-KCl solutions. Urea prills served for comparison. At
Arlington, the experiments were carried out in 1983 and 1984, and at Lancaster in 1984.The
fertilizers were administered at rates of 56 and 112 kg Nlha. N :Ca and N:K ratios in the two
fertilizer solutions were 1:1 and 2:1, respectively. The fertilizers were broadcast on the soil
surface after planting (mid-May) but before plant emergence. Field measurements of NH3
volatilization were made at the higher fertilizer N rate only, and lasted 8-10 days.
Use of the urea-CaCh solution significantly reduced NHJ loss relative to urea at both
locations. Thus, at Lancaster, the 19% N loss as NHJ from the applied urea was reduced to
9% in the urea-CaCh treatment. At Arlington, the corresponding N losses were 8 and 3% (in
1983) and 18 and 5% (in 1984), respectively. Ammonia volatilization from the urea-KCl
solution did not significantly differ from that observed with urea at Lancaster, but
significant reductions in NHJ loss occurred with the urea-KCl treatment at Arlington, in
both 1983 and 1984. Urea-CaCh was more effective than urea-KCI in reduction of NH3
loss.
Lightner et al. (1990) made field measurements to determine the amount of ammonia
lost through volatilization from a series of fertilizers (including prilled ammonium nitrate,
granular urea, and solutions of urea, urea-KCl, and urea-CaCh) surface-applied to a
permanent orchardgrass (Dac(y/is glomerata) sod on a silt loam soil (pH 6.0) in Indiana,
during 1982 and 1983 . Annually, each fertilizer was applied at rates of 200 kg Nlha in the
spring and 100 kg Nlha in late summer. Cationlurea-N equivalency was 0.50 KCI and 0.25
CaCh.2HzO. In both years, volatile NH3 losses from ammonium nitrate were insignificant,
but application of urea granules and of urea and urea-KCl solutions led to high cumulative
NH 3 losses that ranged from 27 to 41 % of the applied N in the spring and from 12 to 37% in
the summer. Ammonia volatilization from the urea-CaCh solution was not significantly
lower than that from urea solution in 1982, but adding CaCh to urea solution resulted in a
significant reduction ofNH 3 10sses in 1983.
Gameh et al. (1990) studied the influence of various urea-KCl mixtures on ammonia
volatilization in two soils (a silty loam and a silty clay loam) from Maryland. Soil sanlples
(each weighing 100 g) in 250-rnl flasks (in which the exposed soil surface equaled 38.5 cmz)
were treated with granular urea (control), granular urea + KCl, urea + KCI in solution and
urea coated with powdered KCl, at rates equivalent to 260 kg urealha and 260 kg KCllha.
The moistened samples were incubated at 26°C for nearly 30 days, during which the amount
of volatilized NH3 was determined. The cumulative NH3 losses showed the following
orders: urea> granular urea + KCl > solution urea + KCI > KCl-coated urea in the silty
loam, and urea ~ granular urea + KCl ~ solution urea + KCI > KCI-coated urea in the silty
29
clay loam. Thus, the only urea-KCl mixture, from which NHJ volatilization was
significantly reduced in both soils, was the KCI-coated urea.
At the International Fertilizer Development Center in Muscle Shoals, Alabama,
Christianson et al. (1995) conducted investigations to detennine if the high pH of some
commercial sources of KCl had an effect on the anunonia loss from three contrasting soils
treated with urea in pots with a surface area of 0.04 m2. Five fertilizer-grade KCl sources
ranging in pH from 6.2 to 9.5 were used, and analytical-grade KCl served for comparison.
Rate of KCl was 100 kg K20lha and that of urea was 100 kg Nlha. Urea and KCl were
applied on the soil surface as granules or in solution. Ammonia loss was measured at 2-day
intervals over 2 weeks, during which soil humidity was maintained at field capacity.
The results showed that in the urea-only treatments the cumulative NH3 losses were high
(approximately 50% of N applied) from both granular and solution urea. In two soils, the
loss was significantly reduced due to the use of KCl: the reduction was 30 and 51 %,
respectively, from granular urea and KCl and even higher from solution urea and KCl. In
the third soil, KCl was ineffective in reducing NH3 loss. pH of KCl sources had no effect on
NH] loss in any of the three soils.
In a field experiment on calcareous soil of clay loam texture in the area of Konya,
Turkey, Gezgin and Bayrakli (1995) treated the plots with urea and with urea and
phosphogypsum (PG), the rate of urea being 200 kg Nlha, while PG was applied at two rates
(1,000 kg/ha, PG 1 and 2,000 kglha, PG2) . The total volatile ammonia losses during 57 days
were: 10.6% (urea), 9.2% (urea + PG1), and 12.0% (urea + PG2), the differences between
losses being significant (p=0.05). Thus, PG at the lower rate reduced, and at the higher rate
increased, NH3 volatilization from the urea-treated soil.
For reducing N loss from urea fertilizer, Avramchuk (2000) patented a mixture
consisting of urea and PG at the ratio of 1:4 (weight per weight) and explained the reduction
of N loss through the following mechanism: Ca(HS04h of PG reacts with (NH4hC0 3
(product of urea hydrolysis) with formation of (NH4)2S04 which is more stable than
(NH4hC0 3 •
It should be mentioned that superphosphate, which contains an acid salt, Ca(H 2P0 4h.
and CaS04, reduced volatilization of ammonia from urea (e.g.. Tomlinson, 1964;
Mahendrappa and Odgen, 1973; Carrier and Bernier, 1976; TyaUi, 1982; Fan and
MacKenzie, 1993a; Sengik and Kiehl, 1995a,b). Moreover, Fenn et al. (1990) found that
Ca(H 2P0 4h enhanced the effect of CaCl 1 to reduce volatilization of NH3 from urea-treated
samples of the two Texas soil studied (a silty clay loam, pH 7.7 and a sandy clay loam, pH
4.5). Ouyang et al. (1998) demonstrated that triple superphosphate, added to urea-treated
soils, is able to reduce N10 emission, too. Samples of three Canadian soils were used: a clay
(pH 5.5), a silty clay loam (pH 6.1), and a sandy clay loam soil (pH 6.1).
The effects of superphosphate to reduce gaseous losses from urea-treated soils are
consistent with the findings that triple superphosphate reduced the rate of urea hydrolysis in
soils due to inhibition of urease activity (Fan and MacKenzie, 1993a,b; Ouyang et aI.,
1998).
30
Addition of KCl and P fertilizers to urea solution is recommended for inhibiting

volatilization loss as NH3 and, thus, for increasing efficiency of urea on sugarcane
(Anonymous, 2(00).
In the literature there are also data, according to which CaCh and other neutral salts did
not reduce or even enhanced volatilization of ammonia from urea. We quote some
examples.
Volk (1961) mixed equal weights of urea pellets and CaS04.2H20, then placed the
mixture and the control pellets (at a rate of 112 kg N/ha) on the surface of samples of a bare
moist soil (fine sand from Florida) and determined the amount of ammonia volatilized
during incubation (7 days). It was found that gypsum addition to urea had no measurable
effect on NH3 evolution.
McClung and Frankenberger's (1985) study on the effect of four concentrations of NaCl,
Na2S04, and CaCh on hydrolysis of urea in three California soils was mentioned on page
23. Volatilization of ammonia from the urea-treated clay loam and sandy loam was also
studied. In the clay loam the two lower concentrations of the three salts did not affect and
their two higher concentrations enhanced volatilization of NH.l from urea. In the sandy
loam, NH3 volatilization was slightly increased by NaCl, strongly increased by Na2S04, and
strongly decreased by CaCb. The effect of NaCl and Na2S04 was dependent on their
concentration, whereas that of CaCl z was nearly the same at each concentration.
Studying the effect of NaCl, Na2S04, and NaHC0 3 on the volatilization of anm10nia
from an Indian sandy loam soil (pH 8.1) treated with urea, Singll and Bajwa (1987) found
that during incubation (16 days at 28°C) the cumulative volatile NH.l losses increased in the
following order: untreated soil (18%), soil treated with NaCI (21 %), with NaHC0 3 (32%),
with Na2S04 (35%).
1.4. BORON COMPOUNDS
Sor (1968, 1969) prepared pellets from urea, borax (Na2B407.1 OH 20) (0.45% by weigllt of
urea, limits of the borax amount being 0.01 and 10%) and a hydrophobic material,
preferentially the san1e asphalt -microcrystalline wax blend also used for preparation of urea
pellets with CUS04 or Pb acetate. The technology of preparation was also the same (see
page 5). Urea prills without inhibitor served for comparison. The pellets and urea prills,
applied at a practical N rate, were incubated on the surface of a sandy loam soil (containing
6.4% moisture) at 25°C for 48 days, during which the volatilized an1l110nia was estimated. A
significant part (approximately 39%) of the urea-N was lost as NH.l in 48 days from the soil
treated with urea priUs, whereas NH3 loss was only 17% when the soil received the pellets
prepared from urea, borax, and hydrophobic material.
According to the descriptions in three inventions patented for preparation of pellets from
urea + borax + hydrophobic material, Sor et al. (1968, 1971) and the Esso Company (1969)
use - in place of the asphalt-microcrystalline wax blend - other hydrophobic materials,
namely primary and secondary amines and diamines having a hydrocarbon chain of 8-22
carbon atoms, preferentially octadccylamine, CHr -{CH 2 )1,NH 2 Diamines having the
o
31
formula H2N-{CH2h-NH-R (where R is a C, - C20, preferably a C IX fatty acid group) may

also be used. Many other hydrophobic substances are also recommended, for example
stearamide, oleates, furfuryl alcohol resins, organosilicones. All the hydrophobic materials
are used in preferred amounts of 0.1-2% by weight of urea, the limits being 0.01 and 5%.
They are added to the urea melt containing the inhibitor or are applied for coating the urea-
inhibitor pellets.
For testing anm10nia volatilization from urea, a loamy sandy soil from New Jersey (pH
about 6.4; initial moisture content = 65-75% of field capacity) was used. Urea was applied
at rates of 0.5 and I glkg soil, respectively. The incubation lasted 4 and II days. It was
found that during both incubation times the volatile NH3 losses from the two urea amounts
presented the following order:
urea> urea + 4% borax> urea + 4% borax + 0.5% octadecylamine (ODA), and urea>
urea + I eyo ODA > urea + 4% borax + 0.1 % ODA, respectively.
In another testing of NH3 volatilization from the same loamy sandy soil, cOl1IDlercially
prepared inhibitor-treated urea prills were applied. Analysis of NH3 volatilized during 3, 7,
10, and 14 days of incubation showed that in this case, too, NH3 volatilization was highest
from urea without inhibitor, intermediary from urea with 4% borax, and lowest from urea
with 4% borax + 0.1% ODA.
Sor et al. (1971) devoted studies also to other aspects related to the diminution of
anmlonia volatilization from urea. Comparison of the effects of three boron compounds,
namely borax, boron oxide (B 20 1), and boric acid (H)BO)), showed that increasing the rate
of percent boron content in the urea granules decreased the amount of NH3 lost by
volatili7,ation regardless of the boron source. Since borax is cheaper than H20) and H3BO),
it remains the preferred source of boron.
In another experiment, NH3 volatilization from urea was estimated by using urea-borax
granules treated with different hydrophobic materials. Besides ODA, other two hydrophobic
amines (Duameen T = N - tallow trimethylene diamine, and Arrneen residue = R-NH2)' as
well as a hydrophobic non-amine (Marcol 72 = white oil, mixed naphthenic/paraffinic
based), a hydrophilic amine (n-butylamine), and a hydrophilic non-amine (Natrosol 250 =
hydroxyethyl cellulose) were tested. All were applied at 1% by weight of urea. The
hydrophobic amines were effective, the hydrophobic non-amine and the hydrophilic amine
were less effective, and the hydrophilic non-amine was ineffective in reducing the volatile
NH3 losses from urea-borax granules. The conclusion may be drawn that the effect of ODA
is due not to its amine radical, but to its hydrophobic nature determined by the C IX
hydrocarbon chain.
When Marcol 72, Orchex 792 (a naphthenic spray oil), Necton 37 (a paraffinic oil) as
well as a vegetable oil (olive oil) were used as hydrophobic materials at 1% by weight of
urea, the NH3 loss from the urea--4% borax granules was reduced. Similarly, the NH.1 loss
decreased following treatment of urea with: 4% borax + 1% p-tolualdehyde, 4% u-
naphthaldehyde, 2% sodium alkyl benzene sulfonate, 2% calcium petroleum sulfonate
(molecular weight of sulfonate = 900--1,0(0), 1% silicone oil (low phenyl dimethyl
32
polysiloxane), 2% Brij 92 (oleyl ether of polyethylene glycol) or 1% Krylon (a 6% acrylic

plastic solution in toluene).
For studying the t-'ffect of boric acid on the anunonia loss from urea applied to a forest
soil, N6mmik (1973) used microplots installed in a Scots pine stand, located 15 km north-
wcst of Stockholm. The microplots received one large urea pellet (2.06-g tablet) or one
2.l7-g tablet containing 95% urea and 5% boric acid (weight/weight). The tablets were
applied to the soil surface at a rate of 200 kg Nlha. The amount of volatilized NH3 was
measured during 28 days (from May 23 to June 20; the average daily mean temperature was
13.3°C). The cumulative NH3 loss from the added urea-N was 20% in the urea-only
treatment and much lower, nanlely 9% in the urea-boric acid treatment.
According to the invention patented by Besekau et al. (1974), boric acid was used as the
boron source, whereas many compounds served as hydrophobic materials, namely primary
and/or secondary aliphatic amines having a hydrocarbon chain of C6-C n or saturated and
unsaturated fatty acids whose hydrocarbon chain has up to 24 carbon atoms or mixtures of
aliphatic amines + alkanoic acids with C1-C 24 • The amount of boric acid was 0.1-5%,
preferably 1-3.5%, and that of hydrophobic material 0.05-4% by total weight of the fertilizer
urea composition. In the mixtures of aliphatic amines and alkanoic acids, the molar ratio
ranges from 10: 1 to 0.5: 1; the preferred ratio is 1: 1. It should be mentioned that the boric
acid in the fertilizer urea composition serves not only as a urease inhibitor, but also as a
trace element (micronutrient).
The fertilizer was prepared by incorporation of boric acid and hydrophobic material into
the urea melted at 130°C, then the mixture was prilled. Besides boric acid, other trace
elements at a total concentration of 0.1-0.5% can also be incorporated into the urea melt.
Effectiveness of the prills was tested on a sandy loam soil (moisture content: 40% of
WHC) at 20D e. Urea was applied at a rate of 100 kg Nlha. During incubation which lasted
18 days the volatilized anlillonia was determined; its amounts showed the following order:
urea> urea + 2.5% boric acid> urea + 2.5% boric acid + hydrophobic material.
Anunonia loss was lowest from the urea-2.5% boric acid prills, in which the
hydrophobic material, used in a 2% amount by weight of priUs, was the aliphatic amine SP
treated with acetic acid at I: I molar ratio.
In the six soils studied by Tabatabai (1977) (see page 8), borax, tested at a concentration
of 5 ~unoles Big soil. caused. during 2-hour incubation at 37°C, the following inhibitions in
urease activity: 98, 27, 18, 15, 14, and 13%, respectively. At 10 times lower borax
concentration only two soils were studied. The degree of inhibition was II and 13%,
respectively.
TIle finding by Abramyan (1982) that borax inhibited urease activity in the three soils
studied has been mentioned on page 21.
Raguotis and Shleinys (1986) installed microplots in a pine forest in Lithuania and
introduced granular urea (180 kg Nfha) containing no or 5% boric acid (on urea weight
basis) under the canopy. Volatilization of anunonia was assessed during 46 days and it was
found that boric acid did not reduce the volatile NHl loss.
33
Xue and Li (1987) showed the inhibitory effect of borax on urease activity of a
periodically water-logged paddy soil. The borax, used at a concentration of 100 ppm by soil
weight, exhibited a 38.92% inhibition of urease activity. TI1e reaction mixtures were
prepared from 5 g of soil and 10 ml of 10% urea solution without or with added borax. The
incubation took place at 37°C and lasted 48 hours.
Zhan et af. (1993) described a technology for cogranu1ation of urea and borax. This urea
fertilizer contains up to 4% borax and consists of 4-6-mm granules. Fan and Ye (1995)
applied the borax-containing urea to paddy field and found that the borax acted as an
inhibitor of soil urease activity, preventing the gaseous N losses from urea.
Another use of boric acid also related to fertilizer urea will be dealt with below. For
preparing controlled-release fertilizer urea granules, Otey et al. (1984) worked out two
technologies, in one of which boric acid is also used. Pregelatinized maize (Zea mays) flour
is dispersed at 25-30°C in a solution of urea (50 ml H20/50 g urea) and concentrated
NH40H (4 mlllOO g of final dry product). Then boric acid (2 gllOO g of final product) is
mixed with the gelatinized flour-urea mixture to form a rubbery mass. Air-dried maize
starch (18 gl100 g of final product) is then added slowly with stirring which causes the
rubbery mass to break into small particles coated by starch. On a dry basis this product
contains 2% boric acid «(U5% B), 18% ungelatinized starch, and 80% urea and gelatinized
flour. Boric acid in this product serves as micronutrient; its effect on soil urease activity was
not tested.
1.5. FLUORIDES
Tomlinson (1964) studied the effect of K and Ca fluorides (KF, CaF 2) on volatilization of
ammonia from urea in two English soils (calcareous clay loan1, pH 8.1 and noncalcareous
sandy loam, pH 6.5). The soil samples were treated with 2,000 ppm urea-N and a
chemically equivalent amount of KF or CaF2 and moistened to about 40% humidity which
was less than the field capacity. Samples not treated with fluoride were the controls.
Incubation took place at 9°C and lasted 5 days. Determination of the NHJ volatilized during
the incubation gave the following percentage NHJ losses in the clay loam: 8.6 (control),
41.5 (KF), and 6.3 (CaF 2), and in the sandy loam: 32.6 (control), 52.3 (KF), and 29.3
(CaF 2 ). In other words, the soluble fluoride (KF) stimulated, while the insoluble salt (CaF 2)
inhibited NHl volatilization from urea in both soils.
Sor (1968, 1969) prepared not only urea pellets with CUS04, Pb acetate, and borax +
hydrophobic material (asphalt + microcrytalline wax) (see page 5), but also urea pellets with
NaF (0.45% by weight of urea) + asphalt-microcrytalline wax blend. In an experiment to
study the volatilization of an1l11onia from urea applied on the surface of samples of a sandy
loam soil, NaF from the urea pellets reduced the cumulative NH3 loss in 48 days to about
12%, while this loss from the control urea prills was 39%.
According to Kozlovskaya et al. (1972), urease activity in peat bog soils was not
inhibited at all by NaF, although the fluoride was applied at high concentrations (16--64 111M
in reaction mixtures).
34
In the pot experiments carried out by Gaponyuk and Kuznetsova (1984), samples taken
from the 0-20-cm layer of a soddy-podzolic soil (PH 7.(5) from Russia were treated with
NaF at rates ranging from 0.1 to 3 g F/kg soil, moistened to 60% ofWHC and preincubated
for I month, then sown with different plants. During the preincubation period, soil urease
activity was measured 20 times. Mean values of this activity showed insignificant changes
at rates of 0.1-0.7 g F/kg soil and significant increases at the 1-3 g F/kg soil rates. Thus, in
these experiments NaF exerted no inhibiting effect on soil urease activity.
Ablizova and Tomina (1997) carried out pot experiments in which samples of a dark-
chestnut soil from Kazakhstan were treated with NaF. In 1991, NaF was applied at rates of
0, 10, and 50 mg F/kg soil, before planting tomatoes. Soil urease activity was determined
several times during the vegetation period. It was found that the activity was inhibited by
the higher NaF rate in spring, by both rates in summer, and was stimulated by both rates in
autumn. In 1992, NaF was applied at higher rates (50 and 500 mg Flkg soil) and the test
plant was onion. Soil urease activity, measured in summer, showed an 11.5% increase at the
lower NaF rate and a 65% decrease at the higher NaF rate. One can state that in these
experiments, in contrast to those of Gaponyuk and Kuznetsova (1984), NaF behaved as an
inhibitor of soil urease activity and the inhibition lasted several months.
1.6. ARSENIC COMPOUNDS
Applying the 5-hour test, Bremner and Douglas (1971) established that arsenic chloride
(AsCl 3 ), arsenic trioxide (As Z0 3), and arsenic pentoxide (As Z0 5) used at a rate of 50 ppm
(soil basis) brought about only negligible inhibitions in the urease activity of a silty clay
loam and a clay loam from Iowa: on average, the degree of inhibition by the three arsenic
compounds was 3, 4, and 3%, respectively.
Tabatabai (1977) treated 5-g samples of six Iowa soils with 1.5 ml of sodium arsenate
(Na zHAs04 ) or sodium arsenite (NaAsO z) solution (at a rate of 5 f.Ulloles As/g soil). 0.2 ml
of toluene, 7.S ml ofO.OS M Tris buffer (pH 9.0) and 1 m1 of 0.2 M urea solution and then
incubated them at 37°C for 2 hours. In two of the soils, the arsenic compounds were used at
a lower rate. too (0.5 Ilmoles As/g soil). Based on the determination of the urea remaining
unhydrolyzed during incubation, it was deduced that none of the two Na zHAs04
concentrations exerted any inhibitory effect on soil urease activity. In contrast, NaAsO z, at a
lower rate, inhibited urease activity in the two soils studied (degree of inhibition: 7 and
14%, respectively), whereas at the higher rate NaAsO z had a urease-inhibiting effect in each
of the six soils studied (degree of inhibition: 98,44,27,24,18, and 9%, respectively).
It should be added that under similar conditions sodium tungstate (Naz W04) behaved
like Na2HAs04, and selenious acid (HzSeO,l) like NaAsO z. The inhibitions caused by
HzSeO] were Sand 9% (at the lower Se rate), and 33,24,24, 19, 16, and 14% (at the higher
Se rate). But Aliev (1988) recorded increased urease activity in samples of a dark-chestnut
soil treated with sodium selenite (Na2SeO]) at rates equivalent to S, IS, and 45 kg'ha. The
increase was inversely proportional 10 the rate of selenite addition.
35
1.7. SULFUR COMPOUNDS
Conrad (1940) compared the antiseptic effect of carhon disu(fide (CS 2) with that of toluene
in the determination of urea hydrolysis in two California soils. The hydrolysis rate was
similar in presence of the two antiseptics. As urea hydrolysis in presence of toluene differed
only to a slight extent from that measured in absence of antiseptics, the conclusion may be
drawn that neither toluene nor CS 2affected soil urease activity.
Using the 5-hour test for studying the effect of 50 ppm of sodium sulfite (Na2S03),
sodium hisu!/ite (NaHSOI ), and lead sur/ide (PbS) on urease activity in two Iowa soils,
Bremner and Douglas (1971) recorded only negligible inhibitions due to these three sulfur
compounds (on average, < 1,4, and 3%, respectively).
The effect of sodium sulfite was also tested with other two soils: an alluvial soil and a
leached chernozen1 (Kiss and Pintea, 1987). The reaction mixtures had the following
composition: 5 g of air-dried soil + 1 ml of 0.6% urea solution + 9 ml of aqueous solution or
suspension of the compound to be tested at 2% rate by weight of urea (i.e.. 0.12 mg of
Na2S01 to 6 mg of urea). Reaction mixtures in which the solution or suspension of the test
compound was replaced by distilled water served for comparison. Incubation was carried
out at laboratory temperature. At 1-2-day intervals, drops were taken from the aqueous
phase of reaction mixtures for detecting the unhydrolyzed urea. 111e drops were placed on
chromatographic paper and, after drying, sprayed with a chromogenic reagent' to visualize
the yellow spot of urea. Na2S0, does not interfere with the detection of urea. If urea
hydrolysis is complete, no colored spot appears. The time (days) necessary for complete
urea hydrolysis is registered.
The results showed that complete hydrolysis of urea in reaction mixtures with or without
Na2S01 required the same time, namely 6 days in the alluvial soil and 9 days in the leached
chernozem. In other words, Na2S01 did not inhibit urease activity in these soils.
In a field experiment conducted on a silty clay loam soil (pH 6.0) in Alberta, Malhi and
Nyborg (1979) determined urea hydrolysis in plots treated with urea (control) or with a
mixture consisting of two parts of urea and one part of calcium sulfide (CaS) or phm,phorus
pentasu(fide (P 2SS; P4 S lO ). Urea and the urea-CaS or urea-P 2 SS mixtures were administered
at a rate of 112 kg N/ha, in bands at a depth of 5 cm. After 5 and 10 weeks, the NH4 + and
NO,- contents in soil were analyzed. The analytical data obtained indicated that after 5
weeks urea hydrolysis was complete in the control plot, but it was only 71 and 68% in the
plots treated with urea-CaS and urea-P 2 S5, respectively; after 10 weeks, urea hydrolysis
became complete in all plots. This means that the inhibitory effect of these inorganic sulfur
compounds on urease activity in the studied soil was not strong and long-lasting, although
they were applied in a considerable amount.
"The reagent is prepared as follows: I g ofp-dimethylaminobenzaldehyde is dissolved in 30 mI of absolute ethanol,

then 7.6 mI of syropous (89%) a-phosphoric acid, 22.4 mI of distilled water, and ISO mI of absolute ethanol are
added.
36
Sodium trithiocarbonate (Na2CS3) is known as a nitrification inhibitor and fumigant. In

soil, Na2CSJ undergoes a rapid chemical decomposition into sodium sulfide (Na2S) and
carbon disulfide (CS2). Its nitrification-inhibiting effect is due just to CS 2 (powlson and
Jenkinson, 1971; Bremner and Bundy, 1974; Malhi and Nyborg, 1982; Maddux et al.,
1985). A study of the effect ofNa2CS J on urea hydrolysis in soil was initiated by Ashworth
et al. (1977) in a laboratory experiment with samples of a silty clay loam at Rothamsted.
The effect of CS 2 was also tested. No experimental details are given in the paper, it is only
mentioned that Na2CSJ and CS2 did not have any effect on urea hydrolysis. In contrast, a
detailed description is given for a field experiment on the same soil. An 18% urea solution
(70 or 100 kg Niha) without or with ammonium trithiocarbonate, (NH4hCS 3 (at a rate
equivalent to 8 kg CS 2iha ) or with CS2 (11 kglha) was injected into the soil of plots. It was
found that urea hydrolysis was complete in all plots in less than two weeks. In other words,
(NH4hCS 3 and CS 2 did not inhibit soil urease activity at allor, if they did, the inhibition did
not last two weeks.
Ashworth et al. (1979) incubated samples of the same Rothamsted soil with urea and
Na2CS3 at 24°C for 5 weeks and analyzed the residual urea weekly. The results showed that
Na2CS3 exerted a very weak inhibitory effect on urea hydrolysis, but even this effect was
evident only in the first week.
The weak inhibitory effect of Na2CS3 on urea hydrolysis, unlike its strong inhibitory
effect on nitrification, was not due to CS 2 released from Na2CS3 in soil, since in separate
tests CS 2 applied in an amount equivalent to that of Na 2CS J showed no urease inhibition.
Working with another soil (clay loam from Alberta), Ashworth et al. (1980) observed
that Na2CS3, used at a rate of 200 mglkg soil in the presence of 400 mg urea-N/kg soil,
inhibited urease activity during the first 17 hours of incubation at 23°C (degree of
inhibition: 40%), but the inhibition decreased very much (up to 6%) after 41 hours. In this
way, the very weak inhibitory capacity of Na2CS3 on soil urease activity was confirmed.
The study of the capacity of ammonium thiosulfate, <NH4)2S20J (ATS) to inhibit soil
urease activity was initiated by Goos (1985a). Under laboratory conditions he tested the
effect of ATS on hydrolysis of urea from the liquid fertilizer urea-ammonium nitrate (VAN;
28-0-0) in two soils (loam, pH 7 and silty clay loam, pH 7.5) from North Dakota. Because
of the hygroscopic nature of ATS, it may have applicaticIDs as a urease inhibitor only with
liquid fertilizers.
Commercial-grade ATS was added to commercial-grade UAN in a proportion of 1,2,5
or 10% volume/volume. From the solution obtained, a droplet (0.1 ml) was taken and placed
on the surface of moistened soil sample (25 g). The next step was incubation at 25°C for 2
or 4 days. As a source of urease, besides the soil samples, a pure jackbean urease
preparation was also used; in this case, the incubation lasted 1 hour.
After incubation, the residual urea was extracted from the soil and from the reaction
mixtures with pure urease and measured. The results indicated that in both soils ATS, even
at its lowest concentration (1 %), inhibited urea hydrolysis (degree of inhibition: between 15
and 35%). The inhibition increased with increasing ATS concentration and reached 40-52%
37
at the highest concentration tested (10% ATS). At the same time, ATS did not have any
effect on activity of jackbean urease.
In another experiment, with the clay loam soil, the effect of sodium thiosulfate
(NaZSZ03, reagent grade) was compared with that of three commercial ATS products,
containing impurities (free ammonia, sulfite, sulfide, sulfate). All commercial ATS products
and reagent sodium thiosulfate gave similar levels of inhibition (21-28%) in urea hydrolysis.
Data concerning the inhibitory effect of ATS on nitrification of NH4 + were also
obtained.
GODS (1985b) studied, under laboratory conditions with 20-g samples of a loamy soil,
the effect of ATS on urea hydrolysis in correlation with its effect on ammonia volatilization
from UAN. The fertilizer solutions in the form offour 0.025-rnl droplets were placed on the
soil surface approximately 2 cm apart from each other. During incubation (at 25°C/5 days),
the NH3 volatilization and, after incubation, the residual urea were determined.
ATS, at each of its rates applied (1-25% volume/volume), decreased urea hydrolysis
which was increasingly inhibited with increasing rates of ATS. At low rates (1, 2, and 5%),
ATS reduced NH3 volatilization, too. But NH3 volatilization increased somewhat at higher
ATS rates. It is possible that at higher levels of ATS there can be NH3 loss from ATS itself.
Under similar laboratory conditions, but using 40-g samples of the same soil, the effect
of ammonium polyphosphate (APP) fertilizer (10-34-0), added to UAN or to UAN + ATS
was also tested. APP was applied at a 20% rate and ATS at a 2% rate by volume ofUAN.
The solutions were applied on the soil surface in the form of a single small (0.1 ml) droplet
("spray" application) or a single large (0.5 m!) droplet ("dribble" application). After 5-day
incubation at 25°C, it was established that, at the O.I-ml droplet size, ATS, APP, and ATS +
APP did not inhibit urea hydrolysis, but reduced, to some extent, volatilization of NH3 from
UAN. At the O.5-rnl droplet size, ATS markedly slowed urea hydrolysis (from 83 to 48%),
APP decreased it to a lesser extent (from 83 to 74%), whereas the two compounds used
together decreased it up to 45%; ATS, APP, and ATS + APP significantly reduced the
volatile NHl loss, by 61, 27, and 72%, respectively.
Based on Goos' (1985a) observation, according to which ATS inhibited soil urease
activity, but did not have such an effect on jackbean urease, Goos ef al. (19800) and Goos
(1987) assume that ATS acts indirectly on soil urease, that is ATS reacts rapidly and
abiotically with Fe(OHh and MnOz from soil, forming tetrathionate anion and Fe 2+ and
Mn2+ cations which inhibit the enzyme by binding to its sulfhydryl (SH) groups:
2Fe(OHh + 2S 20/- + 6H+ -----:>~ 2Fe 2+ + S40t + 6H zO

M1102 + 2S20/- + 4H+ >
Mn2+ + S406 2. + 2H20
SH S
/ / \
Urease + Fe2+ ~ Urease Fe + 2H+
\ \ /
SH S
38
SH S
/ / \
Urease + Mnz+ -----:>::. Urease Mn + 2H+.
\ \ /
SH S
To obtain evidence in favor of this hypothesis, Goos (1987) added Na ZS04 or NaZS203
solution to samples of a silty clay soil and, after 12 hours, removed SO/- and S20/- by
repeated extraction and centrifugation. The soil samples were then analyzed for urease
activity and Fe and Mn contents extractable with 0.1 M HCI. It was found that under the
influence of the thiosulfate treatment - as compared to the sulfate treatment - urease activity
decreased in a proportion of 40%, whereas the amounts of acidosoluble Fe and Mn
increased even in a higher proportion.
The effect of APP to reduce NH3 volatilization was attributed to its capacity to
temporarily moderate the alkalinity produced by rapid urea hydrolysis (Goos, 1985b; Goos
ef al., 1986a,b; Fairlie and Goos, 1986). This means that APP is, essentially, not an inhibitor
of soil urease activity.
Laboratory investigations related to ATS were carried out by Gascho (1986) as well. He
used a loamy sand soil (pH 6.8) from Georgia and determined the amount of NH3 evolved,
at 30°C during 7 days, from 100-g soil samples treated with UAN; UAN + ATS; UAN +
KCl; UAN + KCl + MgCl 2 ; UAN + KCl + MgCIz + CaCh; UAN + KCI + ATS, and UAN
+ KCI + MgCIz + ATS, the N:K 20:Mg:Ca:S ratio being different. Soil samples treated with
urea only or NH 4NO] only were the controls. Nitrogen was applied at the same rate in each
sample, namely 184 kg/ha (based on the area of the soil sample surface). All the solutions
were surface-applied.
No loss of NH3 was detected from the NH 4 NOdreated control, but 48% of the applied
N was lost from the urea-treated control and 20% was lost from the UAN-treated soil. ATS
added to UAN at the N:S ratio of 32: I did not reduce, but at the N:S ratio of 16: 1 reduced,
significantly the NHl loss (from 20% to 19 and 17%, respectively). In soil samples treated
with UAN + KCI + ATS at N:K 20:S ratios of 32:16:1 and 16:8:1, the NH3 loss decreased
very much (to 5 and 4%, respectively). Ammonia loss from the soil treated with UAN +
KCI decreased very remarkably only when UAN and KCl were applied at N:K20 ratios of
1:2,2:5, and 1:3, the loss being 8, 0, and 0%, respectively. Ammonia loss measured at the
N:KzO:Mg:S ratio of 16:8:1:1 and at the N:K2 0:S ratio of 16:8:1 was identical (4%). In
samples treated with UAN + KCI + MgCh + ATS or CaCh (in place of ATS), the losses
were very reduced (1-7%). Therefore it appears that a) MgCI 2, if not needed for Mg
nutrition, can be omitted as it will not bring about much additional NH3 loss reductions in
the presence of both KCI and ATS, and h) CaCl 2 can be substituted for ATS in soils not
requiring sulfur fertilizer.
The laboratory investigations were continued by Goos and Fairlie (1988) with two other
North Dakota soils (silty clay, pH 6.6 and loamy sand, pH 5.8) to study the effect of ATS on
39
hydrolysis of urea from VAN applied in the form of 50 droplets of 0.01 ml or 5 droplets of
0.1 ml or one droplet of 0.5 ml, on the surface of 100-g (oven-dry equivalent) soil samples,
the water content of which was adjusted to field capacity or to wilting point. The soil
surface area was about 50 cm2 and the soil depth was about 2 cm. TIle N rate applied, based
on the soil surface area, was about 30n kglha. The ATS solution (a high quality commercial-
grade product) was used at a single rate (5% by volume of the VAN solution). VAN, to
which water was added in place of ATS, served for comparison. All samples were incubated
at 20°C and the residual urea was determined three times during the course of urea
hydrolysis.
One can deduce from the results obtained (Figure I) that ATS inhibited urea hydrolysis
in both soils and at both water contents. Vrea hydrolysis slowed down with increasing
droplet size in both the presence and absence of ATS. The overall rate of urea hydrolysis
was somewhat slower at wilting point than at field capacity. The inhibitory effect of ATS on
urea hydrolysis was strongest with the larger droplet size and at wilting point. Comparison
of the two soils revealed that ureolytic capacity of the loamy sand was considerably weaker
than that of the silty clay. At the same time, ATS inhibited urea hydrolysis more strongly in
the loamy sand than in the silty clay.
Brenmer et al. (1986a) also carried out laboratory testings for evaluation of ATS as a
soil urease and nitrification inhibitor. Iowa soils were treated with different amounts of ATS
and then incubated at 20, 25, and 30°e. ATS, even when applied at very high rates, up to
1,000 ~lg/g soil (LOO() ppm) had no inhibitory effect on urea hydrolysis in soil.' Nitrification
was inhibited only at rates higher than 50() ~g/g soil. The conclusion that ATS has no
practical value as a soil urease and nitrification inhibitor contradicts the results of the
laboratory experimcnts of Goos (1 985a,b ), Gascho (1986), and Goos and Fairlie (1988) and
also the results of some of the other investigations conducted under field or laboratory
conditions.
Fairlie and Goos (1986) installed microplots on a clay soil (pH 6.9). Polyvinyl chloride
(PVC) cylinders 20 cm long and 12.7 cm in dianleter were forced 5 cm into the soil. The
soil surface was bare or covered with wheat residue at a rate of 2.65 tlha (3.5 g/plot). The
microplots were treated with VAN, VAN + 2.5 or 10'% ATS, VAN + 10 or 20% APP or
with VAN + 2 or 5'% ATS + 20% APP. Rale of VAN was 150 kg N/ha, ATS and APP being
additional N sources. Each of these mixed fertilizer solutions was applied at a rate of 0.817
mllmicroplot, either as a large droplet, as a spot in the center of the microplot (dribble
application), or as a fine spray evenly distributed over the soil surface. Then the ammonia
volatilized from the microplots was estimated cumulatively. The principal results were the
following.
In the case of bare or residue-covered microplots treated with VAN without ATS and
APP less NHl was lost with dribble than with spray application. The inhibitory effect of
'Similar results were obtained. when the eftect of ATS to inhibit soil urease activity was compared with that of
other compounds (see Sections 4.9.1.1 and 4.9.1.5).
40
0.01 ml 100
100 0.1 ml 0.1 ml
0.01 ml
0.01 ml + AlS
O.Sml 10
80 0.1 mI + IITS O.Sml
{
't..
IIN 80 0.5 mt + IITS "'""
N
.?;>
80
0..01 ml + ATS
~,
.@ 0.1 ml + ATS
~
1:l .
>.
"'<!" 40 O.Sml + ATS
'2::"
;:J
;:J
20
100 121i
Incu Dation time (hours., InL'ubati(m time (hours)
100
100
C 0.01 mt
0.1 ml
0.1 ml D 0.01 ml
80
# O.Sml
." O.Srnl ~
i:l
",..
."
80
~,
.?;o
8 0.01 rnl + ATS
2
."
~
0.1 ml + ATS ."
,... 40 0.01 ml + .US
0.1 mi. +
+ All! "'~" ~.T9
~
o.S mt
::> 0.5l1li + IITS
20
0
0 100 200 300 ~oo
100 150 200 Incubalion time (hulII")
IncuhHbon lim,' (hours)
Figure 1. Effect of ammonium thiosulfate (ATS) and liquid fertilizer droplet size on urea hydrolysis in two
soils.
A - Silty clay at field capacity. B - Silty clay at wilting point. C - Loamy sand at field capacity. D -- Loamy
sand at wilting point. IAdapted from Goos and Fairlie (1988), by permission of the Soil Science Society of
America, Inc.!
ATS, APP, and ATS+APP on NHJ volatilization was most evident with dribble application.
For example, after 7 days, the NH3 loss from the bare microplots treated with VAN + 2%
ATS - as compared with micToplots treated only with VAN - was reduced by 66% with
dribble application and by 40% with spray application. There was no additional advantage
to giving more than 2% ATS to UAN. Thus, when 5% ATS + 20% APP were added to
VAN, the NHJ loss after 7 days was reduced by 76% with dribble application and by 41 %
with spray application. In the case of microplots covered with wheat residue, ATS and APP
41
decreased NH3 volatilization for 14 days (dribble application) and, in general, only for the
first 4 days (spray application).
Studying three Minnesota soils (loamy sand, silt loam, and clay loam), Zadak et af.
(1987) applied a urea solution with and without ATS and, then, a lO-mm water layer on the
surface of soil columns. ATS was slightly effective in delaying urea hydrolysis in the loamy
sand, but had no effect on N loss from any of the soils.
Al-Kanani et af. (1990a) used surface (0-5 cm) samples from six eastern Canadian soils
cultivated with maize. For studying the effect of ATS on ammonia volatilization from VAN,
two levels of a 60% ATS solution (namely 1.8 and 3.6% by weight) were added to VAN
solutions. The soil samples moistened to 100% of field capacity were amended with
surface-applied VAN solutions at a rate equivalent to 147 kg N/ha, then incubated at 24°C
for lO days. The volatilized NH3 was measured daily. After incubation, the soil was
analyzed for determination of the urea-, NH/-, N0 3·-, and N02"-N contents. The results
indicated that ATS reduced the NH3 losses, the reduction ranged from 12% (a clay soil) to
23.5% (a sand soil).
Sullivan and Havlin (1992a) found that ATS significantly (p = 0.01) inhibited urea
hydrolysis in VAN-amended samples of each of the studied eight Kansas soils varying
widely in pH, cation-exchange capacity, carbonate and organic C contents, urease activity,
WHC, and clay content. The VAN + 10% ATS solution contained 1.1 M ATS-S/I. Efficacy
of inhibition was greatest for soils low in clay and organic C at high temperatures and low
soil moisture contents. Thus, the inhibition ranged from 18 to 48% in an incubation at 20°C
and -0.1 MPa soil matric potential. Inhibition of urea hydrolysis averaged 29% at 20°C and
37% at 30°C, and 28% at -0.03 MPa and 38% at -0.1 MPa. The cumulative NH3 loss was
also significantly reduced by ATS in seven of the eight soils studied (the reduction on
average was 48% in four noncalcareous soils and 22% in three calcareous soils).
Sullivan and Havlin (1992a,b,c) also demonstrated that sodium tetrathionate
(Na2S406.2H20) applied with VAN inhibited urea hydrolysis in soils to the same extent as
ATS. Since the tetrathionate inhibits, while ATS does not inhibit jackbean urease and as
ATS is oxidized by soil Fe and Mn to tetrathionate, the conclusion was drawn that soil
urease is inhibited not by ATS itself, but by tetrathionate; this is the primary urease inhibitor
which reacts with sulfllydryl (SH) groups of the enzyme. This mechanism of inhibition
differs partly from that suggested by Goos et af. (1986a) and Goos (1987) (see page 37).
In a laboratory experiment carried out in the Plant Production Institute in Prague, Czech
Republic, List'anska (1993) fertilized samples of a grey-brown podzolic soil with either
DAM 390 (a VAN solution) or N-Sol (a urea-formaldehyde solution) at a rate of 300 mg
N/kg soil, then the samples were treated with sodium thiosulfate, STS (4% by weight of
fertilizer N) and incubated at ambient temperature. During the incubation period (June 9,
1988 - May 22, 1989), the samples were analyzed monthly for NH/ and N0 3". The results
indicated that STS was a weak inhibitor of urea hydrolysis and its inhibitory effect on
nitrification was even negligible, especially in the samples fertilized with N-Sol.
Lim and Seo (1994) studied the inhibitory effects of ATS and STS on urea hydrolysis in
a Korean paddy soil. This effect of ATS was slightly lower than that of STS in glucose-
42
amended soil samples, but when glucose was not added, the effects of ATS and STS were
not significantly different.
1.8. OTHER INORGANIC COMPOUNDS
Xue and Li (1987) prepared reaction mixtures from 5 g of paddy soil + 10 rn1 of 10% urea
solution containing 0 or 20-]()0 ppm (soil basis) of potassium ferricyanide (K3 [Fe(CN)6]) or
100 ppm of potassium permanganate (KMn04 ). After incubation (at 37°C for 48 hours), the
reaction mixtures were analyzed for NH4 + produced by urea hydrolysis. The analytical data
indicated that these compounds inhibited soil urease activity. The degree of inhibition at the
100 ppm rate was 37.52 and 24.75%, respectively.
* *
*
The following inorganic compounds will be described together with their organic
derivatives:
phosphorodiamidic acid and thiophosphorodiamidic acid in Subchapter 2.20;
phosphoryl triamide and thiophosphoryl triamide and their thermal polymers in
Subchapter 2.26; and
phosphonitrilic hexamide in Subchapter 2.27.
43
Chapter 2. Organic Compounds Tested for Evaluation of Their Inhibiting Effect on

Soil Urease Activity, Urea Hydrolysis, Ammonia Volatilization, and
Nitrous Oxide Emission
2.1. ORGANIC MERCURY COMPOUNDS
Structural formulas of the organic compounds tested as inhibitors of soil urease activity
are shown in Figure 2.
CI-H9-D-COOH Q-Hg-OOC-CH3
p-Chloromercuribenzoic acid Phenylmercuric acetate

(PCMB) (PMA)
HO-H9-Q-COOH Q-Hg- B02

p-Hydroxymercuribenzoic acid Phenylmercuric borate
(PHMB) (PMB)
Figure 2. Organic mercury cOll1'ounds tested as inhibitors of soil urease activity.
It should be emphasized from the very beginning that these inhibitors present only a
theoretical importance; they can not be recommended as soil urease inhibitors for field
applications because of the possibility of subsequent pollution of soil and groundwater
by mercury.
Mitsui et al. (1960) prepared reaction mixtures from 50 ml of M urea solution and 5
g of air-dried soil (control variant). In the experimental variant, sodium p-
chloromercuribenzoate (PCMB; 0.04% on dry soil weight basis) was added to the
mixture of urea solution + soil. The soil used derived from volcanic ash and was
cultivated with paddy rice. The reaction mixtures, after 1 hour of shaking, were
analyzed for NH/. The analytical results indicated that PCMB brought about a strong
(60%) inhibition of soil urease activity.
Briggs and Segal (1963) mention that activity of the crystalline urease preparation
obtained from a forest soil was markedly inhibited by PCMB (no details on the
inhibition experiment are given in their paper).
In a laboratory experiment, Moe (1967) applied PCMB, at a rate equivalent to 258
kglha, to the surface of a silt loam soil from Indiana. Due to PCMB the urea hydrolysis
rate was approximately halved, but PCMB had no significant effect on total volatile
ammonia losses from soil samples surface-treated with urea (336 kg N/ha) and
incubated at 28°C for 8 weeks, during which time soil moisture content was maintained
at 24%. Values of the total volatile NH, losses from the added urea-N were 4.68%
(untreated soil) and 3.94% (PCMB-treated soil).
Using the 5-hour test, Bremner and Douglas (1971) added 50 ppm (soil weight
basis) of phenylmercuric acetate (PMA) or PCMB to samples of three Iowa soils (silty
clay loam, loam, and clay loam). PMA, in comparison with PCMB, inhibited more
44
strongly the urease activity of the three soils. The degree of inhibition was 64-71 % (on
average, 67%) and 32-38% (on average, 35%), respectively.
The effect of the PMA rate (50, 100, 200, and 300 ppm) on urease activity in the
clay loam soil was also studied and it was established that the inhibition values
increased from 64 to 81 % with increasing PMA rate from 50 to 300 ppm.
In another experiment, samples of the three soils were preincubated with 50 ppm of
PMA or PCMB at 30°C for 3, 7, and 14 days, after which the 5-hour test was applied.
The results showed that the inhibitory effect of both organic mercury compounds on soil
urease activity markedly decreased with increased preincubation time which indicates
partial inactivation ofthese inhibitors in soil.
Some of the results concerning inhibition of urease activity in the clay loam soil
under the influence ofPCMB were also presented by Douglas and Bremner (1971).
Reexan1ining PMA, Bremner and Douglas (1973) determined not only the urea
hydrolysis but also the volatilization of an1monia from urea in five Iowa soils. PCMB
was also reexamined in one of the soils. Reaction mixtures were prepared from 109 of
air-dried soil, 1 ml of a urea solution containing 10 mg N (1,000 ppm), and 1 m1 of
water or 1 ml of a solution containing 0.5 mg inhibitor (50 ppm). Soil moisture content
was adjusted to 50% of WHC. Incubation took place at 20°e. After 3, 7, and 14 days,
the residual urea, exchangeable NIL +, and volatile NH3 were measured. The result.;;
showed that PMA and PCMB retarded urea hydrolysis and reduced NH.l volatilization.
These effects of PMA were strong in the light-textured soils and weak or inexistent in
the heavier soils. For example, during 14 days, NH3 volatilization from a sandy soil was
61.1 % of the added urea-N in the control soil and 23.5% in the PMA-treated soil, but in
the case of a heavier soil (clay loam) PMA reduced NH3 volatilization from 12.8 to
12.7%, i.e.. in this soil PMA had practically no affect on NH3 volatilization.
May and Douglas (1978) have evaluated the effect of PMA on urease activity of an
Australian sandy soil. Urease activity was determined by the method of May and
Douglas (1976). The reaction mixtures were prepared from 3 g of soil + 0.5 ml of
toluene + 12 ml of 1/15 M phosphate buffer (pH 8.8) + 3 ml of a solution containing 3
mg of urea, and were incubated at 31'C for 4 hours. After incubation the ~ + was
extracted and determined. PMA was used at four rates: 10,25,50, and 100 ppm (on soil
weight basis). At the two lower rates, PMA caused 80 and 95% inhibitions respectively,
whereas at the two higher rates, the inhibition was complete.
Shih and Souza (1978) used p-hydroxymercuribenwate (PHMB) as a urease
inhibitor to demonstrate that the 14C02 which evolved from 14C_urea in samples of a
California sandy loam soil was a product of the urease-catalyzed hydrolysis of urea. A
0.1 ml aliquot of 1.9 or 19 mM PHMB in phosphate buffer (25 mM, pH 8.5) or buffer
only (control) was added to 0.2-g soil samples. After 30 minutes, the mixtures received
14C_urea (0.01 /lCi activity) in the same buffer. Incubation took place at 23°C. 14C02
evolved during 2, 4, and 6 hours of incubation was measured. At the higher PHMB rate,
the 14C02 evolved in 2, 4, and 6 hours represented 0.38,0.47, and 0.77%, respectively,
as compared to 14C02 evolution from samples treated with urea only. These results,
corroborated with the finding that there was no 14C02 evolution when 14C_urea had been
added to autoclaved soil samples, prove that release of 14C02 from 14C_urea should be
attributed to soil urease.
Sahrawat (1979) adopted the 5-hour test of Douglas and Brenmer (1971) to evaluate
thc inhibitory effect of PMA (50 ppm) on urease activity of an Indian sandy clay loam
45
alluvial soil (PH 7.6). The inhibition caused by P.MA was 75%. But after I-week
incubation PMA inhibited urease activity to a lesser extent (62%).
Perez Mateos and Gonzalez Carcedo (1982) assayed urease activity in samples of
three Spanish soils (under vegetations dominated by poplar, gramineous, and
leguminous plants, respectively). The samples to which urea was added with or without
50 ppm (soil basis) of PMA or phenylmercuric borate (PMB) were incubated for 4
hours. It was found that the two compounds inhibited urease activity in the same soil to
a similar extent, which means that the anions (acetate and borate, respectively) did not
influence the inhibitory capacity ofPMA and PMB. Inhibition was lowest (62.38% with
PMA and 63.04% with PMB) in the soil richest in clay (the soil under leguminous
plants). In the other two soils 67.82-79.47% inhibitions were recorded. With the poplar
soil, the effect of different rates of PMA and PMB application on urease activity was
also studied and it was established that both compounds at a rate of 75 ppm gave
maximum inhibition (about 80%). Increasing the rate to 150,225, and 300 ppm did not
cause higher inhibitions. The authors drew the conclusion that in this soil the inhibition
exerted by PMA and PMB was of reversible non-competitive type.
For studying the mobility of PMA in soil, Praveen-Kumar et al. (1987) applied the
method of soil thin-layer chromatography. Samples of three Indian soils (acidic,
alkaline, and saline) were ground, sieved «0.15 mm) and coated on glass plates to
obtain thin-layers of soiL 100 11m thick. PMA was dissolved in methanol and the
solution containing 200 I1g of PMA was applied as a single spot on the soil thin-layer
plate. After volatilization of methanol, the plate was run using water as the solvent and
the solvent front was allowed to move 12 cm which took nearly 40 minutes. After
drying the plate was sprayed with a chromogenic reagent to develop color. PMA,
insoluble in cold water, was completely immobile in the thin-layers of each soil studied.
One can deduce from this observation that PMA - even if it would not cause soil
pollution - would not be an effective inhibitor of soil urease activity in agricultural
practice, because urea is very mobile and, therefore, may undergo the action of urease
from different soil depths in which urease is not inhibited since the inhibitor (PMA)
remains at the site of its application.
Studying the kinetics of urea hydrolysis in an alluvial soil (light-textured) and a
leached chernozern (heavy-textured) by determination of Michaelis constant (KM),
maximal velocity (V mIX), and inhibition constant (Ka, Simihaian and Silberg (1999)
found that PCMB exhibited stronger inhibition in alluvial soil than in chernozern; the
inhibition constant was l.8 and 2.9 mM, respectively. The conclusion was drawn that
inhibition of urease activity was competitive in both soils.
2.2. ORGANO BORON ACID COMPOUNDS
Van Der Puy et al. (l984b) patented organo boron acid compounds as inhibitors of soil
urease activity (Assignee: Allied Corporation, Morristown, New Jersey) and selected 1()
such compounds for testing their urease-inhibiting effect in a New York soil (Cazenovia
sandy loam, pH 7.2). Two tests were applied. In both tests the reaction mixtures were
prepared from 20 g of air-dried soil, 0.8 mg of test compound in 5 rnl of water, 42.8 mg
of urea in 1 ml of water, and incubated at 25°C for 3 days. In test A the soil treated with
test compound and urea was incubated, then analyzed for the remaining urea. In test B
the soil treated with test compound was incubated, then received urea and was again
46
incubated, after which the remaining urea was measured. No test compound was added
to the control reaction mixtures.
Four compounds (Figure 3) proved to be able to inhibit urease activity in both tests:
compounds I and II brought about excellent (strong) inhibitions in both tests, whereas
the inhibiting effect of compounds III and IV was partial in test A and excellent in test
B.
~V
B(OH)2 0
II
-6 B(OH)2
1'<:
: :,. . I NH2 .HO -S-OH . H2N A
II
o
3-Aminobenzeneboronic acid hemisulfate (I) 4-Hydroxyaminobenzeneboronic acid (II)
9~'
NH2 . Hel
4-Aminobenzeneboronic acid hydrochloride (Ill) 3-Hydroxybenzeneboronic acid (IV)
Figure 3. Structure of four of the organo boron acid compounds patented and tested by Van Der Puy
et al. (19S4b) for inhibition of soil urease adivity.
The most preferred rate of addition of these inhibitors to urea-based fertilizer

compositions is from about 5 to about 100 ppm (on weight basis).
2.3. FORMALDEHYDE
Urea pellets prepared from molten crystalline urea, 1%, (by weight) of formaldehyde
(CH 20) and 14% of blend of asphalt (90%)-rnicrocrystalline wax (10%) were used at a
practical rate and applied on the surface to samples of a sandy loam soil (pH 7.0;
moisture = 5.5%), then incubated at 25°C for 50 days. The control pellets did not
contain formaldehyde. During incubation, the ammonia volatilized was determined
cumulatively. In 50 days, the CH 2 0-containing urea pellets lost as NH3 8%, whereas the
control pellets lost about 23% of their initial N content. Formaldehyde is recommended
in amounts of (j.O I-I O'Yr, by weight of urea; the preferred rate ranges from 0.8 to 2.0'%.
For the asphalt-microcrystalline wax blend the preferred amount ranges from 3 to 20%
based on the total weight offertilizer pellets (Esso, 1966; Sor, 1968).
Verstraeten el af. (1976) performed two experiments. In the first experiment, 50-g
air-dry soil samples (pH 7.6) were treated with 100 ppm ofurea-N and 70-3,300 ppm of
CH 20, brought to 65'% ofWHC and then incubated under aerobic conditions at 30°C. In
the second experiment, 200 ppm of urea-N and 35-3,300 ppm of CH 2 0 were added to
samples of another soil (pH 5.0). These mixtures were incubated under water-logged
47
conditions. The control mixtures were prepared without CHzO. After 4(5), 7, 14, and 28
days of incubation, the mixtures were analyzed for NH4 +. It was found that under
aerobic and water-logged conditions, CHzO, even at the minimum concentration,
retarded hydrolysis of urea. This effect became more pronounced with increasing
concentrations of CH2 0, and the maximum concentration of CH 20 resulted in complete
st erilization of soil.
The effect of paraformaldehyde on urea hydrolysis was studied with an alluvial soil
and a leached chemozem (Kiss and Pintea, 1987). The reaction mixtures prepared from
5 g of air-dried soil and 10 ml of aqueous phase containing 6 mg of urea and 2% of
paraformaldehyde (on urea weight basis) were incubated at laboratory temperature.
Reaction mixtures without paraformaldehyde served for comparison. At 1-2-day
intervals the aqueous phase of reaction mixtures was analyzed for urea by means of a
chromogenic reagent (see the footnote on page 35). The results showed that
paraformaldehyde did not prolong the time necessary for complete hydrolysis of urea
since urea was completely hydrolyzed in 6 days (alluvial soil) and 9 days (leached
chemozem), irrespective of the absence or presence of paraformaldehyde.
2.4. HEXAMETHYLENETETRAMINE
Hexamethylenetetramine (HMTA; Figure 4) as an inhibitor of soil urease activity was

patented by Neumann and Richter (1976). They found that soil samples lost less
ammonia from urea pellets containing HMTA than from control pellets. In addition,
reduction of NH3 loss was more pronounced when HMTA was used in combination
with other inhibitors. This is why details on these investigations will be presented in
Chapter 3 (page 180).
N
I
1/
He CH2 CH2
~N
2
Figure 4. Structure of hexamethylenetetramine.
2.5. UREA DERIVATIVES
The urea derivatives tested as inhibitors of soil urease activity are specified in Figure 5.
Sor et al. (1966) patented for inhibiting soil urease activity the following urea
derivatives: methylurea, thiourea, N,N-dimethylurea, N,N' -dimethylurea, phenylurea,
t-butylurea, and n-butylurea to be applied in a proportion of 0.01 to 10%, preferably of
0.1 to 3% by weight of urea. The intimate mixtures of urea and inhibitor particles were
48
bound and coated with a hydrophobic material used in an amount ranging from 3 to
25%, preferably from 8 to 15% by weight of urea. For example, urea pellets were
prepared with addition of 0.45% methylurea or 2.44% thiourea + 13% asphalt-
microcrystalline wax blend. Incubation of these pellets, at a practical rate, on the surface
of a sandy loam soil (PH 7.5; moisture content = 6.4%) at 25 QC for 47 days led to
reduced volatilization of urea-N as ammonia. When a significant part (39.5%) of N
volatilized as NH3 from the urea priUs (no inhibitor), the NH3 losses from the
methylurea- or thiourea-containing urea pellets were 16.0 and 25.5%, respectively.
o
o s H3C, II
r-r-c-NH2
II II
H~-HN-C-N~ H;1N-C-N~ H3C""
Melhylurea Thiourea N,N·Dime1hylurea
o o
H3C- HN- c - NH--C~
II
N,N' ·Dimelhylurea
0-
~ A 11
HN-C-N H2
Phenylurea
o
H;>C, CH-CH~ HN-~- NH2
H~"" t-Bu1ylurea n-Bulyturea
NH 0 o
II II II
H;1N-C-HN-C-NH2 H~-C-NH--OH
Guanylurea Hydroxyurea
o o 0
II II II
H;>C- HN- C - NH-QH H~-C-NH--C-NH2
N·Melhyl·N'·hydroxyurea Biurel
Figure 5. Urea derivatives tested as inhibitors of soil urease activity.
In an experiment described by Sor et al. (1971), volatilization of NH3 from pellets

consisting of urea + 4% thiourea + 1% Marcol 72 was, in 3 days of incubation, only
1.90%, whereas 2.65-3.08% of the urea-N volatilized as NH3 from the control pellets
containing no thiourea.
49
Applying the 5-hour test, Bremner and Douglas (1971) recorded weak inhibitions in
urease activity of the three Iowa soils studied, the samples of which were treated with
urea derivatives at a rate of 50 ppm of soil. The average inhibition values were 7%
(thiourea), 4% (phenylurea), 5% (guanylurea sulfate), and 2% (hydroxyurea, biuret).
Kolyada (1970, 1973) described fertilization experiments in vegetation pots with a
soddy-podzolic soil (pH 4.4). The fertilizers (thiourea, ammonium nitrate, urea;
superphosphate; KC1) were applied at a rate of 0.1 g N, P, and K/kg dry soil. The
fertilizer treatments were the following: unfertilized control; PIC; thiourea + PK;
~N03 + PIC; 0.1 parts of thiourea-N + 0.9 parts of NH4N0 3 -N + PK; 0.1 parts of
urea-N + 0.9 parts ofNH4 N0 3-N + PK; exceptionally, only 0.1 parts ofthiourea-N (i.e..
not 0.1, but 0.01 g ofthiourea-N/kg soil) + PK. Moisture content of soil was 60% of
WHC. Oats, barley, onion, and radish were used as test plants. Urease activity was
determined at day 5 after fertilizer application and during the growing season.
The results indicated that thiourea decreased this activity only in the treatment in
which the rate of thiourea-N was 0.1 glkg soil. In this treatment with oats, at day 5 after
fertilization, urease activity was 0.45 mg NH3/g soil/24 hours, whereas in the other
treatments the corresponding activity values were between 1.00 and 1.30. During the
growing season the depressive effect of thiourea on soil urease activity has attenuated.
The effect of biuret on urease activity was the objective of a study by El-Sayed et of.
(1976). Biuret was found not to inhibit activity of pure jackbean urease, but did inhibit
the urease extracted from soil. The extract was obtained from a Giza (Egypt) soil by
shaking the soil suspension (soil:distiIIed water = 1:10) for 24 hours, followed by
filtering through Whatman No.42 paper. In the reaction mixtures containing soil extract
the concentration of urea wa<; 10 mM and that of the biuret ranged from 0.1 to 40 mM.
At concentrations up to 2 mM, biuret manifested a negligible inhibitory effect; the
inhibition increased with increasing biuret concentration and became complete at 40
mM biuret concentration. The inhibition caused by biuret in the urease activity of soil
extract was of the competitive type.
It should be mentioned that biuret can form during melting of urea for production of
priUs: at the melting temperature of urea (132.7!'C), some urea molecules are split into
NH3 and HN=C=O (isocyanic acid), which reacts with urea to form biuret:
Therefore, biuret can be present as an impurity in the urea fertilizer. If this fertilizer
contains more than 2% biuret in the case of soil application or more than 1% in the case
of foliar application, it will exhibit phytotoxicity just because of the biuret (Michaud et
af., 1978).
Gould et al. (1978) established that urease activity in samples of a silt loam soil
from Alberta was weakly inhibited by thiourea, N-methyl-N'-hydroxyurea, and
hydroxyurea, the degree of inhibition being 9, 15, and 16%, respectively. Urease
activity was measured according to the technique described by Gould et of. (1973). The
reaction mixtures consisted of 25 g of soil, 400 ppm of urea-N, 0 or 100 ppm of urea
derivative, 32% moisture (field capacity), and were incubated at 25!!C for 24 hours.
Sahrawat (1979) registered a 10% inhibition in urease activity, when thiourea was
added, at a rate of 50 ppm, to samples of a sandy clay loam soil (PH 7.6). Urease
activity was assayed under the conditions of Douglas and Bremner's (1971) 5-hour test.
so
Malhi and Nyborg (1979) conducted pot and field experiments in Alberta. In the pot
experiments, they studied the effect of thiourea on urea hydrolysis as influenced by soil
moisture levels, size, and method of application of urea and urea + thiourea pellets. Two
soils (loam, pH 7.3 and silty clay loam, pH 6.0) were used, each in an amount of 1
kg/pot. Urea and thiourea were pelleted together in a ratio of 2 parts urea and 1 part
thiourea. The size of the urea and urea + thiourea pellets was OJn or 0.21 g. The pellets
were spread uniformly over the soil surface or mixed into the soil at a rate of 224 kg
urea-N/ha (N in thiourea was not taken into consideration). The control soil was not
fertilized. h1cubation was at 20"C for 20-160 hours and urea hydrolysis was estimated
by measuring the residual urea. The results showed that at the same moisture level, size
and method of application of pellets, urea hydrolysis was significantly slower when the
soil was treated with urea + thiourea than when only urea pellets were applied. Thiourea
suppressed urea hydrolysis by about SO% for approximately 1 week. It was also
established that urea hydrolysis increased with increasing soil moisture levels and
decreased slightly with the increased size of pellets; mixing of urea into soil enhanced
the hydrolysis of urea relative to surface application.
In the field experiments on six soils (one silty clay loam, two clay loams, two loams,
and sandy loam) the effect of thiourea on urea hydrolysis was studied in relation to the
method of application of urea and urea-thiourea pellets (in these experiments, too, two
parts of urea were copelletcd with one part of thiourea). The peliets were placed in
bands (at S cm depth) or mixed into the soil 10 cm deep. The rate of urea application
was S6 kg N/ha. At day 14 after fertilization, the NH/ and NO}- contents in tl1e 0-IS-
cm soil layer were analyzed. The analytical data obtained confirmed tl1e results
registered in the pot experiments: in each soil, thiourea retarded the hydrolysis of urea
(degree of inhibition "" SO%), band placement of pellets-as compared with their mixing
into the soil- diminished the rate of urea hydrolysis.
In another field experiment, on the silty clay loam, urea and urea + thiourea (2:1)
pellets of different size (0.01, 0.21, and 2.26 or 2.S1 g) were tested. The 0.01- and 0.21-
g pellets were banded S cm deep. The larger pellets were applied in a grid pattern at
distance of 30 cm and a depth of Scm. Nitrogen was administered at a rate of 112
kg/ha, taking into account the thiourea-N, too. Urea hydrolysis was studied by
determining the NH4 + and NO} - contents in the 0-IS-cm soil layer. The results obtained
R days after fertilization are presented in Table 6 from which it is evident that thiourea
significantly reduced (by approximately SO%) the rate of urea hydrolysis. Urea
hydrol ysis also decreased with increasing pellet size.
TABLE 6. Effect of thiourea and pellet size on urea hydrolysis in a silty clay loam soil"
Fertilizer Pellet size (g) Urea hydrolysis (%)
Urea (UII 98 a
Urea + thiourea (2:1) (Ull 49 d
Urea 0.21 84 b
Urea + thiourea (2:1) 0.21 36 e
U~ 2~ ~c
Urea + thiourea (2:1) 2.51 25 f
aFrom Malhi and Nyborg (1979), by permission of Kluwer Academic Publishers.
The values are significantly different (p=0.05) when not followed by the same letter.
51
Moawad et al. (1984) treated 20-g samples of an alluvial soil (silty clay, pH 7.4)
from the Nile Delta with 400 ppm N in form of urea or thiourea, then incubated them
under favorable humidity conditions at 30"C. Untreated samples served for comparison.
Urease activity assayed after 1, 3, 7,14,21, and 28 days of incubation was always lower
in the thiourea-treated samples than in the urea-treated and untreated ones. Similarly,
ammonia volatilization from thiourea was more reduced than from urea. For example,
after 28 days of incubation, the volatile NH3 loss was 5.5% from thiourea and 1l.8%
from urea.
Studying the alluvial alkaline soils from Pakistan, Hamid and Ahmad (1987) also
dealt with the effect of thiourea on volatilization of ammonia from urea. Samples of a
calcareous soil (pH S.O) were treated with urea or with urea + 5% thiourea (on urea
weight basis) and then incubated. The NH3 volatilized during incubation was
determined. It was found that in 112 days the cumulative NH3 losses were reduced by
50% under the influence of thiourea.
The German investigators Hartbrich et al. (1978) tested the effect of thiourea on soil
urease activity in comparison with that of various derivatives of thiourea. Two testing
methods were applied.
In the short time test, a black soil with strong adsorptive capacity was used. The soil
samples (30 g) were moistened to 50% ofWHC and then mixed with 214.1 mg of urea
(I 00 mg N) with or without 4 mg of compound to be tested (4% on urea-N basis). The
mixtures were incubated at 30"C for 24 hours, during which the volatilized ammonia
was determined.
In the long time test (see also Iasche et al., 1978), a sandy soil (i.e .. a soil with weak
adsorptive capacity) was used. The 30-g soil samples were moistened to 50% of WHC
and ill1 their surface urea (214.1 mg) with or without 1 mg of test compound was
uniformly distributed. Incubation took place at 20"C. The amount of volatile NH3 was
recorded at 2-3-day intervals.
TABLE 7. Effect of thiourea and its derivatives on volatilization of ammonia from urea-treated soils"
Inhibition (%)
STT LTT
Compound
Incubation time (days)
2 4 6 8 10
Thiourea 13 21 9 0
Bis thiourea 69 85 49 35 12 0
N-Phenylthiourea 29 31 9 ()
N-Phenyl-N' -isopropylthiourea 53 10 2 ()
N-Phenyl-N'-t-butylthiourea 53 18 6 0
N-Phenyl-N'-ethyl-N'-cyclohexylthiourea 9 16 6 ()
N-Phenyl-N'-ethyl-N'-benzylthiourea 7 14 4 ()
N-Phenyl-N',N'-dibenzylthiourea 4 4 ()
Allylthiourea 19 25 IJ ()
N-2-Aminophenyl-N'-allylthiourea 26 31 19 8 ()
"From Hartbrich et al. (1978).
STT - Short time test.
LTT - Long time test.
S2
The data in Table 7 show that the degree of inhibition varied depending on the
nature of thiourea derivatives and on the incubation time. Thus, the inhibiting effect of
bisthiourea was the strongest and evident for 8 days; N-phenyl-N' ,N' -dibenzylthiourea
had a negligible inhibitory effect, detectable only during the first 2 days of incubation.
In these tests, even the thiourea was a weak inhibitor.
Using samples of brown earths from Germany, Germann-Bauer (1987) carried out
several laboratory experiments to study urea hydrolysis and ammonia volatilization
from urea under the influence of guanylthiourea (GTU):
NH S
II II
H2N-C-HN-C-N~ .
For studying urea hydrolysis, reaction mixtures were prepared from SO-g (dry
weight) samples of a loess brown earth + 20 mg of urea-N + 0, 2 or 3 mg of GTU and
submitted to incubation at 4, 8, 12 or 16QC for 1 or 2 weeks, then analyzed for residual
urea. The results indicated that GTU retarded urea hydrolysis, and this effect increased
with increasing rate of GTU and decreased with increasing incubation temperature and
time.
Ammonia volatili7ation from urea was studied with SOO-g samples of a sandy brown
earth. Two experiments were carried out. In the first experiment the reaction mixtures
were prepared from soil samples + 100 mg of urea-N + 0 or 10 mg of GTU and
incubated for 6.6 days. In the second experiment the reaction mixtures, prepared from
soil samples + SO mg ofurea-N + 0 or S mg of GTU, were incubated for 13 days. The
cumulative NH3 loss, assessed during the incubation (6.6 or 13 days, respectively), was
reduced significantly (p<O.OS) (from 17.7 to 13.3%) in the first experiment, but
insignificantly (from 31.8 to 27.3%) in the second experiment.
2.6. DITHIOCARBAMATES
Hyson (1963), assignor to E.!. du Pont de Nemours and Company (Wilmington,

Delaware), nominalized, in his patent, twelve dialkyldithiocarbamates and two
alkyldithiocarbamates (Figure 6) tested for inhibiting soil urease activity. The
dithiocarbarnates are recommended to be used at rates of 3-8,000, preferably S-SOO ppm
by weight of urea. For preparing the fertilizer compositions three procedures are
described, each being exemplified with cupric diethyldithiocarbarnate as an inhibitor.
a) Cupric diethyldithiocarbamate (0.2S g) is ground in a mortar with 10 g of clay
until a homogeneous mixture is obtained. This mixture is distributed on the
surface of SOO g of urea prills by tumbling in a mechanical mixture.
b) Urea powder (98% by weight) is admixed by thorough stirring with 0.2% of
cupric diethyldithiocarbarnate, 2% of Zn stearate, and a trace of green dye
(Pontacyl Green).
c) The cupric diethyldithiocarbamate-clay mixture, obtained as in procedure a, is
treated with 1.6 g of Mg stearate and a trace of Pontacyl Green dye and, after
stirring, is used for coating the urea priUs as in procedure a.
53
A ~Cu2+, Fe3+, Zn2+, Mg2+,

Ca 2+, Na+, Ni 2+, NH4+ or
NH 2 (CHa)2+ (dimethylammonium)
Dimethyldithiocarbamate
r3C-CH2)~~_sJ
In
A
~aC-CH2
Diethyld~hiocarbamate
t~-""-L-j" '
Methyldithiocarbamate
A =Na+ or Zn 2+
n = Valence of A
Figure 6. Dithiocarbamates patented and tested by Hyson (1963) for inhibilion of soil urease
activity.
These compositions were applied by broadcasting to areas of tobacco cropland and

coastal bermudagrass at rates of 28-560 kg of urea/ha (the preferred rates are 56-224 kg
ofurea/ha), Unfortunately, Hyson (1963) gave no further details on the experiments, He
only mentioned that urea-N loss as volatile ammonia was reduced significantly,
evidencing a high stability of these compositions against the action of urease.
The invention patented by Tomlinson (1967), of the Imperial Chemical Industries
(London), relates to ethylene-I.2-bisdithiocarbamates and to other dithiocarbamates
tested as inhibitors of soil urease activity. The use of dithiocarbamates in combination
with halogenated alkanes was also patented. Figure 7 presents the structural formula of
the dithiocarbamates nominalized in the patent. Nabam, zineb, and maneb are important
fungicides. The halogenated alkanes tested were ChBrC-CBrCIz (1,1,2,2-tetrachloro-
1,2-dibromoethane) and Br3C-CBr3 (hexabromoethane). Two testing methods were
applied.
In the first method nine soils were used; each dithiocarbamate was tested in 1-9
soils. The air-dry soil samples (2.5 g) placed in dishes were treated with a urea solution
containing 2,000 ppm N (on soil weight basis), a solution or emulsion containing 10-50
ppm of dithiocarbamate (also ot?- soil weight basis) and sufficient water to bring the soil
to about field capacity. No dithiocarbamate was added to the control samples.
Incubation was carried out at 9°C for 3, 5, and 7 days. During incubation, the volatilized
ammonia was assayed. Taking the amount of NH3 lost from the control samples as
100%, the percentage of NH3 losses from the dithiocarbamate-treated samples was
calculated. Some of the results will be quoted below, Nabam, applied at a rate of 50
ppm in a single soil, manifested a strong inhibitory effect as the NH3 loss in 5 days was
only 1%. The effect of zineb used at rates of 10, 20, 40, and 50 ppm varied in the nine
soils studied: the NH3 losses recorded in 7 days ranged from 5 to 110%. More precisely,
54
S
II
CH2-HN-C-S-Na
I
CH2 -HN - C - S -Na
II
S
Disodium elhylene-l ,2-bisdfthiocarbamale S-Trichloromelhyhhiodfthiocarbamale

(nabam)
S
II
HN-C-S-CH3
I
HN-CO-CH3
Zinc elhylene-l.2-bisdfthiocarbamale S-Melhyl-N-acelamidodfthiocarbamale

(zineb)
-0-
S
II
CI ~ /) HN-C-S-NH4
Manganese e1hylene-l.2-bisdfthiocarbamale Ammonium N- p-chlorophenyldfthioca.rbamale

(maneb)
Figure 7. Dilhiocarbamates patented and tested by Tomlinson (1967) for inhibition of soil urease activity.
zineb reduced NH3 volatilization in parallel with its rates in four soils, irrespective of its
rates in one soil, whereas in the other four soils zineb did not reduce NH3 volatilization
at any of its rates. Maneb and the other three dithiocarbamates, tested at rates of 20 or
40 ppm with two soils, manifested a weak inhibitory effect or no inhibition, the NH3
loss in 7 days varying between 63 and 99%.
In another experiment, zineb and, separately, ChBrC-CBrCh were added to samples
of a soil, at rates of 0, 5, 20, 50, and 200 ppm. Moreover, the two compounds at the
rates mentioned before were also applied in their all possible combinations. In each
case, the urea rate was 200 ppm N. Ammonia losses during 8 days of incubation showed
a tendency to decrease with increasing rates of the two compounds. The decrease was
more marked with ChBrC-CBrCh than with zineb. Ammonia losses decreased from
7.4% (control soil without addition of zineb and C1 2BrC-CBrCIz) to a minimum value
(0.5%) in the soils treated with 0,5 or 50 ppm ofzineb + 200 ppm ofCIzBrC-CBrCl z.
In the second method wheat seedlings are used for testing the inhibitors of soil
urease activity in urea-treated soil samples. This is why this method and the results
obtained will be dealt with in Subchapter 7.2 (on pages 264-265).
55
Pugh and Waid (1969a,b) treated 100-g moist samples of three English soils (sandy
loam, loamy sand, and clay loam) with 6.66 mM of urea and 0.4 mM of sodium
diethyldithiocarbamate, (CHr CH 2 hN-CS-S-Na. The control samples received urea
only. All samples of the three soils were incubated at 20°C for 30, 77, and 148 days,
respectively, during which time the volatilized ammonia was determined. The half-loss
times of NH3 loss from the control and the diethyldithiocarbamate-treated samples were
4 and 5.5 days (soil 1), 11.5 and 14 days (soil 2), and 34 and 59.5 days (soil 3),
respectively, i.e.. sodium diethyldithiocarbamate prolonged half-loss time of NH3 in
each soil. However, the total NH3 losses from the three soils during their whole
incubation period (30, 77, and 148 days, respectively) were similar in the control and
the diethyldithiocarbamate-treated samples (51.2 and 56.8%,39.8 and 45.0%, and 84.0
and 81.9%, respectively).
Under conditions of the 5-hoUf test, Bremner and Douglas (1971) recorded 1, 3 or
7% inhibitions of urease activity in samples of three soils treated with sodium
diethyldithiocarbamate or dimethyl ammonium dimethyldithiocarbamate, (CH3hN-CS-
S-NH 2(CH3h, at a rate of 50 ppm (on soil weight basis).
Lang et al.(l976) patented ferric dimethyldithiocarbamate, [(CH3hN-CS-ShFe (the
fungicide ferbam), as an inhibitor of soil urease activity. Urea with or without 1%
ferbam (relative to urea-N) was surface-applied on soil samples, which were then
moistened to 50% of WHC and incubated at 20°C for 19 days. During incubation, the
volatile ammonia was estimated. The degree of inhibition registered after 5, 10, and 19
days of incubation was 94.7, 81.3, and 24.0%, respectively. This means that the
inhibitory effect of ferbam on the volatilization of NH3 from urea was considerable for
10 days, then decreased very much. Ferbam as an inhibitor of soil urease activity is
recommended for practice at preferred rates of 0.05-5% (relative to urea-N).
Hartbrich et al. (1978), applying the short and long time tests (see page 51),
established that all dithiocarbamates tested, which also comprised important fungicides,
inhibited ammonia volatilization from urea, but the duration of their effect did not
exceed 8 days (Table 8). The results obtained with maneb were also referred to by
Oertel et al. (1978).
TABLE 8. Effect of dithiocarbamates on volatilization of ammonia from urea-treated soils"
Inhibition (%)
STT LTT
Compound
2 4 8 12
Iron dimethyldithiocarbamate (ferbam) 79 100 42 12 0
Zinc dimethyldithiocarbamate (ziram) 82 74 33 14 0
Aluminium dimethyldithiocarbamate 63 82 20 3 0
Sodium di-n-propyldithiocarbamate 72 71 13 0
Sodiumn-butyldithiocarbamate 86 100 72 11 0
Zinc ethylene-I.2-bisdithiocarbamate (zineb) 67 79 17 5 0
Manganese ethylene-I,2-bisdithiocarbamate (maneb) 81 100 51 19 0
"From Ilartbrich et al. (1978).
STT - Short lime test.
L TT - Long time test.
56
In 400-g air-dried samples of an Egyptian clay loam soil treated with urea (70 ppm)
and moistened to 60% ofWHC, zineb at a rate of 3 kglha reduced only to a small extent
urea hydrolysis during the first 3 days of incubation at 25-28 D C (the unhydrolyzed urea
being 3% of the initial amount) and even this weak inhibitory effect disappeared after 7
days of incubation (Moawad et al., 1979a). In contrast, zineb decreased urease activity
in the urea-treated soil samples during the whole (35-day) incubation period
(Moawad et al., 1979b).
Tu (1980) treated samples of a Canadian sandy loam soil (pH 7.6) with maneb at
rates of 0, 100, and 200 ~glg soil and determined their urease activity after 2, 7, and 14
days of incubation. The activity was inhibited significantly (p=0.05) by maneb during
the whole incubation period. As expected, the inhibiting effect was stronger at the
higher maneb rate.
Maneb, added to samples of a clay loam soil (PH 7.2) at two rates (5 and 10 ~g/g
soil) insignificantly inhibited urease activity after 2 days of incubation and did not
influence it at all after 7 days. There was no difference between the effects of the two
maneb rates (Tu, 1981a). In contrast, in an organic soil (pH 7.2), maneb at both rates
significantly inhibited urease activity after 7 days of incubation and significantly
stimulated it after 14 days. The inhibition was stronger and the stimulation was weaker
at the higher rather than the lower rate ofmaneb (Tu, 1981b).
Hartbrich et al. (1978) also published data on testing of four carbamates and drew
the conclusion that these compounds did not show any urease-inhibiting effect.
2.7.THIVRAM DISULFIDES AND SULFIDES
The first data on the inhibitory effect oftetramethylthiuram disulfide, (CH3hN-CS-S-S-

CS-N(CH3h (the fungicide thiram), on soil urease activity were published by
Borchaninova and Filippova (1972). They carried out field experiments on a soddy-
podzolic soil of heavy loam texture (PH 5.6) in Russia. The seeds of the test plant (red
clover) were treated with thiram at a rate of 4 glkg seeds. After 5-6 days the treated
seeds and the untreated (control) seeds were sown (20 kg seeds/ha). At different stages
of plant growth during 2 years, soil was sampled from the rhizosphere for determination
of urease activity. It was found that at each growth stage urease activity was lower in
the rhizosphere of plants developed from thiram-treated seeds than in the rhiwsphere of
control plants. Thiram inhibited soil urease activity in pot experiments, too, in which
Barchaninova and Filippova (1978) used the same soil and applied thiram for treatment
of red clover seeds in the same amount as in the field experiments.
Markert (1974) also found, in laboratory fertilizer trials, that thiram, applied in
amounts equivalent to those used under field conditions in agricultural practice, reduced
the rate of urea hydrolysis in the two German soils studied (silty loam, pH 7.1 and
loamy sand, pH 4.05).
Lang ef af. (1976) and Thieme ef al. (1976) patented tetramethylthiuram disulfide as
an inhibitor of soil urease activity.
Lang ef al (1976) treated soil samples with urea + 0 or 1% thiram (relative to urea-
N), then moistened and incubated them at 20 D C for 18 days, during which the ammonia
volatilized from urea was assayed. The inhibitory effect of thiram decreased depending
57
on incubation time. For example, the inhibition was 97.5% after 4 days, 71.6% after 12
days, and 35.0% after 18 days of incubation. The preferred rates, recommended for
practice, range from 0.05 to 5% thiram relative to urea-No
Thieme et al. (1976) applied the same technique with the same incubation time, but
worked with three thiram concentrations (0.1, 0.5, and 1% relative to urea-N) and
carried out the incubation at 10o e. Determination of the volatilized ammonia indicated
that after the same incubation time inhibition of NH3 volatilization increased with
increasing thiram concentrations, but at each concentration it decreased during
incubation. For example, the inhibitions recorded in samples treated with 0.1, 0.5, and
1% thiram were 70.0, 91.2, and 94.2%, respectively, after 3 days of incubation, and 4.8,
23.4, and 42.0%, respectively, after 10 days. Thiram at 1% concentration after 18 days
of incubation caused only an 11.3% inhibition. Similar results were also presented by
Hartbrich et al. (1978), Oertel et al. (1978), and Jasche et al. (1978), with the difference
according to their data that thiram used at a rate of 1% inhibited NH3 volatilization only
for 12 days.
By applying the short and long time tests, Hartbrich et al. (1978) studied other
thiuram disulfides as well as some thiuram sulfides. Table 9 shows that the inhibitory
effect of tetramethylthiuram disulfide was weaker than that of tetramethylthiuram
sulfide. The other compounds were also weaker inhibitors and their effect was
detectable only during the first 2-8 days of incubation.
TABLE 9. Effect of thiuram disulfides and thiuram sulfides on volatilization of ammonia from urea-
treated soils·
Inhibition(%)
STT LTT
COlT(lound
2 4 8 12 14
Tetmmethylthiuram disulfide 64 83 38 22 12 o
Dimethylthiuram disulfide 28 21 3 0
Di-n-propylthiuram disulfide 40 57 31 11 0
Diisopropylthiuram disulfide 54 78 43 20 0
Diisobutylthiumm disulfide 74 65 36 15 0
Dibenzylthiuram disulfide 31 26 0
Dipiperidylthiumm disulfide 28 35 3 0
Tetmmethylthiuram sulfide 79 93 S6 33 21 4
Dipiperidylthiumm sulfide 33 39 12 3 0
·From Hartbrich et al. (1978).
In the case of some of these compounds, the method of application strongly

influenced manifestation of the inhibitory effect. Thus, when thiram mixed with urea is
uniformly distributed on the top soil layer, urease activity will be inhibited to a larger
extent than when the urea-thiram mixture is applied on the soil surface, which is due to
different solubility and mobility of urea and thiram (Hartbrich et aI., 1978; Muller and
Forster, 1980) .
58
According to the observations by Tu (l981a), thiram, with which samples of a clay

loam soil (pH 7.2) were treated at two rates (5 and 10 I!g/g soil), did not inhibit but
inversely stimulated urease activity after both 2 and 7 days of incubation. The
stimulation was significant at the lower thiram rate and insignificant at the higher rate.
In an organic soil (pH 7.2), thiram at both rates significantly inhibited urease activity
after 7 days and stimulated it after 14 days of incubation. Inhibition and stimulation,
respectively, were more pronounced at the higher thiram rate than the lower rate (Tu,
19 81 b). In other experiments, thiram treatment resulted in significant! y decreased urease
activity ofa sandy loam soil (pH 7.6) after both 7 and 14 days of incubation. The effect
of thiram on urease activity of an organic soil (pH 6.8) was significantly inhibitory after
7 days and stimulatory after 14 days of incubation (Tu, 1990).
In the pot experiments of Khabirov et af. (1992), samples of a leached chernozem
from Bashkiria (Russian Federation) were treated with urea (500 mg urea-Nlkg soil) and
o or 10% thiram (on urea-N basis), moistened to 60% of WHC and analyzed for
determination of urease activity after 1, 2, and 4 weeks of incubation at ambient
temperature. The activity (expressed in mg of NH) produced by 1 g of soil in 24 hours)
gave, after the three incubation times, the following values: 0.99, 1.09, and 0.76,
respectively, in samples treated with urea only, and 0.81,0.81, and 0.57, respectively, in
samples treated with both urea and thiram. In other words, thiram inhibited soil urease
activity.
2.8. XANTHATES
Brenmer and Douglas (1971) mention that, under the conditions of the 5-hour test,
potassium ethyl xanthate (KEtX) applied to three soils at a rate of 50 ppm of soil, gave
less than 4% inhibition of urease activity.
Ashworth et af. (1979) tested KEtX, cellulose xanthate (CX) (Figure 8), and a
mixture in which half of the KEtX amount was replaced by sodium trithiocarbonate
(Na 2CS). As already shown (see page 36), Na2CS] alone has a very weak inhibitory
s s
II II
CH3-CH2-0-c-S-K (cellulose chain) HC-O- C-S - Na
I
Potassium ethyl xanthate Cellulose xanthate
Figure 8. Xanthates tested by Ashworth et al. (1979) for inhibition of soil urease activity.
effect on urea hydrolysis. The tests were performed on a silty clay loam from England.
The reaction mixtures, consisting of 8 g of soil + 1 rnl of a solution containing 5 mg of
urea + 1 rnl of water or 1 ml of solution of test compound (s), were incubated at 24°C
for 5 weeks. Residual urea, NH/, and N0 3- contents were determined weekly.
Inhibition of urea hydrolysis increased in the following order:
KEtX < KEtX + Na2CS) < ex.
59
It should be emphasized that the effect of KEtX and Na2CS3 was synergistic. These
compounds inhibited nitrification more strongly than ureolysis·. Xanthates generate
CS 2, but more slowly than does Na 2 CS 3 , i.e .. xanthates are slowly acting inhibitors of
nitrification. At the same timc, their inhibitory effect on urea hydrolysis was not due to
CS 2 , because in separate tests CS 2 used in amounts stoichiometrically equivalent to
those of KEtX showed no urease inhibition.
Ashworth ef al. (1980) tested l7 xanthates as inhibitors of urease activity in a
Canadian clay loam soil. The reaction mixtures were prepared from 18 g of soil + 0.5
ml of urea solution (400 mg Nlkg soil) + 0.5 ml of water or xanthate solution (200 mg
compound/kg soil). For comparison, Na 2CS J andp-benzoquinone were used. Incubation
took place at 23°C. Residual urea was determined after 17, 24, and 41 hours of
incubation. Table 10 specifies only those xanthates which manifested an evident
inhibitory effect on soil urease activity; the percent inhibitions caused by xanthates,
Na2CS3, and p-benzoquinone are also presented.
TABLE 10. Inhibition of urease activity by xanthates, sodium trithiocarbonate, and

p-benzoquinone in a day loam soil"
Inhibition (%)
Compound Incubation time (hours)
17 24 41
K methyl xanthate 66 74 77
Na methoxymethyl xanthate 55 49 51
K allyl xanthate 51 48 57
K ethyl xanthate 55 29 31
K 2-methoxyethyl xanthate 27 49 33
K isopropoxyethyl xanthate 28 21 14
K ethylene glycol xanthate 25 18 10
K 2-dimethylaminomethyl xanthate 20 29 16
Na 2-nitrilo-2-propyl xanthate 21 19 10
Na trithiocarbonate 40 22 6
p-Benzoquinone 91 82 87
"From Ashworth et ai. (1980), by permission of the Soil Science Society of
America, Inc.
Based on the results obtained with the xanthates specified in the table and with other
xanthates, Ashworth et al. (1980) arrived at the following conclusions: the inhibiting
effect of the xanthates of unsubstituted primary alcohols decreased with increasing size
of the hydrocarbon chain; branched-chain xanthates were less effective than those with
straight-chain; xanthates of substituted alcohols were weaker inhibitors than their
unsubstituted counterparts, except for the xanthatc of 2-nitrilo-2-propanol which,
though more effective than K isopropyl xanthate, still gave weak inhibition; the
This is why Rodgers et ai. (1987) applied the mixture ofKEtX (at 5 kg/hal and Na2CS, (at a rate equivalent
to 10 kg CSz/ha) as a nitrification inhibitor.
60
presence of a carboxyl group diminished effectiveness of xanthates (xanthate of glycolic

acid was an ineffective inhibitor).
In pot experiments performed by Vlek et al. (1980) for studying urea hydrolysis in
flooded soils, urea prills (100 kg N/ha) with or without 2% KEtX (on urea weight basis)
were puddled into samples of a silt loam soil (from Alabama) covered by 5-cm deep
floodwater, and urease activity was estimated by daily analysis of the urea remaining in
floodwater during incubation (1-10 days at 35°C day and 25°C night with 12-hour
days). The amount of residual urea in the floodwater of soil treated with urea without
KEtX after 0, I, 2, 3, and 4 days of incubation was 140, 71, 29, 5, and 1 mg Nil,
respectively. The corresponding values for the urea + KEtX treatment were 123,66,24,
3, and 0 mg Nil, respectively. This means that KEtX did not have practically any effect
on urea hydrolysis.
Gautney el al. (1983, 1984) found that cogranulation of KEtX with urea is not
feasible because KEtX decomposes rapidly in urea melts.
2.9. HYDROXAMIC ACIDS
The inhibiting effect of monohydroxarnic acids on urease actIvIty in soil was the
obj ective of many studies, but this effect of dihydroxamic acids was dealt with, to our
knowledge, in a single work.
2.9.1. Monohydroxamic Acids

The first studies were devoted to the effect of acetohydroxamic acid on urease activity
in an acid loamy sand soil (pH 4.3-4.5) from England (Waid and Pugh, 1967), the effect
of seven monohydroxamic acids (Figure 9) on urease activity of the same acid loamy
sand (Pugh and Waid, 1969a), the effect ofacetohydroxamic acid on urease activity in
H;f--Co--NH-OH
G - c H2-CO-NH-OH
Acetohydroxamic acid
Phenylacetohydroxamic acid
H;f--GH2 - CO ---lIIH-OH
Propionohydroxamic acid Q-cO-NH-OH
Benzohydroxamic acid
H~(CH2l4 - CO--f\lH-OH
Caprohydroxamic acid
q-CO-NH-OH
H;f--(CH2ls - CO-NH-OH
Salicylohydroxamic acid
Caprylohydroxamic acid
Figure 9. Monohydroxamic acids tested by Pugh and Waid (196Qa,b) for inhibition of soil
urease activity.
61
16 soils (13 from England and three from Benin, Nigeria, and Brunei-North Borneo,
respectively), and the effect of the monohydroxamic acids on urease activity in two soils
(sandy loam and clay loam) from England (Pugh and Waid, 1969b; see also Waid,
1975).
Waid and Pugh (1967) treated moist samples (1 kg) of the acid loamy sand with
4,000 ppm of urea (weight/weight) + 0 or 200 ppm of acetohydroxamic acid (AHA),
then incubated them at constant temperature (25°C). During incubation, analyses were
carried out periodically for determination of residual urea and soil pH. AHA was
detectable for 8 days (only traces on the 8th day). In this period, urea hydrolysis was
slower and pH increase was smaller in samples treated with urea and AHA than in those
treated with urea alone. Consequently, complete hydrolysis of urea required about 14
days in AHA-treated samples and 8 days in samples without AHA addition.
In other experiments, Waid and Pugh (1967) followed volatilization of ammonia
from urea (4,000 ppm) in 100-g moist soil samples incubated at laboratory temperature
(fluctuating between 16 and 22°C), and found that this process was delayed by the
addition of AHA (200 ppm). Thus, volatilization ofNH 3 from samples without and with
AHA began after 6 and 13 days of incubation, respectively. Total urea-N losses as
volatile NH3 during 20 days of incubation were about 32 and 25%, respectively.
Pugh and Waid (1969a) also followed volatilization of NH3 from urea for studying
the effect of seven monohydroxamic acids (Figure 9) on urease activity in the acid
loamy sand. Moist 100-g soil samples, to which 4,000 ppm of urea (weight/weight) and
0,50,100,200 or 300 ppm of AHA or 200 ppm of the other six monohydroxamic acids
were added, were then incubated at laboratory temperature (14-23°C) and the amount of
volatilized NH3 was estimated during 29 days. AHA proved to be a more effective
inhibitor than the other monohydroxamic acids: half-loss time of urea-N as NH3 was
prolonged from 10 to 17.5 days in presence of 200 ppm of AHA and from 10 tol 0.5-13
days under the influence of the same concentration of the other monohydroxamic acids;
the weakest inhibitors were phenylacetohydroxamic and caprylohydroxamic acids.
Concerning total loss of urea-N in 29 days, there were no remarkable differences
between these compounds (35.3% in control soil without inhibitor, 30.2% in AHA-
treated soil and 31.3 -36.4% in soil samples treated with the other monohydroxamic
acids). By increasing the concentration of AHA from 50 to 300 ppm, half-loss time of
NH3 was prolonged from 12 to 23.5 days.
In other experiments, in which 0, 0.1, 0.2, and 0.3 mM of benzohydroxamic acid
(BHA) and 6.66 mM of urea were used per 100 g of soil and incubation was carried out
at either laboratory temperature (13-21°C) or constant (20°C) temperature, it was also
observed that half-loss time of NH3 was prolonged proportionately to the concentration
of inhibitor. Total NH3 loss decreased with increasing BHA concentrations at laboratory
temperature, but the reverse was the case at 20°e.
The relationships between the delay in NH3 loss and concentrations of urea and
AHA were also studied. The moist soil samples (100 g) were treated with 1.67, 3.33,
6.66, and 13.33 mM of urea and with 0, 0.2, 0.4, 0.6, 0.8, and 1.0 mM of AHA (1.67
mM of ureall 00 g soil is equivalent to 1,000 ppm of urea, weight/weight, and 0.2 mM
of AHA/I 00 g soil to ISO ppm of AHA, weight/weight). Incubation took place at 20°C
and lasted 29 days. One can deduce from the results presented in Figure 10 that half-loss
time of NH3 was shorter when urea was added at higher concentrations; AHA delayed
62
NH3 loss and the length of the delay was directly proportionate to its concentration at
anyone concentration of urea.
Half-loss time (dAyS)
Figure 10. Relationships between the delay in ammonia loss (expressed in terms ofhaIf-loss
time) and concentration of urea and AHA. IFrom Pugh and Waid (1969a). by permission of
Pergamon Press PLC./
Pugh and Waid (1969a) also carried out an experiment for studying the effect of
AHA on the volatilization of NH3 from either crystalline or prilled urea. The crystalline
urea was mixed evenly throughout the soil or placed on the surface of the soil or at
about 2-cm depth; the prilled urea was placed on the soil surface or at about 2-cm depth.
AHA was applied in or on soil like urea. The urea rate was 4,000 ppm and that of AHA
was 0 or 200 ppm. All samples were incubated at laboratory temperature (16-23°C) for
30 days. In all treatments AHA had a delaying effect on NH3 volatilization. In the
homogeneous mixture of soil + crystalline urea + AHA, the effect of AHA was weaker
than where this compound and urea were placed together on the soil surface or at depth.
Total NH3 losses from both crystalline and prilled urea were smaller with deep than
with surface placement.
It was also established that preincubation of soil samples with 200 ppm of AHA at
25°C before addition of 4,000 ppm of urea and incubation at laboratory temperature
(14-22°C) led to diminution of the inhibitory effect of AHA on volatilization of NH3
from urea. The diminution was evident even after 2 days of preincubation, and in
samples preincubated for 5 days, the inhibitory effect of AHA became negligible. This
behavior of AHA during preincubation may have been due either to its chemical
inactivation and microbial decomposition or to the proliferation of urease-forming
microorganisms tolerant towards AHA.
StUdying the effect of AHA in 16 soils, Pugh and Waid (1969b) treated 100-g moist
samples with 4,000 ppm (6.66 mM) of urea and 300 ppm (0.4 mM) of AHA,
determining, during the incubation at 20°C, the amount of volatile NH3 • The results
indicated an underlying influence of soil properties, especially of soil texture on the
urease-inhibiting effect of AHA. Thus, AHA reduced the maximum rate of NH3 loss
63
and delayed the half-loss time in 10 light-textured soils (3 sands, 4 loamy sands, and 3
sandy loams). In contrast, AHA did not exhibit such an effect in 6 soils with high clay
content. At the same time, AHA brought about even an increase in the rate of NH3 loss
from two of these six soils.
Of the seven monohydroxamic acids tested with two soils (sandy loam and clay
loam), the most effective compound in reducing the rate of NH3 loss was AHA in the
sandy loam and BHA in the clay loam, in which the other six monohydroxamic acids,
excepting salicylohydroxamic acid, accelerated the NH3 loss. The conclusion could be
drawn that monohydroxamic acids offer promise as a means of delaying the hydrolysis
of urea only in the light-textured soils.
By applying the 5-hour test, Bremner and Douglas (1971) registered 16, 14, and
13% inhibitions in urease activity of three soils (silty clay loam, loam, and clay loam),
the samples of which were treated with 50 ppm of AHA. At the same rate, BHA
exhibited smaller inhibitions (12, 10, and 8%, respectively). In the clay loam, the
influence of different rates of AHA on urease activity was also studied and it was
established that inhibition increased from 13 to 24, 33, and 44% when AHA rate was
increased from 50 to 100,200, and 300 ppm, respectively.
Preincubation of soil samples with 50 ppm of AHA at 30°C, before applying the 5-
hour test, led to diminution and, then, to disappearance of the inhibiting capacity of
AHA in each of the three soils. For example, a 3-day preincubation of the clay loam
with AHA diminished the inhibition from 13 to 5%, whereas in samples preincubated
with AHA for 7 days the urease-inhibiting capacity of this compound was not detectable
any more. The results obtained in the preincubation experiment with the clay loam were
also presented in a paper published by Douglas and Bremner (1971).
Continuing the investigations along this line, Bremner and Douglas (1973) evaluated
the effect of AHA on both urea hydrolysis and volatilization of urea-N as ammonia in
five Iowa soils. Ten-g soil samples were treated with urea (1,000 ppm of N) + 0 or 50
ppm of AHA, and water to 50% ofWHC and incubated at 2()°C. After 3, 7, and 14 days
of incubation, the residual urea, exchangeable NH4 +, and volatilized NH3 were recorded.
It was found that AHA inhibited ureolysis and NH3 volatilization in light-textured soils
and had a weak inhibitory effect in heavy-textured soils. For example, in a sandy soil
not treated with AHA, urea was not detectable after 14 days of incubation, whereas in
AHA-treated samples of the same soil, a part (19.4%) of the initial amount of urea
remained unhydrolyzed. In this soil urea-N losses through volatilization of NH3 during
14 days were 61.1 and 25.0%, respectively. In both untreated and AHA-treated samples
of a clay loam, urea was completely hydrolyzed in 14 days, while the volatilized NH3
represented 12.8% (untreated samples) and 12.3% (treated samples) of the initial
amount ofurea-N.
May and Douglas (1978) studied the effect ofBHA on urease activity of a sandy soil
from Australia. BHA was applied at rates of 10, 25, 50, and 100 ppm (on soil weight
basis). The reaction mixtures (3 g of soil + 0.5 rnl of toluene + 12 rnl of 1115 M
phosphate buffer, pH 8.8 + 3 ml of a solution containing 3 mg of urea + BHA) were
incubated at 37°C for 4 hours, then analyzed for ~ + released from urea. Reaction
mixtures without BHA served for comparison. It was calculated from the analytical data
that BHA at its four rates inhibited urease activity in proportions of 4, 14,29, and 46%,
respectively.
64
Gould el al. (1978) and Gould (1979) prepared mixtures from 25-g samples of a silt
loam soil, urea (400 ppm of N), 0 or 100 ppm of AHA and water (up to field capacity)
and incubated them at 25°e for 24 hours. Under these conditions, AHA caused a 17%
inhibition of urease activity.
Matzel et al. (1978) conducted experiments in vegetation pots containing 8-10 kg of
acid sandy soil (pH 3.9-4.8) or black soil (loess, pH 7.2) from Germany. Urea prills
were placed on the soil surface (225 em2 ) at a rate equivalent to 200 kg Nlha and AHA
was incorporated into the 0-I-cm soil layer at a rate of 200 ppm (7% on urea weight
basis). After moistening the soils to 60% ofWHC, the pots were kept at 20-22°e, and at
days 5, 12, and 19 the 0-5-cm soil layer was analyzed for residual urea and NH4 +.
Volatilization of ammonia from urea was studied with the sandy soil in smaller pots
(1.5 kg of soil). Urea (200 kg Nlha) was surface-applied and AHA was administered
together with urea at a rate of 3% on urea weight basis. Incubation was carried out at
20-22°e for 28 days, during which time the volatile NH3 was assessed.
The analyses showed that AHA inhibited urea hydrolysis, especially in the acid
sandy soil. Thus, in this soil when not treated with AHA, urea was present after 5 days
of incubation (the residual urea was equal to 4.1 % of the initial amount), but no urea
was detectable after 12 and 19 days. In the presence of AHA, urea remained
unhydrolyzed in proportions of 38.3% (after 5 days), 13.4% (after 12 days), and 1.1 %
(after 19 days). In the loess, after the three incubation times, the residual urea
represented 31.6, 17.5, and 0%, respectively (soil not treated with AHA) and 40.7, 24.6,
and 0(%, respectively (AHA-treated soil) in comparison with the initial amount of urea.
The NH3 loss was 18.3% in the absence of AHA and 0.4% in its presence after 10
days of incubation. When the incubation was prolonged to 28 days, the effect of AHA
became negligible, as the NH3 loss was 19.3% in the absence of AHA and 18.7% in its
presence.
Matzel et al. (1978) mentioned that AHA was also tested with a series of other soils,
but the inhibitions were weaker than those specified above in the case of the acid sandy
soil (see also Matzel and Heber,1979).
By using the short and long time tests, Hartbrich et af. (1978) examined the effect of
11 monohydroxamic acids on volatilization of ammonia from urea. Caprylohydroxamic
acid and AHA were the most effective: their inhibitory effect, after 8 days of incubation,
was 25 and 6%, respectively (see also Oertel et at., 1978; Jasche et al., 1978). Very
weak inhibitory effects were exhibited by ~-dimethylaminopropionohydroxamic,
bromoacetohydroxamic, and N-phenyl-~-chloropropionohydroxamic acids which
reduced volatilization ofNHJ only by 3,5, and 8%, respectively, after I-day incubation,
and did not have any inhibitory effect after 2 days of incubation. The other hydroxamic
acids occupied, in terms of their inhibitory effect, intermediary positions in the order:
N-phenyl-n-cyclohexylarnino-AHA>chloro-AHA>~-diethylaminopropionohydroxarnic
acid>f\-ethylaminopropionohydroxarnic acid>~-methylaminopropionohydroxamic acid
>N-phenyl-n-pyrroJidino-AHA.
Strumpf et al. (l98Ib) patented seven ro-(naphthoxy)alkanohydroxarnic acids which
inhibited activity of soybean urease at lower concentrations than did AHA,
benzohydroxamic acid, and phenoxy-AHA and reconunended these compounds as
additives to fertilizer urea at preferred rates of 0.1-5% relative to urea-No Another
advantage of these compounds is their capacity to hinder the growth of soil-borne
65
phytopathogenic bacteria and fungi. At the same time, they are well tolerated by the
crops. It should, however, be emphasized that the description of invention does not
contain any data about testing of these compounds as inhibitors of soil urease activity.
Their general structural formula is presented in Figure 11.
o-(CH)n~O-NH---OH
Oj
r l A
~ .&
R = H or low alkyl
n= 1-10(R=H)
Figure 11. General structural formula of Ol-(naphthoxy)alkanohydroxamic acids

patented by Strumpf et al. (l981b).
The inventors Kolc et al. (1985c) (Assignee: Allied Corporation, Morristown, New
Jersey) patented three groups of hydroxamic acid compounds as additives to urea
fertilizers for inhibition of soil urease activity. Twenty-eight such compounds are
nominalized in the patent. We mention a representative compound from each group
(Figure 12).
4-(l'-Maleimido)butanehydroxamic acid S-(3',4'-Dihydroxy-S'-fluorophenyl)hexanehydroxamic acid
7-[N(2'-Nitro-2'-bromovinyl)-N-ethylaminojheptanehydroxamic acid
Fif<ure 12. Structure ofrepresentative hydroxamic acid compounds patented by Kole et al. (1985c).
But it should be mentioned that no data are presented in the description of the
invention on the inhibitory effect of these compounds on activity of soil urease or
jackbean urease.
2.9.2. Dihydroxamic Acids

Eight aliphatic dihydroxamic acids were patented by Strumpf et al. (1981a) for
inhibiting urease activity (Figure 13).
66
Co-HN-OR,
I
(CH-R2ln
ICO-HN-OR,
R1 = H. benzoyl or equivalem of a metallic sah

R2 = H. low alkyl. benzyl or subsl~u19d benzyl
n = 1-6
Figure 13. General structural formula of aliphatic dihydroxamic acids patented by

Strumpf et al. (l981a).
Tested on soybean urease, these compounds proved to be more effective than AHA
and succinomonohydroxamic acid. Moreover, they suppress the growth of soil-borne
phytopathogenic bacteria and fungi and are not toxic for plants. Although they were not
tested as inhibitors of soil urease activity, based on the results obtained with soybean
urease, the inventors consider that these compounds when added to fertilizer urea will
prevent the undesirable effects of the rapid hydrolysis of urea in soil. The preferred rates
at which these compounds are recommended for use range from 0.1 to 5% relative to
urea-No
2.10. MALEIMIDES
Patenting the unsubstituted and halo-substituted alkyl, cycloalkyl, and aryl derivatives
of dichloromaleimide as inhibitors of soil urease activity, Tomlinson (1967)
nominalized four N-alkyldichloromaleimides (N-alkyl-DCMI): N-methyl-, N-ethyl-, N-
n-propyl-, and N-n-butyl-DCMI (Figure 14). Applying the first of his methods already
outlined on page 53, he treated 2.5-g samples of two soils with 2,000 ppm of urea (on
soil weight basis) and 20 ppm ofN-alkyl-DCMI, then incubated them for 3 or 7 days at
9°C. During the incubation the volatilized ammonia was measured. The results showed
that during the first 3 days volatilization ofNH3 from one soil was completely inhibited
by each N-alkyl-DCMI, but in the other soil the volatile N losses as NH3 in the presence
of the four N-alkyl-DCMI represented 26-65% relative to the NH3 losses from the
control soil to which no N-alkyl-DCMI had been added. During the whole 7-day period,
51-77% NH3 losses were registered; with the first soil the minimum loss (57%)
occurred in the N-n-butyl-DCMI-treated samples, whereas with the other soil the NH3
loss was lowest (51 %) from samples treated with N-n-propyl-DCML
Under the conditions of the 5-hour test, Bremner and Douglas (1971) studied
N-ethylmaleimide (N-ethyl-MI) which was added to samples of three soils at a rate of
50 ppm (soil basis). The inhibitions recorded in urease activity of the three soils were
28, 24, and 23%, respectively. In another experiment, soil samples were preincubated
with N-ethyl-MI at 30°C for 3, 7, and 14 days, before applying the 5-hour test. This
experiment showed that during preincubation N-ethyl-MI was partially inactivated.
67
o o o
II II II
CIC-C, CIC-C,
II ..N-R II .,N-R II
HC-C>
-R
CIC-C/ HC-C/ HC-C
II II II
o o o
Dichloromaleimide Chloromaleimide Maleimide
R=H or -CH3
methyl ethyl n-propyl
-CHr-CH2-CH2-CH3
n-butyl
-0 -0 ~
~
~-naphthyl
phenyl cyclohexyl
Figure 14. Maleimides tested as inhibitors of soil urease activity.
Thus, in one of the three soils studied, inhibition of urease activity was reduced from
23% (registered in the soil not submitted to preincubation with N-ethyl-MI) to 21, 19,
and 8% in samples preincubated with N-ethyl-MI for 3, 7, and 14 days, respectively_
The inhibiting effect ofN-ethyl-MI on soil urease activity was attributed to its capacity
to react with SH groups of the enzyme molecule.
Reexamining N-ethyl-MI, Bremner and Douglas (1973) determined both urea
hydrolysis and ammonia volatilization from urea in a sandy soil. Air-dried samples (10
g) were treated with 1 ml of urea solution containing 1,000 ppm N (soil basis) and 1 rnl
of water (control soil) or I rnl of N-ethyl-MI solution (50 ppm compound), then water
was added to bring the soil moisture content to 50% of WHC, and the mixtures were
incubated at 20°C. After 3, 7, and 14 days, the amounts ofresidual urea, exchangeable
NH/, and volatilized NH3 were determined. The results indicated that N-ethyl-MI
inhibited urease activity in the sandy soil studied. Some of the results will be quoted
below. After 14 days of incubation, no urea was detectable in the control soil, whereas
the N-ethyl-MI-treated soil contained 24.2% of the initial urea amount; loss ofurea-N as
volatile NH3 was 61.1 % from the control soil and 23.6% in the presence ofN-ethyl-MI.
Based on the results obtained by Bremner and Douglas (1973) with other inhibitors
administered to heavy-textured soils, one can assume that the inhibiting effect of
N-ethyl-MI on urease activity in such soils would have been much weaker.
Hartbrich et af. (1978) tested the rnaleimides (MI), chlorornaleimides (CMI) , and
dichloromaleimides (DCMI) specified in Table 11 (see also Figure 14). Their inhibitory
68
effect on anunonia volatilization was strong for 4-8 days, but it disappeared, in general,
after 12 days. The effect of N-cyclohexyl-DCMI was the longest lasting (14 days) (see
also Oertel el af., 1978; lasche el af., 1978) and that of N-~-naphthyl-DCMI was the
least durable (4 days). It can also be deduced from the table that the influence of the
number of chlorine atoms on the degree of inhibition depends on the nature of the N-
substituent. Thus, the inhibitory effect of N-phenyl-MI, N-phenyl-CMI, and N-phenyl-
DCMI, being similar, is not influenced by the number of chlorine atoms. In contrast, the
number of chlorine atoms has an influence in the case of other maleimides as the
inhibitory effect increases in the order:
N-cyclohexyl-MI < N-cyclohexyl-CMI < N-cyclohexyl-DCMI,
or presents the following order:
N-~-naphthyl-MI ;:::; N-~-naphthyl-CMI > N-~-naphthyl-DCMI.
TABLE 11. Efiect of maleimides on volatilization of ammonia from urea-treated soils"
Inhibition (%)
STT LTT
Compound
1 2 4 12 14 16
N-Phenyl-MI 99 98 78 9
N-Phenylchloro-MI 92 96 80 16
N-Phenyldichloro-MI 82 96 83 26 0
N-CyC\ohexyl-MI 97 96 74 26
N-Cyclohexylchloro-MI 83 99 98 47 11 0
N-Cyclohexyldichloro-MI 79 94 91 48 19 10 ()
N-~-Naphthyl-MI 97 90 68 26
N-~-Naphthylchloro-MI 92 97 94 23 0
N -~-N aphthy1dichloro. MI 49 76 47 0
Dichloro-MI 88 91 69 14
N- Methyldichloro-MI 88 100 96 23 0
N-Ethyldichloro-MI 95 95 43 18 0
N-n-Propyldichloro-MI 96 98 61 21 0
"From Hartbrich et al. (1978).
STT -- Short time test.
LTT -- Long time test.
MI - maleimide.
In respect to the three N-alkyl-DCMI studied, Table 11 shows that the inhibitory
effect ofN-methyl-DCMI was stronger than that ofN-ethyl- and N-n-propyl-DCMI. In
contrast, in one of the soils studied by Tomlinson (1967), N-n-propyl-DCMI was more
evidently inhibiting than were the other three N-alkyl-DCMI tested (see page 66).
Hartbrich el al. (1978) mentioned that, besides the compounds specified in Table 11,
other N-substituted MI, CMI, and DCMI also manifested an inhibitory effect on soil
urcase activity.
69
Inhibition of soil urease activity by N-ethyl-MI was found to be non-competitive in

both soils (an alluvial soil and a leached chemozem) studied by Simihaian and Silberg
(1999); the inhibition constant was 10.3 and 11.0 mM, respectively.
2.11. MALEIC HYDRAZIDE
The structure of its tautomeric forms (dione and diol) is presented in Figure 15.
o OH
II I
,......c . . . . . ,......c~
HC NH HC N
II
HC
I
NH
II I
HC N
'c""'" , c.?'
~ 6H
Dione form Diolform
1.2-Dihydro-3.S-pyridazinedione 3.S-Dihydroxypyridazine
Figure 15. Structure of maleic hydrazide.
Reaction mixtures prepared from 5 g of soil and 10 ml of aqueous phase containing

6 mg of urea and 0 or 0.12 mg of maleic hydrazide (MH), and incubated at laboratory
temperature were periodically analyzed by a chromogenic reagent (see the footnote on
page 35) to estimate the time necessary for complete hydrolysis of urea. In the presence
of MH, this time was prolonged from 6 to 8-10 days in an alluvial soil and from 9 to 12
days in a leached chemozem. But even this weak inhibitory effect disappeared when
half of the MH anlOunt was replaced by paraformaldehyde (Kiss and Pintea, 1987).
2.12. MUCOCHLORIC ACID
Mucochloric acid (a,t\-dichloro-fonnyl-acrylic acid; Figure 16) as an inhibitor of soil

urease activity was patented by Hartbrich et al. (1 976b). They applied the short and long
time tests (see page 51) with the difference that this compound was used at four rates
CI-C-CHO
II
CI-C-COOH
Figure 16. Structure ofmucochloric acid.
relative to urea-N (0.5, 1,2, and 4% in the short time test, and 0.5, 1, 1.5, and 2% in the
long time test). Determination of ammonia volatilized during 24 hours and 18 days,
respectively, indicated that mucochloric acid brought about complete inhibition of NH3
evolution during the first days of incubation, and then the inhibitory effect decreased
70
slowly. Thus, under the influence of the four mucochloric acid rates, the inhibition
persisted for 12, 14, 16, and 16 days, respectively.
Hartbrich et al. (1978) pointed out that the strong and persistent inhibitory effect of
mucochloric acid, as described in the patent, could not be confirmed in pot experiments.
2.13. BROMO-NITRO COMPOUNDS
The inventors Norden et 01. (1985, 1986) (Assignee: Chernische Werke Hills A.G.,
Marly, Germany) tested and patented 41 bromo-nitro compounds for inhibition of soil
urease activity. For testing, 200-g moist soil samples were treated with urea prills
(usually 0.5 g) and a bromo-nitro compound (BNC) at a rate of 0.25 or 1% by weight of
urea-N, then incubated for 4 or 7 days. Soil samples to which no BNC was added were
the controls. During incubation the ammonia volatilized was determined and expressed
as percentage of the added urea-No
All BNCs reduced NH3 volatilization. The BNCs which were found to be the
strongest inhibitors of soil urease activity are specified in Figure 17.
Tr ft Tr ft
CH3-C -CH2-0-C-NH - CH 3 CH3-CH2-C-CH2-0-C -NH -CH3
~02 1 02
2-Bromo-2-nit ropropy I-N-methy Icarbamate 2-Bromo-2-nitrobuty I-N-methy Icarbamate
2-Bromo-2-nitropropane-l,3-diol 2-Bromo-2-nitropropanol
Figure 17. Stru~ture of the strongest inhibitors among the 41 bromo-nitro compounds patented and tested by
Norden et (I/. (1985, 1986) for inhibition of soil urease activity.
Volatilization of NH3 was 52.8% from the control soil and only 0.3, 0.5, 0.9, and
2.1 % respectively, from the soil treated with these four BNCs, when they were used at
the 0.25% rate and the incubation lasted 4 days. Volatilization ofNH3 was 71.2% from
the control soil and always 0% from the soil treated with each of these four BNCs when
their rate was I % and the incubation lasted 7 days.
These inhibitors are recommended to be added to urea fertilizers at rates ranging
from about 0.05 to 15%, preferably from about 0.4 to 4% by weight ofurea-N.
It should be mentioned that before the patenting of the 41 BNCs as inhibitors of soil
urease activity by Norden et 01. (1985,1986), similar BNCs were already patented as
antibacterial and antifungal agents for use in human and veterinary medicine (Clark et
aI., 1967), for combating phytopathogenic bacteria (Clark et 01_, 1970) and
71
phytopathogenic fungi (Pommer and Wessendorf, 1972), as antimicrobial agents for

washing purposes (Nosier et al., 1971).
2.14. HETEROCYCLIC SULFUR COMPOUNDS
2.14.1. Tetrahydro-l.3.5-thiadiazine-2-thiones
Held et al. (1974) patented tetrahydro-l,3,S-thiadiazine-2-thiones (Figure 18) for
inhibition of soil urease activity. Moist soil samples treated with a mixture of urea and
test compound were incubated and during incubation the volatilized ammonia was
determined. No urea and no test compound or only urea was added to the control
samples. Data on incubation temperature and time are not given in the patent.
Derivative
CH, CH, 3,5-dimethyl-
CH,-CII, CH,-CH2 3,5-diethyl-
CH,--CH=CH2 CH,--CH=CH2 3,5-dialiyl-
C.H, C6H, 3,5-diphenyl-
C 6H'-{:H2 CH, 3-benzy1-5-methyl-
C 6 H,-{:H2 CH,--CH20H 3-benzyl-5-hydroxyethyl-
Figure 18. General structural forl1Ulla oftetrahydro-l,3,S-thiadiazine-2-thiones patented

and tested by Held et al. (1974) for inhibition of soil urease activity, with specification of
some derivatives.
Table 12 presents the results obtained with 3,S-dimethyltetrahydro-l,3,S-thiadiazine-

2-thione, which under the name dazomet* is used, at high concentrations, for
disinfection of soil or as a nematicide or herbicide. One can see from this table that the
inhibitory effect of dazomet increased with its increasing concentration. The pure
dazomet was more effective than the technical-grade product. Dazomet and the other
derivatives are recommended to be used at preferred rates of O.S-IO% relative to urea-No
In the experiments performed by Lang et al. (1976), soil samples were treated with
urea + 0 or 2% dazomet (on urea-N basis), then moistened and incubated at 20°C for 16
days. Volatilization of NHl from urea was inhibited by dazomet. The inhibitions
recorded after 2,4, 8, 12, and 16 days of incubation were 100, 97.8, 77.2, 28.S, and
20.7%, respectively.
'Markert (1974) found that dazomet reduced the rate of urea hydrolysis in each of the two soils studied, but he
did not patent this compound as an inhibitor of soil urease activity.
72
TABLE 12. Inhibition of enzymatic hydrolysis of urea in soil by 3,5-dimethyltetrahydro-I,3,5-

thiadiazine-2-thione (dazomet) applied at different concentrationsO
Concentration ofinhibitor Inhibition (%)
(% relative to urea-N) Pure cO!IJlound Technical-grade compound
0.125 40.4 30.6

0.25 43.5 32.7
0.5 53.5 51.7
1.0 66.5 57.0
2.0 71.6 67.2
4.0 74.6 74.1
8.0 77.8 75.8
°FromHeld et al. (1974).
Hartbrich et al. (1978), Oertel et al. (1978), and Jasche et al. (1978), applying the
long time test, obtained, after 2, 4, 8, 12, and 14 days of incubation, similar inhibitions,
namely 100, 95, 64, 28, and 11 %, respectively, under the influence of dazomet added to
urea-treated samples at a rate of 1% relative to urea-No
In Winiarski's (1990) experiments, 2-kg samples of a light-textured and a heavy-
textured Polish soil, the pH of which was adjusted to 5 (with 1 N H2 S04 ) or to 7 (with
CaC03), were treated with 328 mg of urea and 0 or 0.5, 1 or 2% of dazomet or 2% of
"thione" (tetrahydro-l,3,5-thiadiazine-2-thione) (on urea weight basis), then moistened
to 50% ofWHC and incubated at 22°C for 7 or 14 days.
Loss ofurea-N as volatile ammonia from the inhibitor-treated samples was reduced.
The inhibitory effect of dazomet tended to increase with its rate and to decrease with
incubation time in both soils. The inhibition was stronger in the light than in the heavy
soil. The maximum inhibition (64.1 %) was registered in the light soil (at pH 5; 2%
dazomet) after 7-day incubation, while the minimum value (16%) was recorded in the
heavy soil (at pH 7; 0.5% dawmet) after 14-day incubation. The inhibitory effect of
thione was less pronounced at pH 5 than at pH 7 and showed a tendency to decrease
with incubation time in both soils. Thus, at pH 7, the reduction ofNH3 loss during 7 and
14 days of incubation was 31.2 and 13.6% (in the light soil) and 23.2 and 17.0% (in the
heavy soil), respectively.
The effect of dazomet on soil urease activity was studied under field conditions as
well (Tu et al., 1995). A Canadian loamy sand was treated with 0 or 56 kg of
dawmetlha ridging it into the soil to depth of 14 cm on May 10, 1978. A 3-18-24
tobacco fertilizer was applied and tobacco seedlings were planted on May 23. For
determination of urease activity, soil was sampled on May 23 (before fertilization and
planting) and on June 27. This activity was found to be insignificantly (p > 0.(5) lower
on May 23 and insignificantly higher on June 27 in the dawmet-treated soil than in the
untreated one.
2.14.2. 1.3.4-Thiadiazoline-2-thiones
In the patent of Held et al. (1976a), the 1,3,4-thiadiazoline-2-thiones as well as their
alkali metal and substituted ammonium salts are described as inhibitors of soil urease
activity. The tautomeric (thione and thiol) forms of these compounds and the derivatives
specified in the patent are presented in Figure 19.
73
5-R-l,3,4-thiadiazoline-2-thione 2-Mercapto-5-R-l ,3, 4-thiadiazole

(5-R-l,3,4-thiadiazole-2-thiol)
R Derivative
C.Hs 5-phenyl-
CH)-CH2-O 5-ethoxy-
(CHlhCH-O 5-isopropoxy-
HS 5-mercapto-*
CH)(CH2hClh-NHtS- 5-mercapto- (amylammonium salt)
CH)-S 5-methylmercapto-
CH)-CH2-S 5-ethylmercapto-
CI b-CHrOh-S 5-n-propylmcrcapto-
(CH,hCH-S 5-isopropylmercapto-
C.Hs-Clh-S 5-benzyhncrcapto-
H2N 5-amino-
C.Hs-NH 5-phenylamino-
'2,5-dimercapto-1 ,3,4-thiadiazole (I ,3,4-thiadiazole-2,5-dithiol)
Figure 19. General structural formula of 1,3,4-thiadiazoline-2-thiones (2-mercapto-I,3,4-

thiadiazoles) patented and tested by Held et al. (I 976a) for inhibition of soil urease activity, with
specification of some derivatives.
Soil samples, to which a mixture of urea + test compound was added, were
moistened and incubated at 30°C for 24 hours. Samples treated with urea alone served
for comparison. During incubation, the evolved ammonia was determined. Table 13
shows the results obtained with seven derivatives, each containing an unsubstituted or
substituted mercapto (sulfhydryl; SH) group in position 5. It is evident from this table
that the degree of inhibition increased with increasing concentration of test compounds,
but not linearly. In general, the increase in inhibition was most marked when the test
compound concentration was increased from 0.5 to 1%. The alkylated derivatives were
more effective than the aralkyl (benzyl) derivative. 2,5-Dimercapto-l,3,4-thiadiazole
was more effective than its amylammonim salt, except when it was used at 0.5%
concentration. The preferred rates, at which these compounds are recommended for the
practice, range from 0.5 to 10% relative to urea-No
Hartbrich et al. (1978) presented data on the inhibitory effect of a single derivative
(2-mercapto-5-methylmercapto-l,3,4-thiadiazole) which, under the conditions of the
long time test, caused, after 2,4, and 8 days of incubation, 100,61, and 19% inhibitions,
respectively, in volatilization ofNH3 from urea-treated soil samples.
Gould et al. (1978) and Gould (1979) studied three compounds whose structure
corresponds to the general formula in Figure 19 (2,5-dimercapto-l,3,4-thiadiazole,
2-mercapto-5-amino-l,3,4-thiadiazole, and potassium salt of 5-mercapto-3-phenyl-
1,3,4-thiadiazoline-2-thione) as well as three other compounds (Figure 20), two of
which are disulfides prepared through oxidation (with H20 2) of 2-mercapto-5-amino-
1,3,4-thiadiazole and 5-mercapto-3-phenyl-l,3,4-thiadiazoline-2-thione, whereas the
third compoud is a sulfur-containing imidazole derivative.
74
TABLE 13. Inhibition of enzymatic hydrolysis of urea in soil by 2-mercapto-l,3,4-thiadiazoles applied at

different concentrations·
Inhibition (%)
Concentration of compound
Compound
(% relative to urea-N)
0.5 2 4
2,5-Dimercapto-l,3,4-TO 31.4 59.8 59.8 62.9

Amylammonium salt of 2,5 -dimercapto-l ,3,4-TO 39.4 40.1 42.0 50.0
2-Mercapto-5-methylmercapto-1,3 ,4-TO 32.6 51.0 60.0 75.0
2-Mercapto-5-ethylmercapto-l,3,4-TO 30.1 46.9 58.1 64.6
2-Mercapto-5-n-propylmercapto-l,3,4-TO 23.9 48.2 57.3 63.2
2-Mercapto-5-isopropylmercapto-l,3 ,4-TO 27.8 36.9 53.1 62.9
2-Mercapto-5-benzylmercapto-I,3,4-TO 12.1 30.8 45.1 42.5
"Adapted from Held et al. (1976a).
m - thiadiazole.
The effect of these compounds was estimated by analyzing the residual urea
extracted from 25-g samples of a silt loam soil which had been treated with urea (400
ppm N, on soil weight basis), test compound (0 or 100 ppm), and water (to field
capacity), then incubated at 25°C for 24 hours. It is evident from the results obtained
(Table 14) that, of the six compounds tested, 2,5-dimercapto-l,3,4-thiadiazole had the
most pronounced inhibitory effect. Potassium salt of 5-mercapto-3-phenyl-I,3,4-
thiadiazoline-2-thione did not inhibit soil urease activity.
CsHs-N N N---N-CsH5
=~L-&-I~..,
2.2·-Di(5-amino-l.3.4-thiadiazole) d~u~ide 5.5·-Di(3-phenyl-l,3,4-thiadlazoline-2-thione) disuWlde
HC---N
I
HC" /C-SH
I
N
I
CH3
2-Mercapto-l-methylim idazole
Figure 20. Structure of three of the heterocyclic sulfur compounds tested by Gould et al. (1978) and Gould
(1979) for inhibition ofsoi! urease activity.
The effect of different concentrations of 2,5-dimercapto-I,3,4-thiadizole on soil

urease activity was also studied, and it was found that inhibition of soil urease activity
was linear with the concentration of this compound up to 40 ppm, but leveled off at
higher concentrations_
75
TABLE 14. Effect of heterocyclic sulfur compounds on soil urease activity"
Compound Inhibition (%)
2,5-Dimercapto-1,3 .4-thiadiazole 46
2-Mercapto-5-amino-l,3.4-thiadiazole 27
2-Mercapto-I-methylimidazole 13
2,2'-Di(5-amino-I,3.4-thiadiazole) disulfide II
5,5 '-Di(3 -phenyl-I.3,4-thiadiazoline-2-thione) disulfide 6
5-Mercapto-3-phenyl-l,3.4-thiadiazoline-2-thione, K salt 0
"From Gould et al. (1978) and Gould (1979). by permission of the Soil Science Society of
America, Inc.
It was also established that moisture content above field capacity of the silt loam
studied had no effect on the inhibition of soil urease activity by 2,5-dimercapto-l,3,4-
thiadiazole used at 50 ppm, but the inhibition decreased somewhat at lower moisture
contents.
It should be mentioned here that, according to Gem1ann-Bauer (1987) and Amberger
(1989), the inhibition of urease activity in guanylthiourea (GTU)-treated soils (see page
52), is caused not by the GTU itself, but by 3,5-diamino-l,2,4-thiadiazole, which is
produced in soil due to abiotically catalyzed oxidation of GTU. The next, abiotically
catalyzed reaction is the reduction of this thiazole into dicyandiamide which is a potent
nitrification inhibitor (Figure 21).
- -
N - - - G -NH2 NH
NH
II
H:<N-C-NH -c-NH2
S
II I
H:<N-C, / N
I II
H:<N-C-NH -c =N
S
Guany Ithiourea 3,5-Diamino-1,2,4-thiadiazole Dicy andiam ide
Figure 21. Transformation of guanylthiourea in soil. IFrom Gennann-Bauer (1987); Amberger (I 989)./
2.14.3. Rhodanine-5-acetic Acid and Its Derivatives

Schroth et al. (1974) patented rhodanine-5-acetic acid and its 3-substituted derivatives
as well as their heavy metal salts for inhibition of soil urease activity. The patent
nominalizes nine compounds (Figure 22). The reaction mixtures had the following
composition: 10 g of of soil + 214 mg of urea (100 mg ofurea-N) + 0 or 10 mg of test
compound (e.g. 3-methylrhodanine-5-acetic acid) + water to 50% of WHC. They were
incubated at 30°C for 24 hours, during which the volatilized an1ffionia was determined.
76
Rhodanine-5-acetic acid
O=C---N-R
I
HOOC-CH2--HC, /G=S
I
S
R Derivative
CH, 3-methyl-
CH,-CH2 3-ethyl-
CH,-CH2-CH2 3-n-propyl-
(CH,hCH- 3-isopropyl-
CH,-CH2-CH2-CHz 3-n-butyl-
(CH,J:,-CH-CH2 3-isobutyl-
C.H" 3-cycIohexyl-
C.H 5-CHz 3-benzyl-
Figure n. Structure ofrhodanine-5-acetic acid and of its derivatives patented and tested
by Schroth e/ al. (1974) for inhibition of soil urease activity.
One can see from Table 15 that the inhibitory effect increased in the order:
rhodanine-5-acetic acid ~ l:ycloalkyl derivative < aralkyl derivative ~ alkyl
derivatives.
TABLE 15. Effect ofrhodanine-5-acetic acid and ofits derivatives on soil

urease activity"
Rhodanine-5-acetic acid 35
3-Methylrhodanine-5-acetic acid 97
3-Ethylrhodanine-5-acetic acid 95
3-n-Propylrhodanine-5-acetic acid 92
3-Isopropylrhodanine-5-acetic acid 92
3-n-Butylrhodanine-5-acetic acid 91
3-Isobutylrhodanine-5-acetic acid 90
3-Cyclohexylrhodanine-5-acetic acid 38
3-Benzylrhodanine-5-acetic acid 93
"Adapted from Schroth et al. (1974).
Of the alkyl derivatives, the methyl derivative was the most effective. Cu, Cd, Zn,
Co, Pb, and Fe salts of 3-ethylrhodanine-5-acetic acid gave 82-93% inhibitions. In the
case of the Ni salt the inhibition was 56%, whereas the Sn and Hg salts did not inhibit
77
volatilization of NHJ from urea. The inhibitory compounds are recommended for the
practice at rates of 0.1-5%, preferably at a rate of 1% relative to urea-No
Oertel et al. (1978) found that in reaction mixtures prepared from 30-g samples of
moistened sandy soil + 214 mg of urea + I mg of surface-applied 3-methylrhodanine-5-
acetic acid (1 % relative to urea-N) and incubated at 20°C, volatilization of NH3 from
urea during 2, 4, 8, and 12 days was reduced in proportions of 95, 60, 8, and 0%,
respectively, under the influence of 3-methylrhodanine-5-acetic acid.
Muller and Forster (1980) confirmed the inhibitory effect of 3-methylrhodanine-5-
acetic acid on soil urease acivity. They worked with a humous sandy loam (PH 7.2)
from Germany. Urea (40 mg of N) was distributed uniformly in an 8-cm high soil
column (40 g) or applied on its surface. The test compound (2% on urea-N basis) was
introduced at 2 or 4 cm depth in the soil column. After moistening and incubation at
laboratory temperature for 6 days, the residual urea and NH4 + + NO) - contents were
determined. In all treatments, 3-methylrhodanine-5-acetic acid caused at least 50%
inhibitions of urea hydrolysis.
2.14.4.2-Mercaptobenzothiazole
Urease activity was not inhibited in reaction mixtures consisting of 5 g of soil + 10 ml
of aqueous phase containing 6 mg of urea and 0 or 0.12 mg of 2-mercaptobenzothiazole
(Figure 23) and incubated at laboratory temperature; in both the absence and the
Figure 23. Structure of2-mercaptobenzothiawle.
presence of this compound, complete hydrolysis of urea occurred in 6 days in an

alluvial soil and in 9 days in a leached chernozem (Kiss and Pintea, 1987).
2.14.5. 2- Thiocarboxamidothiazoles
Simihilian and Silberg (1996) and Simihaian et al. (1999) tested 2-thiocarboxamido-5-
aminothiazole (crysean) and 2-thiocarboxamido-5-benzamidothiazole (benzoyl cry sean)
(Figure 24) for inhibition of soil urease activity. Samples were taken from the O-lO-cm
Ht-J-ClI~s/-
s
"~NH-C-o~ 0
II -
2-Thiocarboxamido-5-aminothiazole 2-Thiocarboxamido-5-benzamidothiazole
(cry sean)
(benzoyl cry sean)
Figure 24. Structure of cry sean and benzoyl crysean.

layer of an alluvial soil (pH 8.30), a leached chemozem (pH 7.l2), and a brown forest
soil (pH 4'<)0), and amended with urea and test compund (at a rate of 0.12% relative to
weight of urea), then incubated at 37°C for 5 hours, followed by determination of the
residual urea.
The results showed that both crysean and benzoyl crysean behaved differently in the
three soils studied: they had no significant effect on urease activity of the alluvial soil,
caused a 3-4% stimulation of this activity in the chernozem, and an about 5% inhibition
of the activity in the forest soil.
In another experiment, urease activity was determined not only in the surface layer
(0-10 cm) but also in deeper ones (10-30, 30-40, and 40-70 cm) of the brown forest soil
and it was found that urease activity of the deeper layers as compared to that of the
surface layer was inhibited to a larger extent. Crysean as compared to benzoyl crysean
was a stronger inhibitor, which means that substitution of a H atom by the benzoyl
group at the 5-NH2 of crysean molecule caused a diminution of the urease-inhibiting
capacity.
2.15. MONOHYDRIC PHENOLS
The monohydric phenols that were tested for evaluation of their inhibitory effect on soil
urease activity comprise: phenol (hydroxybenzene), a series of its derivatives, including
some derivatives of methylphcnols (cresols), as well as a-naphthol (I-naphthol) and one
6
OH OH OH
OCH' O-CH'
Phenol a-Cresol m-Cresol
OH
(J()
p-Cresol a-Naphthol
FiKure 25. Structure of some monohydric phenols.
of its derivatives. Figure 25 presents the structure of phenol, cresols, and a-naphthol.
Their derivatives tested will be specified during the description of investigations.
79
Rotini (1935), studying hydrolysis of urea in presence and absence of antiseptics,

prepared reaction mixtures from 50 g of soil + 30 ml of 0.1 % urea solution + 1 ml of
water or 5% phenol solution. After incubation (42 D C/4 hours), the residual urea was
determined. Its amount was found to be nearly equal in the untreated and phenol-treated
soil. The conclusion was drawn that phenol stopped the growth of microorganisms, but
did not lyze them and, thus, did not lead to release of urease from the microbial cells;
urease activity measured in the presence of phenol was due to those urease molecules
that existed in soil in a free state even before the addition of phenol.
Under conditions of the 5-hour test, Bremner and Douglas (1971) evaluated the
effect of 14 monohydric phenols, applied at a rate of 50 ppm (soil weight basis) on
urease activity in three Iowa soils. The results presented in Table 16 show that only
phenol and 4-chiorophenol had an evident inhibitory effect on soil urease activity.
The other phenols caused inhibitions of less than 1%. The inhibition exhibited by
a-naphthol was less than 2%.
With the most effective inhibitors (i.e.. phenol and 4-chiorophenol), the influence of
preincubation on the inhibitory capacity was also studied. Water or an aqueous solution
containing 500 J.lg of phenol or 4-chiorophenol was added to lO-g soil samples . After 0,
3, 7, and 14 days of preincubation at 30D C, the 5-hour test was applied for determination
of urease activity. It was found that during preincubation the inhibiting capacity of both
phenols decreased. For example, in the clay loam soil, after 14 days of preincubation,
the inhibition caused by phenol decreased from 41 to 8% and that brought about by 4-
chlorophenol was reduced from 30 to 3%.
TABLE 16. Efl"ect of monohydric phenols on soil urease activity"

Compound
Silty clay loam Loam Clay loam
Phenol 43 41 41
4-Chlorophenol 37 37 30
Other compounds" <l <I <I
"Adapted from Bremner and Douglas (1971). by permission of Pergamon Press PLC.
b2,5-Dichlorophenol, 2-nitrophenol, 4·nitrophenol, 2A-dinitrophenol, 2,4,6-trinitro-
phenol (picric acid), 4-chloro-2-nitrophenol, 4-hydroxybenzoic acid, 2-aminophenol,
4-aminophenol, 2-methoxyphenol (guaiacol), 4,6-dinitro-2-methoxyphenol.
Gould et al. (1978) tested 25-g samples of a silt loam soil with 400 ppm of urea-N, 0
or 100 ppm of phenol or a-naphthol and water to field capacity, then incubated them at
25 D C for 24 hours. Phenol exhibited a 10% inhibition, whereas a-naphthol gave a 1%
Mishra and Flaig (1979) prepared mixtures from 200-g soil samples (a brown earth
and a black earth from Germany), 0 or 5 ml of urea solution containing 20 mg ofN (100
ppm in soil), 0 or 5 ml of solution containing 10 mg of2, 4-di-t-butylphenol (50 ppm in
soil) and water to 60% of WHC. After 0, 14, and 28 days of incubation at 30D C, soil
urease activity was assayed by the method of Tabatabai and Bremner (1972) (see page
8). 2,4-Di-t-butylphenol inhibited urease activity at 0 time and stimulated it after 14 and
80
28 days of incubation in the case of brown earth, and constantly stimulated it in the case
of black earth.
Xue and Li (1987) studied the effect of phenol and methylnaphthol" on urease
activity in a paddy soil. The reaction mixtures contained 5 g of soil + 10 ml of 10% urea
solution without or with 100 ppm of phenolic compound. Incubation took place at 37°C
and lasted 48 hours. Degree of inhibition was 22.35% with phenol and 30.33% with
methylnaphthol.
Rodgers (l984b) tested 5 aminocresols with three English soils (silty clay loam,
sandy clay loam, and loamy sand). Air-dried soil samples (8 g) were treated with 1.2 ml
of 0.4% aqueous urea solution (280 Ilg N/g soil) + 0.8 m1 of 60% methanol or 0.8 m1 of
aminocresol solution in 60% methanol (concentrations: 5-100 Ilg of aminocresol/g soil).
During incubation (at 30°C for about 100 hours), the amounts of residual urea and of
NH4 + were determined periodically. Table 17 presents the results obtained with one of
the three soils studied after 20 hours of incubation. One can see from this table that only
two aminocresols (4-amino-o-cresol and 4-amino-m-cresol) inhibited significantly the
urease activity; degree of inhibition increased as their concentrations increased.
TABLE 17. Inhibition of urease activity in a sandy clay loam soil by aminocresols applied at
different concentrations"
Compound Concentration of compound (/lWg soil)
5 10 20 50 100
2-Amino-p-cresol I I I 8 14
3- Ami no-o-cresol <I <I <I <I <1
4-Amino-o-cresol 8 26 24 53 74
4-Amino-m-cresol 9 24 21 40 62
6-Amino-m-cresol <1 <1 <1 <1 <1
"From Rodgers (1984b), by permission of Kluwer Academic Publishers.
Since 2-,3-, and 6-aminoderivatives had practically no inhibitory effect, it is evident

that the presence of the amino group in position 4 is essential for the inhibiting capacity.
The presence of methyl group is also necessary, because the urease-inhibiting effect of
2- and 4-aminocresols is negligibly low (see Table 16).
The most inhibitory aminocresol (4-amino-o-cresol) acted more strongly on the silty
clay loam which is characterized by a weaker urease activity, than on the other two soils
whose urease activity is higher. For this reason, Rodgers (1984b) arrived at the
conclusion that this aminocresol also fails to present practical importance as a soil
urease inhibitor, because it is less effective in precisely those soils in which inhibition of
urease activity would be necessary.
Kole et al. (1985b), of the Allied Corporation (Morristown, New Jersey),
nominalized in their patent 45 aminophenols as inhibitors of soil urease activity.
Samples of a New York soil (Cazenovia silt loam, pH 7.0) and of a Wisconsin soil
(Plano silt loam, pH 5.4) were used for testing. The reaction mixtures, prepared from 20
g of air-dry soil, 0.8 mg of test compound in 5 ml water or only 5 rnl of water, and 42,8
mg of urea in I ml of water, were incubated at 25°C for 3 days, then analyzed for
'Positions ofthe OR and CH, groups are not specified in the paper.
81
determination of the remaining urea. According to the patent description, only eight
aminophenols were tested with the Cazenovia soil and only five of these eight
compounds were used for testing with the Plano soil.
In both soils, the strongest urease-inhibiting compounds were N-(4-hydroxyphenyl)
glycine and 4-(N-methylamino)phenol (Figure 26), both causing a 82% inhibition in the
Cazenovia soil and 61 and 55% inhibitions, respectively, in the Plano soil, whereas the
weakest inhibitor was 2-aminophenol, causing an 8% inhibition. But the degree of
inhibition by 4-aminophenol was 46% which is in contrast with the <1% inhibition
mentioned in Table 16.
H3C-H~H
N-(4-Hydroxyphenyl)glycine 4-(N-Methylami no)phenol
Figure 16. Structure of the strongest urease inhibitors among the aminophenols tested
by Kolc et al. (1985b).
The most preferred aminophenol amount recommended by the inventors to be

applied to soil is from about 1 to about 500 ppm relative to soil weight.
A thiophenol derivative, zinc mercaptide of 2-chloro-4-aminothiophenol (Figure 27)
did not inhibit urease activity in two soils studied by Kiss and Pintea (1987). The
reaction mixtures had the following composition: 5 g of air-dried soil + 10 ml of
Figure 17. Structure of zinc mercaptide of2-chloro-4-aminothiophenoI.
aqueous phase containing 6 mg of urea and 0 or 2% mercaptide relative to urea weight.

Incubation took place at laboratory temperature. Complete hydrolysis of urea needed, in
both absence and presence of mercaptide, 6 days (alluvial soil) and 9 days (leached
chemozem).
82
According to Shen et al. (1997), nitrophenor, added to samples of a Chinese soil at

rates of 50, 100, and 200 ppm (soil weight basis), caused 30.9, 44.5, and 68.2%
inhibitions, respectively, in soil urease activity. These data are in contrast with those in
Table 16.
2.16. POLYHYDRIC PHENOLS AND QUINONES
These two classes of organic compounds will be dealt with together, because they
inhibit soil urease activity by similar mechanisms, which was demonstrated in the case
(JcH
Q
OH OH
Or
OH OH
O-aH ~ H
Catechol Resorcinol Hydroquinone Pyrogallol
COOH COOH
4
OH
~H Q~
Phloroglucinol Protocatechuic acid Gallic acid
Figure 28. Structure of some polyhydric phenols.
of hydroquinone and p-benzoquinone by Mulvaney and Bremner (1978). The structure

of some of these compounds is presented in Figures 28 and 29. Their derivatives tested
will be specified during the descriptions of the investigations.
Comad (1940) studied the effect of two dihydric phenols (hydroquinone and
catechol) on urease activity in two California soils (a loam and a fine sandy loam). Soil
samples (1 kg) were moistened with about 350 ml of 0.5 M hydroquinone (HQ) or 0.2
M catechol (CT) solution, incubated under a toluene layer at 30°C for 3 days, then dried
at laboratory temperature. Urea solution was added to 75-g portions of the preincubated
soils and incubation continued under toluene for 7 days. Afterwards the residual urea
and NH4 + contents were analyzed. Samples of the clay soil that had been previously
'Position of the nitro group is not specified in the paper.

83
Q 0 CQ
0 0 0
p- or 1,4-Benzoquinone 0- or 1,2-Benzoquinone p- or 1,4-Naphthoqulnone 0- or 1,2-Naphthoqulnone
Quinhydrone Anthraquinone Phenanthrenequinone
2,6-Dichloroquinone-4-chloroimide
Figure 29_ Structure of some quinones_
heated to 85°C as well as untreated samples of both soils served for comparison. The
analyses showed that in the case of the clay soil urea hydrolysis was complete in 7 days
in the untreated samples. It was about 17% in the heated samples and had much lower
values in the HQ- or CT-treated samples; in the case of the other soil, urea hydrolysis
was nearly complete in the untreated samples and almost completely inhibited in the
treated ones.
Paulson and Kurtz (l969b) carried out a laboratory experiment with a silty clay
loam soil from Illinois. Reaction mixtures were prepared from 20 g of air-dry soil +
pelleted urea or pelleted urea-stearic acid complex (200 ppm N) + p-benzoquinone (BQ)
(0 or 0.2% relative to urea-N) + water to 30% moisture (about field capacity). After 1
and 7 days of incubation at 22°C, the mixtures were analyzed for residual urea, NiLJ +,
and (N03-+N02-). The inhibitory effect of BQ on hydrolysis of pelleted urea was
evident after 1 day of incubation but not after 7 days. When the reaction mixtures
contained pelleted urea-stearic acid complex and BQ, a very small amount of urea
remained unhydrolyzed after 7 days also.
Anderson (1969,1970). assignor to the Imperial Chemical Industries. London,
patented catechol. hydroquinone, and pyrogallol. p-benzoquinone and a series of its
derivatives, p- and o-naphthoquinone and one derivative of p-naphthoquinone (Figure
30) as inhibitors of soil urease activity. The inhibitors were added to urea at rates of
0.05-5%, preferably at rates of 0.1-2% by weight of urea; the inhibitor and urea were
84
p-Benzoquinone
Rl R2 Derivative
CI H chloro-
CI CI 2,5-dichloro-
Cl CI 2,6-dichloro-
CH 3 H methyl-
CH 3 CH, 2,5-dimethyl-
CH, CH,-CHz 2-methyl-5-ethyl-
CH3-CHz CH,-CHz 2,5-diethyl-
CHz-OH CHz-OH 2,s-dihydroxymethyJ-
O-CHz-CH 3 O-CHz-CH, 2,5-diethoxy -
S--CHz-COOH H carooxymethylthio-
C6 HS H phenyl-
C.H"NH-DC-CH3 H 3 '-N-acetylaminophenyl-
p-Naphthoquinone
R Derivative
HO 5-hydroxy-
Figure 30. p-Benzoquinone and p-naphthoquinone derivatives tested by Anderson (1969, 1970).
cogranulated. Use of inhibitor mixtures was also recommended. In these mixtures the
amount of polyhydric phenol used was generally between 10 and 300% molar of the
quinone present; conveniently, substantially equimolecular quantities may be used.
For evaluation of the inhibitory effect, 2.5-g soil samples were moistened with a 3%
urea solution to approximately one third of WHC. The urea solution did or did not
contain inhibitor(s) (40-50 ppm relative to soil weight). During incubation which took
place at laboratory temperature, the amount of volatile ammonia was assayed. In the
control soil (treated with urea alone), 50% of the initial urea-N amount was lost as
volatilized NH3 in 3 days. In the presence of inhibitors, total inhibition of volatilization
required a number of days beyond the control period of 3 days. In Table 18, which
presents the results obtained with four soils, the inhibition period indicates the number
of these days. It is evident from the data in this table that catechol and hydroquinone
acted synergistically with 2,5-dimethyl-p-benzoquinone.
85
TABLE 18. Effect of catechol, hydroquinone, and 2,5-dimethyl-p-benzoquinone and of their ruixtures on
volatilization ofammonia fiom urea-treated soils"
Inhibition period (days)
Concentration
Sandy Loamy Clay Calcareous
COflllound of compound
loam sand loam loam
(ppm of soil)
pH 6.0 pH 6.5 pH 7.1 pHS.5
Catechol ICT) 50 2-3 N.D. N.D. N.D.
Hydroquinone (HQ) 50 3 N.D. N.D. N.D.
2.5-Dimethyl-p-benzo-
quinone (DBQ) 50 7 N.D. N.D. N.D.
CT+HQ 25+25 5 N.D. N.D. N.D.
CT+DBQ 25+25 10 17 J 3-4
HQ+DBQ 25+25 10-12 17 3 2-3
CT+HQ+DBQ 25+25+25 14 20 4-5 4-5
"From Anderson (1969,1970).

N.D. - Not deterruined.
Evaluating the effect of seven polyhydric phenols and six quinooes, used at a rate of
50 ppm (soil weight basis), on the urease activity in three soils, Bremner and Douglas
(1971) obtained, by applying the 5-hour test, the results specified in Table 19. Of the
polyhydric phenols, only catechol and hydroquinone, and all of the quinooes except
TABLE 19. Effect of polyhydric phenols and Quinones on soil urease activity"
COflllound
Silty clay loam Loam Clay loam
Polyhydric phenols:
Catechol 77 71 73
Hydroquinone 69 63 60
Pyrogallol 6 5 2
"",0";'01 }
Phloroglucinol
Protocatechuic acid <4 <3 <2
Gallic acid
Quinones:
p-Benzoquil1one 68 61 58
2,5-Dichloro-p-benzoquinone 68 62 56
2,5-Dimethyl-p-benzoquinone 35 33 29
2,5-Dihydroxy-p-benzoquinone 7 6 6
2,6-Dichloro-p-benzoquinone 63 60 52
o-Naphthoquinone 48 42 42
a Adapted from Bremner and Douglas (1971). by permission of Pergamon Press PLC.
2,5-dihydroxy-p-benzoquinone, manifested a remarkable inhibitory effect on soil urease

activity. By increasing the amount of inhibitors from 50 to 300 ppm, the inhibition
increased but did not become total at the maximum amount, either. Table 20 presents
the results registered with the clay loam.
86
TABLE 20. Effect of different amounts of poly hydric phenols and quinones on soil urease activity"
COl1l'ound Amount of compound (ppm of soil)
50 100 200 300
Catechol 73 81 89 94
Hydroquinone 60 75 86 93
p-Benzoquinone 58 74 86 93
2,5-Dichloro-p-benzoquinone 56 75 81 87
2.6-Dichloro-p-benzoquinone 52 66 81 87
2,5-Dimethyl-p-benzoquinone 29 33 39 42
"Adapted from Bremner and Douglas (1971), by permission of Pergamon Press PLC.
The compounds specified in Table 19 and o-naphthoquinone lost, during

preincubation at 30°C for 3, 7, and 14 days, a considerable part of their inhibitory
capacity. The only exception was 2,5-dimethyl-p-benzoquinone, whose inhibitory effect
increased during preincubation. For example, in the clay loam the degree of the
inhibition caused by tbis compound after 0, 3, 7, and 14 days of preincubation was 29,
66,71, and 74%, respectively.
The inhibiting effects of hydroquinone, catechol, and p-benwquinone on urease
activity in the clay loam as well as the influence of their preincubation are also referred
to in a paper published by Douglas and Bremner (1971).
Bremner and Douglas (1973) studied the effects of catechol, hydroquinone, p-benzo-
quinone, and 2,5-dimethyl-p-benzoquinone on hydrolysis of urea and volatilization of
ammonia from urea in five soils. Soil samples (10 g) were treated with 1 ml of urea
solution (10 mgofN) + 1 ml of water or 1 ml of solution containing 0.5 mg ofinbibitor
+ water up to 50% of WHC, then incubated at 20°C. The amounts of residual urea,
exchangeable NH/, and volatilized NH3 were recorded after 3, 7, and 14 days of
incubation. The conclusion was drawn that the inhibitory effect manifested in the
decrease of both urea hydrolysis and NH3 volatilization. The effect was more evident in
light-textured than in heavy-textured soils. In these experiments 2,5-dimethyl-p-benw-
quinone was the most effective inhibitor, reducing NH3 losses from 61.1 to 0.3% in a
sandy soil treated with urea and incubated for 14 days.
Bundy and Bremner (1973a) evaluated the effect of 34 p-benzoquinone (BQ)
derivatives on urease activity in three Iowa soils. Each compound was used at a rate of
50 ppm by weight of soil. Results of the urease activity determination by the 5-hour test
are shown in Table 21. One can deduce from this table that the inhibitory effect of the
substituted BQs depended largely upon their substituent groups. Methyl-, chloro-,
bromo-, and fluoro-substituted BQs usually had a marked inhibitory effect on soil
urease activity, whereas phenyl-, (-butyl, and hydroxy-substituted BQs had very little if
any effect. The effect of methyl-, chloro-, bromo-, and isopropyl-methyl-substituted
BQs on soil urease activity was reduced or eliminated by the introduction of hydroxy
groups into these compounds. The effect of methoxy-substituted BQs depended on the
number and position of the methoxy groups. BQ was considerably more effective than
most of the substituted BQs. Only methyl- and chloro-substituted BQs gave more than
50% inhibition of soil urease activity.
87
TABLE 21. Etl'ect of various p-benzoquinones on soil urease activity'
Inhibition of urease activi!:!:: (%2

Sandy Clay Clay loam
Compound
loam loam pH7.3 Average
EH6.7 EH 7.6
p-Benzoquinone (BQ) 53 77 81 70
Methyl-BQ 52 61 48 54
2,3-Dimethyl-BQ 53 53 55 54
2,5-Dimethyl-BQ 30 27 26 28
2.6-Dimethyl-BQ 25 :1I 34 30
Trimethyl-BQ () () () ()
Tetramethyl-BQ 0 0 0 ()
2-Isopropyl-5-methyl-BQ (thymoquinone) 9 12 23 15
2,5-Di-t-butyl-BQ 0 0 0 0
2.6-Di-t-butyl-BQ 0 0 0 0
Phenyl-BQ 0 0 0 0
2.5-Diphenyl-BQ 0 0 0 0
Methoxy-BQ 35 32 19 29
2,5-Dimethoxy-BQ 0 0 0 0
2,6-Dimethoxy-BQ 10 8 12 10
Tetramethoxy-BQ 0 () 0 ()
2-Methoxy-3,5-dimethyl-BQ () 0 0 0
2-Methoxy-3.6-dimethyl-BQ () 0 0 0
2-Methoxy-trimethyl-BQ 0 0 0 0
2,5-Dichloro-BQ 69 70 67 69
2,6-Dichloro-BQ 61 68 54 61
Trichloro-BQ 44 44 37 42
Tetrachloro-BQ 50 44 51 48
2,3-Dichloro-5,6-dicyano-BQ 16 7 9 11
2,6-Dibromo-BQ 32 38 47 39
Tetrafluoro-BQ 44 36 33 35
2.5-Dibromo-3.6-dihydroxy-BQ 0 () () 0
2.5-Dichloro-3.6-dihydroxy-BQ 0 0 0 ()
2.5-Dihydroxy-BQ 0 2 3 2
Tetrahydroxy-BQ 0 () () 0
3-Hydroxy-2-isopropyl-5-methyl-BQ 0 0 () 0
6-Hydroxy-2-isopropyl-5-methyl-BQ 9 5 7 7
3.6-Dihydroxy-2-isopropyl-5-methyl-BQ 0 0 0 ()
3-Hydroxy-2,5-dimethyl-BQ 0 0 0 0
2-H~drox~-3.6-diEhen~I-BQ 0 0 0 0
"From Bundy and Bremner (1973a). by permission of Pergamon Press PLC.
In the case of the clay loam (PH 7.6), the effect of preincubation at 30°C for 7 and
14 days on the inhibitory capacity of these compounds added to 10-g soil samples at a
rate of 50 ppm was also studied. The 5-hour test was applied after preincubation. It was
found that preincubation usually led to a marked increase in the inhibitory capacity of
the methyl-substituted BQs and to a decrease in this capacity of the other BQs. A series
of BQs that were ineffective before preincubation remained ineffective after
preincubation. The inhibitory effect of dimethyl-BQs after 14 days of preincubation was
greater than that of the other BQs. Presence of a hydroxy group besides two methyl
groups eliminated the inhibitory capacity of the dimethyl-BQs. Thus, the inhibition
caused by 2,5-dimethyl-BQ increased from 27% (no preincubation) to 75% (after 7-day
88
preincubation) and to 82% (after 14-day preincubation); in contrast, the inhibitory effect
of 3-hydroxy-2,5-dimethyl-BQ remained zero after preincubation as well.
The increase in inhibiting capacity of methyl-substituted BQs during preincubation
is not attributed to their decomposition in soil with the concurrent formation of more
potent urease inhibitors, because the inhibitory effect of BQ (the only potential
decomposition product of methyl-substituted BQs shown to be an effective urease
inhibitor) was less after 7 or 14 days of preincubation than that of dimethyl-substituted
BQs. The alternative explanation is that the process whereby substituted BQs inactivate
soil urease is much slower with methyl-substituted BQs than with other substituted
BQs. This explanation is supported by the finding that the effect of methyl-substituted
BQs on soil urease activity after 3 hours of preincubation was similar to that observed
after 7 days of preincubation.
p-Benzoquinone and 10 of its derivatives were selected to study their effect on urea
hydrol ysis and ammonia volatilization from a sandy soil (pH 6.7) treated with urea. The
reaction mixtures (10 gofsoil + 10 mg N as urea + 0.5 mg of test compound + water to
50% of WHC) were incubated at 20°C for 14 days. The results of the determination of
NH3 volatilized during incubation and of residual urea and exchangeable ~ + in
incubated soil showed that the compounds that inhibited urease activity also reduced
the rate of urea hydrolysis and volatilization ofNH3 from urea. Thus, 2,3-dimethyl-BQ,
2,5-dimethyl-BQ and, 2,6-dimethyl-BQ reduced NH3 losses from 62.8 to 0.1 %. At the
same time, NH3 losses were reduced under the influence of 2 ,5-dihydroxy-BQ,
tetrahydroxy-BQ, and 2,5-diphenyl-BQ, from 62.8 only to 62.0, 62.6, and 59.4 %,
respectively.
The investigations performed by Bundy and Bremner (1973a) are also cited in detail
by Flaig (1976).
Applying the short and long time tests (see page 51), Hartbrich et al. (1976b) studied
the effect of p-benzoquinone (BQ) used at rates of 0.5, 1.0, 2.0, and 4.0% relative to
urea-N on the volatilization of ammonia from a heavy- and a light-textured soil, whose
30-g samples were treated with 100 mg ofurea-N. In both tests the inhibitory effect of
BQ on NH3 volatilization from urea increased with its amount and decreased during
incubation. For example, BQ at the four rates used caused 86, 100, 100, and 100%
inhibitions respectively, after 2-day incubation, 52, 61, 70, and 75% inhibitions
respectively, after 4 days, and 9, 19,20, and 21 % inhibitions respectively, after 8 days.
At day 10, the degree of inhibition became zero regardless of the BQ rate.
Thieme et al. (1976) dealt with the effect of hydroquinone (HQ), p-benzoquinone
(BQ), and quinhydrone (QH) and their mixtures with tetramethylthiuram disulfide
(thiram) on the volatilization of ammonia from urea-treated soil samples. Their rates
relative to urea-N were 2% (HQ) and 1 and 2% (BQ and QH). In mixtures the molar
ratio between thiram and the other compounds was 1:1 or 1:0.5 or 1:0.1, whereas the
rate of mixtures was 0.5 or 1% relative to urea-N (the rate may range from 0.01 to
10%). Incubation took place at 10°C. It resulted from the determination of the
volatilized NH3 that HQ, BQ, and QH were effective inhibitors. Thus after 4 and 12
days of incubation, they exhibited, when used at 2% rate, the following inhibitions: 99.2
and 22.6% (HQ), 97.2 and 14.8% (BQ), and 96.0 and 37.8% (QH), respectively. In
mixtures with thiram the inhibitory effect was more intense and long-lasting. For
example when thiram and the other compounds were used at 1:0.1 molar ratio, rate of
mixtures was 1% and incubation lasted 4 and 31 days, the following inhibitions were
89
registered: 100 and 52.5 1yo (HQ), 100 and 56.2% (BQ), and lOO and 68.8% (QH),
respectively.
The percent inhibitions caused by polyhydric phenols and quinones in urease
activity of a silt loam soil studied by Gould et al. (1978) and Gould (1979) are specified
in Table 22. The most effective inhibitors were p-benzoquinone and hydroquinone.
TABLE 22. Effect of polyhydric phenols and quinones on soil urease activity"
p-Benzoquinone 88
Hydroquinone 88
2.5-Dimethyl-p-benzoquinone 50
Catechol 47
2,6-Dimethyl-p-benzoquinone 45
2,6-Dichloroquinone-4-chloroimide 30
Tetrachloro-o-benzoquinone 6
Pyrogallol 2
Resorcinol 0
"From Gould et al. (1978) and Gould (l979). by permission of the Soil Science
Society of America. Inc.
Pyrogallol was a weak inhibitor, whereas resorcinol lacked inhibitory capacity. The
reaction mixtures consisted of 25 g of soil + 400 ppm of urea-N + 0 or 100 ppm of test
compound + water to field capacity and were incubated at 25°C for 24 hours.
In the experiments described by Hartbrich et al. (1978), three chloro-substituted
hydroquinones proved to be weaker inhibitors than was the parent compound. The most
effective inhibitor was 2,5-dimethyl-p-benzoquinone; its effect was evident for 14 days,
whereas the inhibitory effect of the parent compound lasted 8 days (Table 23) (see also
Oertel et at., 1978; lasche et al., 1978).
TABLE 23. Effect of polyhydric phenols and quinones on volatilization of ammonia from urea-
treated soils"
Inhibition (%)
STT LTT
Compound
Incubation time (dai:s2
2 4 8 12 14 16
Hydroquinone (HQ) 96 99 96 34 15 2 0
Chloro-HQ 93 91 69 12 0
2,5-Dichloro-HQ 69 49 17 0
Tetrachloro-HQ 72 45 15 0
p-Benzoquinone (BQ) 100 99 67 16 0
2.6-Dichloro-BQ 88 99 57 7 0
Methyl-BQ 95 96 55 16 6 0
2,5-Dimethyl-BQ 91 100 96 83 33 12 0
Tetramethyl-BQ 38 61 12 ()
Quinhydrone (QH) 96 53 12 4 0
Tetrachloro-QH 81 78 3~ 6 0
"From Hartbrich et al. (l978).
90
Urease activity in a sandy soil from Australia was assayed by May and Douglas
(1978) who used reaction mixtures prepared from 3 g of soil + 0.5 ml of toluene + 12 ml
of 1/15 M phosphate buffer, pH 8.8 + 3 ml of solution containing 3 mg of urea and a
TABLE 24. Inhibition of urease activity in a sandy soil by polyhydric phenols and quinones
applied at different concentrations'
Inhibition (%)
Compound Concentration of compound (ppm of soil)
\0 25 50 100
Catechol 74 85 93 100
p-Benzoquinone 75 91 92 97
2,5-Dimethyl-p-benzoquinone b 0 14 31 45
2,5- Dimethy l-p-benzoquinone' 82 90 92 93
Hydroquinone 45 64 87 96
o-Naphthoquinone 15 26 42 51
a Adaptedfrom May and Douglas (1978).

h Assayedwith no preincubation.
'Assayed after preincubation of soil + inhibitor at 3{)'C for 3 days.
test compound. No test compound was added to the control mixtures. Incubation took
place at 37°C and lasted 4 hours, after which the NH/ content was determined. The
compounds tested, their concentrations and the results obtained are presented in Table
24. One can see from this table that preincubation of 2,5-dimethyl-p-benzoquinone in
soil led to an increase in its inhibitory capacity. At higher concentrations (50 and 100
ppm), catechol, hydroquinone, p-benzoquinone, and preincubated 2,5-dimethyl-p-
benzoquinone manifested nearly the same inhibitory effect.
Similar results were obtained by Mulvaney and Bremner (1978) concerning
p-benzoquinone (BQ) and hydroquinone (HQ). They compared the inhibitory effect of
these two compounds on urea hydrolysis in 25 soils selected to obtain a wide range in
pH (4.6-8.0), texture (2-90% sand, 1-72% clay), organic C content (0.30-6.73%), urease
activity (4.7-84.9 Ilg of urea hydrolyzed/g soillhour at 37°C). and in other soil
properties. The reaction mixtures (5 g of air-dried soil + 2 ml of aqueous solution
containing 10 mg of urea or 10 mg of urea and 25 or 50 ~lg of BQ or HQ) were
incubated at 20°C for 24 hours, and then the residual urea was determined.
In each soil, the inhibition caused by BQ was almost identical to that brought about
by HQ. The degree of inhibition in air-dried and field-moist samples of the same soil
was the same. Simple correlation analyses showed that the inhibition of urea hydrol ysis
by BQ or HQ was very highly significantly (0.1 % level) correlated with organic C
content (r=-0.76), total N content (r=-0.74), urease activity (r=-0.70), and cation-
exchange capacity (r=-0.62), highly significantly (1 % level) correlated with sand
content (r=0.57) and significantly (5% level) with silt content (r=-0.42), clay content
(r=-0.49), and surface area (r=-0.49), but was not significantly correlated with pH
(r=0.36) or CaC03 equivalent (r=O.27). Multiple-regression analyses indicated that the
91
effectiveness of BQ and HQ for retardation of urea hydrolysis in soils tends to increase

with the decrease in soil organic matter content.
The inhibitory effect increased markedly with the amount of BQ or HQ applied (5-
50 ppm) and decreased markedly with incubation time (2-20 days) and with increase in
incubation temperature (l0-40DC).
Since the inhibitory effect of HQ was nearly identical to that of BQ and since HQ is
considerably less expensive than BQ, it was concluded that HQ has greater practical
potential than BQ as a soil urease inhibitor.
Mishra and Flaig (1979) evaluated the effect of six anthraquinones (anthraquinone,
1,2,4,5,8-pentahydroxy-, l-chloro-, and 2-methylanthraquinone, sodium 1,2-dihydroxy-
anthraquinone-3-sulfonate and disodium anthraquinone-l,5-disulfonate), two benzo-
quinones (2-isopropyl-5-methyl-p-benzoquinone and 4,6-di -t-butyl-a-benzoquinone),
two naphthoquinones (p-naphthoquinone and 2-methyl-p-naphthoquinone), two hydro-
quinones (2,3-dichloro- and 2,5-di-t-butylhydroquinone), and four catechols (3-methyl-,
4-methyl-, 4-t-butyl-, and 4,6-di-t-butylcatechol) on urease activity of two soils (brown
earth, pH 5.9 and black earth, pH 7.0) from Germany. Soil samples (200 g) were treated
with 5 ml of solution containing 10 mg of test compound (50 ppm relative to soil) + 5
ml of water or 5 ml of urea solution containing 20 mg of N (100 ppm) + water to 60%
of WHC. After 0, 14, and 28 days of incubation at 30DC, urease activity was estimated
by the method ofTabatabai and Bremner (1972).
The results indicated that the anthraquinones did not significantly inhibit urease
activity except 2-methylanthraquinone which inhibited it in the brown earth. Anthra-
quinone and l-chloroanthraquinone increased urease activity in both soils. 2-Isopropyl-
5-methyl-p-benzoquinone had an inhibitory effect only at zero time in the brown earth
and always in the black earth. The other benzoquinone constantly inhibited urease
activity in both soils. The two naphthoquinones also inhibited urease activity in both
soils, but in the brown earth the inhibition decreased with time. 2,3-Dichloro-
hydroquinone inhibited constantly, whereas 2,5-di-t-butylhydroquinone stimulated
urease activity in both soils. The four catechols had significant inhibitory effects in both
soils during the entire period of incubation.
Although urease activity was inhibited by some of the compounds tested, the total
contents ofNH/-N and N0 3--N after 14 and 28 days of incubation were similar in the
untreated samples and those treated with the test compounds (excepting 1-
chloroanthraquinone and 2,5-di-t-butylhydroquinone). The explanation of this finding is
that the residual urease activity made possible the complete hydrolysis of the small
amount of urea (100 ppm of N) in soil samples. But in respect of the ratio between
NH/-N and N0 3--N there were differences between samples treated with different
compounds due to the fact that some of the compounds (both benzoquinones and
naphthoquinones, 2,3-dichlorohydroquinone, 4-t-butyl- and 4,6-di-t-butylcatechol) also
inhibited nitrification, mostly in the brown earth. Two of the compounds manifesting a
stimulating effect on urease activity (l-chloroanthraquinone and 2,5-di-t-butylhydro-
quinone) also caused N losses.
Besides the 16 compounds specified above, Mishra and Flaig (1979) studied other
compounds, including six anthraquinones (l,2-dihydroxy-, 1,2,5,8-tetrahydroxy-, 1-
amino-2-chloroanthraquinone, anthraquinone-I-sulfonate, anthraquinone-2,6-disulfo-
nate, and l-aminoanthraquinone-2-sulfonate) which did not inhibit mineralization of
92
urea, and two polyhydric phenols (4,6-di-t-butylresorcinol and 4,6-di-t-butylpyrogallol)

which did not inhibit urease activity and nitrification.
Using the soils and methods specified above, Mishra et al. (1980) reexamined the
most effective compounds which, were applied at lower concentrations (10 and 20
ppm). The compounds studied again were: p-naphthoquinone, 2-methyl-p-naphtho-
quinone, 4,6-di -1-butyl-o-benzoquinone, 2,3-dichlorohydroquinone, 4-t-butyl- and 4,6-
di -t-butylcatechol.
At zero time the compounds at both rates inhibited urease activity in both soils,
except 4,6-di-t-butyl-o-benzoquinone which at 10 ppm concentration did not inhibit
urease activity in black earth. As expected the inhibition was more pronounced at 20
than at 10 ppm and, in general, was greater in brown earth than in black earth. During
incubation (7, 14, and 28 days) the inhibition decreased; maximum inhibitions occurred
after 14 days only with 4,6-di-t-butylcatechol. It should be emphasized that this
compound was the most effective urease inhibitor in both soils.
Each compound at both concentrations inhibited urea hydrolysis in both soils for 7
days. Maximum inhibitions of urea hydrolysis (50% in brown earth and 53% in black
earth) were caused by 4,6-di-t-butylcatechol, i.e.. by the most effective urease inhibitor.
After 14 days inhibition of urea hydrolysis diminished or disappeared, but nitrification
remained inhibited.
In another experiment. inhibitor-coated urea granules were used. For the coating,
500 mg of test compound was dissolved in 2 rnl of acetone, and I rnl of this solution
was added to 5.25 g of urea granules and mixed thoroughly. Acetone was then allowed
to evaporate at 70°C for 4 hours. The inhibitors in form of coatings on urea granules -
as compared to their solutions - inhibited more markedly both urease activity and urea
hydrolysis and nitrification, as well.
Hydroquinone (HQ) is a photosensitive compound. Therefore, prevention of its
photodegradation is a necessity for a fertilizer constituted of urea with added HQ. Hera
et al. (1980) recommended shadowing of HQ by addition of fmely powdered lignite to
the urea + HQ mixture. HQ and powdered lignite are used as (XJ.ual parts, in proportions
ranging from 1 to 5%, the preferred proportion being 2% relative to weight of urea. In
an experiment, 109 of urea granules were mixed with 0.2 g of HQ and 0.2 g of
powdered lignite. The mixture was used for preparing an aqueous suspension containing
0.6 mg of urea, 0.012 mg ofHQ and 0.012 mg oflignite/ml. Ten rn1 of this suspension
was added to 5 g of air-dried soil (a heavy-textured soil - leached chernozem and a
light-textured - alluvial soil). The control reaction mixtures contained soil + urea and
soil + urea + lignite. Incubation took place at laboratory temperature. Daily analysis of
residual urea showed that complete hydrolysis of urea in leached chernozem required 7
days in the absence ofHQ and 30 days in HQ-treated samples. The corresponding times
in the case of alluvial soil were 5 and 10 days, respectively.
Under experimental conditions identical to those described on page 77, Muller and
Forster (1980) also studied the effect of HQ, applied at a rate of 2% relative to urea-No
It was found that HQ inhibited urease activity and maintained a significant part of the
urea unhydrolyzed in all treatments.
In samples ofa clay loam soil, treated withp-benzoquinone (50 Ilg/g soil), inhibition
of urease activity was 41.6% after 2 days of incubation and 35.3% after 7 days (Tu,
1981a). In the same concentration, p-benzoquinone inhibited insignificantly the urease
93
activity in samples of an organic soil after 7 days of incubation, but had a significant
stimulating effect after 14 days (Tu, 1981 b).
Tan (1982) studied the effect of hydroquinone (HQ) and p-benzoquinone (BQ) on
hydrolysis of urea in two Malaysian soils (sandy soil, pH 4.6 and clay soil, pH 4.7) from
rubber (Hevea brasiliensis) plantations. Reaction mixtures were prepared from 50 g of
soil + 2 ml of solution containing 0, 0.5, 1.25 or 2.5 mg of HQ or BQ (i.e.. 0, 10, 25 or
50 J1.g ofinhibitor/g soil) + 5 ml of urea solution containing 20 mg ofN (i.e.. 400 J1.g of
urea-N/g soil) + water to 50% of WHC. After 1, 2, 4, 7, and 14 days of incubation at
32°C the residual urea was determined.
In the absence of HQ and BQ urea hydrolysis was complete in 7 days in the sandy
soil and in 2 days in the clay soil. Both HQ and BQ were equally effective in retarding
urea hydrolysis. The inhibitory effect increased with their rate and was more marked in
the sandy than in the clay soil. For example after 4 days of incubation, HQ and BQ, at
the 10, 25, and 50 J1.g/g soil rates applied, exhibited the following percent inhibitions: 87
and 85, 95 and 90, and 100 and 100%, respectively, in the sandy soil, and 0, 40, and
45%, respectively, in the clay soil. The inhibitory effect decreased with the time of
incubation. For example under the action of HQ and BQ at the rate of 50 J1.glg soil, the
degree of inhibition registered after 4, 7, and 14 days of incubation was 100, 98 (96),
and 95%, respectively, in the sandy soil, and 45, 10, and 5%, respectively, in the clay
soil.
Reddy and Mishra (1983) determined urease activity and volatilization of ammonia
from urea in an alkaline sandy loam (PH 8) from India. The soil samples were treated or
not with p-benzoquinone (BQ). Urea prills were surface-applied at a rate of 100 kg Nlha
with or without 1% BQ (relative to weight of urea). Urease activity measured at 30°C
and expressed as mg of hydrolyzed urea/kg soillhour decreased from 11.8 (soil treated
with urea alone) to 5.2 (soil treated with urea + BQ). Ammonia volatilization
commenced on the day urea was applied, but from urea+BQ-treated soil no such loss
was detected for the first 4 days. In the presence of BQ, cumulative losses of urea-N as
NH3 during 16 days decreased by 60%.
Gorelik et al. (1983) prepared urea granules coated with 1, 5 or 10% of
hydroquinone (HQ) relative to urea weight. HQ was dissolved in diethyl ether. The urea
granules were introduced into the solution, then diethyl ether was allowed to evaporate
at room temperature. Three soddy podzols (sandy, sandy-loamy, and loamy texture,
respectively) and two heavier-textured soils (chemozem and sierozem) were used.
Water was added to 10-g air-dried soil samples to 50% ofWHC, then uncoated or HQ-
coated urea granules were placed on the soil surface. Rate of urea application was 1 mg
of N/g soil. Incubation was carried out at 20°C. The amounts of volatilized ammonia
and residual urea were recorded after 3, 5, 7, and 14 days of incubation.
In the very urease-active sandy-loamy podzol and chemozern, all of the added urea
was hydrolyzed in 3 and 5 days, respectively, whereas in the presence of HQ complete
hydrolysis of urea required 7 days. In the other three soils, hydrolysis of urea was
slower, it became complete in 7 days in samples treated with urea alone and in 14 days
in samples treated with urea containing 1% HQ; in the presence of higher amounts of
HQ, complete urea hydrolysis needed more than 14 days. Thus after a 14-day
incubation, cumulative losses as volatile NH3 from urea alone or with 1 and 10% HQ
were the following (in percentages of the added urea-N):
94
sierozem (2, 2, and 1%) < chernozem (3,3, and 2%) < loamy podzol (10, 10, and
3%) < sandy-loamy podzol (51,49, and 44%) < sandy podzol (75, 67, and 50%).
It is evident that HQ inhibited more effectively urea hydrolysis than ammonia
volatilization.
Rodgers et at. (1984) studied the effect of hydroquinone (HQ) over three years
(1981-1983) in field experiments on a clay loam (in 1981) and on a silty clay loam (in
1982 and 1983), covered by ryegrass leys at Rothamsted. Urea prills were applied
annually at a rate of 375 kg N/ha as a single dressing or as three equally divided
dressings. HQ was used at an annual rate of 5 kg/ha as an addition to the single dressing
in each year or to the three dressings in 1982 and 1983. Analyses were carried out for
residual urea, volatilized NH1, NH4 +, and NO)-.
Complete hydrolysis of urea always occurred in 14 days in both the absence and
presence of HQ, but HQ reduced total ammonia volatilization losses during the 4 weeks
after urea application as a single dressing in 1981 and had little effect in 1982 and 1983.
Reduction of NH3 losses was substantial: from 17 to 8 kg of NHrN/ha. However, it
should be added that never more than 5% of the applied urea-N was lost by NH3
volatilization in any year, irrespective of treatment.
In similar field experiments conducted on a silty clay loam (PH 7.5-8.0) in 1983-
1984 and on another silty clay loam (PH 7.8-7.9) in 1984-1985, both soils having been
cropped with winter oil-seed rape, Rodgers et at. (1986) used urea priUs uncoated or
coated with HQ (HQ was dissolved in acetone, then the solution was poured over urea
prills; finally, acetone was allowed to evaporate at room temperature). Three days
before sowing in August 1983 and September 1984, respectively, the seed bed was not
or was fertilized with uncoated or HQ-coated urea priUs (50 kg of N and 3 kg of
HQ/ha). In March 1984 and 1985, some plots were fertilized with urea (150 kg of Niha)
without or with HQ (10 kg/ha), whereas the other plots were treated with half of these
amounts in February 1984 and 1985 and with the other half in March 1984 and 1985.
Urea, NH4 +, and N0 3- in soil were systematically analyzed in the February-June 1984
and 1985 periods. Ammonia volatilization was determined during the first 3 weeks after
spring fertilization. The plants were harvested in July 1984 and August 1985,
respecti vel y.
Urea hydrolysis during the first 2 weeks after spring fertili7..ation was inhibited by
HQ (10 kg/ha) (degree of inhibition: 50%), but in 3 weeks urea hydrolysis became
complete. HQ significantly reduced volatilization of NH3 from urea in all treatmens in
both years. It should however be emphasized that in these experiments even the highest
losses were less than 3% of the added urea-No
The inhibitors tested by Sen and Bandyopadhyay (1986) were catechol (CT) and p-
benzoquinone (BQ). The experimental field was on a coastal saline soil (PH 7.5) in
India and was cropped with rice under submerged conditions. Prilled urea at a rate of
100 kg of Niha without or with an inhibitor (25% of CT or BQ relative to urea-N) was
applied to 3-cm deep floodwater. In a treatment, urea at the same rate, but in the form of
3-g briquettes without any inhibitor was placed at 5-cm depth in the soil after draining
off the excess floodwater from the field. In each treatment, pH and NH/content in
floodwater and the amount of ammonia volatilized were determined at 2-day intervals
for 16-18 days following fertilization.
Under the influence of inhibitors, pH and NHJ volatilization were lower for 6 days,
whereas NH4 + concentration remained lower for 8 days. Cumulative NH3 losses during
95
18 days were reduced by 30.3% due to CT and by 28.6% due to BQ. However, the most
marked reduction (71.3%) was recorded in the treatment in which urea was placed at 5-
cm soil depth.
Abdel Magid and El Mahi (1986) studied the effect of p-benzoquinone (BQ) on urea
hydrolysis in five Sudan soils (three clays, a silty clay loam, and a sandy loam). Urea
granules (at a rate of 90 kg of N/ha) were placed on the surface of 1O-g soil samples
with or without 0.05 mg of BQ/g soil. After moistening soil samples up to 50% of WHC
and incubation at 35°C for 1,3, 7, and 14 days, the residual urea was estimated. It was
found that BQ inhibited urease activity in each soil. From the analytical data obtained
after 14 days of incubation the following inhibitions could be calculated: about 60% (in
the three clay soils), about 55% (in the silty clay loam), and 100% (in the sandy loam).
Preliminary data on these investigations were published by Abdel Magid et al. (1983)
who also tested catechol, but later this compound was abandoned because its urease-
inhibiting capacity was much weaker than that of BQ. Catechol, like BQ, was applied at
the rate of 0.05 mg/g soil, whereas the rate ofurea-N was 22.5 kg/ha.
One can see from Table 25 that hydroquinone (HQ) and chlorohydroquinone
prolonged complete hydrolysis of urea in a light-textured (alluvial) soil by 4 days. In a
TABLE 25. Effect of some poly hydric phenols and tetrachloro-p-benzoquinone on soil
urease activity"
Time required for complete hydrolysis of urea (days)

Compound
Alluvial soil Leached chemozem
Control 6 9
Hydroquinone 10 16
Resorcinol 8 12
Pyrogallol 8 12
Chlorohydroquinone 10 14
Tetrac hI oro-p- benzoguin on e 6 9
"Adapted from Kiss and Pintea (1987).
heavier-textured soil (leached chernozem), HQ acted more markedly than did chloro-
hydroquinone, with complete hydrolysis of urea prolonged by 7 and 5 days,
respectively. In both soils, resorcinol and pyrogallol were weak inhibitors, whereas
tetrachloro-p-benzoquinone did not manifest any inhibitory effect.The reaction mixtures
(5 g of air-dried soil + 10 ml of aqueous phase containing 6 mg of urea + 0 or 0.12 mg
of test compound) were incubated at laboratory temperature (Kiss and Pintea, 1987).
The Chinese inventors Cao et al. (1987) patented the technology of hydroquinone-
containing urea fertilizer production.
Xue and Li (1987) tested HQ and seven other polyhydric phenols and quinones
(catechol, resorcinol, gallic acid, p-benzoquinone, quinhydrone, anthraquinone, and
phenanthrenequinonc; see Figures 28 and 29) on urease activity of a paddy soil. The
reaction mixtures were prepared from 5 g of dry soil + 10 ml of a 10% urea solution
(1 % solution for p-benzoquinone and anthraquinone) containing (on soil weight basis)
0, 20, 40, 60, 80, and 100 ppm test compound (only 100 ppm, with gallic acid).
Incubation took place at 37°C and lasted 48 hours.
96
The degree of inhibition depended on the nature of the test compounds and, as
expected, increased with their concentration. Thus, at the 100 ppm concentration, the
inhibitions showed the following order: HQ (68.26%), quinhydrone (64.07%), p-
benzoquinone (58.87%), catechol (55.88%), phenanthrenequinone (44.71 %), gallic acid
(26.34%), resorcinol (19.36%), and anthraquinone (5.60%). The same decreasing order
was also observed at lower concentrations, HQ, quinhydrone and p-benzoquinone
always being the most effective inhibitors.
With these three compounds and with catechol (at 100 ppm), the inhibitory effect
was followed for 12 days. The inhibition decreased during this period but remained
evident even at day 12.
In the case of HQ (at 100 ppm), the influence of urea concentration (1-10%) and that
of incubation temperature (22, 37, and 45°C) were also studied. It was established that
the urease-inhibiting effect of HQ decreased with increasing urea concentration. In
respect of the influence of temperature, the following situation was observed:
inhibition at 22°C < inhibition at 37°C;::: inhibition at 45°C.
The data in Table 26 show that the inhibiting effect of HQ increased with the
increase in pH from 4.7 to 6.6, but remained practically unchanged in the pH zone of
6.6-8.0. HQ was used at a concentration of 100 ppm.
TABLE 26. Effect of hydro quinone on soil urease activity at different pH values"
Urease activity (mg NH3-N/g dry soil)
pH Inhibition (%)
Control soil Soil treated with hydroquinone ( 100 ppm)
4.7 2.15 1.90 11.60
5.5 2.45 1.66 32.24
6.2 3.25 1.79 44.95
6.6 4.54 2.01 55.72
7.0 4.79 2.07 56.78
7.5 4.42 2.09 52.71
s.o 4.79 2.07 56.78
"From Xue and Li (1987).
Zhou et 01. (1988) studied the influence of HQ concentration and incubation time on
urease activity in a brown soil from China. Soil samples (250 g) were treated with 10 ml
ofa 10% urea solution, HQ (0, 25, 50,100 or 200 ppm on soil weight basis) and water
to 65% of WHC, then incubated at 30°C. After I, 7, 15, 30,45, 60, and 75 days of
incubation, the residual urea was determined. The results presented in Figure 31 prove
that the inhibition increased with increasing HQ concentration and decreased during
incubation. This decrease was very marked at lower HQ concentrations and less marked
at higher ones. For example, the inhibitory effect of HQ at 25 ppm after I day of
incubation (about 62%) disappeared after 45 days; at 100 ppm the disappearance of the
inhibitory effect took 75 days.
97
70
60
10 ZO 3n 40 50 60 10 80
Figure 31. Inhibition of urease activity in a brown soil by different concentrations of

hydroquinone as a function of the incubation time./FrornZhou et ai. (\988)./
In a pot experiment carried out by Zhou et at. (1988), HQ added to urea, at rates of
10 and 20 mg of HQ to 0.9 g of urea-N/pot, reduced volatilization of urea-N as
ammonia, and when HQ was added at a higher rate (40 mg to 0.9 g ofurea-N/pot) NH3
volatilization remained at a level similar to that recorded in the control soil (treated with
urea alone). At the same time, the amount of residual urea-N increased with the increase
in HQ concentration.
Samples (100 g) of a soddy-podzolic soil and a calcareous chemozem were treated
with 20 mg ofurea-N + 0 or 5% hydroquinone (HQ) (relative to urea-N), moistened to
60% ofWHC and incubated at 28°C. After 6, 12,24,48, and 72 hours of incubation the
residual urea was determined. In the soddy-podzolic soil, the residual urea represented
25% (control sample) and 80% (HQ-treated sample) of the initial amount after 24 hours
of incubation, and 0 and 25%, respectively, after 48 hours. In the calcareous chernozem
no residual urea was detected in the control sample after 12 hours of incubation,
whereas in the HQ-treated samples after 12 and 24 hours a part (35 and 8%,
respectively) of the initial urea amount remained unhydrolyzed (Pisareva and Muravin,
1988; Pisareva, 1989).
Zhou el al. (1992) and Zhao el al. (1992a) compared the effect of HQ on urea
hydrolysis and ammonia volatilinltion in normal (nonfumigated) and fumigated samples
of a Chinese brown meadow soil (silty loam, pH 6.55). Fumigation was carried out with
chloroform at 30°C for 1 week. The soil samples (l00 g) were treated with 0 or 480.8
mg of urea and with 0, 0.96, 2.0, 4.81, and 9.61 mg of HQ. The mixtures were
moistened to 40% ofWHC and incubated at 30°C. Residual urea was determined after 3
and 10 days of incubation and NH/-N content and NH3 evolution were periodically
assessed during 88 days of incubation. All the experimental procedures with the
fumigated soil were performed under sterile conditions.
Urea hydrolysis in all normal and fumigated soil treatments was complete after 10
days of incubation. On day 3, urea hydrolysis in nomwl soil was inhibited by each HQ
rate (the degree of inhibition increased with increasing HQ rates, from about 20 to about
44%), but in the fumigated soil only the two higher rates of HQ were effective (degree
98
of inhibition: 21.5 and 74.0%). One can see that at the highest HQ rate, the degree of
inhibition was higher in the fumigated than in the normal soil.
In normal soil, NH/-N content during 0-10 days of incubation was higher in the
HQ-treated samples than in the untreated control, and the decreasing content was
positively related to HQ dosage. The same trend was maintained during the 10-25-day
period, but the differences between control and HQ treatments were very small. In the
25-88-day period, NH/-N content in the HQ treatments increased and exceeded that
measured in the control. The increase was positively related to HQ dosage. Ammonium-
N in the fumigated samples showed the same pattern as that of the normal soil, but its
amounts in all treatments were significantly higher than those found in the normal soil,
which was attributed to lack of microbial activity in the fumigated soil.
Volatile loss of NH3 during the first 3 days of incubation was higher in the control
than in the HQ-treated normal or fumigated soil samples. In most of the treatments the
highest loss occurred on day 6. After day 25, the loss became constant in all treatments
with only small amounts of volatile NH3 loss. Cumulative NH3 losses in 88 days were
higher in the normal soil than in the fumigated one. At the two higher rates of HQ,
cumulative losses were higher in the normal soil and lower in the fumigated soil than in
the control. Thus, the nonnal soil lost about 38 flg N/g soil from the control (urea-only
treatment) and about 50 flg N/g soil from the sample treated with urea and the highest
HQ rate. The corresponding values for the fumigated soil were about 29 and about 27
flg N/g soil. respectively.
Xu et at. (1994) preincubated soil samples treated with 0, 2.5, 5 or 15 flg HQ/g soil
for 45 days and determined their urease activity at 5-day intervals. Inhibition of urease
activity increased with the rate of HQ. In a slightly acid soil (PH 6.15), the inhibition
was maximum (- 50, 65, and 70%, respectively) on day 5, then decreased and became
negligibly low on day 45. In another, more acid soil (pH 5.58), the maximum
inhibitions (-45, 70, and 90%, respectively) were registered on day 10, then the
inhibition decreased and disappeared on day 30.
The effect of HQ on volatile losses of ammonia from urea-treated soil (pH 6.15) was
also studied, in four experimental variants: urea alone; urea + HQ; urea + wheat straw;
and urea + wheat straw + HQ. The rates of additions per 100 g of soil were: 230 mg
urea-N, 5 mg HQ, and I g straw. The volatile NH3 was measured during 30 days. The
cumulative NH3 losses in the four variants were the following: -50% (urea alone); 45%
(urea + straw); 40% (urea + straw + HQ); and 30% (urea + HQ). These data mean that
addition of straw reduced the inhibitory efficiency ofHQ.
Effect of polyphenol and quinone urease inhibitors on energy barriers of urease
activity in soils were dealt with by two research groups.
Tomar and MacKenzie (1984) investigated the effects of temperature, catechol (CT)
and p-benzoquinone (BQ) on urease activity in five Quebec soils. Reaction mixtures
were prepared from air-dry soil equivalent to 109 of oven-dry soil + 1 ml of water or 1
ml of aqueous solution containing 50 or 100 flg of CT or BQ/g soil + 1 ml of urea
solution (1 mg of N/g soil) + water to field capacity. After 3 days of incubation at 5, 10,
15,20, and 25°C, the mixtures were analyzed for residual urea, NH/, and N0 3-. The
results showed that CT and BQ reduced hydrolysis of urea in each soil. BQ was
consistently more effective than CT. The degree of inhibition increased significantly
with the amount of inhibitors applied and decreased significantly with increase in
temperature of incubation. The inhibition was strong in a loamy sand (poor in organic
99
matter) and very weak in two high clay soils (rich in organic matter) in which complete
hydrolysis of urea occurred in 3 days at 10 or 15°C and at higher temperatures.
Consequently, for these soils it was impossible to calculate thermodynamic values.
The temperature coefficient (QJO) had values ranging in general from about 1.50 to
2.60, the inhibitors manifesting a tendency to diminish the effect of temperature on urea
hydrolysis. The other thermodynamic values are presented in Table 27 from which one
can see that the inhibitors tended to increase energy of activit ion (Ea ), enthalpy of
activation (.JH\ and entropy of activation (-1S") of urease, excepting BQ in the first
soil, in which these values decreased. The effect of inhibitors was evidently influenced
TABLE 27. Energy of activation (K). enthalpy of activation (L1H'J. entropy of activation (L1S"). and free
energy of activation (L1n") of soil urease in absence and presence of catechol (eT) and p-benzoquinone
(BQ)"
Rate of
E" L1H" lIS" Llct
Soil Inhibitor inhibitor
(kcal/mo Ie) (kcalllllOle) (kcal/mole) (kcai/mole)
(I:!!l~ soi'2
0 9.2 8.6 -21.4 14.8
CT 50 10.2 9.6 -18.5 14.9
Silty clay 100 11.2 10.7 -15.2 15.0
loam 0 9.1 8.6 -21.5 14.8
BQ 50 7.7 7.1 -28.7 15.4
100 7.0 6.4 -31.5 15.5
0 6.6 6.0 -30.6 14.8
CT SO 7.S 7.0 -27.7 IS.O
100 8.S 7.0 -24.4 IS.O
Clay
0 6.9 6.3 -29.6 14.8
BQ 50 7.7 7.1 -28.8 15.4
100 7.9 7.4 -28.1 15.4
0 9.3 8.7 -22.0 15.1
CT 50 10.7 10.2 -18.1 15.4
Loamy 100 10.7 10.1 -18.5 15.5
sand 0 9.4 8.8 -21.6 15.1
BQ 50 15.8 15.2 -1.6 15.6
100 14.6 14.0 -5.8 15.7
"From Tomar and MacKenzie (1984).
by the nature of soil, but very little by their concentration. At the same time, free energy
of activation (JG*) of the urease activity had practically the same value in all soils in
both absence and presence of inhibitors, which suggests that the energetic requirement
of the urea-urease system is independent of the nature of the soils used.
The conclusion arrived at was that an increase in the activation energy of soil urease
as a result of inhibitor use is related to an increase in the effectiveness of the inhibitor.
In other words, determination of the energy of activation of urease in soil samples
treated with different compounds may prove useful in evaluating the relative
effectiveness of these compounds as soil urease inhibitors, the effectiveness being
indicated by increase in energy of activation.
In a similar study, Praveen-Kumar et al. (1990) used samples of an acidic, an
alkaline, and a saline soil from India. Eight polyphenol and quinone urease inhibitors,
namely catechol (CT), 4-methylcatechol (MeT), hydroquinone (HQ), p-benzoquinone
(BQ), p-naphthoquinone (NQ), 2-methyl-p-naphthoquinone (MNQ), 2,6-dichloro-
100
quinone-4-chloroimide (DC C), 2,6-dibromoquinone-4-chloroimide (DBC) as well as

phenylmercuric acetate (PMA) were tested. The reaction mixtures, consisting of 10 g of
air-dried soil + 0.5 ml of water or solution of inhibitor at a concentration equivalent to
0.125 or 0.375 ppm (on soil basis) + urea solution (100 ppm of N) + water to field
capacity, were incubated at 13,21,30,39,49, and 55°C for 4 hours, then analyzed for
unhydrolyzed urea and the analytical data were used for calculation of different
thermodynamic parameters.
The Arrhenius plot (i.e .. plot of urease activity versus temperature) was linear in the
acidic soil and nonlinear in the other two soils. In nonlinear plots, a break-up point
(inflexion) appeared at 300C (critical temperature) which can bc attributed to the sudden
change in conformation of enzyme in these soils. Two values of Ea in nonlinear plots
corresponding to each segment of the Arrhenius plot, one below and another above the
critical temperature, were observed. In the absence of inhibitors, Ea was higher in the
acidic than in the alkaline soil, but the highest value was recorded in the saline soil.
These values were considerably changed in the presence of inhibitors, but the basic
linear or nonlinear nature of the Arrhenius plot was not affected. In the acidic soil, all
inhibitors, excepting NQ and MNQ, increased the Ea value as compared with the
control. In the alkaline soil, only HQ, BQ, and PMA increased Ea below the critical
temperature, whereas above it all inhibitors, excepting NQ and MNQ, increased Ea. In
the saline soil, none of the inhibitors increased Ea below the critical temperature,
whereas MCT, DCC, DBC, and PMA increased Ea above the critical temperature.
Increasing the inhibitor concentration from 0.125 to 0.375 ppm resulted in no
change or in an increase in Ea values. Thus, in the alkaline and saline soils, Ea values
remained unchanged on increasing the concentration of all inhibitors below the critical
temperature, but above this temperature Ea increased with the increasing concentration
of HQ, BQ, DCC, DBC, and PMA, whereas Ea was the same at both concentrations of
CT, MCT, NQ, and MNQ. In the acidic soil, the influence of inhibitor concentration on
Ea was similar to that observed in the other two soils. These results show that the
structurally related inhibitors CT and MCT, HQ and BQ, DCC and DBC changed Ea in
a similar or comparable manner in all the soils.
In absence of inhibitors, with an increase in temperature from 21 to 39°C, the i1,'{'
values decreased slightly in the acidic soil, but increased considerably in the other two
soils. In the presence of inhibitors, ,d,,{, behaved like Ea.
Values of the free energy of activation were nearly constant irrespective of the
presence of inhibitors.
Simihaian and Silberg (1999) have found that HQ caused a stronger inhibition of
urease activity in an alluvial soil than in a leached chemozem; the inhibition constants
were 1.9 and 3.0 mM, respectively. The results indicated that the inhibition of urease
activity by HQ was competitive in both soils.
Mobility ofpolyhydric phenols and quinones in soils was also studied. The soil thin-
layer chromatography, applied by Praveen-Kumar et af. (1987) for studying mobility of
phenylmercuric acetate with three diverse soils (acidic, alkaline, and saline) was
described on page 45. These investigators used the same method and soils also for
studying mobility of eight polyhydric phenols and quinones. Three of them, p-
naphthoquinone, 2,6-dichloroquinone-4-chloroimide, and 2,6-dibromoquinone-4-
chloroimide were immobile (RFO), not being soluble in cold water. The other five
compounds had in the acidic: alkaline, and saline soils the following R f values:
101
hydroquinone 0.87, 1.00, and 0.88; catechol 0.80, 0.74, and 0.46; 4-methylcatechol
0.74, 0.73, and 0.83; p-benzoquinone 0.93, 0.93, and 0.90, and 2-methyl-p-
naphthoquinone 0.54, 0.54, and 0.57, respectively. Thus, hydroquinone and p-
benzoquinone were the most mobile compounds in each soil, whereas the least mobile
compounds were 2-methyl-p-naphthoquinone (in acidic and alkaline soils) and catechol
(in saline soil). Urca was very mobile (RF1.00) in each soil.
For inhibition of urease located at different depths in soil, it is a requirement for the
inhibitor to move along with urea. This is why hydroquinone and p-benzoquinone
appear more suitable as inhibitors of soil urease than the other six compounds tested.
Zhao et al. (1991) also found that hydroquinone is mobile in soil. In their
experiment, hydroquinone applied on the surface of a soil column moved down to the
30-60-cm layer.
Finally, it should be mentioned that Zhao et al. (1992b) elaborated methods for
determination of hydroquinone and biuret in hydroquinone-containing urea fertilizers.
2.17. THIOPYRIDINE-N-OXIDES, THIOPYRIDINES, AND THIOPYRIMIDINES
Some dimeric and monomeric compounds from these classes (Figure 32) were tested as
inhibitors of soil urease activity by Radel e( al. (1989). The testings were also described
in a patent by Radel and Crenshaw (1990) (Assignee: Tennessee Valley Authority,
Muscle Shoals, Alabama). Phenylphosphorodiamidate (PPDA) served as a reference
compound.
~_s-O
! !J
0," I
22'-Di1hiobis-pyridne-N-oxide
"
2-Mercaptopyndne-N-oxide
(DTPNO) (MPNO)
~ ~N'"
~_s-O ~>-s-s~)
2,2'-Dilhiobis-pyridine (Aldrithiol)
(An
2 -Mercaptopyndine 2-Mercapto-3-pyridnol
(MP) (MPOl)
~" 0-0
~ ~ 0 I
0
2-Mercaplopyrmiline 2,2··Dipyridl PYricine-N-oxide

(MPM) (OP) (PNO)
Figure 32. Structure of the compounds tested by Radel et al. (1989) and Radel and Crenshaw (1990)
for inhibition of soil urease activity.
102
A banded soil procedure was developed for the testings. The sample of a silt loam
soil was moistened to a water content of 20% (dry weight basis) and preincubated at
room temperature for 2 days. In the next step, plcxiglass containers (6x6x6 cm) were
one-half filled with the preincubated soil and packed to a bulk density of 1.1 glcm3 •
Urea or urea + test compound (thoroughly mixed) was distributed in a narrow band
approximately 6 cm long and about 0.5 cm wide on the surface of the soil (rates of
additions per band were 410 mg of urea and 41 mg of test compound). The containers
then were filled with soil and again packed to a bulk density of 1.1 glcm 3 , followed by
incubation at 25°C for 3 and 6 days, when the soil from each container was thoroughly
mixed and a 10-g subsample was extracted and analyzed for determination of the
unhydrolyzed urea. From the analytical data, the percent inhibition of soil urease
activity was calculated.
The results are summarized in Table 28. Two compounds (DTPNO and PPDA) were
compared in two test series.
TABLE 28. Effect of two thiopyridine·N-oxides. four thiopyridines. one

thiopyrimidine. and three other compounds on urease activity in a silty loam soil"
Inhibition (%)
Compound Incubation time (days)
6
DTPNO 92.2; 92.6 46.4; 38.9
MPNO 76.8 22.2
AT 42.9 n.7
DTNP 8.6 n.s
MP 80.2 37.8
MPOL 40.1 a
MPM 72.8 44.6
DP 12.0 0.2
PNO 11.3 n
PPDA 95.6; 92.1 59.6; 54.2
'Adapted from Radel and Crenshaw (1990). by permission of Tennessee Valley
Authority. Muscle Shoals. Alabama.
It is evident from this table that DTPNO was nearly as effective a soil urease
inhibitor as PPDA. Strong inhibitions were produced also by MPNO, MP, and MPM,
i.e .. by compounds containing S-S or SH group and pyridine or pyrimidine moiety,
whereas the weakest inhibitors were DP and PNO, in which no S is bound to the
pyridine moiety. These findings indicate that in the monomeric compounds the key
functional group responsible for the inhibition is the mercapto group and N-oxide
function adds little to the inhibitory power, whereas in the dimeric compounds the N-
oxide moiety increases the solubility and weakens the strength of the disulfide bond,
thus producing a more active inhibitor.
The most preferable rate, at which the inhibitors are recommended as additives to
fertilizer urea, ranges from about 0.5 to about 2% relative to weight of urea.
103
2.18. N,N' -DIHALO-2-IMIDAZOLIDINONES AND N-HALO-2-0XAZOLI-

DINONES
Gautney et at. (1990), assignors to Tennessee Yalley Authority, Muscle Shoals,

Alabama, patented these two classes of N-halamines as dual purpose, urease and
nitrification inhibitors. Four of these compounds (Figure 33) were tested for inhibition
of soil urease activity. Phenylphosphorodiamidate (PPDA) served as a reference
compound.
HaC, T'
HaC- C -
HaC_I
HaC ..... ' N /
'=0
N
I
CI
l-Bromo-3-chloro-4.4.5,5-tetramethyl- 1,3-Dibromo-4.4,5.5-tetramethyl-
2-imidazolidinone (ABC) 2-imidazolidinone (AB)
3-Chloro-4,4-dimethyl- 3-Bromo-4.4-dimethyl-
2-oxazolidinone (9 2-oxazolidinone (8)
Figure 33. Structure of the N-halamine compounds tested by Gautney et al. (I990) for
inhibition of soil urease activity.
The same silt loam soil and banded soil procedure were used for testing as those also
applied by other collaborators of Tennessee Yalley Authority (see the preceding
subchapter). Rates of additions per soil band were 410 mg of urea and 0 or 41 mg of test
compound. The incubation was carried out at 25°C for 3 and 6 days, then the
unhydrolyzed urea was determined.
The amounts of unhydrolyzed urea were expressed as percentages of the added urea
amount. The values recorded after 3- and 6-day incubations in the different treatments
were the following: 4.9 and 1.0% (urea-only treatment); 84.7 and 53.7% (ABC); 70.6
and 15.0% (AB); 82.4 and 56.5% (I); 65.5 and 10.0% (IB); and 94.3 and 58.7%
(PPDA), respectively. In other words, the urease-inhibiting capacity increased in the
order:
IB < AB < I;::: ABC < PPDA.
The effectiveness of ABC and 1 as inhibitors of soil urease activity approached that
ofPPDA.
For use in practice, the N-halamine compounds are recommended in the most
preferred amounts of 0.5-2% relative to weight of urea.
104
2.19. y-L-GLUTAMYL NITROANILIDES
The m-nitroacetanilide and 0-, m-, and p-nitroanilines, together with other (13)
compounds, all patented as nitrification inhibitors by different companies, were studied
by Bremner and Douglas (1971) for evaluation of their effect on urease activity in three
Iowa soils. The 5-hour test was applied and it was found that the urease-inhibiting effect
of m-nitroacetanilide like that of the three nitroanilines was very weak: less than 1%.
Simihiiian et al. (1992) studied three y-L-glutamyl nitroanilides (Figure 34) to
evaluate their effect on urease activity in a heavy-textured soil (leached chernozem) and
a light-textured (alluvial) soil. The free nitroanilines and aniline were also tested, and
hydroquinone served as a reference compound.
COOH COOH
H2N-!H H2N-!H
I
CH2
I
CH2
!H2 N02 !H2
bO_HN-O b O - H N - 0 - - N02
y-L-Glutamyl m-nitroanilide y-L-Glutamyl p-nitroanilide y-L-Glutamyl 2-l1lethoxy-p-nitroanilide
Figure 34. Structure of the y-L-glutal1lyl nitroanilides tested by Sil1lihaian et al. (1992) for inhibition of soil
urease activity.
The reaction mixtures had the following composition: 5 g of air-dried soil + 2 ml of

toluene + 1 ml of 0.6% urea solution + 9 ml of distilled water or 9 ml of aqueous
solution of glutamyl nitroanilide or hydroquinone at 2% rate by weight of urea or 9 ml
of aqueous solution of free nitroaniline or aniline in an amount equimolecular to that of
glutamyl nitroanilide. Reaction mixtures without soil were also prepared. The
incubation took place at laboratory temperature. At 1-2-day intervals, the aqueous phase
TABLE 29. Etfect ofy-L-glutal1lylnitroanilides and free nitroanilines on soil urease activity
as compared with the etlect of aniline and hydro quinone"
Time necessary for complete
Compound
hydrolysis of urea (days)
No (control) 11
y-L-Glutamyl m-nitroanilide 18
m-Nitroaniline 15
y-L-Glutamyl p-nitroanilide 18
p-Nitroaniline 15
y-L-GlutamyI2-l1lethoxy-p-nitroanilide 18
2-Methoxy-p-nitroaniline 15
Aniline 25
Hydroquinone 32
"From Simihiiian et al. tl992).
105
of the reaction mixtures was analyzed by means of a chromogenic reagent (see the
footnote on page 35) for detecting the unhydrolyzed urea. The time (days) necessary for
complete hydrolysis of urea was registered. Prolongation of this time, relative to
control, indicates inhibition of soil urease activity.
Surprisingly, the two soils used behaved similarly, in this experiment, in regards to
their urease activity. Therefore, the data presented in Table 29 are valid for both soils.
One can see from this table that each compound tested had an inhibitory effect on
soil urease activity. Hydroquinone was the strongest inhibitor prolonging the time
necessary for complete urea hydrolysis from II to 32 days. The three y-L-glutamyl
nitroanilides tested brought about a 7-day delay in complete urea hydrolysis. The delay
caused by equimolecular amounts of the free nitroanilines was shorter, lasting only 4
days. This means that the glutamyl moiety in glutamyl nitroanilides enhanced the
urease-inhibiting effect of nitroanilines. The inhibitory effect of aniline exceeded that of
nitroanilines. Consequently, the presence of nitro group or of nitro and methoxy groups
in nitroanilines attenuated the inhibitory effect of their aniline moiety.
This experiment was also referred to by Simihlhan et al. (1999).
2.20. PHOSPHORO(MONO)AMIDATES, PHOSPHORODIAMIDATES, AND THIO-

PHOSPHORODIAMIDATES
The phosphoroamides constitute a very important class of the inhibitors of soil urease
activity. They are derivatives ofphosphoro(mono)amidic acid [(mono)amidophosphoric
acid]; phosphorodiamidic acid (diamidophosphoric acid); thiophosphorodiamidic acid
(diamidothiophosphoric acid); phosphoryl triamide and thiophosphoryl triamide (Figure
35).
o s
II/NH2 II/NH2
Ho--P, Ho--P,
NH2 NH2
Phosphoroamidic acid Phosphorodiamidic acid Thiophosphorodiamidic acid
s
II/NH2
H~-P
'NH2
Phosphoryl triamide Thiophosphoryl triamide
Figure 35. Basal structure of phosphoroamides.
One of tl}c most widely studied phosphoroamidcs is phenylphosphorodiamidate

(PPDA; phosphorodiamidic acid phenyl ester; diamidophosphoric acid phenyl ester). A
106
comprehensive bibliography on this compound was compiled in 1981 (Anonymous,

1981) (Figure 36).
Figure 36. Structure of phenyl phosphorodi amidate (PPDA).
2.20.1. The Patented Compounds and the First Studies on Their Inhibitory ~tfect on
Soil Urease Activity
Phosphoro(mono)arnides and phosphorodiamides as inhibitors of soil urease activity
were patented by the German investigators Hartbrich et al. (1976b), Held et al. (1976b),
and Lang et al. (1976). They studied the compounds presented in Figure 37. These
compounds were recommended to be added to urea at rates ranging from 0.01 to 50%,
the preferred rates being ()'05-5% relative to urea-N, for fertilization of light- and
medium-textured soils as well as of grasslands.
RI Ro R, R. Derivative
CH, Nih H H phenylposphorodiamidate (phenyl-PDA)
2-Cl-C,JI, Nih H H 2-chlorophenyl-PDA
4-CI-C,H4 NH, H H 4-chlorophenyl-PDA
(',H, NH-CH, CH, H phenyl-N.N·-dimethyl-PDA
J-CII,-C,H. NH-CH, CH, II m-cresyl-N,N'-dimethyl-PDA
4-CI-C,H. NH-CH, CH, H 4-chlorophenyl-N,N' -dimethyl-PDA
C,H, NH-C,H, C,H, H phenyl-N,N'-di-n-propyl-PDA
4-C1-C.H4 NH-C,H, C,H 7 H 4-chlorophenyl-N,N' -di-n-propyl-PDA
e.H5 NH-CH(CH,), CH(CH,h H phenyl-N,N'-diisopropyl-PDA
4-Cl-C,H4 NH-CH(CH 3 )o CH(CH,h H 4-chlorophenyl-N,N'-diisopropyl-PDA
C,H 5 NH-n-C4H9 n-C.H9 H phenyl-N,N'-di-n-butyl-PDA
4-CI-C,1I4 NH-n-C 4 H9 n-C4 H 9 H 4-chlorophenyl-N,N'-di-n-butyl-PDA
4-CI-C.H4 NH-C,H, C,H, H 4-chlorophenyl-N,N'-diphenyl-PDA
C,H, NH-CHo-C.H< CH,-C,H, II phenyl-N,N'-dibenzyl-PDA
C,H5 NH-C,H, C,H, H ethyl-N,N'-diphenyl-PDA
C,H, O-C,H, H H diphenylphoshoroamidate (diphenyl-PA)
4-CI-C.H4 0-4-CI-C .H 4 H H di-(4-chlorophenyl)-PA
C,H, O-C,H, CH, H diphenyl-N-methyl-PA
C 9H I9 0-C9HJ9 C,Hs C,Hs dinonyl-N ,N' -diethyl-PA
Figure 37. General structural formula of compounds patented and tested by Hartbrich et al. (1976b), Held et
al. (1976b), and Lang e/ al. (1976), with specitication of some derivatives.
107
By applying the short and long time tests (see page 51)*, Held et a1. (1976b)
obtained the results given in Tables 30 and 31. They show that both compounds
inhibited urease activity in soil at both 30 and 10°C; the inhibitory effect increased with
TABLE 30. EfJect of different concentrations of two phosphoroamides on volatilization of ammonia

from urea-treated soil samples incubated at l00C for 24 hours"
Inhibition (%)
Compound Concentration of compound (% relative to urea-N)
D.D5 D.I D.2 D.l D.4 D.5 1.0 2.0 4.0
4-Chlorophenyl-
pbosphoro- 46.9 68.8 79.1 86.0 86.0 94.1 95.5 100 100
diamidate
Diphenyl-
phospboro- 29.7 35.9 40.6 48.3 57.8 80.0 78.5 86.2 93.8
ami date
"From Held et al. (l976b).
inhibitor concentration and decreased or disappeared with prolongation of incubation

time; 4-chlorophenylphoshorodiarnidate (4-chloro-PPDA) was more effective than was
diphenylphosphoroarnidate (di-PPA).
TABLE 31. Effect of difTerent concentrations of two phosphoroamides on volatilization of ammonia

from urea-treated soil samples incubated at IODC for 2-24 days"
Inhibition (%)
Concentration
of compound
Compound Incubation time (days)
(% relative to
urea-N)
2 4 12 16 20 24
2 IDO 100 100 100 98.1 89.1 32.8

4-Cblorophenyl-
100 100 100 100 87.9 57.5 23.6
phosphoro-
0.5 100 100 100 100 64.4 32.5
diamidate
0.1 100 100 85.4 49.2 2D.O 10.6
2 100 100 100 94.4 39.5 13.0
Oiphenyl- 100 100 99.1 85.6 42.0 25.3
phosphoro- 0.5 IO() 100 98.8 84.6 30.8 12.3
ami date 0.1 100 68.2 12.9
"From Held et al. (1976b).
Lang et al. (1976) found, by applying the long time test, that phenylphosphoro-
diamidate (PPDA) was more effective than were 4-chloro-PPDA and di-PPA.
Held et af. (1978) norninalized63 phosphoroarnides and specified the percent
inhibitions registered in soil urease activity under the influence of these compounds
which comprise: phosphorodiamidic acid, 5 alkyl esters, 14 phenyl esters, and 2
'In the long time test, incubation was carried out not at 20. but at lOOC.
108
naphthyl esters of phosphorodiamidic acid, 22 N-alkyl derivatives of phosphoro-

diamidic acid phenyl esters, 6 diphenyl esters of phosphoroamidic acid, and 13 phenyl
esters ofthiophosphorodiamidic acid (Table 32). The inhibitory effect was studied with
the short time test; the test compound" were used at a rate of 4% relative to urea-No
TABLE 32. Effect of 63 phosphoroamides on volatilization of ammonia from urea-treated soil samples
incubated at 30°C for 24 hours"
Substituents Inhibition
COlI1'ounds Structural formula
X RorR' (%)
2 3 4 5
Pho!!horodiamidic acid HO-~O~(NH2l2 20
Phosphorodiamidic acid alkyl R-O-P(O)(NH2)2 CH, 38
esters CzHs 5
n-C,H7 3
n-C4 H9 23
n-C 6 H 13 10
Phosphorodiamidic acid phenyl X-C_H.-O-P(O)(NH2h H 100
esters 2-CI 95
3-CI 93
4-CI 100
4-Br 93
2-CH, 96
3-CH, 90
4-CH, 93
2-0CH, 99
4-COOC,H7(n) 94
3-N02 87
4-NO z 55
2-CH,-4-CI 93
2-CH,-6-CI 93
Phosphorodiamidic acid n- C JOH7-n-O-P(O)(NH2)2 93
nal!hth~1 and ~-nal!hth~1 esters ClOH7-~-O-P(Ol~H2l2 96
N-A1kyl derivatives of X-C6 H.-O-(O)(NHR·h H CH, 42
phosphorodiamidic acid phenyl H C2Hs 34
esters H n-C,H7 30
H i-C,H7 42
H n-C4 H9 34
H C6H U 0
H CH2-C6 H, 0
2-CI CH, 0
4-CI CH, 33
4-CI CzHs 23
4-CI n-C,H7 27
4-CI i-C,H7 19
4-CI n-C.H9 14
2A-CI CH, 0
2,5-CI CH, 25
2-CH, CH, 44
3-CH, CH, 26
4-CH, CH, 24
2-CH,-4-CI CH, 29
2-CH,-6-C1 CH, 71
3-NO z ClI, 32
2-0ClI, CH, 24
lO9
TABLE 32. - continued -

2 3 4 5
Phosphorodiarnidic (X -C.H.-O )2P( O)NH2 H 91
acid diphenyl esters 2-CI 81
4-CI 20
2-CH3 31
3-CH3 24
4-CH3 49
Thiophosphorodiamidic H 92
acid phenyl esters 2-CI 89
3-CI 98
4-CI 83
4-Br 95
2-CH3 90
3-CH3 94
4-CH3 94
2-OCH, 89
2-CH,-4-CI 92
2-CH3 -6-CI 46
3-CH3-4-CI 93
2,4-CI 77
"Adapted from Held et al. (1978).
One can see from this table that phosphorodiamidic acid and its alkyl esters are
weak inhibitors, with their degrees of inhibition being between 3 and 38%. At the same
time, the phenyl esters (excepting the 4-N02 derivative) and the naphthyl esters of
phosphorodiarnidic acid are characterized by a strong inhibitory capacity (inhibition
degree: 87-100%). The strongest inhibitors are PPDA and 4-CI-PPDA (inhibition
degree: 100%). The ~-naphthyl ester ofphosphorodiamidic acid is a little more effective
than its a-naphthyl ester (inhibition degree: 96 and 93%, respectively). Alkylation of
NH2 groups leads to diminution or even to disappearance of the inhibitory capacity of
phosphorodiamidic acid phenyl esters. Introduction of substituents to phosphoro-
dilimidic acid phenyl esters also determines diminution of inhibitory capacity. Phenyl
esters of thiophosphorodiarnidic acid are nearly as effective as those of phospho-
rodiarnidic acid, excepting 2-CH3-6-Cl-phenylthiophosphorodiarnidate which is a weak
inhibitor when compared to 2-CH3-6-CI-PPDA.
PPDA was also found to be the most effective inhibitor in the long time test, with
1% PPDA relative to urea-N; incubation temperature: 20°C. Thus, the inhibition caused
by PPDA was 100% for 7 days, 50% after about 10 days, and 25% after about 12 days
of incubation (Held et aI., 1978). Other data published by Held et af. (1978) and
Hartbrich et af. (1978) indicate even stronger inhibitions, e.g. a 33% inhibition by 1%
PPDA after 12 days of incubation at 20°C. After the same time period of incubation at
the same temperature and PPDA concentration, Oertel et af. (1978) and Jasche et af.
(1978) registered a 94% inhibition.
These results are in agreement with those of Heiseler et af. (1980), who applied the
long time test and 1% inhibitor on urea-N basis and found that after 13 days of
incubation at 15°C urea hydrolysis was inhibited by the seven phosphorodiarnidate
(PDA) and phosphoroamidate (PA) compounds tested, in the following proportions:
PPDA (99.6%), o<resyl-PDA (59.6%), phenylthio-PDA (58.7%), di-p<resyl-PA
(15.6%), di-phenyl-PA (13.5%), di-o<resyl-PA (0%), and ethyl-PDA (0%).
110
Concerning duration of the inhibitory effect, PPDA and its chIoro derivatives and
those having a substituent in the same position (4) presented the following orders:
PPDA > 4-Cl-PPDA > 3-Cl-PPDA > 2-Cl-PPDA, and
PPDA > 4-CH 3-PPDA > 4-CI-PPDA > 4-COOC3H7 (n)-PPDA;:::: 4-Br-PPDA > 4-
NOrPPDA, respectively.
It was also established that duration of the inhibitory effect of PPDA (used at
concentrations of 0.25, 0.5, and 1% relative to urea-N) decreased with increasing
incubation temperature (from 10 to 15, 20, and 30DC). For example, PPDA, at 1%
concentration, completely inhibited volatilization of ammonia from urea for 16 days at
lODC, for 8 days at 15 DC, for 4 days at 20DC, and only for 2 days at 30 DC. At the same
concentration, PPDA was less effective at constant 20 DC temperature than under natural
conditions where the temperature varied between 10 DC (by night) and 28 DC (maximum
temperature by day) (Held et al., 1978).
TIle inhibitor and enzyme substrate, i.e.. PPDA and urea contain a similar amide
stmcture (Figure 38), which allows the attachment of PPDA to the active site of
enzyme. The hydrophilic group, O=P(NH 2h determines the recognition of receptor and,
thus, the specificity of inhibition, whereas the hydrophobic aryl residue (phenyl ring)
binds PPDA to the receptive site of the enzyme and, consequently, is decisive for the
(> 0
Figure 38. Comparison of the structure ofPPDA with that of the urea.
strength of inhibition. The inhibitor is bound with a higher affinity than urea, because
inhibition occurs even with an excess of urea. Derivatives of PPDA also act as tight-
binding inhibitors (Barth et al., 1978, 1980).
Jasche et al. (1978) and Oertel et at. (1978) underline the advantageous properties of
PPDA. PPDA has a remarkable urease-inhibiting effect at low concentrations (0.5-1 %
relative to urea-N). PPDA is not toxic for man. In soil, PPDA is degraded to nontoxic
products.
PPDA can easily be synthesized from phenol, phosphoms oxychloride, and
ammonia on a technical scale (Jasche et al., 1977, 1978; Oertel et al., 1978; see also
Kurze and Richter, 1978; Heiseler et al., 1980; Anonymous, 1985a).
PPDA is soluble in ethanol, methanol, acetone, but its solubility in water is low
(0.6643 gllOO ml of water at 25 D C) (Oertel et al., 1978).
For detennination of PPDA in urea fertilizers, soil extracts and other solutions,
colorimetric methods are available (Wenzel et al., 1981; Martens and Brenmer, 1983a).
A method based on high-performance liquid chromatography for analysis of PPDA was
also developed (Austin et al., 1984).
111
Elbe et al. (1979) patented a procedure for coating urea priUs with PPDA by means
of a mineral oil-bitumen mixture used as anticaking agent. The following example is
described in the patent: 25 kg of urea priUs are blended at 20°C, with 75 g of mineral
oil-bitumen mixture (80% of oil + 20% of bitumen), then 116 g of finely ground PPDA
(diameter of particles: 0.1 mm) is added and mixed again. The conditioned urea
fertilizer prepared contains 0.46% ofPPDA.
Kurze (1981) performed laboratory experiments, in which PPDA was used with
inhibitors more soluble in water: diammonium and calcium salts of imidodiphosphoric
acid diphenyl ester (Figure 39). Thus, solubility of the diammonium salt in water at
25°C is 46 ~100 mI, whereas that ofPPDA, as mentioned above, is only 0.6643 ~100
m!.
0-
o o
o- O-~-ONH4 _ 0-,-0\
II
- I NH NH Ca
0-
J 0 -1i - ONH4 O-o-l-!
Figure 39. Structure of diammonium and calcium salts of imidodiphosphoric acid diphenyl ester tested by
Kurze (198\).
Under the conditions of the long time test, these compounds, used at a rate of 1%
relative to urea-N, inhibited volatilization of ammonia from urea-treated soil samples to
a lesser extent than did PPDA, but acted synergistically with PPDA. For example, in an
experiment the mixture of PPDA + diammonium salt of imidodiphosphoric acid
diphenyl ester (at a molar ratio of 1: 1.5) produced a nearly complete (96%) inhibition of
the volatilization ofNH 3 from urea-treated soil samples during their 14-day incubation.
Kurze et al. (1985) evaluated, using the long time test, the inhibitory capacity of
seven heavy metal salts and the Al salt of the imidodiphosphoric acid diphenyl ester
(IPP) and of two heavy metal salts of the amidophosphoric acid monophenyl ester
(APP) on the volatilization of ammonia from urea-treated soil samples incubated for 6-
16 days. These compounds are more soluble in water than PPDA. The ~ salts of IPP
and APP served for comparison. The results obtained are reproduced in Table 33.
One can see from this table that the heavy metal salts (excepting the Fe salt) and the
Al salt of IPP were more inhibitory than was the NH4 salt. Most inhibitory were the Hg,
Cu, and Ni salts.
Cu salt of APP was more inhibitory but the Ni salt was less inhibitory than the ~
salt.
The Cu salt of APP was a stronger inhibitor than the Cu salt ofIPP, but the reverse
was true for the Ni salts.
112
TABLE 33. Effect of salts of imidodiphosphoric acid diphenyl ester (IPP) and amidophosphoric acid
monophenyl ester (APP) on volatilization of ammonia from urea-treated soil samples incubated at
200C for 6-16 da:z:s·
Inhibition (%l
Salts Incubation time (days)
6(7l 9 11(12 l 14(15l 16
Salts ofIPP
Hg 100 99.7 (99.2) 97.4 96.4
Cu 98 90 82 68 47
Ni 98 86 75 55 44
Co 94 74 61 39 17
Mn 98 81 (50) 29 17
Zn 83 36 (9.9) 6.9 4.6
Fe 74 22 (16) 4.6 1.9
Al 80 30 (16) 7.7 6.9
Nil. 72 10.4 (9.2) S.4 2.4
Salts ofAPP
Cu 100 99.5 98.6 93.4 70
Ni 96 56 36 27 8.8
NH. 98 88 51 32 10.8
·Adapted from Kurze et al. (1985).
Swerdloff et at. (1984, 1985a) and Anello et al. (1985), of the Allied Corporation
(Morristown, New Jersey), patented a great number of phosphorodiamidates and
thiophosphorodiamidates as inhibitors of soil urease activity and tested some of them
with samples of one or two soils. The soils and the testing method were the same as
those used by Kole et al. (1985b) (see page 80). Briefly, 20-g of air-dry samples of a
New York soil (Cazenovia sandy loam, pH 7.2) or a Wisconsin soil (Plano silt loam, pH
5.4) + 0.8 mg of test compound in 5 rnl of water or only 5 rnl of water + 42.8 mg of urea
in 1 ml of water were incubated at 25°C for 3 days, then analyzed for remaining urea.
Swerdloff et al. (1984) and Anello et al. (1985) patented 88 phosphorodiamidates
(PDAs) and 18 thiophosphorodiamidates (TPDAs), but testing of only 12 PDAs
(Cazenovia soil) and 6 PDAs (Plano soil) is described in the patents. The results
obtained are presented in Table 34.
The table shows that in both soils the most inhibitory PDAs were 2,2,2-
trifluoroethyl-PDA and 2,2,2-trichloroethyl-PDA, and the weakest inhibitors were
TABLE 34. Inhibition of soil urease activity by phosphorodiamidate compounds, R-O-P(O)(NH2 l2"
Inhibition (%)
R
Cazenovia soil Plano soil
2,2,2-Trifluoroethyl 97 90
2,2,2-Trichloroethyl 97 87
2-BrolJX)ethyl 85
Cyclohexyl 85
2-Chloroethyl 84
1,3-Dichloro-2-propyl 72 49
Benzyl 69 20
Allyl 65 38
3-Chloropropyl 41 22
2,2,6,6-Tetrachlorocycloheocyl 38
Methyl 34
Propyl 15
" Adapted from Swenlloff et al. (1984) and Anello et al. (1985).
113
propyl-PDA (Cazenovia soil) and benzyl-PDA (Plano soil). All PDAs tested in both
soils were more inhibitory in the Cazenovia soil than in the Plano soil. It should also be
mentioned that propyl-PDA was less inhibitory than were 1,3-dichloro-2-propyl-PDA
and 3-chloropropyl-PDA; contrarily, cyclohexyl-PDA was more inhibitory than 2,2,6,6-
tetrachlorocyclohexyl-PDA.
Swerdloff et af. (1985a) patented 74 aryl phosphorodiamidates and 18 aryl thio-
phosphorodiamidates, for inhibition of soil urease activity. However, testing of the
inhibitory effect was only described for two compounds: 4-aminophenylphosphoro-
diamidate and 3-(1', l' -dimethylethyl)-4-hydroxyphenylphosphorodiamidate (Figure
40); only the Cazenovia soil was used in testing. The two PPDAs produced 91 and 87%
inhibitions, respectively, in soil urease activity.
4-Amino-PPDA 3-(1',1 '-Dimethytethyl)-4-hydroxy-PPDA
Figure 40. Structure of the aryl phosphorodiamidates tested by Swerdloff eI al. (1985a) for inhibition of soil
urease activily.
The use of these compounds is recommended at the following (most preferred) rates:
1-500 ppm on soil weight basis, 0.28-16.8 kglha or 0.01-20% relative to weight of urea
in liquid and solid fertilizers.
Kolc et al. (1985c) patented 10 phosphorodiarnidates and 2 thiophosphoro-
diamidates as urease inhibitors. However, it results from the patent description that none
of them were tested for evaluation of their inhibitory effect on soil urease activity.
Neither was jackbean urease used for testing of inhibitory capacity.
2.20.2 Further Studies on Phenylphosphorodiamidate (PPDA) and Some Other

Phosphoroamides
Matzel et af. (1978) and Matzel and Heber (1979) tested the effect of PPDA on
ammonia volatilization from urea in 168 light -, medium-, and heavy-textured soils from
Germany. Urea prills (at a rate equivalent to 100 kg NIha) with or without 1% PPDA
(on urea-N basis) were applied to the surface (14 and 28 cm2) of soil samples (50 and 80
g, respectively). After moistening, the samples were incubated at 8°C for 28 days,
during which the volatile NH3 was determined. The results showed that during the first
14 days the inhibitory effect of PPDA was strong in all soils, but after this period the
effect was still strong only in light- and medium-textured soils and was weak in the
heavy soils. It should be emphasized that PPDA, like other inhibitors, acted more
strongly in light soils, i.e.. in those soils from which urea-N losses as NH3, in the
absence of inhibitors, are the greatest.
114
Matzel and Heber (1979) obtained similar results with samples of 140 other German
soils, as well. They also established that reducting the PPDA rate from 1 to 0.5%
relative to urea-N as well as replacing chemically pure PPDA (0.5%) with technical-
grade PPDA (0.5%) did not lead to a marked diminution of the inhibitory capacity of
this compound on volatilization ofNH3 from urea.
Heber et al. (1979) described experiments on microplots (700 cm2 ) located on a
permanent grassland. Urea (at a rate of 100 kg N/ha) with or without 1% PPDA was
surface-applied after each harvest (cut) in the 1974-1976 period. The volatile ammonia
was continuously assayed. Under the influence of PPDA, volatilization of NH3 began
with a delay of about 1-4 days and total NH3 losses were significantly reduced.
Muller and Forster (1980) carried out two experiments. The first was carried out
under conditions identical to those described on page 77; the inhibitor tested was
diphenylphosphoroamidate (di-PPA) which was applied, like the other inhibitors, at a
rate of 2% relative to urea-N (40 mg of urea-N!40 g of soil). It was found that when
urea was surface-applied and di-PPA was introduced at a 2 or 4 cm depth in the soil
column, urea remained unhydrolyzed during 6 days of incubation at laboratory
temperature, in proportions of about 60 and 80%, respectively. The effect of di-PPA
was weaker when urea was distributed uniformly in the soil column.
In the second experiment, urea (40 mg of N) alone or together with 2% PPDA or
with 1% di -PP A + 1% 4-chloro-PPDA was applied on the surface of soil columns (each
consisting of 40-g dry sample of a humous sandy loam, pH 7.2). The soil columns were
moistened up to 60% of WHC by adding water in three portions, namely at the
beginning of the experiment and after 24 and 48 hours. Incubation took place at
laboratory temperature and lasted 3 days. The soil was analyzed daily for residual urea,
NH/, and N0 3-. The results showed that after 3 days the residual urea represented about
60 and 80% of the initial amount in the urea + PPDA and urea + di-PPA + 4-chloro-
PPDA treatments, respectively. This proves the urease-inhibiting capacity of the
compounds tested.
Kampfe et al. (l982c) reviewed the results of 25 experiments carried out in
Bulgaria, in the former Czechoslovakia and (East) Germany, as well as in Hungary,
Poland, and Romania, within an international fertilization project, in the 1976-1978
period. The effect of PPDA on volatilization of ammonia from urea was evaluated in
Mitscherlich pots. Light- and heavy-textured soils were used in mixtures with sand (2
parts soil:l part sand) and moistened to 40% of WHC. Urea prills alone or urea prills
coated with 1% PPDA (on urea-N basis) were applied on the soil surface at a rate of 0.5
g of N/pot in 1976 and at a double rate in 1977 and 1978. TI1en the amount of NH3
volatilized in 5, 10, and 15 days was determined. One can deduce from Table 35, which
presents the mean values of NH3 volatilized at different mean air temperatures, that
PPDA manifested an inhibitory effect even at temperatures higher that 14°C, at which
NH3 losses in the absence of PPDA, were greater especially when the incubation time
was prolonged to 15 days.
Linke et at. (1982) studied volatilization of ammonia from urea and from urea-
PPDA under field conditions on three German soils (sand, loamy sand, and sandy
loam), cropped with wheat, in the 1976-1979 period. Conditioned urea (urea in the form
of prills coated with a mineral oil-bitumen mixture - 0.2% relative to weight of urea -
115
TABLE 35. Influence of air temperature on volatilization of ammonia from soils fertilized with
urea and urea-PPDN'
Ammonia losses
(% relative to applied urea-N)
Temperature (0C) Fertilizer
5 10 15
8-10 Urea 1.7 6.8 11.4
Urea + 1% PPDA 0.5 1.3 3.9
> 14 Urea 5.4 11.5 15.2
Urea + 1% PPDA 0.6 3.4 7.8
"From Kiimpfe ef al. (1982c).
,with or without 1% PPDA on urea-N basis) was applied on soil surfaces annually twice
at the following rates: 50 + 40 kg ofurea-N with 15N label for two soils (loamy sand and
sandy loam) or 50 + 50 kg (in 1976) and 100 + 100 kg of unlabeled urea-N (in 1977-
1979) for the third soil (sand). After each fertilizer application, volatilization of NH3
was determined during 14 and 21 days. The experiments were carried out under natural
weather conditions or under conditions of simulated drought after fertilization.
According to the results obtained, the inhibitory effect of PPDA was more evident
during the first 14 days than between days 14 and 21. After the first fertilization, PPDA
had a greater effect than after the second one. The conclusion was drawn that, under
natural weather conditions, mean NH3 losses from urea were 30%, whereas losses from
urea-PPDA were not lower than 20%. Under conditions of simulated drought, urea-
PPDA was more effective than urea alone: in the presence of PPDA, more urea
remained unhydrolyzed, less NH4+ was produced, and less NH3 was lost by
volatilization.
The effect of PPDA on urea hydrolysis in a grey-brown podzolic soil from the
former Czechoslovakia was studied by List'anska (1982). Dry soil samples (350 g) were
treated with urea or with the liquid fertilizer DAM 390 (urea-ammonium nitrate) at a
rate of 300 mg N/kg soil. No PPDA or 1% PPDA (on N basis) was added to the
fertilizers. The samples were moistened (to 15-17% moisture content) and incubated at
28°C. During incubation (13 weeks) the NH/ and N03' contents were determined. The
analytical data indicated that PPDA inhibited hydrolysis of urea from both fertilizers for
2 weeks and did not retard nitrification. Similar results were published in another paper,
also (List'anska, 1984).
In a pot experiment, Byrnes et al. (1983) measured volatilization of ammonia from
flooded samples of an Alabama silt loam soil (oven-dry weight of each sample: 1,620
g). The floodwater of samples received a urea solution, containing 0.4193 g of 4.814
atom% excess urea- 15N, (equivalent approximately to 50 kg N/ha on area basis) and a
PPDA solutions to provide 1, 2 or 5% PPDA relative to weight of urea. The floodwater
depth was then adjusted to 2 cm and maintained at this level throughout the experiment.
The soil in pots was incubated at 35°C by day and at 25°C by night (12-hour days)
during 14 days. Daily analyses were performed for volatilized NH3, residual urea, NH/
and pH in the floodwater. At the end of the experiment, total N contents in floodwater
and soil were determined. The isotope ratio in total N contents and in volatilized NH3
was also determined.
Figure 41 shows that in the absence of PPDA cumulative NH3 volatilization losses
amounted to 31.4% of the added urea- 15N, while losses from the PPDA treatments were
116
3.9-5.3%. Effects of the three PPDA concentrations were not significantly different (at
p=0.05).
35r----------------------------,
- Nt, PPDA
·..····1% PPDA
-·,·,2% PPDA
._- 5% PPDA
o 2 4 6 B 10 12 14
Incubatioll time (days)
Figure 41. Cumulative ammonia volatilization losses ofurea- 15 N with time. !From Byrnes e/ al. (1983), by
pennission of the Soil Science Society of America, Inc.!
It was also found that NH.l losses from the treatment without PPDA occurred
concurrently with the rise in NH4 + concentration and pH of the floodwater during the
first few days. The final 15 N balance showed total recoveries of 96.7-99.2%. There was
no significant difference (p=0.05) between total recoveries in the absence or presence of
PPDA. Reduction of NH3 volatilization under the influence of PPDA was accompanied
by an increased recovery ofN in floodwater (algae) and in soil.
In another experiment with the same silt loam soil used in the first experiment, the
effect exerted by different amounts of PPDA in urea pellets on hydrolysis of urea was
studied. Soil samples (l00 g on oven-dry basis) placed in cylinders formed
approximately 9-cm high soil columns covered by l-cm deep floodwater. Each column
then received a single 15-mg urea pellet containing 0, 0.1, 0.25, 0.5, 1, 2, and 5% of
PPDA (relative to weight of urea). Afterwards, the soil columns were incubated in the
dark at 300C for 16 hours. Following incubation, the soil-floodwater systems were
extracted and the extracts analyzed for urea and NH4 +. It was found that PPDA at
concentrations 2: 1% in pellets completely inhibited hydrolysis of urea. The inhibition
was about 92% at 0.5% concentration of PPDA in the pellet (which corresponds to a
concentration of approximately 4 ppm of PPDA dissolved in the floodwater). At 0.25
and 0.1 % PPDA concentration in the pellets, degrees of inhibition were about 80 and
50%, respectively.
In a short report, O'Connor et al. (1983) quote a laboratory experiment carried out
for studying the influence of soil temperature (m the effectiveness of PPDA. Soil
samples treated with 1 mg of urea-N and 20 Ilg of PPDAIg soil were incubated at 5-
35°C for various time periods. PPDA prevented urea hydrolysis and associated
ammonia losses for 1,2,4,8, and 17 days at temperatures of 35,25, IS, 10, and 5°C,
117
respectively. Since surface soil temperatures during the growing season often exceed
30°C, the agronomic utility ofPPDA is very much reduced.
In another experiment, dried maize (Zea mays) leaves were moistened with a urea
solution containing 0 or 2% PPDA (relative to urea-N) and incubated at 25°C.
Ammonia losses were reduced from 82% (no PPDA) to 20% (PPDA). The same
treatments applied to bare soil resulted in cumulative NH3 losses of 26 and 12%,
respectively. These results indicate a greater potential for NH3 losses from no- or
reduced tillage systems and the role of urease inhibitors in keeping these losses to a
minimum.
Another short communication, by Ornholt and Hendrickson (1983), refers to field
experiments conducted to evaluate the ability of PPDA to inhibit urea hydrolysis and
reduce ammonia volatilization losses. A 50% urea solution (95 kg Nlha) with or without
2.2 kg of PPDAIha was applied in mid-August to bare plots and to plots amended with
oat straw. In both systems, PPDA extended the persistence of urea by only 2-3 days.
Nonetheless, PPDA reduced NH3 losses from 33 to 23% of the added urea-N on bare
plots and from 42 to 6% on straw-amended plots.
Martens and Bremner (1984a) devoted a complex study to factors influencing
effectiveness of PPDA to retard urea hydrolysis in 15 Iowa soils selected to obtain a
wide range in pH (4.6-8.0), texture (5-57% sand, 11-55% clay), organic carbon content
(0.30-6.73%), urease activity 04.2-84.9 /lg of urea hydrolyzed/hour/g soil at 37°C), and
other soil properties. The reaction mixtures, consisting of 5 g of air-dried soil + 2 ml of
aqueous solution containing 10 rug of urea without or with 1-125 (usually, 25) /lg of
PPDA, were incubated at 20°C for 1-21 days, then analyzed for residual urea.
The inhibition caused by PPDA (5 /lg!g soil) in urea hydrolysis in the 15 soils
studied, ranged, after a 7-day incubation, from 37 to 93%. The percent inhibition
correlated Significantly with organic C content (r=-O.68), total N content (r=-0,74),
cation-exchange capacity (r=-0.65), sand content (r=0.66), clay content (r=-O.64), and
surface area (r=-0.60), and insignificantly with urease activity (r=-0.29), silt content (r=-
0.46), pH (r=0.29), and CaC0 3 equivalent (r=O.03). One can see that most correlations
are negative. Multiple-regression analyses indicated that the effectiveness of PPDA to
retard urea hydrolysis in soils tends to increase with a decrease in soil organic matter
content.
Table 36 shows that the average percent inhibition increased markedly with PPDA
concentration and decreased markedly when incubation time was prolonged from 2 to
10 days.
The influence of incubation temperature on effectiveness ofPPDA (5 /lg!g soil) was
studied with six soils. The following average values of percent inhibitions were
registered at 10, 20, 30, and 40°C: 91, 90, 59, and 35%, respectively, after 3 days of
incubation, and 85, 76, 28, and I %, respectively, after 7 days. Thus, effectiveness of
PPDA decreased proportionately with an increase in temperature and this decrease
became more pronounced with prolongation of incubation time.
To study the effectiveness of PPDA, Broadbent et al. (1985) conducted a field
experiment on a silty loam soil from California. The experiment included two variants:
plots not covered and plots uniformly covered with chopped wheat straw (at a rate of
4.93 t/ha). Maize (Zea mays) was sown in all plots at a population density of 86,000
plantslha. As soon as the maize seedlings emerged, urea labeled with an excess of 15N
118
TABLE 36. Effect of different amounts of PPDA on urea hydrolysis in 15 soils after
two incubation times·
Amount ofPPDA Incubation time
Average value of inhibitions (%)
(p.gIg soil) (days)
0.2 2 60
10 18
0.5 2 68
10 24
2 76
10 30
3 2 84
10 36
5 2 91
10 45
10 2 93
10 58
25 2 96
10 71
•Adapted from Martens and Bremner (1984a), by permission of Pergamon Press PLC.
was surface-applied in form of solution with or without PPDA. Rates of application

were: 180 kg of urea-Nlha and 2.5 kg of PPDAlha. Immediately after fertilization and
three times during the first week after fertilization, samplings were made from the 0-10-
cm soil layer to determine total N extractable with 1 M KCI solution and of inorganic N.
The 15N contents in the total N and in inorganic N were also determined. Soluble
organic N, primarily urea, was calculated from the difference between total KCl-
extractable N and inorganic N. After the maize was harvested, the same analyses were
performed in soil sampled at 30-cm increments to a depth of 120 cm. Overground parts
of maize plants were also analyzed for total N and lSN.
Results of the analyses performed during the first week after fertilization are
presented in Figure 42. They show that in plots treated with urea alone, a greater
amount of urea)5N was recovered from the bare soil than from the straw-covered soil
which should be attributed to a more intense urease activity associated with straw (in
which urease-producing microorganisms could develop). The effect of PPDA on the
inhibition of urea hydrolysis was weak in the bare soil and considerable in the straw-
covered soil.
Analyses ofN in soil and overground parts of maize plants after harvest (N in roots
and N nonextractable from soil organic matter with 1 M KCI were not included)
indicated that the added urea-N (180 kg/1m) was only partially recovered from soil and
plants. Thus, the amounts of N recovered in the urea and urea-PPDA treatments were
85.0 and 94.2 kg Nlha, respectively, in the case of bare plots, and 76.5 and 76.9 kg
Nlha, respectively, in the case of straw-covered plots. One may conclude that volatile
NH3 losses from urea were high, especially in the straw-treated plots, in both presence
and absence ofPPDA.
119
160
A
120
80
i 40
UreA alone
I!.
OUreA+PPDA
Z 0 -..J._-'----'-_-'---'_-'----'"----'-
<1:1'60
5
120
eo
40
o 2 4 6
Days
Figure 42. Labeled urea-N in soil samples taken fiom fertilizer band during the first week after fertilizer
application.
A - Bare soil. B - Soil + straw. Bars show ± standard deviation. IFrom Broadbent et al. (1985), by
permission of the Soil Science Society of America, Inc.!
Simpson et al. (1985) also studied the effectiveness ofPPDA under field conditions,
on an Australian clay soil (PH 8.2) cropped with flooded rice. At the stage at which the
rice plants reached 20 cm above the -11 cm deep floodwater, microplots were installed
within the rice field. They were enclosed by metal frames (54 x 54 em) inserted to
contain three rice rows and buried in the soil to a 15-cm depth so that 5 cm projected
above the floodwater surface. Prilled urea (80 kg NIha) with or without PPDA (1 %
relative to weight of urea) was administered in form of solution in the floodwater of
microplots. Some microplots received a solution of urea labeled with 15N at 1.48 atom%
excess (at a rate of 80 kg of N/ha) with or without 1% PPDA. On days 0 to 11 after
fertilization, residual urea, NIL +, and NO)' contents, pH and temperature in floodwater
as well as the volatilized ammonia and wind speed were determined. Total N and 1~ in
floodwater, soil, and plants were analyzed three times: on days 11,47 (early heading),
and 130 (maturity).
Figure 43 shows that in the absence of PPDA, urea concentration in floodwater
decreased continuously and at day 8 after fertilization it became negligible. PPDA
completely inhibited hydrolysis of urea for about 4 days (urea concentration having
been maintained at the initial value of about 80 g of N/m3 ), after which the inhibitory
effect of PPDA decreased steadily until day 11 when no urea was detectable in the
floodwater. In concordance with these results PPDA delayed the appearance and
120
accumulation of NH4 + in floodwater; maximum NH4 + concentration was reached after 3

days in the urea-only treatment and after 9 days in the urea+PPDA treatment.
,. B
"
Z l , I • 7 • • 10 '1 U
DRy"l after fertilizer application
Figure 43. Effect ofPPDA on concentration of urea CA) and ammoniacal (E) nitrogen in floodwater. (Adapted
from Simpson e/ al. (1985). by permission of the Soil Science Society of America. Inc.!
Concentration ofN0 3" in floodwater was low during the entire period of observation
and was not influenced by PPDA. In urea+PPDA treatment, pH increased slightly on
days 3 to 8, probably due to a more intense development of algae. In the urea-only
treatment, more ammonia volatilized during the first 3 days (when NH4 + concentration
in floodwater was higher) and smaller amounts of NH3 were lost on the next days.
Contrarily, in the urea+PPDA treatment, volatilization of NH3 was reduced for 6 days
and then increased. The cumulative NH3 volatilization losses during 11 days in the urea
and urea+PPDA treatments were 20.6 and 12.5% of the applied urea-N, respectively.
The greater NH3 losses in the urea-only treatment were due partly to wind speed which
was higher during the first 3 days than during the last 3 days.
Analyses of total N and 15N in floodwater, soil, and plants on day 11 after
fertilization showed that total loss of N from the applied urea-N was 40.2% in the urea-
only treatment (i.e.. N recovery was about 60%) and 18.5% in the urea+PPDA treatment
(i. e.. N recovery was over 80%). As under the conditions of these experiments no N was
lost by leaching, run-off or in form of N20, it was deduced that N was also lost as N2
(i.e .. through denitrification of nitrates resulted from oxidation of NH/ released from
urea). The loss in form of N2 represents the difference: 40.2-20.6=19.6% in the urea-
121
only treatment and 18.5-12.5=6.0% in the urea+PPDA treatment. It was concluded that
the major inhibiting effect of PPDA on soil urease activity was to reduce those N losses
which are conditioned by release of urea-N in form of NH4 + followed by oxidation of
NH4 + to N0 3- and then by denitrification ofN03- up to N 2.
In a field experiment on a Quebec silty clay loam soil, Tomar et al. (1985) studied
the effect of PPDA and chopped timothy (Phleum pratense) straw on urea hydrolysis
and ammonia volatilization from urea. Bare plots and plots on the surface of which
timothy straw (4.6 t/ha) had been spread, were fertilized with a urea solution (75 kg
N/ha) without or with PPDA (0.25,0.5, 1,2 or 3.74 kg/ha). All plots were also fertilized
with triple superphosphate and potassium chloride at rates of 63 kg of P and 100 kg of
Kiha, respectively. Residual urea, NH4 +, and (N03- + N0 2-) were extracted with 1 M
KCl solution and measured at days 0, 1,2,3,4, 8, and 14 after fertilization. Amounts of
NH3 volatilized during 1, 2, 3, 4, 14, and 17 days following fertilization were also
assessed,
PPDA at rates :s 1 kg/ha had no significant effect on urea hydrolysis. However, urea
hydrolysis was significantly retarded at 2 and 3.74 kg PPDAIha rates on days 4 and 8. A
reduction in urea hydrolysis with increased PPDA concentration was also observed
when the data were pooled across straw treatments. The inhibitory effect of PPDA was
not long-lasting: almost all the added urea had disappeared by day 14. On days 2 to 14
after fertilization, more urea was hydrolyzed in straw-amended soil than in bare soil, but
this influence of straw was significant at day 4 only.
Ammonia volatilization was negligible during the first 4 days in all treatments and
attained higher values after 14 days. Between days 15 and 17, volatilization of NH3
declined. PPDA reduced volatilization of NH3, especially at its highest rate, The straw
enhanced ammonia losses in treatment with PPDA at rates:S 1 kg/ha had no affect at
higher PPDA rates. In this experiment, volatilization NH3 losses within 17 days never
exceeded 1% of the applied urea-No
Nitrogen recoveries as residual urea-N + NH/-N + (N03' + N02')-N measured on
days 1, 2, 3, 4, 8, and 14 after fertilization gave the following values across all
treatments: 92, 89, 82, 68, 62, and 48% of the applied urea-N, respectively. This proves
that a part of urea-N was progressively incorporated into N compounds nonextractable
with 1 M KCI and/or lost mostly through nitrification-denitrification, not through NH3
volatilization.
The experiments performed by Mai and Fiedler (1986) intended to study the
influence of weather conditions on the effectiveness of PPDA, In a German spruce
forest estimated to be about 90-year-old on an acid loamy soil (PH 4.3), urea hydrolysis
was compared in plots fertilized (in 1979 and 1980) with surface-applied urea (150 kg
N/ha/year) with or without 1% PPDA. In 1979, the fertilization was followed by a
drought period, whereas in 1980 the soil was very moist at time of fertilization. In 1979,
complete hydrolysis of urea needed 17 days in urea-only treatment and 23 days when
urea and PPDA were applied. In 1980, the corresponding durations were 1 and 8 days,
respectively.
The influence of meteorological conditions on the effectiveness of PPDA was also
evidenced in the field experiments described by Fillery and De Datta (1986) and Fillery
et al. (1986a), The experimental field, on a silty clay loam (PH 5.8) in the Philippines,
was cropped with flooded rice. Urea (58 kg N/ha) alone or with PPDA (1% relative to
weight of urea) was applied to floodwater on day 18 after the transplanting of (20-day-
122
old) rice seedlings. (Triple superphosphate, 13 kg P/ha and KCl, 25 kg Klha were
incorporated into soil immediately before transplanting.)
Determinations of N~ + in floodwater and of volatile NH3 showed that PPDA
retarded accumulation of ~ + and volatilization of NH 3. Thus, NH3 volatilization
losses from urea and urea-PPDA over 8 days were 36 and 22% of the applied N,
respectively, i.e.. PPDA reduced NH3 losses by about 40%. This reduction, however,
was not attributable only to the inhibitory effect of PPDA, but also to the decline in
wind speed over the period (days 6 and 7) in which the maximum concentrations of
NH4 + were detected in the floodwater amended with urea-PPDA. Had the wind speed
increased, larger amounts ofNH3 could have volatilized and in this case PPDA may not
have reduced NH3 losses as effectively.
The investigations by Fillery et at. (I 986b ) draw attention to the importance of
timing of urea-PPDA application for inhibition of urea hydrolysis and reduction of
NH/ concentration in floodwater of rice, i.e.. for diminution of N losses caused by
volatilization of NH3. The experimental plots were installed on two soils in the
Philippines: a clay (PH 6.7) at Los Banos and a silty clay loam (PH 5.8) at Munoz. The
soils were fertilized with triple superphosphate (13 kg P/ha) and KCl (17 kg Klha)
immediately before transplanting of rice seedlings. At Los Banos, prilled urea was
administered in the dry season (90 kg N/ha) and in the wet season (60 kg N/ha) in 1982
and in the dry season (87 kg N/ha) in 1983. In this year, rnicroplots fertilized with 15 N_
labeled urea (= 5 atom% excess) were also installed. At Munoz, urea fertilization (87 kg
N/ha) was carried out in the dry season of 1983 and rnicroplots fertilized with urea- 15 N
were also installed.
Two-thirds of the urea with or without 2 or 5% PPDA (relative to weight of urea) in
1982 and with or without 1% PPDA in 1983 were incorporated into the soil or added to
floodwater immediately before or after transplanting of the seedlings or between days
18 and 26 after transplanting. The remaining third of urea and urea-PPDA was applied
later (50 days after transplanting or 5-7 days before panicle initiation). Depth of
floodwater was 4-6 cm. Daily, at the same hour, floodwater was sampled for analysis of
residual urea and NH4 +.
In both soils, PPDA inhibited urea hydrolysis and reduced accumulation of NH4 + in
floodwater only for 1 day, when it was administered together with urea immediately
before or after transplanting. The inhibition lasted 3 days, when urea-PPDA was added
between days 18 and 26 after transplanting.
In a laboratory experiment, Rao and Chai (1986b) used an alkali sandy loam soil
(pH 9.0) from India. Air-dried soil samples (50 g) were moistened with 10 ml of water
and allowed to equilibrate overnight, and then were treated with 10 ml of water or
PPDA solution,S m1 of urea solution and, finally, with 25 m1 of water to form an
aqueous surface layer. Urea was added at rates of 150, 100, 75, 50, and 37.5 ppm N,
whereas the amount of PPDA was 5% relative to weight of urea. During incubation
(35°C/20 days), the floodwater level was maintained and periodically analyzed for ~ +
and (N03- and NOz-). One can see from Figure 44 that, under the influence of PPDA,
NH4 + appeared in floodwater with a delay of 1 day (at maximum urea-N rate), of 3 days
(at 100 and 75 ppm urea-N) or of 4 days (at 50 and 37.5 ppm urea-N). Moreover, the
peak NH4 + concentration had significantly lower values in the presence than in the
absence of PPDA.
123
40.-------------------__.-__________________--.
i
,eo 30
-rrt'.1l
--. Urea+PPDA
!:l A
OJ
~
"" 20
~
.S
g
01)
.~
]
J
'S
10
4 6
Figure 44. Effect of PPDA on ammoniacal nitrogen concentration in floodwater of an alkali soil, as
influenced by incubation time and urea-N application rates (ppm): A - 150; B - 100; C - 75; D - 50;
E - 37.5. IF rom Rao and Chai (198Gb), by permission of Kluwer Academic Publishers.!
The same soil was also used to study the effect of PPDA on volatilization of
ammonia from urea. Soil samples (20 kg) placed in pots were saturated with water, then
urea (7S, SO or 37.S ppm N) or urea + PPDA (S% relative to weight of urea) were mixed
into a few centimeters of the top soil layer. Finally, the soil was flooded to 3.S cm
which was maintained throughout the 24-day incubation in greenhouse. Each day,
amounts of NH3 volatilized and of Nf4 + in floodwater were measured. The results
indicated that in urea-only treatments volatilization of NH3 began on the first day after
fertilization and continued until the 12th day, with similar trends at the three urea-N
rates. In the presence of PPDA, volatilization ofNH3 started after S days and percentage
inhibitions of NH3 volatilization from the three urea-N amounts were not significantly
different. On an average, 4S% of the applied urea-N was lost in the absence of PPDA,
but only 8.S% in its presence, the difference between the two values being very
significant.
Reviewing the investigations on N fertilizers for rice, Youngdahl et of. (1986)
present data obtained in pot experiments with rice grown under submerged conditions.
Urea without or with 1% PPDA was administered as basal fertilizer on day 21,30 or 42
after transplanting of rice seedlings. PPDA always reduced ammonia losses, regardless
of the timing of application, although the relative effectiveness varied. Thus, minimum
effectiveness was recorded when urea was applied as basal fertilizer (NH3 losses in urea
and urea-PPDA treatments were about 41 and 30% of the applied urea-N, respectively).
The maximum effectiveness of fertilization was on day 30 after transplanting (NH3
losses in urea and urea-PPDA treatments were about 28 and 10%, respectively).
A laboratory experiment performed for studying the influence of temperature on
effectiveness ofPPDA was quoted in a short communication by O'Connor et af. (1983)
(see page 116) and described in detail by O'Connor and Hendrickson (1987) who
pointed out that 20-g samples of silt loam soil (pH 7.S) from New York were used and
reiterated the conclusion that effectiveness of PPDA (20 l1g/g soil) in inhibition of
124
hydrolysis of urea (l mg N/g soil) and of ammonia volatilization from urea decreased
with increase in temperature from S to 3SoC.
Using the same soil, O'Connor and Hendrickson (1987) also performed other
laboratory experiments (A, B, C), through which the effects of urea and PPDA rates and
soil depth (of the height of soil column) on ammonia volatilization were studied. The
urea solution and then the water with or without PPDA were applied to the surface of
completely moistened soil samples. Then the mixtures were incubated at 2S C for 14-IS
D
days, during which, at 1-3-day intervals, the amount of volatilized NH3 was assessed.
In experiment A, the variable factor was the urea rate (S, 10, and 20 mg N120 g soil),
while soil depth (height of soil column) of 8 mm, corresponding to 20 g of air-dried
soil and PPDA rate (0.2 mg/20 g soil) were constant. Cumulative NH3 losses from three
urea-N amounts during incubation (14 days) were IS, 22, and 38%, respectively, in the
absence of PPDA and 7, 18, and 38%, respectively, in its presence. This means that
PPDA was most effective at the lowest urea-N rate and became ineffective at the
highest rate ofurea-N. It should be added that during the fust days of incubation PPDA
reduced volatilization ofNH3 even from the highest urea amount.
In experiment B, soil depth (height of soil column) was the variable factor (8, 16,
and 32 mm, corresponding to 20, 40, and 80 g of soil). Rate of urea (20 mg N) and that
ofPPDA (0.2 mg) were the same, irrespective of the soil weight, i.e. these rates relative
to I g of soil were I, O.S, and O.2S mg of urea-N, respectively, and 10, S, and 2.S Ilg of
PPDA, respectively. Ammonia losses decreased with an increase of soil depth. PPDA
delayed the onset of NH3 volatilization in each treatment, but reduced the cumulative
NH3 losses during incubation (IS days) only when soil depth was 16 or 32 mm.
Effectiveness of PPDA was more marked with the 32-mm deep soil sample than with
the 16-mm deep sample.
Experiment C, like experiment B, included three soil depths (8, 16, and 32 mm),
corresponding to 20, 40, and 80 g of soil, but the urea rates were 20, 40, and 80 mg of
N, respectively, and the PPDA rates were 0.2, 0.4, and 0.8 mg, respectively, i.e., the
rates relative to I g of soil were constant (1 mg of urea-N and 10 Ilg of PPDA,
respectively). The results obtained are similar to those registered in experiment B. Thus,
in the case of the 32-mm soil depth, PPDA reduced cumulative NH3 losses during IS
days from 20 to 12% of the applied urea-No
Extrapolating the results of experiments Band C to field conditions, one can assume
that conditions that promote diffusion of the surface-applied urea and PPDA more
deeply into the soil will reduce the potential for NH3 losses.
In a laboratory experiment, Hendrickson et al. (1987) studied the influence of plant
residue and PPDA on the volatilization of NH3 from urea-treated soil samples. The
same silt loam soil (pH 7.S) from New York was used as in the experiments described
above. There were three experimental variants: 20 g of air-dried soil; 0.3 g of coarsely-
ground, dried maize (Zea mays) stover; and 0.3 g of stover intimately mixed with 20 g
of soil. Each variant was treated with 1 ml of solution containing 20 mg of N as urea
and S ml of water or S ml of solution containing 0.4 mg of PPDA, and sufficient
additional water to adjust the final water potential to approximately 30 kPa. Incubation
took place at 2S C. The ammonia evolved was periodically assessed during 17 days of
D
incubation. Urea and pH were also periodically determined during II days.

PPDA retarded urea hydrolysis in soil and soil + residue mixture and also in the
residue-only treatment. The resultant pH closely paralleled urea hydrolysis. In the
125
absence of PPDA, cumulative NH3 volatilization losses were approximately 20% in

both soil and soil + residue mixture after 4 days and 38 and 28%, respectively, after 17
days. PPDA prevented the NH3 losses from both soil and soil + residue mixture for 4
days, after which the inhibitory effect of PPDA gradually declined and beginning with
the 11 th day the NH3 losses in the presence and absence of PPDA became equal.
When urea was applied to residue alone, very rapid NH3 losses were observed, with
over 30% of the applied N lost within 3 days and over 80% volatilized within 7 days.
PPDA essentially eliminated NH3 losses for the first 4 days, but subsequent losses
occurred at a reduced, but consistent rate throughout the rest of the incubation time, and
after 17 days the cumulative losses in the residue + urea + PPDA treatment (about 80%)
approached those registered in the residue + urea treatment (about 90%).
Rodgers et al. (1987) studied the effect of PPDA on volatilization of ammonia from
prilled urea applied on a light sandy soil under a grass (timothy and meadow fescue) ley
sown in 1979 and cut each year, at Woburn Experimental Farm, Bedfordshire. The
fertilization experiment was carried out in 1984 and 1985. Each plot was 2.4 m wide
and 12.2 m long. In both years, the rate of prilled urea wa<; 375 kg N/ha as single
application or as divided applications (3 x 125 kg N/ha). PPDA was applied as a 1% by
weight coating on urea prills. The control plot received urea prills without PPDA.
Ammonia volatilization was determined during 12 days following each urea application.
It was found that in 1984 the total NH3 losses (expressed in kg NHrN/ha) in the
urea and urea-PPDA treatments were 13.2 and 0.3, respectively (single application), and
32.8 and 17.4, respectively (divided applications). In 1985, the corresponding values
were 28.7 and 0.8, respectively (single application), and 31.5 and 13.8, respectively
(divided applications). The reduction of NH3 losses by PPDA was always very
significant.
Studying soils from Cuba, Knappe and Carrion (1987) found that PPDA produced a
100% inhibition of urea hydrolysis over periods of 10-15 days in sandy soils, but only
medium inhibitions were measured in base-saturated soils.
Pedrazzini et al. (1987) performed a pot experiment using an Italian sandy loam soil
(pH 5.8). Deionized water was added to the soil samples (6 kg) to provide a 2-cm layer
of standing water. After 1 week rice was planted and at day 14 after planting
(transplanting) urea was broadcast at a rate of 100 kg N/ha either alone or with PPDA
(13 kglha). There were six different treatments (no plants; no plants + urea; no plants +
urea + PPDA; plants; plants + urea; plants + urea + PPDA). The samples, all water-
logged, were incubated at 20'C. During 30 days, urease activity and ~ + content in
floodwater were determined at 24-hour intervals, whereas the NH3 volatilized was
assessed three times each day for a total period of 1 hour, i.e., not continuously.
In the controls (no plants and plants only) urease activity in floodwater was very low
throughout the whole incubation period. In the urea-only treatments, a lag of 3 days
appeared, after which urease activity increased and reached a peak value at days 6 and
lOin the presence and absence of rice plants, respectively. The increased urease activity
is explained by microbial synthesis of new urease molecules. However, these molecules
were short-lived as after 6 and 10 days, respectively, urease activity markedly decreased
and approached the values of control at the end of the incubation period. In the urea +
PPDA treatments, after a 4-day lag, urease activity increased, but to a much lesser
extent than in the urea-only treatments, and remained almost constant up to the end of
the incubation period; the rice plants exerted no influence on urease activity.
126
Ammoniacal-N concentration in floodwater was low in the controls, reached a

maximum at day 8 in the urea-only treatments, remained low for 8 days and became
highest at day 22 in the urea + PPDA treatments. Volatilization of ammonia followed a
similar pattern. These observations mean that in the presence of PPDA increases in
NH4 + concentration in floodwater and in NH3 volatilization were delayed but not
eliminated.
Buresh (1987) tested PPDA on a flooded rice field in the Philippines (location:
Maahas). The clay soil (PH 6.6) was fertilized with urea (30 kg Nlha) without or with
1% PPDA (on urea weight basis). Besides urea, other fertilizers were also tested, for
example urea phosphate, 17-44-0; urea-urea phosphate, 34-17-0, also at a rate of 30 kg
Nlha. The fertilizers were applied by two methods: 1. broadcast and incorporation into
the soil without standing floodwater 1 day before transplanting of rice seedlings, and 2.
broadcast into 5-cm deep floodwater at day 18 after transplanting. Ammonia loss was
not directly measured; instead, relative susceptibility to NH3 loss among fertilizers was
assessed from equilibrium NH3 vapor pressure (pNH3) which at a given wind speed is
directly related to NH3 volatilization from floodwater.
The results obtained with the first method have shown that at day 2 after
transplanting pNH3 values recorded with different fertilizers increased in the following
order:
urea phosphate < urea-urea phosphate < urea-PPDA < urea,
but at day 4 pNH3 became equal to zero with each of these fertilizers.
By using the second method it was found that at day 19 after transplanting pNH3
increased in the order:
urea-PPDA « urea phosphate ~ urea-urea phosphate ~ urea,
but disappeared at day 22 with each of these fertilizers. One can deduce from these
findings that PPDA inhibited hydrolysis of urea and volatilization of NH3 only during
the first days after fertilization.
Buresh et al. (l988a) tested PPDA on a clay soil (PH 6.2-6.4) at another locality in
the Philippines (Pila) in the 1985 dry season. The test plant was flooded rice. Urea,
applied at rates of 30, 60, and 120 kg Nlha without or with 2% PPDA relative to weight
of urea, was broadcast into 5-cm floodwater with two-thirds of the dose at day 18 after
transplantation of seedlings and one-third of the dose at day 5 to 10 after panicle
initiation. During 8-10 days after both fertilizations, the floodwater was analyzed daily
for residual urea and ~ +; pNH3 was also assessed. The results proved that in this soil,
too, the retarding effect of PPDA on urea hydrolysis, ~ + accumulation, and pNH3
lasted only a few days after both first and second applications of urea at each rate.
Phongpan et al. (1988) performed a pot experiment using a wetland soil (clay, pH
5.1) from Thailand. Pots filled with soil to reach a total dry weight of 4 kg/pot were
flooded with demineralized water, puddled and allowed to stand overnight. The
floodwater covering the soil surface was I cm deep. Urea (460 mg N/kg soil) was then
surface-applied without or with 2% PPDA (on urea weight basis). In another treatment,
urea without PPDA was placed at 10 cm depth in the soil. Following urea application,
sufficient water was added to form a 5-cm layer on the soil surface and this level of
floodwater was maintained throughout the experiment. The control soil received no urea
and no PPDA. On days 1, 3, 7, 14, 21, and 28 after urea application, analyses were
made to determine residual urea, ~ +, pH, total alkalinity in floodwater and the
volatilized ammonia.
127
The analytical data presented in Figure 45 prove that PPDA effectively inhibited
urea hydrolysis. Complete hydrolysis of urea occurred in 14 days in both urea-only
treatments and in 28 days in the soil treated with urea + PPDA. When urea was placed
deeply into the soil, concentration of urea in floodwater was highest not on the first, but
on the 3rd day, which shows that the upward movement of urea from the site of
placement to floodwater was relatively slow because urea is weakly adsorbed by soil
colloids.
600
0\3 7 14 21 28
I)(lV~ <tiler urea anplu.:ahon
Figure 45. Urea-N concentration in floodwater after application of urea and urea-PPDA. a -
Unfertilized control. b - Deep placement of urea. c - Surface-applied urea. d - Surface-
applied urea +2% PPDA.lFrom Phongpan et al. (1988).1
Concentration ofNa + in floodwater during the first 14 days presented the following
order of the treatments:
surface-applied urea + PPDA < deeply placed urea < surface-applied urea alone.
In all treatments, NH4 + concentration in floodwater decreased after 14 days and by
days 21 to 28 the values were approximately the same. pH and total alkalinity showed a
similar pattern. Volatilization of ammonia started on day 3 after urea application and
increased in the next period. However, the cumulative NH3 losses were small in all
treatments even after 28 days: 2.9% of the applied urea-N (surface-applied urea alone),
1.3% (deeply placed urea), and 1.2% (surface-applied urea + PPDA).
In this experiment, the eventual N losses through nitrification-denitrification were
not studied.
Based on the finding that in unsaturated soil columns, an increase of bulk density by
compacting led to a marked increase in hydrolysis of surface-applied urea (Savant et af..
1987a), Savant et al. (1988a) initiated studies concerning the influence of soil bulk
density on the effectiveness of PPDA as a soil urease inhibitor. The experiments were
carried out with clay and loam soils from the U.S.A.
In the first experiment, samples of four unsaturated soils (0.16 g water/g soil),
equivalent to 100 g of oven-dried soil, were placed into cylinders (internal diameter: 4
em; height: 6-12 cm). The soil columns were then hand-compacted to bulk densities that
ranged from 0.69 to 1.70 g/em3 • Three preweighed urea granules (about 20 mg of
128
urea/granule) not containing or containing 1% PPDA were placed on the surface of the
columns. The column systems were covered with Parafilm to prevent evaporation of
water. After incubation at 30°C, the residual urea was extracted and determined. The
results presented in Table 37 were obtained after 2-day incubation of the clay soil and
after 3-day incubation of the three other soils. It is evident from the table that inhibition
of urease activity by PPDA Significantly decreased with the increase in soil bulk
density; this effect was more apparent in the clay and clay loam soils than in the loam
and silt loam soils.
TABLE 37. Effect of soil bulk density on urease inhibition after surface application of urea
granules containing 1% PPDA in four unsaturated soils"
Average bulk density Urease inhibition (%)
(g/cmJ) Clay Loam Silt loam Clay loam
0.71(0.69-0.72) 48a 74a 78a 56 a
1.I1(O.95-1.2I) 23b 44b 35b 19b
1.47 (1.36-1.70) 19 c 36 b 38 b 4c
"From Savant et al. (1988a), by courtesy of Marcel Dekker, Inc.
Values followed by different letters in the same colwnn are statistically different (p = ()'05).
In another experiment, 1 ml of a 0.06% aqueous solution of PPDA and 60 mg of

powdered urea were uniformly applied on the surface (12.6 cm2) of precompacted
columns of five unsaturated soils. The next steps of the experiment were the same as in
the first experiment. Analysis of the residual urea showed that PPDA was significantly
more inhibitory in this experiment than in the first one.
In the first experiment, in which PPDA-containing urea granules were surface-
applied on unsaturated soil, there was a spatial separation of urea and PPDA because of
the differences in diffusive transports in unsaturated soils caused in part by differences
in their solubilities in water (0,6643 g of PPDA in 100 ml of water at 25°C; 108 g of
urea in 100 ml of water at 20cC). In the second experiment, in which a PPDA solution
and powdered urea were surface-applied, the spatial separation of urea and PPDA was
almost completely avoided and, therefore, PPDA could manifest its strong inhibitory
effect on soil urease activity.
Winiarski (1990) studied PPDA under conditions identical to those, under which he
studied dazomet and thione (see page 72). The rate of PPDA addition to urea was 0.5, I
or 1.5% (relative to weight of urea). Its inhibitory effect on volatilization of ammonia
from urea was stronger in light rather than heavy soil and was most pronounced at 1%
PPDA, and was not significantly different at rates of 1 and 1.5% PPDA. The inhibition
decreased with prolonging the incubation time from 7 to 14 days. However, even the
lowest inhibition, which was recorded in the heavy soil (at 0.5% PPDA, after 14-day
incubation), may be considered high (38.2%). In other pot experiments with both light
and heavy soils, PPDA was found to cause decreasing inhibitions when incubation
temperature increased from 10 to 20 and 30°C.
To study the influence of soil properties on the effectiveness of PPDA in reducing
ammonia volatilization from surface-applied urea, Watson (1990) used samples of 20
soils of widely differing physical and chemical properties from Northern Ireland. Urea
granules with or without PPDA were applied evenly to the soil surface to give 50 mg
129
N/sample (equivalent to approximately 100 kg Nlha). PPDA was incorporated within

the urea granules at a rate of 0.5 or 1% (on urea weight basis). During incubation, at
10DC, the volatilized NH3 was measured daily over 14 days.
The total cumulative loss of NH3 from samples without PPDA ranged from 0.37 to
29.2% (of the urea-N applied), depending on soil type, whereas the reduction of this
loss by PPDA varied between 0 and 91 %, depending on soil type, the average over all
20 soils being 30 and 36% for 0.5 and 1% PPDA, respectively.
The most important soil properties influencing the effectiveness of PPDA were soil
pH (in H2 0) and titratable acidity (TA): PPDA was less effective on soils with a high
pH and low TA. The conclusion was that a urease inhibitor should be more effective
than PPDA on soils with a high pH and low TA to be successful in reducing high
ammonia losses.
In Kucharski's (1992) laboratory experiments, 4-kg samples of a brown soil of
loamy sand texture (PH 6.3) from Poland were treated with urea granules (30 mg N/kg
soil) without PPDA or with PPDA impregnated into the granules at a rate of 1%
(relative to weight of urea). The samples were moistened to 60% ofWHC and incubated
for 20 days, during which the volatilized ammonia was measured. The cumulative NH3
losses represented 6.74 and 1.82% of the added urea-N in the urea and urea-PPDA
treatments, respectively. In other words, PPDA was efficient in reducing N losses from
urea.
In other laboratory experiments, Kucharski (1994) determined urease activity in 50-
g samples of the brown soil untreated or treated with urea (30 mg Nil 00 g soil) or with
urea + 1% PPDA and incubated at 10, 20 or 30°C for 2 up to 25 days. After each
incubation time, the total mineral N (NH/-N + N0 3--N) contents were determined.
Decrease of these contents in the urea+PPDA-treated samples as compared with those
treated with urea indicates the inhibition of the urease-catalyzed hydrolysis of urea.
Thus, after 25-day incubation the following total mineral N contents (mg/l00 g soil)
were found in the untreated, urea- and urea+PPDA-treated samples: 2.08, 22.65, and
17.11, respectively (at 10°C incubation temperature); 2.48, 25.1 0, and 19.02,
respectively (at 20°C), and 3.57, 13.20, and 16.50, respectively (at 30°C). These data
prove that PPDA inhibited urease activity at 10 and 20DC, but not at 30°C.
In a pot experiment, Fernandes da Mata and Drauschke (1996) compared
volatilization of ammonia from samples of a loamy soil (pH 6.4) fertilized with
granulated urea, S-coated urea or urea-PPDA. Following surface application, the
volatile NH3 loss from urea (14.4%) was reduced to 8.2% due to the S coating, but it
was not significantly affected by PPDA. When the fertilizers were introduced into the
soil (at 7.5-cm depth), the volatile NH3 losses were 13.4% (from urea), 1.4% (from S-
coated urea), and 10.6% (from urea-PPDA). The losses were further reduced by
applying urea supergranules.
2.20.3. Effect ofPhenylph osphorodiamidate (PPDA) on Immobilization ofUrea-N in

Soils
In a pot experiment with a German loamy sand soil sown with spring wheat and
fertilized with 15N-labeled urea with or without 1% PPDA, Matzel et al. (1979a,b)
found, by analyzing total N and 15N in soil and the plant at maturity stage, that PPDA
increased immobilization of urea-N in soil by 4% of the applied N and decreased
volatile losses of urea-N as ammonia by 10% (see page 269).
130
The effect of PPDA on immobilization of urea-N was also dealt with by

Hendrickson et 01. (1987), who used a silt loam soil from New York. Air-dried soil
samples (20 g) were mixed with 0.3 g of coarsely ground, dried oat straw. Other
samples were not amended with straw. All samples were then treated with 1 ml of water
or 1 ml of a solution containing 20 mg of N as urea, (NH4hS04 or KN0 3 and 5 ml of
water or 5 ml of a solution containing 0.4 mg of PPDA. The straw-amended samples
received additional water. Water potential in all samples was maintained at
approximately 30 kPa. Incubation took place in closed vessels for preventing volatile
ammonia losses, at 25°C. After 0, 3, 7,14, and 21 days of incubation the samples were
analyzed for urea, NH4 +, (NO}- + N0 2-), and pH. The added N sources not recovered as
mineral N were considered as immobilized N.
In samples not treated with PPDA, urea hydrolysis in unamended and straw-
amended soil was 70 and 63%, respectively, after 3 days and 100% in both samples
after 7 days. PPDA significantly delayed urea hydrolysis with only 11 and 13% of the
urea hydrolyzed after 3 days in the unamended and straw-amended soil, respectively.
Even after 7 days, only 42 and 81 % of the applied urea had been hydrolyzed in the
corresponding PPDA-treated samples. Urea hydrolysis resulted in an increase in pH
from 7.2 to 8.1-8.4. PPDA prevented this increase for 3 days in both unamended and
straw-amended soil, but after 7 days its effect was weak in unamended soil (PH 7.6) and
inexistent in the straw-amended soil (pH 8.3).
Figure 46 shows that immobilization ofN from the three N sources increased in the
order: KNO)«NH4hS04<urea. Much more N was immobilized in the straw-amended
than unamended soil. PPDA prevented immobilization ofurea-N in the unamended soil
for 7 days, after which immobilization proceeded as in the absence of PPDA. In the
straw-amended soil, PPDA eliminated immobilization for the first 3 days and markedly
reduced the extent of immobilization throughout the experiment. Immobilization can be
attributed to chemical and/or biological processes. In the straw-amended soil, the
biological fixation (microbial uptake of N~ + released from urea) made a greater
contribution to immobilization than the chemical fixation of NH4 +.
The first data on these investigations were published by Hendrickson et at. (1983).
2.20.4. Stability ofPhenylphospho rodiamidate (PPDA)
2.20.4.1. Stability o/PPDA in Solutions (Including Urea Melt) and in Solid State
Jasche et al. (1978) emphasized that to obtain urea-PPDA, addition of PPDA to urea
melt before the prilling process is not possible, because at the temperature of molten
urea PPDA undergoes a decomposition to form polyphosphates and other products
noninhibiting to urease activity. Therefore, for preparing fertilizer compositions
consisting of urea and PPDA, coating of urea prills with powdered PPDA was
recommended.
Martens and Bremner (l983a) studied the stability of PPDA in solutions. For
determination of PPDA a colorimetric method was elaborated. Water and isopropanol
were used as solvents. The solutions that contained 50 !!g PPDA/ml were refrigerated
(4-6°C) for 7 days and analyzed periodically. It was established that PPDA was stable in
isopropanol, but in aqueous solution only 43 !!g PPDA/ml could be evidenced after 7
days. Diminution of the initial concentration was attributed to partial hydrolysis of
PPDA.
131
~O.-----------------------------,
iZ 25
20 ~. ....
:....
.
....
·····0
"5
a? 15 :' l.rell-PPDA
o 3 T \4 2.\
60r-----------------------______ ~
B
=
.9
1< 20 .··0
;l;;I
~ •. 0·· . lJrea-PPIlA
j ~
0 ::::::::..... ..:::::;:::" •• ~::.::: •• -•.....•••~~?~....•
o 3 7 \4 2\
Incubation time (day.)
Figure. 46. Apparent immobilization of nitrogen applied to unamended soil (A) and straw·
amended soil (B). IAdapted from Hendrickson e/ al. (l987), by permission of the Soil
Science Society of America, Inc.!
Indirect evidence was also obtained concerning decomposition of PPDA dissolved

in water (Martens and Bremner, 1984a). Aqueous PPDA solution (50 Ilg PPDNml) was
stored at 20 or 30"C for 0-28 days. Reaction mixtures were prepared from 5 g of air-
dried soil + 1.5 ml of solution containing 10 mg of urea + 0.5 ml of stored PPDA
solution. Incubation took place at 20°C and lasted 2 days, after which the residual urea
was assayed. The results obtained (Table 38) show that effectiveness ofPPDA to inhibit
hydrolysis of urea in soil decreased with prolongation of storage time and following
TABLE 38. Effect of • to ring an aqueous solution ofPPDA on its effectiveness to inhibit
urea hydrolysis in soil"
Storage Inhibition of urea hydrolysis (%)
Soil temperature Time of storage (days)
(0C) 0 7 14 21 28
Silty clay loam 20 90 88 85 81
95
30 89 84 80 75
Loam 20 92 89 86 82
97
30 91 85 82 79
"From Martens and Bremner (1984a), by permission ofPergarnon Press PLC.
132
increase in storage temperature from 20 to 30o e, which proves that PPDA underwent a
partial decomposition during the storage.
Working with phosphate buffer solutions (pH 2-12) at 25, 35, and 45°e and
developing a method based on high-performance liquid chromatography for
determination of PPDA and of its organic hydrolytic products, and using other
analytical methods, too, Austin et al. (1984) elucidated the mechanism of acidic and
basic hydrolyses of PPDA. In strongly acid solutions, hydrolytic products of PPDA are
phenylphosphoroamidate (PPA) and ammonia, whereas in strongly basic solutions
hydrolysis of PPDA results in phenol and phosphorodiamidic acid. At intermediary pH
values, both hydrolyses occur, i.e .. both PPA and phenol are formed; at pH 6, phenol
predominates over PP A (Figure 47).
+ NH3
Phe nylphosphoroam idate

H20 (PPA)
+
~
Phenylphosphorodiamidate
(PPDA) o-OH
Phenol Phosphorodiamidic acid
0-
o
pH 2 II . . . . . . OH
O-P + NH3
+ H20 -
"'-OH
Phenylphosphoroamidate Phenylphosphoric acid
(PPA)
Figure 47. Mechanism of acid and basic hydrolyses ofPPDA and of acid hydrolysis ofPPA. IFrom Austin et
ai. (1984), by permission of the American Chemical Society.!
At pH 2, PPDA is very labile: approximately 4 mg of PPDA in 100 ml of buffer

solution completely hydrolyzed in about 20 minutes. At this pH, PPA is less labile, but
its hydrolysis also occurs although less rapidly: PPA was still at 74% of its maximum
obtained value after 138 minutes of reaction time. Hydrolytic products of PPA are
phenylphosphoric acid and ammonia (Figure 47). Increasing temperature led to
increased rate of both acid and basic hydrolyses of PPDA. However, the basic
hydrolysis rate was affected more than the acidic rate. For example, at 25°e the rate
constant at pH 12 was approximately 5 times greater than the rate constant at pH 2.
Gautney et al. (1983, 1984) (see also Anonymous, 1983), aiming at the elaboration
of a technology for cogranu1ation of urea and PPDA, determined the decomposition of
PPDA added to urea melt (140°C) in a proportion of 3 .2% relative to the weight of urea.
It was found that decomposition of PPDA obeyed first-order reaction kinetics;
calculation of reaction half-life at 1400 e for PPDA gave a value of 34 minutes. The
133
decomposition products were identical to those of the basic hydrolysis of PPDA, i.e ..
phenol and phosphorodiamidic acid. Weight loss of volatiles from a mixture of urea
melt (135°C) + 0.5, 2 or 4% PPDA was greater than from urea melted at the same
temperature. Thus, approximately 1% of the weight of mixture was lost in 20 minutes.
The loss of PPDA increased when the time of contact between urea melt and PPDA was
prolonged from 20 to 120 minutes, but remained under 10%. The loss was due to
volatilization of phenol and ammonia.
Since PPDA dissolves faster in the urea melt (140°C) than in concentrated urea
solutions and retention time for the complete urea granulation process is less than 1
minute, it was concluded was drawn that cogranulation of urea and PPDA leads to the
smallest losses when PPDA (0.5-4%) is added to the urea melt (l40°C) before
cogranulation. However, with this technology, the volatilized phenol, depending on its
concentration in the off-gas, could be an environmental problem as a source of air
pollution in the immediate area of the granulation plant.
Preparation of urea-0.5% PPDA granules is a 24% more expensive than the cost of
urea alone.
Gautney et at. (1985) evidenced the decomposition, at 25°C, of PPDA in seven fluid
fertilizers containing urea and other compounds (sources of N, P, K, and S) and being of
the following grades: 31-0-0, 36-0-0, 29-0-0-5S, 14-14-14,24-8-0,18-9-9, and 21-7-7.
In the first three fluid fertilizers, having a pH ranging from 7.08 to 7.65, PPDA
decomposed slowly, but its decomposition in the other fertilizers (pH 5.12-5.97) was
rapid. The mechanism of decomposition was basic or acidic hydrolysis as shown in
Figure 47. To minimize decomposition losses, PPDA should be added to the first three
fluid fertilizers not sooner than 1-2 days before their application on field. However,
some loss of PPDA would still occur, and it would be best to minimize the time
between PPDA addition and fertilizer application. The other four fertilizers should be
applied immediately after PPDA addition (see also Anonymous, 1983).
Gautney et al. (1983, 1984) mentioned tests which showed that dry solid PPDA and
urea-PPDA mixtures decompose very slowly to phenol at room temperature, indicating
that PPDA decomposition and evolution of phenol may be a problem during bulk
storage ofPPDA and granular urea-PPDA mixtures.
These tests were described in detail by Gautney et al. (1986). The following
preparations were studied:
-pure PPDA;
- pure PPDA to which impurities usually present in technical-grade PPDA
were added, namely (C 6HsO)zP(O)NH2 (diphenylphosphoroamidate, DPPA) and
NH4 Cl, in the proportions: 900 g ofPPDA + 100 g of DPPA or NH4 Cl; 900 g ofPPDA
+ 50 g of DPPA + 50 g ofNH 4 Cl;
- urea-PPDA mixtures, containing pure urea and pure PPDA in the proportions:
960 g of urea + 40 g of PPDA; 860 g of urea + 40 g of PPDA + 100 g of DPPA or
NH4 Cl; 860 gofurea +40 g ofPPDA + 50 gofDPPA + 50 gofNH4 Cl.
Samples (0.25 g) were weighed from each preparation, then placed in a forced draft
oven and heated at different temperatures (50-100°C) for various time periods (up to
120 hours). The effect of relative humidity (0-75%) on the decomposition of PPDA
alone at 90°C during the different times (up to 220 hours) was also studied. Then, each
sample was analyzed for residual PPDA by high-performance liquid chromatography.
134
The analyses showed that, under the influence of heat treatment, PPDA in each
preparation decomposed with the formation of phenol. The decomposition rate was
much higher in the urea-PPDA mixtures than in pure PPDA, and always increased with
the increase of temperature. The impurities (DPPA and NRtCl) alone and in
combination accelerated decomposition of pure PPDA, but did not significantly affect
decomposition of PPDA in urea-PPDA mixtures. The decomposition rate of PPDA
increased very much with an increase in relative humidity.
The decomposition of pure solid PPDA obeys first-order reaction kinetics, whereas
decomposition of PPDA in urea-PPDA mixtures obeys zero-order kinetics. The
calculations gave a value of 254 years as reaction half-life at 25°C for pure solid PPDA
and 56% decomposition per year at 25°C for PPDA in urea-PPDA mixtures. These data
suggest that the time between preparation (manufacture) and testing (field application)
ofurea-PPDA mixtures should be minimized.
The decomposition of solid PPDA at high relative humidities, where solid PPDA
liquefies, occurs by acidic and/or basic hydrolysis (see Figure 47). The mechanism of
PPDA decomposition at low relative humidities was not clarified; it was assumed on the
basis of some analytical data that in this case decomposition of PPDA proceeds
primarily via a combination hydrolysis-condensation reaction with the formation of a
diphosphorus compound containing NRt + and phenoxy (C6 HsO) substituents.
Byrnes et af. (l989a) studied the decomposition of PPDA in buffered solutions
made of KH 2P04 , H3 B03 , and CH 3COOH at concentrations of 0.024, 0.075, 0.15, and
0.30 M. The pH was adjusted to 5.0 by the addition of KOH. pH 7.0 solutions were
prepared with K2 HP04 , H3 B03 , and CH3COOK. At pH 9.0, a carbonate buffer made
from K2 C03 was used instead of acetate. Recrystallized PPDA was added to each buffer
solution to produce a concentration of approximately 25 ppm PPDA. The solutions were
kept at room temperature (- 20°C) and analyzed for PPDA at hourly intervals.
The decomposition of PPDA followed first-order reaction kinetics, in accord with
the results of similar studies performed by Austin et af. (1984) (see page 132). The rate
of PPDA decomposition increased linearly with increasing buffer concentrations.
Increasing the salt concentration shortened the half-life of PPDA to as little as one-
eighth its half-life without a buffer. PPDA was also greatly affected by the nature of
buffering salts: at pH 5.0, the effect of phosphate and acetate was strong, while borate
had no effect; at pH 7.0, phosphate and borate had little effect and acetate had no effect;
at pH 9.0, borate had a greater effect than phosphate and carbonate had an intermediary
effect.
The finding that decomposition of PPDA depends not only on pH, but also on
concentration and nature of buffering salts presents importance for studies on PPDA
decomposition in floodwaters overlying soils in which the presence of carbonate and
NH4 + due to urea hydrolysis may enhance the decomposition ofPPDA.
2.20.4.2. Stability ofPPDA in Soils

Metzel et al. (1978, 1979a) and Matzel and Heber (1979) considered that decomposition
of PPDA in soil can be deduced from weakening and then from the disappearance of its
inhibitory effect on urease activity. The moment of disappearance of this effect
corresponds to complete decomposition of PPDA which, in the soils studied by these
investigators, occurs in 2-3 weeks.
135
Following the fate of PPDA added to samples of 25 Iowa soils at a rate of 100 !!g!g
soil, Martens and Bremner (l983b) determined PPDA and inorganic N during
incubation of soil + PPDA mixtures at 20 D C for 5 days, and established that PPDA is
sorbed and decomposed in soils. The capacity of soils for sorption of PPDA was
positively and significantly correlated with clay content and surface area, but was not
significantly correlated with pH and organic matter content.
Continuing these investigations with a variety of soils, Bremner and Martens (1987)
found that the rate of decomposition of PPDA was not significantly affected when the
soils were steam-sterilized or treated with antimicrobial agents before addition of
PPDA. All these findings indicate that PPDA is decomposed in soils by nonbiological
processes. The conclusion that these processes are catalyzed by clay minerals was
supported by experiments with kaolinite, illite, and montmorillonite.
Investigations carried out at the International Fertilizer Development Center (Muscle
Shoals, Alabama) also showed that biological factors do not play any major role in the
decomposition of PPDA, since the decomposition rate was not much different in heat-
sterilized and unsterilized soils (Anonymous, 1986).
Observing that PPDA (added to urea in 1% proportion) did not exert toxic effects on
spruce, Mai and Fiedler (1986) advanced the idea that this water-soluble compound
decomposes in soil through pH-dependent hydrolysis with release of a-phosphate.
Because of the rapid decomposition of PPDA in flooded rice soils, Fillery and Vlek
(1986) express the opinion that more effective inhibitors are needed to completely
eliminate the accumulation of NH4 + in floodwater and thereby drastically reduce the
potential for ammonia volatilization after the application of urea.
Youngdahl et af. (1986) also emphasize the need for continuation of research to
identify new compounds that have a longer lasting effect than PPDA. Also, compounds
that are lower in cost should be sought.
From the findings by O'Connor et af. (1983) and O'Connor and Hendrickson (1987),
according to which in the soil studied (silt loam, pH 7.5) PPDA (20 !!g!g soil) prevented
urea hydrolysis and volatile NH3 losses for 1, 2, 4, 8, and 17 days at 35, 25, 15, 10, and
5°C, respectively, one can deduce that decomposition of PPDA in soil at low
temperatures was so slow that this compound exerted a remarkable retarding effect on
urea hydrolysis. With an increasing temperature, the rate of PPDA decomposition
increased to a large extent and at 35°C the inhibitory effect on soil urease activity was
largely diminished.
Continuing these investigations, Hendrickson and O'Connor (1987) added, to 20-g
samples of the same soil used in the investigations mentioned above, 6 rnl of water or 6
ml of a solution containing 0.4 mg of PPDA. TIle next step was incubation (more
precisely, preincubation) of samples at 5, 10, 15, 25, and 35°C for 0-28 days, after
which 1 rnl of urea solution (20 mg N) was added to each sample. TIle reaction mixtures
prepared in such a way were incubated at 25°C for 3 days, then the residual urea was
determined to evaluate the inhibitory effect ofPPDA on soil urease activity.
Preincubation of soil samples at 5-25°C gave different re~mlts from those obtained
after preincubation at 35°C.
With preincubation at 5-25°C, the inhibitory effect of PPDA decreased to 45-60% in
samples preincubated for 7 days, remained at this level in samples preincubated for 14-
21 days, then decreased again in samples preincubated for 28 days, in which the
136
inhibition was minimal (about 2S%) when preincubation took place at 2SoC and
maximum (about SO%) at preincubation temperatures ofS-lOoC.
After preincubation at 3SoC, the inhibitory effect of PPDA decreased to about 40%
in samples preincubated for 7 days, then increased markedly in samples preincubated
for 14 and 21 days, attaining about 80% in samples preincubated for 28 days.
To explain these unexpected results it is assumed that at 3SoC, during the fIrst 7 days
of preincubation, part of PPDA decomposed at a sufficiently high rate and the
decomposition products caused the increase of the inhibitory effect following
prolongation of preincubation time to 14-28 days.
In order to verify this assumption, soil samples (20 g) were preincubated with the
two hydrolytic products of the basic hydrolysis of PPDA (phenol and
phosphorodiamidic acid) as well as with two dihydric phenols (catechol and
hydroquinone). Preincubation and then incubation with urea were carried out as with
PPDA, but there were some differences, namely phenol was added to the soil samples at
the rate of 0.22 mg; preincubation temperatures were 2S and 3SoC (phenol), 10 and
2SoC (phosphorodiamidic acid), and 2SoC (catechol and hydroquinone).
Figure 48 shows that at 3SoC PPDA and phenol, applied at comparable molar rates,
brought about inhibitions to the same degree, but the inhibition caused by PPDA
appeared with a delay of about 3 days as compared to the inhibition given by phenol,
which suggests that the phenol released from PPDA did determine the increase in the
inhibitory effect of PPDA after prolongation of preincubation time. At 2SoC, PPDA was
a more effective inhibitor than phenol, but effectiveness of both compounds weakened
with prolongation of preincubation time.
100r-----------=:-~=:_:
.~·····-··~!:'!~~~.3.;;l
80 .......-- PPDA
.~/ )1'" we
\ i / r - ........
\... I PPDA
/
/\ \. I I
,,;1.. . 'ri
20 .D ..
Preincubation tUne (days)
Figure 48. Inhibition of urease activity in soil samples preincubated with PPDA (20 J.lglg soil) and phenol (11
J.lglg soil) at 25 and 35°C./From Hendrickson and O'Connor (1987), by permission ofPergamm Press PLC}
Phosphorodiarnidic acid in samples not submitted to preincubation exhibited only a

20% inhibition, whereas PPDA completely inhibited urease activity in these samples
and was also more effective in the preincubated samples. The inhibitory effect of
phosphorodiamidic acid was also more evident at 10 than at 2SoC. Thus,
137
phosphorodiamidic acid did not contribute to the increase in the inhibitory effect of
PPDA in samples preincubated for 14-28 days.
Inhibitory capacity of catechol and hydroquinone on urease activity manifested a
tendency to decrease in soil samples, in which these hydroxylated phenols were
preincubated at 25°C for 28 days.
In a field experiment on a silt loam soil (PH 5.7) from Indiana, Beyrouty et al.
(1988a) have applied urea prills or urea prills coated, by means of paraffin oil, with
PPDA (urea: 200 kg Nlha; PPDA: 4 kg/ha) on the surface of microplots not covered or
covered (in a proportion of approximately 60%) by maize (Zea mays) residue left from
the previous year's crop. The unhydrolyzed urea was determined periodically during 24
days after fertilization. The results showed that PPDA retarded urea hydrolysis for 19
days in each microplot. It was deduced from this finding that there was no deactivating
interaction between maize residue and PPDA, i.e.. stability of PPDA was not reduced
by the maize residue. The PPDA molecules diffused into the residue material may even
be protected from the deactivating action of higher temperatures. It should be added
that, under similar conditions, trichloroethylphosphorodiamidate was unable to retard
urea hydrolysis which suggests some type of deactivating interaction of this compound
with the plant residue.
Results of a laboratory experiment performed by Beyrouty et al. (l988b) point out
the importance of soil pH for stability of PPDA. Samples of an Indiana soil, collected
from plots submitted to long-term liming, had pH values of 5.6, 6.5, and 7.2, and their
texture was silty clay loam, silt loam, and silty clay loam, respectively. Reaction
mixtures were prepared from 20-g air-dry soil samples, 4.8 mJ of water or 4.8 ml of urea
solution (20 mg N) with or without 0.4 mg of PPDA, and incubated at laboratory
temperature for 14 days, during which time the evolved ammonia was assessed. At pH
5.6, cumulative NH3 losses from the applied urea-N represented 5.6% in urea-only
treatment and 0.3% in the urea-PPDA treatment. At pH 6.5 the corresponding values
were 25.0 and 15.2%, respectively. At pH 7.2 the cumulative NH3 losses were similar,
31.2 and 33.1 %, respectively. It is evident from these data that the inhibitory effect of
PPDA, which was nearly total at pH 5.6, decreased very much at pH 6.5 and
disappeared at pH 7.2. Reduction or disappearance of the effect of PPDA can be
explained by partial or total decomposition of PPDA at these pH values.
Savant et af. (l988a), who proved that inhibitory effectiveness of PPDA on urea
hydrolysis decreases with increasing bulk density (see page 127), also studied the
influence of soil bulk density on stability (and decomposition) of PPDA.
Columns of an unsaturated clay soil (0.16 g water/g soil), installed in cylinders were
compacted to an average bulk density of 0.70,0.99 or 1.56 g/cm3 • One ml of aqueous
solution containing 0.5 mg of PPDA was applied to the surface (12.6 cm 2) of the
columns. After having been covered by Parafilm for preventing evaporation of water,
the columns were preincubated at 30°C for 0, 7, 14, and 30 days, then 50 mg of
powdered urea was uniformJy placed on their surface. Subsequently they were again
incubated for 16 hours, extracted, and the unhydrolyzed urea was determined.
The results confirmed the finding that the inhibitory effect of PPDA on urea
hydrolysis decreased with increasing soil bulk density. In addition, they showed that the
inhibitory effect of PPDA decreased during the 30-day preincubation at each bulk
density. It was also established that the negative slopes of the three plots presenting
percent inl1ibition versus preincubation time for the three bulk densities were nearly the
138
same, which suggests that the rate of decomposition of PPDA was not influenced very
much by soil bulk density.
Dealing with the degradation of urease inhibitors in soils, Lee and Radel (1988) pay
special attention to the literature data on degradation of PPDA, and present a brief
characterization of the new and improved methods for trace analysis of urease inhibitors
and their degradation products in soils.
***
Decomposition in soil of the PPDA derivative 3-(1 ',1 '-dimethylethyl)-4-hydroxy-
PPDA was demonstrated during studies of its effect on nitrification (Swerdloff et al..
1985a; see page 239).
2.21. 2-AMINE-2-0XIDE-1,3,2-BENZODIOXAPHOSPHOLE AND ITS

PHOSPHORODIAMIDIC ACID ESTERS
For inhibition of urease activity, Swerdloff et al. (1985b) patented 2-amine-2-oxide-

1,3,2-benzodioxaphosphole and 2-amine-2-thio-l,3 ,2-benzodioxaphosphole (which are
derivatives of phosphoroamidic acid and thiophosphoroamidic acid, respectively) as
well as their derivatives, including their phosphorodiamidic acid and thiophosphoro-
diamidic acid esters. There are nominalized in the patent 34 substituted 2-amine-2-
oxide-l,3,2-benzodioxaphospholes and two phosphorodiamidic acid (2-arnine-2-oxide-
1,3 ,2-benzodioxaphosphole)esters, and 15 substituted 2-amine-2-thio-1,3 ,2-benzodioxa-
phospholes and two thiophosphorodiamidic acid (2-amine-2-thio-l,3,2-benzodioxa-
phosphole)esters.
2-Amine-2-oxkle- Phosphorodiamidic acid 4-(2-amine-2-oxide-

1,3,2-benzodioxaphosphole 1,3,2-benzodioxaphosphole)esler
Phosphorodiamidic acid 5-(2-amine-2-oxide-

1,3,2-benzodioxaphosphole)esler
Figure 49. Structure of 2-amine-2-oxide-l,3,2-benzodioxaphosphole and of its two phosphoro-

diamidic acid esters tested by Swerdloffet al. (l985b) for inhibition of soil urease activity.
139
For the three compounds considered the most effective, the patent also describes the
testing of their inhibitory effect on urease activity in a New York soil (Cazenovia silt
loam, pH 7.0). The testing method was the same as that used by Kolc et af. (1985b) (see
page 80).
The three compounds tested (Figure 49) produced the following percent inhibitions:
54,5, and 21, respectively. Thus, the compound having the smallest molecule was the
most effective soil urease inhibitor.
The most preferred rates, at which these compounds are recommended for practical
applications, range from 1 to 500 ppm relative to soil weight, from 0.28 to 16.8 kg/ha,
from 0.01 to 10% relative to weight of urea in liquid or solid fertilizers.
2.22. POLYPHOSPHORODIAMIDES
In their patented invention, Swerdloff et al. (1985c) nominalized 191 polyphosphoro-

diamide compounds, of which 155 are diphosphorodiamides, 22 dithiophosphoro-
diamides, 10 triphosphorodiamides, 3 trithiophosphorodiamides, and 1 tetraphosphoro-
diamide. In the case of six especially efficacious compounds for use in the practice,
their testing as inhibitors of urease activity in a New York soil (Cazenovia silt loam, pH
7.2) was also described. These six compounds are diphosphorodiamides (Figure 50).
The testing method was identical to that applied by Kole et af. (l985b) (see page 80).
>o-
o
o 0 H:!N U 0
H:!N II II/NH2 -"""P- HN
H:!N/
yH 3 II NH2
C-NH-p(
-...... p--HN-(CH:!)6-NH - P
H:!N/ 'NH 2 IiJC tH3 NH2
N,N'-Bis-(diaminophosphinyl)- N,N'-Bis-(diaminophosphinyl)-
1,6-diaminohexane 1,8-diamino- p-menthane
N,N'-Bis-(diaminophOsphinyl)- Phosphorodiamidic acid

piperazine 1,4-phenylene ester
Phosphorodiamidic acid Phosphorodiamidic acid (l-methylethylidene)-

1,3-phenylene ester di-4, l-phenylene ester
Figure 50, Structure of the diphosphorodiamide compounds tested by Swerdloff et al. (l985c) for
inhibition of soil urease activity.
140
The inhibitions caused by the six diphosphorodiamide compounds were 86, 84, 76,
90, 81, and 70%, respectively. It is evident that the most inhibitory compound was
phosphorodiamidic acid 1,4-phenylene ester.
The most preferred amounts in which these urease inhibitors are recommended to be
used range from I to 500 ppm of soil weight, from 0.56 to 16.8 kg/ha, and from 0.1 to
20% of weight of urea in liquid and solid fertilizers.
2.23. OXlMATED O-DIAMINOPHOSPHINYL DERIVATIVES
Of the 33 O-diaminophosphinyl and 7 O-diaminothiophosphinyl derivatives of oximes

nominalized as inhibitors of soil urease activity in the patent of Swerdloff et al. (1986a),
only one derivative was tested, according to the patent description, for evaluation of the
urease-inhibiting capacity. Two soils were used, a New York soil (Cazenovia silt loam,
pH 7.2) and a Wisconsin soil (Plano silt loam, pH 5.4). The testing method was the
same as that applied by Kole et af. (1985b) (see page 80).
Fif;ure 51. Structure of O-(diaminophosphinyl)-2-propanone oxime tested by Swerdloff et

al. (I 986a) for inhibition of soil urease activity.
The tested compound (Figure 51) induced 84 and 53% inhibitions in the urease
activity of the Cazenovia and Plano soils, respectively.
For use in practice, the oxirnated O-diaminophosphinyl derivatives are
recommended in the following most preferred amounts: I to 500 ppm of soil weight,
0.28 to 16.8 kg/ha, 0.01 to 20% of weight of urea in liquid and solid fertilizers.
2.24. OXIDIZED DIAMINOPHOSPHINYL SULFUR DERIVATIVES
In the patents of Swerdloff et al. (l986b, 1987), 84 oxidized diaminophosphinyl sulfur

derivatives are nominalized as N-(diaminophosphinyl)sulfonamides (25), -sulfarnides
(15) and -sulfinamides (11), and as N-(diaminothiophosphinyl)sulfonamides (9),
-sulfamides (13) and -sulfinamides (11). Testing of urease-inhibiting capacity of only
two compounds (Figure 52) is described in the patents. Two soils (Cazenovia silt loam,
pH 7.3; Plano silt loam, pH 5.3) were used. and the testing method was the same as
141
N-(Oiaminophosphinyl)benzenesulfonamide N-(Oiaminophosphinyl)-p-loluenesulfonamide
Figure 52. Structure of the two oxidized diaminophosphinyl sulfur derivatives tested by Swerdloff et al.
(l986b. 1987) for inhibition of soil urease activity.
that applied by Kolc et al. (1985b) (see page 80).

Both compounds inhibited urease activity in both soils. The inhibitions induced by
N-(diaminophosphinyl)benzenesulfonamide were 91 and 23%, and those caused by N-
(diaminophosphinyl)-p-toluenesulfonamide were 86 and 51%, in the Cazenovia and
Plano soils, respectively.
For use in practice, the oxidized diaminophosphinyl sulfur derivatives are
recommended in the same most preferred amounts as the oximated O-diamino-
phosphinyl derivatives (see the preceding subchapter).
2.25. DIAMIDOPHOSPHOROTHIOLATE AND DIAMIDOTHIOPHOSPHORO-

THIOLATE COMPOUNDS
In their patents, Van Der Puy et al. (1984a, 1985a) nominalized 74 diamidophosphoro-
thiolates [R-S-P(O)(NH2)2l and 71 diamidothiophosphorothiolates [R-S-P(S)(NH2hl as
inhibitors of soil urease activity and tested the urease-inhibiting capacity with five S-
alkyl and one S-cycloalkyl diamidophosphorothiolates and, in comparison, with two
alkyl phosphorodiamidates [R-O-P(O)(NH2)2l. Two soils (Cazenovia silt loam, pH 7.3;
Plano silt loam, pH 5.4) were used. The testing method was the same as that applied by
Kole et af. (l985b) (see page 80), but the amount of most test compounds was reduced
from 0.8 to 0.2 mg/20 g soil. The results are reproduced in Table 39.
TABLE 39. Effect of some diamidophosphorothiolate (DAPT) and phosphorodiamidate

(PDA) compounds on soil urease activity"
Inhibition (%)
Compound Amount (flglg soil)
Cazenovia soil Plano soil
S-Methyl-DAPT 40 93 93
S-(n-Butyll-DAPT 20 74 82
S-(sec-Butyl)-DAPT 20 74 84
S-(iso-Butyl)-DAPT 20 80 83
S-{n-Hexyl)-DAPT 20 52 66
S-Cyclohexyl-DAPT 20 66 58
Methyl-PDA 40 34 N.D.
n-Propyl-PDA 40 15 N.D.
"Adapted from Van Der Puy el al. (l984a, 1985a).
N.D.-· Not determined
142
Table 39 shows that each DAPT compound inhibited urease activity in both soils.
The S-butyl-DAPTs were more inhibitory than the S-hexyl-DAPTs. The inhibitory
effect of S-methyl-DAPT exceeded by nearly 2.8 times that of the methyl-PDA.
The most preferred amounts recommended for practical use of diamidophosphoro-
thiolate compounds range from about 10 to about 500 ppm of soil weight and from
about 0.0 I to about 20% of weight of urea in liquid and solid fertilizers.
2.26. PHOSPHORIC TRIAMIDES AND THIOPHOSPHORIC TRIAMIDES
2.26.1. The Patented Compounds and the First Studies on Their Inhibitory Effect on
Soil Urease Activity
The number of N-aliphatic + N-aryl + N-aliphatic-N-aryl phosphoric triamide (PTA)

compounds [R-HN-P(O)(NH2h; R\R2N-P(O)(NH2h] and that of N-aliphatic
thiophosphoric triamide (TPT A) compounds [R-HN-P(S)(NH2)2] nominalized as urease
inhibitors in patent descriptions are 125 and I (Swerdloff et ai., 1984), 66 and 0
(Swerdloff et al., 1985d), and 60 and 1 (Van Der Puy et al., 1985b), respectively.
Eight of the 125 PTA compounds nominalized by Swerdloff et al. (1984) and three
of the 66 and 60 PTA compounds nominalized by Swerdloff et at. (1985d) and Van Der
Puy ef at. (l985b), respectively, were tested to evaluate their inhibitory capacity on
urease activity in a New York soil (Cazenovia sandy loam, pH 7.2). The same testing
method was used as that applied by Kole et al. (1985b) (see page 80), but the test
compounds were applied at two rates: 0.8 and 0.02 mg/20 g soil. The results are
summarized in Table 40.
TABLE 40. Effect of some N-aliphatic, N-aryl, and N-aliphatic-N-aryl phosphoric

triamide (PTA) compounds on soil urease activity"
Inhibition (%)
Compound Amount of compound (/lg/g soil)
40 I
N-(2-Chlorocthyl)-PTA 100 77
N-(3-Brol11opropyl)-PT A 99 83
N.N-Bis-(2-chloroethyl)-PT A 30
N-Phenyl-PTA 74 19
N-( 4-Nitrophcnyl)-PTA 81 30
N-Methyl-N-( 4-nitrophenyl)-PT A 95 59
N-Methyl-N-(4-hydroxyphcnyl)-PT A 86
N-Methyl-N-( 4-mcthoxyphcnyl)-PT A 47
"Adapted from Swcrdlolf el at. (1984, 1985d) and Van der Puy el at. (1985b).
It is evident from Table 40 that N-(2-chloroethyl)-PT A and N-(3-bromopropyl)-PT A

were the strongest inhibitors and N,N-bis-(2-chloroethy1)-PT A was the weakest one; the
inhibitory effect of N-methyl-N-(4-nitrophenyl)-PTA was stronger than that of N-(4-
nitrophenyl)-PT A.
There are nominalized in the patents of Kole et al. (1984, 1985d) 85 N-aliphatic
and N,N-aliphatic phosphoric triamide (PTA) and 81 N-aliphatic and N,N-aliphatic
143
thiophosphoric triamide (TPTA) compounds, including one of the most studied

inhibitors of soil urease activity, namely N-(n-butyl)thiophosphoric triamide (nBTPTA).
The majority ofTPTA compounds differs from the PTA compounds only by containing
a sulfur atom, instead of an oxygen atom, bound to the phosphorus atom.
The urease-inhibiting capacity was tested for 13 PTA and 9 TPTA compounds, by
using samples of two soils (Cazenovia silt loam, pH 7.0 or 7.3; Plano silt loam, pH 5.3)
and the same testing method as that applied by Kolc et al. (1985b) (see page 80). The
amount of 15 test compounds was 0.8 mg and that of 7 test compounds was only 0.2
mg/20 g soil.
Of the compounds tested at the rate of 40 !!g/g soil, the most potent urease inhibitors
were N-(n-butyl)-TPTA, N-(n-butyl)-PTA, and N-(sec-butyl)-PTA which caused 98
and 95%, 99 and 52%, and 96 and 66% inhibitions of urease activity in the Cazenovia
soil (pH 7.3) and Plano soil, respectively.
At the rate of 10 !!g test compound/g soil, N-(n-hexyl)-TPTA and N-cyclohexyl-
TPTA were the most inhibitory compounds, producing 97 and 52%, and 84 and 58%
inhibitions in urease activity of the Cazenovia soil (pH 7.0) and Plano soil, respectively.
Swerdloff et al. (1985a) nominalized, in the description of their patent, 76 aryl-PTA
and 7 aryl-TPTA compounds as urease inhibitors, but testing of the inhibitory effect on
soil urease activity is described in the patent only in the case of N-methyl-N-(4-
hydroxyphenyl)-PTA and N-(4-aminophenyl)-PTA. The two PTA compounds, used at a
rate of 0.2 mg/20-g soil sample treated with 42.8 mg of urea, caused 87 and 55%
inhibitions, respectively, in the urease activity of the Cazenovia silt loam (PH 7.2)
during 3-day incubation at 25°C.
In the patent description of Kole et at. (1985a) there are nominalized 46 N-acyl
phosphoric triamide (PTA) and 36 N-acyl thiophosphoric triarnide (TPTA) compounds
as inhibitors of soil urease and/or nitrification. Four of these compounds were tested
with the Cazenovia silt loam soil (pH 7.2) by using the same method as that also applied
by Kole et al. (1985b) (see page 80). The tested compounds and their urease-inhibiting
capacity are specified in Table 41.
TABLE 41. Effect of some N-acyl phosphoric triamide compounds on soil urease activity"
Compound Structural formula Inhibition (%)
N-(DAP)-2-chloroacetamide CHzCI-CO-NH-P(O){NHzh 83
N-(DAP)-2,2-dichloroacetamide CHCb-CO-NH-P(O)(NHz)2 73
N-(DAP)-2,2,2-trichloroacetamide CCh-CO-NH-P(O)(NH2)2 64
N-(DAP)-2,2,2-trifluoroacetamide CF3-CO-NH-P(O) (NH212 52
"Adapted from Kolc et al. (l985a).
(DAP) - (Diaminophosphinyl).
The data of Table 41 indicate that the increase in number of the CI atoms in the
acetamide moiety of the tested compounds led to a decrease in their urease-inhibiting
capacity.
The first three compounds specified in Table 41 and two other N-acyl PTA
compounds were also tested with jackbean urease and it was found that at 10-Ji M
concentration they induced 56-100% inhibitions in the activity of this urease.
144
In another patent, Kole et al. (1985c) nominalized 14 PTA and 2 TPTA compounds
as urease inhibitors, but none of them was tested for evaluating their effect on soil
urease activity. The only compound tested with jackbean urease (Figure 53) proved to
be a potent inhibitor of this urease.
N-(Diaminophosphinyl)-4-(l'-maleimido)benzamide
Figure. 53. Structure of the compound tested as a urease inhibitor by Kolc et al. (l985c).
The PTA and TPTA urease inhibitors patented by Swerdloff et al. (1984, 1985a,d),
Kole et af. (1984, 1985a,c,d), and Van Der Puy et al. (1985b) are recommended for use
in practice in the most preferred amounts ranging from about 1 to about 500 ppm of soil
weight, from 0.28 to 16.8 kglha, from 0.01 to 20% of weight of urea in liquid and solid
fertilizers.
Omilinsky et af. (1997) invented improved solvent systems which enable the
preparation of stable concentrated solutions of N-alkyl thiophosphoric triamide urease
inhibitors (including nBTPTA) for their storage, transportation, impregnation onto solid
urea fertilizers and incorporation into liquid urea fertilizer compositions. These solvent
systems are based on glycols and glycol derivatives.
Sulzer et al. (1998) patented an improved technology which makes it possible to
efficiently produce large scale commercial quantities of N-hydrocarbyl thiophosphoric
triarnides, including nBTPTA at high yields, by means of a continuous process.
Investigations performed at the National Fertilizer Development Center (Tennessee
Valley Authority, Muscle Shoals, Alabama) showed that thiophosphoryl trian1ide, H2N-
P(S)(NH 2h (TPTA) inhibits soil urease activity (Anonymous, 1985a; Radel et al.,
1987). Based on these and further investigations, TPTA was patented as an inhibitor of
soil urease by Gautney (1987) and as a dual purpose, soil urease and nitrification
inhibitor by Radel (1990).
TPTA was tested with two reference compounds (PPDA and phosphoryl trian1ide,
PTA) with two testing techniques.
In the first technique (testing in "banded configuration"), the powdered mixture of
urea and test compound (10% on urea basis) was applied in narrow bands in the soil,
and after 3- and 6-day incubations the remaining urea was extracted and determined. It
was found after 3 and 6 days that the added urea remained unhydro1yzed in the
following proportions: 72 and 33% (under the influence of TPTA) , 63 and 10%
(PPDA), and 54 and 3% (PTA), respectively.
In the second technique (testing in "well-n1ixed configuration"), solutions or
suspensions of the compounds are well-n1ixed throughout the soil, then the mixtures are
submitted to preincubation at 30°C for 0, 3, 7, 14, 21, and 36 days. Urea is added to the
preincubated mixtures and they are then incubated again, for 16 hours. The incubation is
145
followed by determination of the unhydrolyzed urea and calculation of the percent

inhibition of urease activity. The results showed that in the mixtures not preincubated,
the most effective inhibitor was PPDA (100% inhibition) with TPTA and PTA
inhibiting only 59 and 29%, respectively. But in the preincubated mixtures the strongest
inhibitions were caused by TPTA. We quote only the inhibition values registered in the
mixtures preincubated for 3, 7, and 21 days: 53,53, and 26% (TPTA), 35, 28, and 0%
(PPDA), and 4, 12, and 0% (PTA), respectively (Anonymous, 1985a; Radel et aI.,
1987).
TPTA was also tested by using as a substrate not urea, but urea-ammonium nitrate
(DAN) fluid fertilizers: VAN suspension of grade 36-0-0 and VAN solution of grade
31-0-0. PPDA served as a reference compound. The testings were performed in banded
configuration. Containers were one half-filled with soil (silt loam), then samples of fluid
fertilizers without or with TPTA or PPDA at a rate of about 11 % (on urea basis) were
distributed evenly in a narrow band 6 cm along the soil surface. In the next step, the
containers were filled with soil and incubated at 25°C for 3 and 6 days. After
incubation, the residual urea was determined. In the case of 36-0-0, urea remained
unhydrolyzed after 3 and 6 days in proportions of 88 and 48% under the influence of
TPT A, and 84 and 11 % under the influence of PPDA, respectively. In the soil not
treated with inhibitors the unhydrolyzed urea represented 31 and 0.6%, respectively. In
the case of 31-0-0, the corresponding values were 98 and 48% (TPTA), 83 and 13%
(PPDA), and 39 and 0.7% (no inhibitor), respectively. Consequently, in VAN fluid
fertilizers, TPTA was more effective than PPDA (Anonymous, 1987; Radel et aI., 1987;
Gautney, 1987).
The concentration limits, between which TPTA is recommended to be added to
urea-containing fluid fertilizers, are 0.01 and 10% (on urea basis) for fluid fertilizers of
pH 5.5-10. The 0.5-5% TPTA concentrations are the preferred ones for fluid fertilizers
of pH 6.5-9 and the most preferred ones for fluid fertilizers of pH 7.4-8.5 (Gautney,
1987).
The linear thermal polymers of TPTA were also considered highly effective
inhibitors of soil urease activity (Gautney, 1987), and testing of the linear thermal
pol ymers of PTA and TPTA was described by Radel et al. (1994).
The polymers of PTA were tested in banded configuration and those of TPTA in
both banded and well-mixed configurations according to the techniques published by
Anonymous (l985a) and Radel et al. (1987) (see page 144). PPDA and the monomers
served as reference compounds. The soils used were silt loams.
The polymers of PTA were weaker urease inhibitors than was PPDA, but nearly
equal or slightly better ones than PTA itself. The polymers ofTPTA were significantly
better inhibitors than those of PTA and some of them had greater persistence and thus
exhibited stronger urease-inhibiting capacity than PPDA and TPTA. The polymers
consisting of 2 to 4 monomers were most inhibitory.
The problem encountered in the production of these polymers was that wide
variation in the product mixtures may preclude their use as practical urease inhibitors.
In light of the investigations proving the soil urease-inhibiting capacity of TPTA, it
is understandable that Radel's (1990) patent, presenting TPTA as an inhibitor of both
soil urease activity and nitrification, deals nearly exclusively with its nitrification-
inhibiting capacity (see page 240).
146
2.26.2. Further Studies on Phosphoric Triamides and Thiophosphoric Triamides

Bremner and Chai (1986) described experiments aimed at establishing the influence of
various factors on the effectiveness of N-(n-butyl)thiophosphoric triamide (nBTPTA).
Thirteen Iowa soils were studied; they were selected to obtain a wide range in
propertics. The reaction mixtures (5 g of air-dried soil + 2 ml of solution containing 10
mg of urea without or with 50 ~g of nBTPTA) were incubated at 20°C for 7 days, then
analyzed for residual urea. The analytical data served for calculation of percent
inhibitions in urease activity.
The inhibitions recorded in urease activity of the 13 soils studied ranged from 40 to
90%. Simple correlation analyses showed that percent inhibition of urea hydrolysis by
nBTPTA was significantly correlatcd with organic e content (r=-0.70), total N content
(r=-0.76), cation-exchange capacity (r=-0.67), sand content (r=0.61), clay content
(r=-0.76), and surface area (r=-0.66), but was not significantly correlated with pH
(r=O.21), silt content (r=-0.38), Caeo 3 equivalent (r=0.24), and urease activity (r=-0.29)
in soils. Multiple regression analyses indicated that organic matter content is the most
important factor for the effectiveness of nBTPTA in the sense that this effectiveness
tends to increase with a decrease in soil organic matter content.
Another experiment was carried out with six Iowa soils. The reaction mixtures,
containing nBTPTA amounts between I and 125 ~gl5 g soil, were incubated at 20D e for
2 and 10 days. Percent inhibition of urea hydrolysis increased with the increase in
inhibitor concentration and the effect of higher concentrations was considerably more
marked after 10 days of incubation than after 2 days.
The same six soils were also used for studying the influence of time of incubation
(1-21 days) at 200 e on the inhibitory effectiveness of 50 ~g of nBTPTAJ5 g soil. It was
established that percent inhibition decreased markedly with incubation time. On the
average, the inhibition registered after 21 days was only half of that recorded after I
day.
The influence of incubation temperature (I 0-40°C) was the objective of another
experiment also performed with six soils. nBTPTA rate was 50 ~gl5 g soil. Incubation:
3, 7, and 14 days. Effectiveness of inhibition decreased very much with increase in
temperature and the decrease became more pronounced following prolongation of
incubation time.
Joo et at. (1987) studied the effect of nBTPTA on volatilization of ammonia from
urea under field conditions on a fine loamy soil (pH 7.5) from Iowa. Microplots covered
by Kentucky bluegrass (Poa pratensis) turf were fertilized monthly in June through
September in 1986, with liquid urea (495 llha, containing 49 kg Nlha) with or without
0.5, 1 or 2% nBTPTA relative to urea-No Ammonia volatilized during 0.25, 1, 2, 3, and
4 days after each fertilization was measured. The cumulative NH3 losses in 4 days
represented 18.5% of the urea-N applied (in control without inhibitor) and only 6.9% in
treatments with 0.5 and 1% nBTPTA and 5.6% in treatment with 2% nBTPTA.
Carmona et at. (1988, 1990) pcrformed several laboratory experiments related to
nBTPTA.
In the first experiments, the effect of different concentrations of nBTPTA on
anunonia volatilization from urea-treated samples of a Guthrie soil (pH in H20 6.2; clay
28 1}!o; silt 61 %; organic matter l.2%) was studied. In the apparatus used, sand (6.5 kg)
was placed in 8.5-1 pots and saturated with water, then sufficient air-dried soil was
placed on the wet sand to result in a lO-cm thick soil layer, which was wet to field
147
capacity by capillarity. The soil samples were limed (1% on soil weight basis) to raise
pH to 7.5, then amended with urea (at a rate equivalent to 100 kg Nlba on a surface area
basis) containing 0, 0.005, 0.01, 0.05,0.1 or 0.5% nBTPTA (on urea weight basis) and
incubated at laboratory temperature (20-23°C) for 14 days, during which time the
volatile NH3 was measured.
The cumulative NH3 losses decreased with increasing nBTPTA concentrations, but
these effects of 0.1 and 0.5% nBTPTA were not significantly different. By day 14, the
losses were 32.2, 14.7, and 6.1 % of applied urea-N for the 0.005, 0.01, and 0.05%
nBTPTA, respectively, compared with a 52.0% loss from urea without nBTPTA.
In other experiments, the limed Guthrie soil and a Savannah soil (PH in H20 5.7;
clay 17%; silt 21 %; organic matter 0.9%) were used and both ammonia volatilization
and urea hydrolysis were studied. The urea rate was the same as in the first experiments,
but concentrations of nBTPTA were only 0.01, 0.05, and 0.1 %. The incubation was
carried out at laboratory temperature. The volatilized NH3 was measured during 12-14
days, and the unhydrolyzed urea was determined four times during 10 days. These
experiments were repeated using the same two soils, into which 109 of finely ground
soybean straw had been mixed in the surface 2 cm. These soils were kept moist for 30
days to establish a stable microbial population.
Ammonia volatilization losses from urea without nBTPTA ranged from 47 to 73%.
nBTPTA at all concentrations resulted in significant reductions in NH3 loss and the
losses decreased with increasing concentration of nBTPTA during the first days of
incubation. Thus, after 6-day incubation, 54.3% of urea-N had been lost from the
Savannah soil treated with urea only, whereas losses of NH3 were 9.4, 3.4, and 2.7%
from urea containing 0.01, 0.05, and 0.1% nBTPTA, respectively. As the incubation
continued, the NH 3 losses began to increase from urea-nBTPTA, but at 12-14 days, they
were still much less than those from urea alone.
The effectiveness of nBTPTA was reduced by amending the soils with straw. Thus,
NH3 losses from the straw-amended Savannah soil after 6-day incubation was 25.2, 3.5,
and 2.5% for the 0.01, 0.05, and 0.1 % nBTPTA added to urea, respectively, and 54.0%
from urea alone.
Urea hydrolysis was greatly inhibited by nBTPTA in both soils and the degree of
inhibition increased with the rate of nBTPTA. For example, after 10 days of incubation,
in the Savannah soil without straw, 96.2% of urea had been hydrolyzed, whereas
hydrolysis of urea containing 0.01, 0.05, and 0.1% nBTPTA was 66.6, 51.2, and 44.6%,
respectively. The corresponding values in the straw-amended Savannah soil were 100,
82.1, 50.8, and 34.8%, respectively. In other words, the straw amendment caused some
reductions in the urease-inhibiting capacity of nBTPTA.
Experiments were also carried out to study the effect of temperature on the
effectiveness ofnBTPTA as an inhibitor of ammonia volatilization and urea hydrolysis.
The Guthrie soil was used. The rate of urea application was the same as in the first
experiments, whereas the rate of nBTPTA was 0.01, 0.05 or 0.1 % relative to weight of
urea. The incubation was carried out in growth chambers at 18, 25, and 32°C and lasted
12 days (NH3 volatilization experiments) or 10 days (urea hydrolysis experiments).
The inhibitory effect of nBTPTA on NH3 volatilization decreased with increasing
temperature. Thus, the cumulative volatile NH3 losses at 18, 25, and 32°C in 12 days
were 42.3, 49.4, and 55.5% (urea alone); 16.8, 23.5, and 47.8% (urea + 0.01%
nBTPTA); 11.7, 17.8, and 42.4% (urea + 0.05% nBTPTA), and 4.3, 11.2, and 28.9%
148
(urea + 0.1% nBTPTA), respectively. The inhibitory effect of nBTPTA on urea

hydrolysis also decreased with increasing incubation temperature.
In conclusion, the inhibitory effect of nBTPTA on urease activity manifested itself
in a similar manner in the two, largely different, soils studied; only the extent of
inhibition showed some differences between the two soils.
Bronson et al. (1989) confirmed the effectiveness of nBTPTA in diminishing
ammonia losses from urea, under field conditions on an Alabama soil (loamy sand, pH
6.3). The experimental plots were planted with maize (Zea mays). At week 6 after
planting, urea, urea + 0.25 or 0.50% (on urea-N basis) nBTPTA and NH4N0 3 at a rate
of 134 kg N/ha were surface-applied. Volatilization of NH3 was determined during 9
days. The cumulative N losses as volatile NH3 were the following: 20.5 kg/ha (urea-
only treatment), 1.5 kg/ha (urea + 0.25% nBTPTA), 1.4 kg/ha (urea + 0.50% nBTPTA),
and 0.25 kg/ha (NH4 N0 3). These values show that nBTPTA at the lower rate was nearly
as effective as at the higher rate.
It is mentioned in a short report by Goodroad and Wilson (1989) that laboratory
studies indicated the effectiveness of nBTPTA in decreasing urea hydrolysis in soils
from Georgia (U.S.A.).
Hongprayoon et al. (1991) and Bouldin et al. (1991) studied urea transformations in
flooded soil columns. A silt loam soil (pH in H20 5.8) collected from the Rice Research
Station in Crowley (Louisiana) was used.
In studying urea diffusion and hydrolysis, I-kg soil samples were packed into
plexiglas columns (30-cm length by 7.5-cm i.d.). In one set of 12 columns, the soil
sample used was first sterilized (by autoclaving at 120°C for 2 hours), then amended
with 10% nBTPTA (relative to weight of urea) and saturated with sterilized water. (It
was found in a previous experiment that nBTPTA at this concentration effectively
stopped all urea hydrolysis in sterilized soil for 6 days). In another set of 12 columns,
the soil and water were nonsterile. In both sets a 2-cm floodwater depth was established.
All columns were incubated at 30° for 3 days, then the floodwater was removed and
replaced by a urea solution to a depth of 2 cm. The concentration of urea solution (750
mg Nil) corresponds to a soil application rate of 150 kg N/ha. The soil columns were
again incubated in the dark at 30°C for 6 days, during which the residual urea in
floodwater and soil layers was analysed at l-2-day intervals.
Besides urea diffusion and hydrolysis, the urea adsorption and amnlOnium diffusion
were also studied.
The results obtained made it possible to conclude that the rate of urea hydrolysis,
i.e., urease activity in the floodwater and soil is the most important factor influencing
the potential for ammonia volatilization. If rates of urea hydrolysis are very high,
incorporation of urea in the soil and shallow floodwater are the indicated management,
while, with low hydrolysis rates, deep floodwater and no incorporation of urea are the
indicated management.
Phongpan and Byrnes (1990) conducted a field experiment during the 1988 wet
season (July-November) on a clay loam soil (PH in H2 0 5.4) at the Suphan Buri Rice
Experiment Station in Thailand. The plots (3 by 5 m) were PK-fertilized with triple
superphosphate (24 kg/ha) and KCl (33 kg K/ha) incorporated into the drained soil
before transplanting of rice seedlings. Following transplanting, the plots were
maintained under continuous flooding with 5-7 em floodwater depth until the final
harvest.
149
At day 10 after transplanting, urea with or without nBTPTA was broadcast into the
floodwater. Rates of urea were 0, 25, 50, and 75 kg Nlha. The rate of nBTPTA was
constant at 0.536 kglha by addition of urea containing 1% nBTPTA by weight. The
control plots received no urea and no nBTPTA.
During 10 days after N fertilization, floodwater samples were taken twice daily (at
8-9 and 13-14 hours) from the plots that received 75 kg urea-Nlha with or without
nBTPTA and from the control plots. The residual urea, ammoniacal-N, pH, and
temperature were measured and the vapor pressure of ammonia (pNH 3) was calculated
from ammoniacal-N, pH, and temperature values.
nBTPTA delayed urea hydrolysis only for 1 day and at day 9 no unhydrolyzed urea
was detected in floodwaters of either urea- or urea+nBTPTA-treated plots.
The ammoniacal-N content increased during the first 6 days, then decreased. The
increase was higher in the urea than in the urea + nBTPTA treatments, but the decrease
was similar and at day 10 the ammoniacal-N content represented about 4% of the
applied urea-N in both treatments.
pH raised appreciably to 8.3 on the 3rd day, then gradually decreased to less than 7.0
over 10 days. nBTPTA suppressed pH rise and maintained lower values than those of
urea alone for 4 days, after which no difference in pH was observed.
Floodwater temperature was not affected by any treatment.
The values of pNH3 were negligibly low in the morning hours due to low floodwater
pH and moderate temperature. The maximum pNH3 was recorded in the early afternoon
at day 4 after N fertilization and the afternoons 3-5 days after fertilization provided the
greatest potential for NH3 loss. nBTPTA significantly reduced the pNH3 indicating that
the urea + nBTPTA treatment was less prone to NH3 volatilization loss than was that
with urea alone.
In an experimental variant, microplots (1.2 by 1.2 m) were installed within the main
plots. 15N-labeled urea (4.9 atom% excess) without or with nBTPTA at a rate of 50 kg
Nlha was broadcast into the floodwater of microplots. Analyses of floodwater, plant,
and soil for 15N at day 37 after N fertilization indicated that the losses of 15N from the
floodwater-plant-soil system were 50.17 and 49.33%, i.e., almost 50% of the applied
15N in the urea and urea + nBTPTA treatments, respectively. The losses were attributed
to ammonia volatilization in absence of nBTPTA and to denitrification in its presence.
A similar field experiment was conducted by Phongpan and Byrnes (1993) during
the 1989 dry season (March-June) on a clay soil (pH in H2 0 4.7) at the same Rice
Experiment Station in Thailand, and similar results were obtained concerning nBTPTA.
Urea applied at a single rate (100 kg Nlha) without or with 1% nBTPTA was broadcast
into the floodwater of rice plants. Total N loss during 41 days after fertilization was
26% of applied urea-N from the plots treated with urea alone and it was insignificantly
lower, 24% from the urea+nBTPTA-treated plots. Again, the losses were attributed to
ammonia volatilization from the urea-treated plots and to denitrification in plots treated
with urea + nBTPTA.
Mullen et af. (1991) prepared reaction mixtures from 10-g samples of a silt loam
soil, 40 mg of urea, 0 to 25 ).lg of nBTPTA, and 0 or 50 mg of wheat residue, and
incubated them at 25°C for 1 week. Analysis of the remaining urea indicated that
effectiveness of nBTPTA decreased with its decreasing rate. At the 25 ).lg nBTPTA rate,
80% of the added urea remained unhydrolyzed without wheat residue and only 47%
150
with residue. In another experiment, at 0.5 mg nBTPTAJkg soil, 60 and 8% of the added
urea remained unhydrolyzed after 4 and 7 days, respectively.
Mullen and Ravelle (1992) and Howard et at. (1992) evaluated the effects of
nBTPTA (1.12 kglha) on hydrolysis of urea and volatilization of ammonia from urea
and urea-ammonium nitrate (UAN), applied as solid and liquid fertilizcr, respectively,
to field microplots on a silt loam soil amended with wheat straw. The results indicated
that nBTPTA was more effective in urea- than in UAN-fertilized microplots. Thus, after
8 days, 17 and 1% of the added urea-N was recovered from the urea+nBTPTA- and
urea-treated microplots, respectively, and 3.1 and 1.3% of the added urea remained
unhydrolyzed in the UAN+nBTPTA- and UAN-treated microplots, respectively.
Ammonia volatilization from urea and UAN was reduced by 73 and 20%, respectively,
by nBTPTA over 10 days.
To study the effect ofnBTPTA on N transformations at the microsite of urea granule
placement, Christianson et al. (1993) used two contrasting soils, a silt loam (PH 5.2)
and a clay (pH 8.2). Uniform cylindrical small (12-mg) urea granules containing 0, 0.5
or 0.05% nBTPTA (weight/weight) were placed on the soil surface in plastic cups (a
single granule in exact center of each cup) for up to 6 days. Soils were then frozen in
liquid N2 and a 0.9-cm wide vertical slice was cut through the fertilizer placement site.
A section of this slice was cut into 45 squares (004 by 004 cm) and analyzed for soil pH,
extractablc ammonium, nitrate, and urea concentrations at the microsite where the
fertilizer had been placed.
In the silt loam soil, nBTPT A lowered soil pH and NH4 + concentrations at the
placement site as compared to urea alone and allowed more diffusion of urea away from
the fertilizer microsite. In the clay soil, nBTPTA was less effective in reducing NH/
concentrations from the zone of high soil pH associated with urea hydrolysis.
The conclusion was that effectiveness of nBTPTA depends on the capacity of the
soils to permit diffusion of urea and ammonium.
Watson et at. (l990a,b) carried out a field experiment at the Agricultural Research
Institute, Hillsborough, County Down, Northern Ireland, to compare the effects of mid-
summer applied urea, urea + 0.5% nBTPTA (relative to weight of urea), and calcium
ammonium nitrate (CAN) on ammonia volatilization and on herbage dry matter yield of
an established perennial ryegrass (Lolium perenne) sward on a clay loam soil (pH in
H20 6.3). The fertilizers were surface-applied, at a rate of 100 kg N/ha, to (6 by 2 m)
plots. The plots also received P and K fertilizers. No fertilizers were added to the
control plots. Ammonia volatilization was measured daily during 13 days after
fertilization.
The maximum NH.l volatilization occurred on day 1 in the urea treatment and with
an approximately 5-day delay in the urea + 0.5% nBTPTA plots. Delaying the
maximum volatilization loss increases the chance of rain falling to move the urea below
the soil surface and hence lower NHJ volatilization.
Cumulative NH3 volatilization loss expressed as a percentage of urea-N applied was
8.1 from urea, 1.89 from urea + 0.5% nBTPT A, and 0.09 from CAN. The reduction of
volatile NHjloss by nBTPTA was very significant.
Watson et al. (l994a) studied the effect of nBTPTA added to urea granules at five
rates (0, 0.01, 0.05, 0.1, 0.25, and 0.5%) on ammonia volatilization. The site was again
an established perennial ryegrass sward on a clay loam soil at the Agricultural Research
Institute, Hillsborough. As the pH of this soil was SA, it was limed (2.5 t finely ground
151
CaC03/ha) 3 weeks prior to commencing the experiment to increase the surface pH (0-
2.5 cm) to approximately 6.0. The experiment was carried out in five repetitions on
different plots (I by 2 m) and in different time periods over the 1992 growing season to
cover a range of weather conditions.
The first experimental time started at the end of April 1992 and it was repeated at
approximately 4-week intervals (noted as time periods 2-5), moving to new plots each
time. Prior to fertilizer application the grass was cut to a height of 2 cm. The plots at
each time period received 100 kg N/ha together with 8.7 kg P/ha as single
superphosphate and 67 kg KIha as muriate of potash. The plots designated for the later
time periods 3-5 received, approximately 8 weeks before their use, one basal dressing of
a compound fertilizer containing 50 kg N, 5.5 kg P, and 21 kg Klha (plots for time
periods 3 and 4) or two basal dressings (plots for time period 5). All fertilizers were
surface-applied.
Volatilization ofNH3 was measured daily for 9 days following fertilizer application
to plots for time periods 1,2,4, and 5 or for 8 days in the case of plots for time period 3.
Over all of the time periods, the mean value of the times at which maximum rate of
NH3 volatilization loss occurred was 2.59 days in the case of urea-only treatment and
increased to 3.17-4.73 days in parallel with increasing level of nBTPTA from 0.01 to
0.5% relative to weight of urea.
Over all of the time periods, the inhibition of NH3 volatilization was 50.4, 82.8,
89.0, 96.5, and 97.0% at the O.oI, 0.05, O.l, 0.25, and 0.5% nBTPTA levels,
respectively.
Over all levels of nBTPTA, inhibition of NH3 volatilization loss from plots for time
period 1 to those for time periods 2-5 was 83.8, 81.2, 77.0, 88.3, and 85.3%,
respectively. There was no significant difference in these values and no significant time
x treatment interaction, suggesting that the effectiveness of nBTPTA was not dependent
on climatic conditions.
Based on the percent inhibition values and different nBTPTA levels, it was
calculated that the optimum level of nBTPTA at the site studied would appear to be
0.1 % which would be predicted to give 93% inhibition ofNH3 volatilization loss.
Watson et al.(1994b) studied, under laboratory conditions, the influence of soil
properties on the relationship between ammonia volatilization and nBTPTA
concentration. Surface samples (0-15 cm) of 16 soils of widely differing chemical and
physical properties, collected from grassland sites in Northern and Southem Ireland,
were used.
Fresh soil samples were placed in cylindrical screw-top plastic jars (85 mm height,
80 mm diameter) to a depth of 50 mm. Urea granules (50 mg N equivalent
approximatively to 100 kg N/ha) were applied to the surface of soil samples. The urea
granules contained 0, 0.01, 0.058 or 0.28% nBTPTA. The samples were incubated at
13°C. The NH3 volatilized was measured daily for 9 days after fertilizer application.
Total NH3-N loss over 9 days (as % ofN applied) had the following mean values in
the 16 soils: 20.67 (samples treated with urea only), 15.94, 7.15, and 2.79 (samples
treated with urea + 0.01, 0.058, and 0.28% nBTPTA, respectively), indicating that the
NH3 volatilization decreased with increasing nBTPTA concentration. Similarly, the
average times of the maximum loss ofNH3 from the 16 soils were 2.99,3.86,6.43, and
6.49 days at the 0, 0.01, 0.058, and 0.28% nBTPTA levels, respectively.
152
The % inhibition of NH3 loss by nBTPTA was highly dependent on soil type; on
some soils nBTPTA was effective even at the 0.01 % concentration. nBTPTA was most
effective in reducing NH3 loss from soils with a high pH and low organic matter
content, i. e.. from soils very prone to NH3 loss from urea containing no urease inhibitor.
Simple correlation analysis proved that NH3 volatilization from samples of the 16 soils
treated with urea only was most significantly correlated with pH in H20 (r=0.896) and
pH in KCI (r=O.920).
Modeling the relationship between total NH3 loss and nBTPTA concentration
showed that the % nBTPTA required to achieve a given % decrease in NH3
volatilization was constant for all soils. For example, 0.092% nBTPTA was predicted to
lower total NH3 loss by 90% from any given soil. Little additional benefit can be
expected by using nBTPTA concentrations above 0.1 % relative to weight of urea.
Watson et al. (1998) studied the longer-term effect of repeated application of
nBTPTA-amended urea on ammonia volatilization and urease activity as well as on
herbage dry matter yield and N uptake by perennial ryegrass.
A 3-year field experiment was carried out at the Agricultural Research Institute,
Hillsborough. Plots (6 by 2 m) were installed under a three-cut silage system. The soil
received one, two or three applications of urea, CAN or urea amended with nBTPTA at
either 0.1 or 0.5% level, during each of the 1994, 1995, and 1996 growing seasons.
There were ten treatments.
In each year, the plots were fertilized three times: in March/April (120 kg Nlha, 13
kg Plha as single superphosphate, and 75.5 kg K/ha as muriate of potash); in May (100
kg N, 8.7 kg P, and 75.5 kg K/ha), and in June/July (80 kg N, 8.7 kg P and 75.5 kg
K/ha).
Ammonia volatilization was studied at the end of the first growing season (early
September 1994). Soil was collected (0-5 cm depth) from each plot. Fresh soil samples
(250 g each) were treated with urea or urea + 0.1 % nBTPTA at a rate of 50 mg N
(equivalent to -100 kg Nlha) and incubated at 15°C. The volatilized NH3 was measured
daily for 9 days after fertilizer application.
It was found that 0.1 % nBTPTA not only lowered NH3 loss but substantially
delayed the time at which maximum NH3 loss occurred; the average value over the ten
treatments was 1.6 days (urea) and 4.9 days (urea + 0.1% nBTPTA). It was concluded
that previous fertilizer application had no significant effect on the subsequent efficacy
of nBTPTA in lowering NH3 volatilization from urea.
Soil urease activity (in the 0-5-cm layer) was measured four times: at the end of the
three growing seasons (September 1994, October 1995 and 1996) and at the beginning
of the growing season of the last year (March 1996).
In September 1994 and October 1995, urease activity was significantly lower in
plots fertilized with urea + 0.5% nBTPTA than in the other plots, including those
fertilized with urea + 0.1 % nBTPTA. But in March and October 1996 the activity was
similar in all plots which indicates that 0.5% nBTPTA had no cumulative inhibitory
effect on soil urease.
Ma et al.(l995) performed a laboratory experiment to study the effects of nBTPTA
and the percolation rate on hydrolysis and movement of urea under simulated flooded
soil conditions. A silt loam soil, typical in south-west Louisiana rice fields, was used.
Soil samples (1.13 kg) were packed into PVC columns, then saturated and covered by a
2-cm floodwater layer, which was maintained during the preincubation of columns in
153
the dark at room temperature for 3 months to develop reduced soil layers. The columns
were then percolated at a rate of 0, 0.5 or 1 cm/day, the floodwater layer being replaced
by a urea solution containing 0 or 10% nBTPTA (on urea weight basis). Total urea-N
added to each colunm was 70.4 mg. In the next step, the colunms were incubated in the
dark at 30°C for 0.5, 1 or 2 days, then the floodwater was collected and the soil colunms
were sectioned into eight layers at depths of 0.5, 1, 2, 3, 5, 7.5, 10, and 12.5 em. The
soil sections were analyzed to determine residual urea.
The analytical data showed that more than 70% of the urea was hydrolyzed after 2
days of application without nBTPTA, whereas only 40% was hydrolyzed with
nBTPTA. Consequently, more urea was distributed in the soil profile in nBTPTA-
treated columns. High percolation rates caused substantial urea movement downward
into the soil when urea was not hydrolyzed.
In studying the effect of nBTPTA on ammonia volatilization from urea and on the
maize yield under the climatic conditions of the Mediterranean regions, Palazzo et al.
(1995, 1996) conducted 3-year field experiments (1989-1991) on a sandy clay soil and
on three clay soils (PH 7.87-8.25) at Metaponto and Monterotondo. Urea was surface-
applied yearly at a rate of 150 kg N/ha without or with 0.1 or 0.25% nBTPTA (relative
to weight of urea), one day after irrigation. Single superphosphate and potassium sulfate
were also applied. Ammonia volatilization was determined during 6 weeks after
fertilizer application.
Cumulative NH3 losses from urea without nBTPTA averaged 43.3% in the sandy
clay soil and 11.3% in the clay soils, whereas the average losses from the nBTPTA-
amended soils were 24.2 and 3.1% respectively. There was no significant difference
between the effects of the 0.1 and 0.25% nBTPTA concentrations.
Vittori Antisari et al. (1996) evaluated the effects of different concentrations of
nBTPTA on ammonia volatilization, urea hydrolysis, and evolution of mineral N in
surface (0-20 cm) soil samples collected at the locations Ozzano, Rimini, and Carpi. Of
the three soils, the Rimini soil has the highest pH (8.5 in H2 0), contents of sand (81 %),
organic C (2.3%), total N (2.287 glkg) and is the most urease-active. Urea was added to
the samples at a rate of 1 mg N/g dry soil. The rate of nBTPTA relative to weight of
urea ranged from 0.01 to 0.5%. The amount of the volatilized NH3 and the
concentrations ofresidual urea, exchangeable NH/, N0 2-, and N03- were determined
periodically during incubation at 23°C for 15 days.
nBTPTA reduced NH3 volatilization proportionately with its rate. The extent of
reduction was smallest in the Rimini soil.
Similar results were obtained concerning urea hydrolysis. For example, the urea
applied with 0.5% nBTPTA was hydrolyzed during 1 and 15 days of incubation in
proportions of 0 and 79.5%, respectively, in the Carpi soil and of 30 and 87%,
respectively, in the Rimini soil.
nBTPTA slowed down the formation of exchangeable ~ + due to inhibition of
urea hydrolysis and decreased the concentration of N0 2- due to reduction of NH/
formation. Both effects were weakest in the Rimini soil.
In contrast, the concentration of NO) - was increased in each soil by increasing the
nBTPTA rate and/or incubation time.
In a laboratory experiment performed by Murphy and Ferguson (1997), air-dry
samples of a silt loam soil were packed to a depth of 6 cm in acrylic chambers (3.4 cm
diameter). Urea-ammonium nitrate (DAN) solution (28% N) with or without nBTPTA
154
was placed 1.8 cm deep in the soil to simulate a subsurface band application. The rate of
N addition was 224 kglha and that ofnBTPTA was 1.12 kglha. The residual urea, NH/,
and N03 - concentrations were determined over a 14-day period. Urea-N recovery was
significantly greater and urea hydrolysis rate significantly less (p=0.05) only on day 1,
but by day 14 the urea was completely hydrolyzed and recovery ofNH/-N and that of
NO) --N became the same in both UAN-treated and UAN+nBTPTA-treated soil.
The Argentinean investigators Sainz Rozas et al. (1997) conducted a 2-year field
experiment (1994/95 and 1995/96) studying ammonia volatilization from surface-
applied urea to no-till maize as a function of urea rate (0, 70, 140, and 210 kg N/ha),
presence or not of nBTPTA and fertilization time (at seeding and 6-leaf stage). In both
years, the NH3 loss increased with an increasing rate of urea and was higher following
fertilization at 6-leaf stage tllan at planting; nBTPTA reduced the NH3 loss even when
the no-till maize was fertilized later than at planting time.
nBTPTA was submitted to a laboratory test at the South African Sugar Association
Experiment Station at Edgecombe (Anonymous, 1998). The commercial nBTPTA
Agrotain, recommended for field application at a rate of 5 1/t urea granules, was used.
The test showed that anlfllonia volatilization losses from urea applied with Agrotain
before rain could be reduced by up to 14% on bare soil and up to 10% on trash. The
strongest suppression of N losses «1 %) was, however, derived from a simulated
irrigation of 10 mm. The results also indicated that Agrotain has a limited useful life of
about 2 weeks on both bare soil and trash, so that the absence of rain or irrigation after
urea application for periods longer than 2-3 weeks would greatly reduce the N savings.
The reason for this continued loss is that the urea which was initially protected from
hydrolysis by Agrotain is still susceptible to losses after the Agrotain is depleted, unless
water additions from rain or irrigation incorporate the urea into the soil.
Gill et al. (1999) studied the effect of green manure and wheat straw amendment of
a flooded alkaline sandy loam soil from India on the effectiveness on nBTPTA to
reduce urea hydrolysis and anlfllonia volatilization from urea. Hydrolysis of the urea
applied was complete in 12, 8, and 6 days in unamended, green manure- and wheat
straw-amended soil, respectively, and as a consequence, the NHJ loss through
volatilization was enhanced by the organic amendments.
nBTPTA applied at rates of 0.5, lor 2% (on urea weight basis) was more inhibitory
on urea hydrolysis and NHJ volatilization from the unamended soil than from the soil
amended with green manure or wheat straw. For example, by applying 0.5% nBTPTA,
complete hydrolysis of urea was delayed up to 16 days in the unamended soil, whereas
in wheat straw-amended soil urea was completely hydrolyzed by the 12th day even
when it was treated with 2% nBTPTA.
***
A great number of studies, in which the effect of phosphoric triamide (PTA) and
thiophosphoric triamide (TPTA) compounds on soil urease activity, urea hydrolysis,
ammonia volatilization, and on immobilization of urea-N in soils as well as their
stability were compared with those of other inhibitors, will be dealt with in Chapter 4.
155
2.27. CYCLOTRIPHOSPHAZATRIENE DERIVATIVES
The inorganic derivative 2,2,4,4,6,6-hexaaminocyclotriphosphazatriene (also called

phosphonitrilic hexamide) (Figure 54) occupied an intermediary position between
phosphoryl triamide (PTA) and thiophosphoryl triamide (TPTA) concerning both the
Figure 54. Structure of 2,2,4,4,6,6-hexaaminocyclotriphosphazatriene (phosphonitrilic

hexamide).
inhibitory effect on soil urease activity and stability in urea melt at 140°C. Besides its
reduced inhibitory capacity, this compound also has another disadvantage: its
production cost is significantly higher with respect to the two triamides as well as
PPDA (Anonymous, 1985a; Radel et al.. 1987).
The inventors Medina and Sullivan (1986, 1987) tested a series of cyclotri-
phosphazatriene (CTP AT) derivatives, also called phosphonitrilic derivatives. Some of
these compounds, namely the hexaamino derivative mentioned above and three organic
(phenoxy) derivatives (Figure 55) were found to be able to inhibit soil urease activity.
P3N3(NH2)S(OCsHs) P3N3(NH2)4(OCsHs)2
2-Phenoxy-2,4,4,6,6-pentaamino-CTPAT 2,4-Diphenoxy-2,4.6,6-tetraamino-CTPAT
2,
H N /OC 6Hs
N...... P~N
C6 HSO, II I ,.........OCsH s
/P'N PP ,
H2N NH2
P3N3(NH2b(OCsHsb
2,4,6-Triphenoxy-2,4,6-triamino-CTPAT
Figure 55. Structure of the phenoxy derivatives of cyclotriphosphazatriene (CTPAT) found

by Medina and Sullivan (1986,1987) to be inhibitors of soil urease activity.
156
In testing the inhibitory effect, samples of two silt loam soils (from Alabama and
Louisiana, respectively) were used. Several procedures were applied. PPDA and PTA
served as reference compounds.
a) Small containers (6x6x6 cm) were one-half filled with soil containing 20%
moisture. Urea powder or urea powder + powdered test compound (thoroughly mixed)
was distributed in a narrow band, 6 cm long on the soil surface. The urea rate was 410
mg (6.72 mmoleslband) and equivalent to 100 kg Niha. The test compounds were used
in an equimolar amount (0.25 mmoleslband). Then the containers were filled with soil
and submitted to incubation at 25°C. After 3, 6, and 9 days, the residual urea was
determined.
It was found that after 3 days of incubation PPDA was a more effective inhibitor
than 2,2,4,4,6,6-hexaamino-CTPAT, with the unhydrolyzed urea representing 68.3 and
48.1 % of the added urea, respectively. However, after 6 days, the reverse was true, with
the corresponding values ofunhydrolyzed urea being 12.9 and 16.8% respectively. In 9
days, the urea was completely hydrolyzed in all treatments. It also established that after
both 3 and 6 days of incubation the inhibitory effect of 2,2,4,4,6,6-hexaamino-CTPAT
on soil urease activity was more marked than that of PTA.
b) This procedure is similar to the first one, but each test compound was applied at a
rate of 10% relative to urea weight (which remained 410 mglband). By using this
procedure it was found that PPDA was a more effective inhibitor than 2,2,4,4,6,6-
hexaamino-CTPAT not only after 3 days of incubation, but also after 6 days. PPDA and
the four urease-inhibiting CTPAT derivatives presented the following decreasing order
of their inhibitory capacity:
PPDA > hexaamino ~ phenoxy ~ diphenoxy > triphenoxy.
The compounds specified below were unable to inhibit soil urease activity:
P3N3C1 6 , 2,2,4,4,6,6-hexachloro-CTPAT;
P3N3(NH2h(NHCH3)4, 2,2,4,4-tetra( methylarnino)-6,6-diamino-CTPAT:
P3N3(NH2h[N (CH3 h]4, 2,2,4,4-tetra(dimethyl amino)-6,6-diamino-CTPAT;
P3N3(NHCH3)6, 2,2,4,4,6,6-hexa(methylamino)-CTP AT;
P3N3[N(CH3)2k 2,2,4,4,6,6-hexa(dimethylamino)-CTPAT;
P3N3CI4 [N(CH3hh. 2,2,4,4,-tetrachloro-6,6-di(dimethylarnino)-CTPAT;
P3N3Cls(OC6 Hs),2-phenoxy-2,4,4,6,6-pentachloro-CTPAT;
P3N3C4(OC6 Hsl2. 2,4-diphenoxy-2,4,6,6-tetrachloro-CTPAT;
P3N3Ch(OC6Hs)3,2,4,6-triphenoxy-2,4,6-trichloro-CTPAT.
c) Soil samples (100 g) were mixed with 40 ml of water, 410 mg of urea powder
without or with 41 mg of powdered test compound. The mixtures were incubated at
25°C for 1, 3, and 6 days, after which the unhydrolyzed urea was assessed. PPDA
proved again to be a more effective inhibitor than were the CTPAT derivatives tested
which showed the following order of inhibitory capacity:
phenoxy> diphenoxy > hexaarnino > triphenoxy.
One can see that the order obtained with procedure c (well-mixed system) differs
from that recorded with procedure a (banded system).
d) Moist soil samples (120 g) were well mixed with 1 ml of aqueous solution or
suspension containing 7 ~oles of test compound. The mixtures were preincubated at
30°C for 0, 3, 7, 14, and 21 days before adding urea (50 mg in 1 ml solution). After a
new mixing and incubation at 30°C for 16 hours, the residual urea was determined.
Under these conditions, the three phenoxy derivatives of CTPAT proved to be superior
157
not only to the hexaamino derivative and PTA, but also to PPDA. Thus, after 0, 7, 14,
and 21 days of preincubation, the inhibition had the following values: 22, 74, 84, and
82% (triphenoxy derivative), 75, 71, 69, and 65% (diphenoxy derivative), 95, 49, 42,
and 44% (phenoxy derivative), 46, 20,4, and 4% (hexaamino derivative), 100, 27, 4,
and 0% (PPDA), 29, 12, 0, and 0% (PTA). In other words, without preincubation,
PPDA strongly inhibited soil urease activity, but as a result of preincubation its
inhibitory effect decreased and then disappeared. Phenoxy derivatives of CTPAT
excelled in persistence of their urease-inhibiting effect.
Based on the results mentioned above, binary and tertiary mixtures of phenoxy
CTP AT derivatives were suggested in a model to predict urease inhibitions of various
intensity and duration to meet the demands of a wide variety of agroclimatic, soil, crop,
and management conditions and practices.
According to the model (Figure 56) it is possible, by using different mixtures, to
regulate the inhibition of soil urease activity between about 25 and 80% for the first 3
days after urea fertilizer application, and between 50 and 80% for periods of 3-21 days.
For use in practice, the CTPAT derivatives are recommended at a rate ranging from
about 0.01 to about 10% relative to weight of urea fertilizer.
100 A
o.---a lS"/.M 'S"IoT
¢----o
-I-
':!C·... M + 'jJo... T
.-._ •• 7S-'.M+ 25"1.T
~'~~........--"'---
----<)------------.-- ~
....._._,_._._.- _.....
....... ~.'¢-- --- --_ .. _--
....... -._.-._._._._
~
~
~ 2S
°0 '0 20
().....--o 5",'.1-1+5"1.0 .9O"'.T

100 8 0-----0 10'''' M +30'"1.0 C ..
..--... l)"oM"3Jo,.O" )4·I.T
50~/. T
i 7S /r~"'-·"':.J(;-:;::;::~:~:~:-:-;;-=-=-~~~~--:~----<;,
~ l.'
.: " .I
~ 50 / /
!l I
~
" 2S
°DL------~----~,O-------~1~------~~~---
""yo
Figure 56. Model for prediction of the effect of different mixtures prepared from phenoxy derivatives of
cyciotriphosphazatriene on urease activity in soil.
A - Influence ofM + T compounds awlied at different rates. B - Influence of M + D + T compounds awlied
at different rates. M - 2-(Mono)phenoxy-2.4,4.6.6-pentaaminocyclotriphosphazatriene. D - 2,4-Diphenoxy-
2,4.6.6-tetraarninocyclotriphosphazatriene. T - 2,4.6-Triphenoxy-2,4.6-triaminocyclotriphosphazatriene.
IAdapted from Medina and Sullivan (1986, 1987), by permission of Tennessee Valley Authority.!
158
Savant et af. (1988b) performed investigations that are similar to those described in
the patents of Medina and Sullivan (1986, 1987). The same Alabama silt loam soil and
reference compounds (PPDA and PTA) were used. Four cyclotriphosphazatriene
(CTP AT) derivatives were studied: 2-phenoxy-2,4,4,6,6-pentaamino-CTPAT, cis-2,4-
diphenoxy-2, 4,6, 6-tetraamino-CTP AT+trans-2,4-diphenoxy-2,4, 6, 6-tetraamino-CTPAT
(cis/trans ratio 2.60), cis,trans-2,4,6-triphenoxy-2, 4,6 -triamino-CTP AT + cis,cis-2,4,6-
triphenoxy-2,4,6-triamino-CTP AT (cis/trans ratio 0.55), and 2,2,4,4,6,6-hexaamino-
CTPAT.
For testings, essentially the procedure d (see page 156) was applied. The 120-g
moist soil samples were well mixed with 7 ~moles of test compound or mixture of test
compounds in 1 ml solution or suspension and were pre incubated at 20, 30 or 40°C for
0,3,7, 14,21, and 36 days. Following preincubation, the mixtures were treated with 50
mg of urea in 1 ml solution and after a new mixing were incubated at 30°C for 16 hours
and then analyzed for residual urea.
After O-day preincubation at 30°C and 16-hour incubation at 30°C, the six test
compounds inhibited soil urease activity in the order: triphenoxy (22% inhibition) <
PTA < hexaamino < diphenoxy < phenoxy < PPDA (100% inhibition). After 14 days of
preincubation the order was: PTA = hexaamino = PPDA (0% inhibition) < phenoxy <
diphenoxy < triphenoxy (75% inhibition). After 36-day preincubation, the urease-
inhibiting order phenoxy < diphenoxy < triphenoxy was maintained. Thus, the results
obtained by Medina and Sullivan (1986, 1987) were confirmed.
After O-day preincubation, the urease-inhibiting capacity of the phenoxy and
diphenoxy derivatives was higher and that of the triphenoxy derivative was lower at 30
and 40°C than at 20°C, but the inhibition by each derivative was lowest during
incubation at 40°C.
To verify the prediction model presented in Figure 56, a separate experiment was
conducted. The (mono)phenoxy (M) derivative and its three combinations with the
diphenoxy (D) and triphenoxy (T) derivatives were used in proportions expressed as
percentages. The preincubation and incubation lasted 25 days and 16 hours,
respectively.
The urease-inhibiting capacity of the M derivative and its combinations presented
the order:
100% M > 50% M + 50% T > 33% M + 33% D + 33% T ;.::: 25% M + 75% T, after
O-day preincubation;
50% M + 50% T;.::: 33% M + 33% D + 33% T > 25% M + 75% T > 100% M, after
14 days of preincubation, and
25% M + 75% T;.::: 33% M + 33% D + 33% T ;.::: 50% M + 50% T > 100% M, after
25 days of preincubation.
It can be deduced from these orders that after shorter preincubations the urease
inhibition depended mostly on the M derivative and later on the T derivative.
The Czech investigators Minar et al. (1990) studied the effect of hexaamino-CTP AT
on soil urease activity and ammonia volatilization from soil treated with urea solution or
with the urea-ammonium nitrate liquid fertilizer DAM 390. Samples of a garden soil
(pH 7.08) were used.
In the urease activity study, 100-g soil samples were sprayed with urea solution or
DAM 390 at a rate of 100 mg N. Hexaamino-CTPAT was applied at rates of 0, 0.5, 1,
and 2% (relative to fertilizer N). The mixtures were preincubated at 20°C and after 1, 2,
159
3, 4, 7, and 14 days of preincubation, urease activity in subsamples was assayed (5-g

subsamples were treated with 2.5 ml of 0.08 M urea solution and incubated at 37°C for
2 hours).
The results showed that in samples treated with urea or DAM 390 and preincubated
for 2 days urease activity was markedly inhibited by each hexaamino-CTP AT rate.
After 3 days of preincubation, inhibition occurred only in the samples treated with urea
and with one of the two higher rates of hexaamino-CTPAT. After 4-14 days of
preincubation, urease activity was rather enhanced than inhibited in the hexaamino-
CTP AT-treated soil samples.
In the NH, volatilization study, 100-g soil samples received 100 or 300 mg N in urea
solution or DAM 390 with or without 0.5, 1, and 2(% hexaamino-CTP AT (relative to
fertilizer N). During incubation (at 22°C for 15 days) the evolved NH3 was assessed.
Cumulative NH, losses in 15 days were higher from urea than from DAM 390. The
losses increased with an increase in fertilizer N rate. Hexaamino-CTP AT reduced the
losses and this effect was more marked at the higher hexaamino-CTP AT rates. Thus, the
cumulative NH, losses from urea (100 mg N) in the samples treated with 0,0.5, 1, and
2% hexaamino-CTP AT were about 51, 35, 26, and 22% of the added N, respectively.
The corresponding losses from DAM 390 (100 mg N) were about 23, 15, 8, and 6%
respectively (see also Zehmilek et aI., 1990).
The postulation by Medina and Sullivan (1986,1987) that perhaps many derivatives
containing the PN cyclic structure, especially the cyclotetraphosphazatetraene
derivatives, are also potentially effective inhibitors, has been verified by Honza and
Minar (1990) in the case of octaaminocyclotetraphosphazatetraene. They studied the
inhibition of ammonia volatilization from urea and DAM 390 by using three
preparations: pure hexaamino-CTP AT, nonpurified hexaamino-CTP AT containing
residual ammonium chloride, and a mixture of pure hexaamino-CTPAT and
octaaminocyclotetraphosphazatetraene. Eighty-g soil samples (with 12-13% moisture
content) were placed in Petri dishes (1 dm2) and treated with 5 ml of urea solution or
DAM 390 containing 100 mg of Nand 0 or 2% inhibitor (relative to weight of
fertilizer), then incubated at 23-25°C for 17 days. During incubation the evolved NH3
was measured. Each preparation reduced the volatile NH3 losses, especially during the
first days of incubation. However, cumulative losses in 17 days were also reduced from
about 45% of the added N (urea-only treatment) to about 10-30% (urea + inhibitors),
and from about 32% (DAM 390-only treatment) to about 15-20% (DAM 390 +
inhibitors). The pure hexaamino-CTPAT was a little more effective than the other two
preparations.
Stability (decomposition) of CTPAT derivatives in soil was studied in the case of
hexaamino-CTPAT.
Dick and Tabatabai (1978) studied the decomposition of this compound
[P3N 3(NH 2kH 20] in three Iowa soils (a loam and two clay loams). The compound
containing 37.3% P was added to 2-g soil samples in an amount of 500 ppm on soil
basis. Incubation took place under aerobic or water-logged conditions at 20°C and lasted
7 days. Hydrolytic decomposition of the compound, as evaluated by determination of
the released a-phosphate, was very reduced: during 7 days of incubation, the rate of
hydrolysis under aerobic conditions in the three soils studied ranged from 7 to 13%; on
average, it was 10%. Under water-logged conditions the decomposition averaged only
3%.
160
The results of a maize pot experiment, in which Calancea et al. (1986) used labeled
hexaamino-CTPAT, P3Nl 5NH 2 )6 or P3 15 N3(NH2)6, indicated that hexaamino-CTPAT is
decomposed slowly in the soil. At the beginning the compound is deaminated which is
followed by the decomposition of the cycle (see page 351).
2.28. PHOSPHORYLATED 2-0XIMINOPHENYLACETONITRILE COMPOUNDS
Simihliian (1998) and Simihaian et al. (1999) tested the effect of three phosphorylated
2-oximinophenylacetonitrile compounds (Figure 57) on soil urease activity.
Hydroquinone served as a reference compound. Weakly urease-active samples of an
alluvial soil (light-textured) and a chernozem (heavy-textured) were used.
Bis·( 2·oximinophenylacetonitrile) Bis·(2·oximinophenylacetonitrile) 2·(Oximinophenylacetonitrile).2·oxo-

ester of cyclohexylthiophosphonic acid ester of phenylphosphonic acid 5,5-dimethyl-l,3,2·dioxaphosphorinane
(A) (8) (C)
Figure 57. Structure of the phosphOIylated 2-oximinophenylacetonitrile cOlJ1lounds tested for inhibition of
soil urease activity.
The reaction mixtures, prepared from 5 g of air-dried soil, 2 ml of toluene, 1 ml of

0.6% urea solution, and 9 ml of distilled water or 9 ml of aqueous solution containing a
test compound at 2% rate relative to weight of urea, were incubated at laboratory
temperature and analyzed periodically by means of a chromogenic reagent (see the
footnote on page 35) to detect the presence of urea and establish the time (in days)
necessary for complete hydrolysis of urea. The difference between the number of days
necessary for complete urea hydrolysis in inhibitor-treated samples and in untreated
(control) samples (prolongation of time for complete urea hydrolysis) indicates the
intensity of inhibition. The results obtained are summarized in Figure 58.
Figure 58 shows that each compound tested inhibited urease activity in both soils.
Hydroquinone was a stronger inhibitor than were compounds A, B, and C in both soils,
whereas these three compounds were more inhibitory in the alluvial soil than in the
chemozem. There was no difference in the inhibitory capacity of compounds A, B, and
C in the alluvial soil, but compounds A and B were more inhibitory than compound C in
the chernozem. These findings mean that the bis-(2-oximinophenylacetonitrile) esters of
cyclohexylthiophosphonic acid and phenylphosphonic acid, respectively (compounds A
and B) behaved similarly as inhibitors in both soils, and compound C, in which the
161
HQ A B C
Figure 58. Inhibition of soil urease activity by hydroquinone and three phosphorylated 2-
oximinophenylacetonitrile compounds.
HQ - Hydroquinone. A B, C - Compounds A, B. and C (see Figure 57). IFrom Simihllian, 1998./
2-oximinophenylacetonitrile moiety is bound to a 1,3,2-dioxaphosphorinane structure,

was less effective as an inhibitor in the chemozem.
2.29. 2-THIONO-5,6-DIMETHYL-l ,3,2-DIOXAPHOSPHORINANE COMPOUNDS
Three such compounds were tested by Sirnihaian (1998) and Simihaian el al. (1999) for
evaluation of their effect on soil urease activity. Their structure is shown in Figure
59.
CH3,C " CH 2- 0 ,-r:tI'S
CHI 'CH2-0/"NH--S02-0-CH3
2-Benzenesulfonamido-2-thiono- 2-(p-Methyl}-benzenesulfonamido-2-thiono-
5,5-dimethyl-l,3.2-dioxaphosphorinane 5,5-dimethyl-l.3.2-dioxaphosphorinane
(I) (II)
CH 3,C " CH 2-0 "r:tI'S
CHI 'CH2-0/"NH--802-o-0-CH3
2..(p-Methoxy)-benzenesulfonamido-2-thiono-
5,5-dimethyl-l.3 ,2-dioxaphosphorinane
(III)
Figure 59. Structure of the 2-thiono-5,6-dimethyl-l,3,2-dioxaphosphorinane compounds tested for inhibition

of soil urease activity.
The reference compound, soils, and methods used were identical to those specified
in the preceding subchapter .
Figure 60 comprises the results obtained.
162
t! 50
iii. 45
E., 40
8i;'
15~ 35
-."
~.~ 30
+=025
-a
'0c,.. 20
.2 -; 15
~~
c
10
.2 5
£ 0
HQ II III
Figure 60. Inhibition of soil urease activity by hydroquinone and three 2-thiono-5,6-dimethyl-l,3,2-
dioxaphosphorinane compounds.
HQ - Hydroquinone. I, II. III - Compounds I. II. and III (see Figure 59). IFrom Simihiiian. 1998.1
It is evident from this figure that the compounds tested inhibited urease activity in
both soils. Hydroquinone was a more potent inhibitor than were compounds I, II, and III
in both soils. Compounds II and III were more inhibitory than compound I in the
alluvial soil, but these three compounds were similarly weak inhibitors in the
chemozem. The presence of p-methyl or p-methoxy substituent in benzene ring
(compounds II and III) increased the inhibitory effect, to the same extent, in the alluvial
soil, but no increased inhibitory effect was observed in the chernozem.
2.30. ANTIMETABOLITIES
These compounds were tested for their ability to suppress growth of urease-producing
soil microorganisms, i.e.. not for the inhibition of the activity of urease accumulated in
soil.
2.30.1. Sulfanilamide
Sulfanilamide (amide of sulfanilic acid; Figure 61) was tested by Pugh and Waid
(1969a,b) to evaluate its inhibitory effect on volatilization of ammonia from urea in four
Figure 61. Structure of sulfanilamide.
soils from England. In the first experiment, 100-g moist samples of an acid loamy sand
were treated with 6.66 mM of urea, 0.582 mM (0.1 g) of sulfanilamide (SA) or 0.4 mM
of acetohydroxarnic acid (AHA) or with 0.582 mM SA + 0.4 mM AHA. Soil treated
only with urea was the control. During incubation which lasted 77 days at 20°C, the
evolved NH3 was measured. Half-loss time of NH3 increased form 11.5 days (control
163
soil) to 12.5 days (SA-treated soil), 28.5 days (AHA-treated soil), and to 38.5 days
(SA+AHA-treated soil). In other words, SA had a very weak inhibitory effect. In
contrast, SA + AHA exhibited a synergistic effect. However, total NH3 losses during the
77-day incubation period were very similar in all the samples.
In other experiments, in which three other soils were also used, amounts of soil
sample, urea, SA, and AHA were the same as in the first experiment, and the results
obtained with two of these soils (a sandy loam and a loamy sand) were also similar to
those registered in the first experiment. However, in the third soil (a sandy loam),
neither SA nor SA + AHA delayed volatilization of NH 3; in fact, they increased total
loss ofNH3 •
In the three Iowa soils studied by Bremner and Douglas (1971) under the conditions
of the 5-hour test, SA at a rate of 50 ppm (on soil basis) caused only negligible «1%)
inhibitions in urease activity.
2.30.2. Other Antimetabolites

Peterson and Walter (1970), assignors to Allied Chemical Corporation, New York,
patented five antimetabolites as inhibitors of urea hydrolysis in soil, of losses of urea-N
in form of volatile ammonia. These compounds, in the order preferred by the inventors,
are: pyridine-3-sulfonic acid (PSA), desthiobiotin (DTB) (5-methyl-2-oxo-4-imidazo-
linecaproic acid), oxythiarnine chloride (OTC) [5-(2-hydroxyethyl)-3-(4-hydroxy-2-
methyl-5-pyrimidinylmethyl)-4-methylthiazolium chloride], y-benzene hexachloride (y-
BHC) (cis-l,2,4,5-trans-3,6-hexachlorocyclohexane; the insecticide lindane), and 0-
chloro-p-aminobenzoic acid (CABA) (Figure 62).
They are recommended for practice at the following preferred and most preferred
rates: 4.5-450 g and 27-54 g/ha, respectively, for 336 kg or less urea-Nlha. For example,
PSA applied at rates of 27 and 54 g/ha together with 112 kg urea-Nlha reduced total
losses ofNH3 during 2 weeks by about 30 and 42%, respectively, as compared to losses
recorded in the soil treated with urea alone. When the rate of urea was increased to 336
kg Nlha, PSA at the rate of 54 g/ha reduced NH3 loss by about 33% over a period of 2
weeks. After 11 weeks PSA still showed the inhibitory effect.
The antimetabolite and urea were applied separately on the soil surface, then were
leached into the soil with water until the total moisture content was about 10%.
Fatty acid-kaolin-coated PSA-urea peliets were also prepared and tested. The pellets
contained PSA at 0.02, fatty acid at 0.3, and kaolin at 0.3% of the urea. The control
pellets lacked only the PSA. The pellets (at a rate of 118 kg urea-N/11a) were not washed
into the soil but were covered with a thin soil layer to simulate field dispersal. The
cumulative losses of NH3 during 4, 12, and 21 days were 4.8, 9.7, and 10.6%,
respectively (from the control pellets) and 2.0, 4.7, and 5.5%, respectively (from the
PSA-urea pellets).
Mulvaney and Bremner (1977) performed complex investigations to evaluate the
effects of three of the five antimetabolites patented by Peterson and Walter (1970)
(PSA, DTB, and OTC) on a) urea hydrolysis in soil; b) volatilization of ammonia from
urea-treated soil, and c) production of urease by soil microorganisms. The
antimetabolites were always applied at much higher rates than those recommended by
Peterson and Walter (1970).
164
O-S~
Pyridine-3-sulfonic acid Desthiobiotin
a.c
CI
y"'
CI
I
CI
"0
y-Benzene hexachloride o-Chloro-p-aminobenzoic acid
Figure 62. Structure of the antimetabolites patented and tested by Peterson and Walter (1970) for inhibition of
a) Three Iowa soils were used. Samples of field-moist soil containing 5 g of oven-
dry material, to which sufficient water was added to bring the total volume of water to 2
ml, were treated with I ml of aqueous solution containing 2 mg of urea with or without
50 or 250 f.!g of antimetabolite. An inhibitor of soil urease activity, hydroquinone (HQ),
was included for comparison. HQ was applied at the same rates as the antimetabolites.
Incubation took place at 20D e. After 24 and 48 hours, the residual urea was analyzed.
The analytical data showed that in each soil the inhibitory effect of PSA, DTB, and
OTe on urea hydrolysis was zero at their both rates and after both incubation times. In
contrast, the inhibitory effect ofHQ was strong. Thus, in the three soils incubated for 48
hours, HQ at the lower rate brought about 48, 36, and 24% inhibitions, respectively,
whereas at its higher rate the inhibitions were 68, 100, and 51 %, respectively.
b) The effect of antimetabolites and HQ on volatilization of NH3 from urea was
studied with a sandy soil. Field-moist samples (10 g oven-dry material) were treated
with 2 ml of aqueous solution containing 10 mg of urea-N with or without 0.5 mg of
antimetabolite or HQ. Water was then added to bring the total volume of water to 3 m!.
Ammonia evolved during incubation (14 days at 20De) and urea and exchangeable NH/
contents in incubated samples were determined. It was found that in contrast to HQ
which markedly retarded urea hydrolysis and greatly reduced volatilization ofurea-N as
NH3 , PSA, DTB, and OTe had no effect on these processes.
165
c) To promote proliferation of microorganisms and, thus to enhance microbial

production of urease, field-moist samples (5 g of oven-dry material) of two soils were
treated with 1 ml of aqueous solution containing 10 mg of glucose with or without 50
J.lg of antimetabolite. Water was added until total water volume was 2 ml. After 0, 24,
and 48 hours of incubation at 30°C, the samples were submitted for determination of
urease activity. Urease activity increased in glucose-treated samples to the same extent
in either the absence or presence of PSA, DTB, and OTC, which proves that glucose
promoted microbial production of urease and this process was not inhibited by any of
the three antimetabolites studied.
A practical aspect should also be emphasized. Even if the antimetabolites studied
had proved effective for retardation of urea hydrolysis in soil, practical use of these
compounds would be precluded by their very high costs (DTB and OTC are 300-1,000
times more expensive than HQ).
2.31. NATURAL PRODUCTS
2.31.1. Coal and Peat

Dobrotvors'ka (1956) studied the effect of brown coal on the urease activity in samples
of a chemozemic soil from the Ukraine. The reaction mixtures contained 5 g of soil, I
ml of toluene, I g of urea in 25 ml phosphate buffer (PH 7.2), and coal powder (its
amount is not specified in the paper). Reaction mixtures with heat-sterilized soil and
without coal were the controls. After incubation (25-28°C/40 hours), the NH4 + formed
from urea was determined. It was found that addition of coal powder caused a 37%
It is mentioned in a short abstract that the long-lasting composite fertilizers invented
by Zhao (1993) and patented in China comprise plant nutrients and soil urease inhibitors
selected from weathered coal and turfy soil. These inhibitors, as shown by Zhao et al.
(l993c), are humic acids, whose capacity to inhibit soil urease activity was
demonstrated by these investigators in a laboratory experiment.
In the laboratory experiments of Siva et al. (1999), pellets prepared from both urea
and peat and urea pellets were surface-applied on an alkaline soil (PH in H2 0 7.96) and
an acid sulfate soil (PH 2.96) from West Malaysia. The peat (PH 3.64) also originated
from this region and was oven-dried at 60°C for 3 days and then finely ground to pass a
I-mm screen. The amount of peat in pellets was 40, 50, and 60% (weight/weight) in
relation to 0.52 g of urea. Volatilization of ammonia from the alkaline and acid soils
was measured during 8 and 14 days, respectively.
Less NH3 volatilized from the urea+peat pellets than from the urea pellets. The
effect of peat was attributed to NH/ adsorption. Inhibition of urease activity by peat
was ruled out based on the finding that the pH increase was higher at the microsite of
the urea+peat pellets than at the urea pellets (the microsite was assigned to soil within
15 mm radius of the pellet).
It should be mentioned that urease activity in soils and peat was assayed only before
starting the experiment and it was found that the activity (expressed in J.lg NH/-N/g
soil/hour) gave nearly identical values: 86.905 in alkaline soil, 86.899 in acid soil, and
86.906 in peat.
It should also be emphasized that peat addition to soil followed by surface-
application of urea may greatly increase the N losses through volatilization of ammonia.
166
Thus, in the laboratory experiments of Sengik and Kiehl (l995a), peat increased these
losses by 843%. A reddish-brown podzolic clay soil from the State of Sao Paulo, Brazil
was studied. The rate of peat addition was equivalent to 10 tlha and that of urea to 72 kg
Nlha. TIle peat was mixed with the top centimeter of soil and urea in solution was
uniformly distributed on the soil surface. Volatilization of NH3 was measured over 23
days.
2.31.2. Humic Substances. Lignins. and Tannins

The first data on the effect of these substances on soil urease activity were published by
Bremner and Douglas (1971). They applied the 5-hour test in evaluation of the effect of
three humic acid preparations (isolated from Iowa soils by alkali extraction and acid
treatment), two commercial lignin preparations, and five commercial tannin
preparations on urease activity of two soils (silty clay loam and clay loam). All
preparations were used in solid state at a rate of 50 ppm relative to soil weight. The
results have shown that none of these substances gave more than 4% inhibition of soil
urease activity.
The effect of humic substances and lignosulfonates on soil urease activity was also
dealt with by several research groups.
Humic suhstances. Ceccanti et al. (1978) described the fractionation of the hurnic-
urease complexes extracted from an Italian alpine podzolic soil of clay loam texture (pH
3.9). 0.1 M sodium pyrophosphate (pH 7.1) was used for extraction and ultrafiltration
and gel chromatography techniques were applied for fractionation. Five fractions of
different molecular weights were obtained, each exhibiting urease activity. Removal of
the humic molecules from the complexes led to increased urease activity. This finding
indicates that urease is present in soil in a state in which some of the enzyme molecules
are inhibited by humic substances.
Gonzalez Carcedo and Perez Gonzalez (1980) obtained three humic acid (HA)
preparations from each of the four alpine soils studied (Sierra Demanda, Spain), through
sequential extraction with solutions of sodium tetraboratc O.l M (PH 9.7), sodium
pyrophosphate 1% (pH 9.8), and sodium hydroxide 0.1 M (PH 13.0), followed by acid
precipitation and centrifugation. The inhibitory effect of the extracts was tested with
two calcareous soils (one from a white poplar, Populus alba, plantation and the other
from a garden). The reaction mixtures, composed of 109 of soil + 1 ml of toluene + 5
ml of urea solution (10 mg N) + 5 ml of water or 5 ml of a solution containing 0.5 mg
ofHA-carbon, were incubated at 37°C for 5 hours, then analyzed for residual urea.
The analytical data showed that the NaOH-extracted HA from each alpine soil
inhibited more effectively the urease activity in both calcareous soils than did the HA
from the other two extracts. For urease activity in the poplar soil, the pyrophosphate-
extracted HA was a more effective inhibitor than was the tetraborate-extracted HA. For
urease of the garden soil, the inhibitory effect of the pyrophosphate-extracted HA was
equal to or weaker than that of the tetraborate-extracted HA. However, the mean value
of the inhibitions caused by the three HA preparations was equal to approximately 4%
in both calcareous soils, this value being similar to that recorded by Brenmer and
Douglas (1971 ).
Patti et al. (1992) prepared nitrohumic acid (NHA) by single nitric acid oxidation of
a low rank brown coal sampled in the Loy Yang mine in Victoria, Australia.
Ammoniated nitrohumate (ANH) was also prepared by adjusting the pH of an aqueous
167
slurry of NHA to 7 with NH 4 0H. The effect of NHA and ANH on urea hydrolysis and
nitrification was studied with three soils: an alkaline (PH 7.7) sand, an acid (PH 5.4)
sandy loam, and an acid (pH 5.0) sand. N-(n-butyl)thiophosphoric triamide (nBTPTA)
was used as a reference compound. The reaction mixtures contained 109 of soil,S mg
of urea-N, 0 or 0.75 mg of NHA or ANH or 0.1 mg of nBTPTA, and water to field
capacity, and were incubated at 25°C for 1, 2, 7, 14,21 or 28 days. During incubation
the volatilized ammonia was measured, and after each incubation time the reaction
mixtures were analyzed for ammonium, nitrite, and nitrate.
The results showed that NHA and ANH had little effect on the rate at which
ammonium was produced by urea hydrolysis in the three soils studicd. In contrast,
nBTPTA inhibited urea hydrolysis; the inhibition was stronger in the alkaline soil than
in the two acid soils.
Ammonia volatilization from the alkaline soil was insignificantly affected by NHA
and ANH and significantly reduced by nRTPTA, whereas the NH3 loss from the two
acid soils was negligible (0-2 j.lglg soil).
The nitrate content was also very low (0-3 ~Lglg soil) in all reaction mixtures in any
timc pcriod. In the alkaline soil, but not in the two acid soils, production of nitrate was
rctarded by NHA and particularly by ANH during the second week of incubation and by
nBTPTA during the first two weeks of incubation. This effect of NHA and ANH was
attributed to inhibition of nitrification, whereas the effect of nBTPT A was explained by
inhibition of urease activity and thus, by the decreasing amount of ammonium available
for nitrification.
The conclusion drawn was that NHA and ANH did not inhibit urease activity in any
soil, but inhibited nitrification in one of the three soils studied.
Ablizova and Tomina (1997) studied the effect of sodium and ammonium hlU11ates
on the urease activity in a dark-chestnut soil from Kazakhstan. Two experiments were
done, in 199 I and 1992, respectively. In the first experiment, sodium humate (SH) was
applied at rates of 0.5, 1, and 1.5 t/ha before the planting of tomatoes. In the second
experiment, the soil was treated with ammonium humate (A H) (0.3 and 0.6 t/ha) before
the planting of onions. Soil urease activity was measured seasonally in the first
experiment and recorded only in summer in the second experiment. The results have
shown that each rate of SH decreased urease activity in spring and summer and
increased it in autumn. The decrease was more pronounced in sumnler than in spring.
The lower rate of AH caused a 25% inhibition of urease activity, whereas the inhibition
was 42.6% at the higher AH rate.
Albiach et af. (2000) conducted as-year (1989-1994) field experiment on a
horticultural soil at Moncada, Valencia, East Spain to study the effect of organic
amendments, including a commercial humic acid (HA) product (a solution of hurnic
acids containing 20% organic matter), on soil enzyme (including urease) activities and
microbial biomass. The HA solution was applied at a rate of 100 l/ha/year on plots (4.8
by 12.0 m), which also received mineral fertilizers at aru1Ual rates of250 kg of Nih a (as
ammonium sulfate), 120 kg of P20;Jha (as calcium superphosphate), and 250 kg of
K2 0/ha (as potassium sulfate).
Soil enzyme activities and microbial biomass were determined only in the 4th and
5th years. Urease activity was not significantly affected by HA addition in any of these
years.
168
Lignosu!fimates. Lignosulfonate is a waste product of the pulp and paper industry

produced from the acid sulfite pulping processes. The effect of calcium lignosulfonate
(CaLS) on urea hydrolysis and nitrification in soil was evaluated by Xie et al. (1993).
Two commercial forms of CaLS were used: the bulk-digested CaLS (BD) contains a
high level (20%) of reducing sugars, and the nearly sugar-free CaLS (SF) is a product of
desugaring treatment (it contains 4.7% reducing sugars). BD is a light-brown powder
and SF is a brown powder, and both are highly soluble in water. BD is more acid (PH
3.6) than SF (pH 5.7).
Ten-g samples (on oven-dry basis) of a Canadian clay soil (PH 5.7) were mixed with
BD or SF at rates ofO, 5,10, and 15% of soil mass, then urea solution was added (1,000
mg N/kg dry soil) and water to 80% of field capacity. The mixtures were incubated at
2(tC for 6 up to 96 hours, then analyzed for urea, ammonium, and nitrate as well as for
total N contents.
The half-life times of urea in the soil samples treated with 0, 5, 10, and 15% CaLS
had the following values: 36.3, 14.4, 44.1, and 79.7 hours, respectively, in the BD
treatments, and 39.4, 3l.4, 169, and 165 hours, respectively, in the SF treatments. These
values show that at the 5% rate both BD and SF stimulated urea hydrolysis, whereas at
the higher rates their effect was inhibitory. The stimulatory effect was stronger and the
inhibitory effect was weaker in the BD treatments than in those with SF.
The increase in ammonium-N content was less than the difference between the
added (1,000 mg N/kg soil) and the recovered urea-N contents, but the total N content
indicated that there was no loss of N during the whole incubation period (96 hours).
Therefore, it can be stated that the unaccounted N was present as microbial and/or
organic soil N.
Nitrification was studied in a separate experiment without use of urea. Air-dried
samples (100 g) of the clay soil were treated with BD or SF (0 or 5, 10 or 20% of soil
mass), then water was added to 80% of field capacity. The mixtures were incubated at
20°C for 40 days under aerobic conditions and then analyzed for NH4 +, NO)', and N0 2 -.
The nitrate contents in the BD-treated mixtures were 0.5-1.2% and those in the SF-
treated mixtures were l.5-lO.1 % of the nitrate content in the control. In other words, BD
was more effective than SF in reducing nitrate contents. At the same time, the
ammonium contents were higher and the nitrate plus ammonium contents were lower in
the BD- and SF-treated mixtures than in the control. It was deduced from these findings
that BD and SF enhanced microbial immobilization of mineral N, inhibited nitrification,
and possibly induced denitrification of N0 3- by sugars added with CaLS, particularly
BD.
Meier et al. (1993) performed a similar experiment on a Canadian silty clay loam
soil with ammonium lignosulfonate (ALS) (containing 17% reducing sugars). Air-dried
soil samples (100 g) were weighed into containers (7.5 cm diameter).
Several treatments were applied, including treatments with urea (U) only (10.1 mg
N/g soil); U+diammonium phosphate (DAP) (7.4 + 5.9 mg N/g soil); U+ALS;
U+DAP+ALS. The rate of ALS was always 26.7 mg/g soil. U+ALS and U+DAP+ALS
were well mixed and pressed into pellets, which were then placed in bands across the
containers 1 cm below the soil surface. The samples were moistened to 80% of field
capacity.
Two series of containers were prepared: Series I for measurement of CO 2 evolution
and NH3 volatilization and Series 2 for analysis of urea, ammonium, and nitrate. In
169
Series 1, the incubation was carried out at 24°C and lasted 69 days. In Series 2, the soil
was analyzed eight times during 3 8-day incubation.
CO 2 evolution from the treated soil increased in the order: U<U+ALS<
U+DAP+ALS during the first 25 days of incubation, then reached a steady state. The
high initial CO 2 evolution in the ALS-treatments was attributed to the stimulated
microbial activity by the sugars contained in the ALS. However, only 10.4-22.2% of the
ALS-C was evolved as CO 2 over the 69 days of incubation, indicating that microbial
degradation of ALS is a slow process.
Urea remained in the soil longer in the U+ALS treatment than in the U treatment,
proving that ALS inhibited urease activity. Urea hydrolysis was not reduced in the
U+DAP+ALS treatment, which was attributed to increased microbial production of
urease in this treatment.
The volatilized NHrN loss during the 69-day incubation, expressed as the
proportion of the added N, was significantly (p<0.05) lower from the U+ALS than from
the U treatment and insignificantly lower from the U+DAP+ALS than from the U+DAP
treatment.
Ammonium-N content after the 38-day incubation was significantly higher in the
U+ALS than in the U treatment and in the U+DAP+ALS than in the U+DAP treatment.
The reverse was true for the nitrate-N content. These results suggest that nitrification
was inhibited by ALS.
It was concluded that ALS has the potential to increase fertilizer N efficiency.
In the laboratory experiments of Xie et al. (1994), urea and ALS (PH 4.5) were
applied at several rates. Fresh samples of a Canadian clay soil equal to 30 g dry weight
were mixed with ALS (0, 25, 50, 100 or 150 glkg soil). Urea solution was added to the
mixtures (0, 500 or 1,000 mg N/kg soil). The moisture content was 70% of field
capacity. The incubation took place at 25°C and lasted 60 days.
It was found that addition of ALS enhanced the transformation ofurea-N into NH4 +-
N and organic N fraction and reduced the production of N03--N. This means that in this
experiment ALS stimulated urea hydrolysis, but inhibited nitrification.
2.31.3. Plant Residues and Extracts
2.31. 3.1. Residues and Extracts Containing Poiyphenois

Fernando and Roberts (1976) studied plant residues rich in oxidized polyphenols
(tannins): waste tea (Camellia sinensis) (stalk and fluff discarded from factories), bark
of Acacia decurrens, and seed coat ofinknut (Terminalia chebula) from Sri Lanka. The
dry and powdered residues (25 g) were extracted with 100 ml of boiling water for 5
minutes; the extract was filtered and aliquots were used for the urease inhibition studies.
Solid urea-extract mixtures were also prepared by evaporating to dryness a saturated
solution of urea in filtered extract. Samples of a tea soil were used for testing.
In a short incubation (16 hours), the solid tea waste added to 5-g soil samples in
amounts of 0.25, 0.5, 1.0, and 1.5 g caused 44, 48, 52, and 58% inhibitions in urease
activity, respectively. The plant extracts were more effective in the order:
Acacia extract = inknut extract> tea extract.
In longer incubations (6-24 days), ammonia losses from urea were stimulated by
solid tea waste, but the losses were reduced when solid urea-extract mixtures were
170
added to the soil. In this case as well, the mixtures prepared with Acacia and inknut
extracts were more effective than the mixture prepared with tea extract.
These observations were explained by assuming that the tea waste served as
substrate for the microorganisms which proliferated and produced new amounts of
urease, whereas the polyphenols present in extracts inhibited not only the soil urease but
also the proliferation of microorganisms.
Wickremasinghe et at. (1981) presented indirect data which confirm that the tea
residues inhibit urease activity in soil. Studying four acid tea soils (PH 4.0-4.5) in Sri
Lanka, these investigators identified a negative relationship between polyphenol content
and urease activity in soils, demonstrated by the following values: 17, 24, 30, and 68 Ilg
polyphenols/g soil and 6.7, 4.5,2.5, and 1.6 Ilg urea-N hydrolyzed/g soillhour. It should
be mentioned that the tea leaf litter contains 15-20% polyphenols.
The relationship between urease activity and polyphenol content in soil was also
studied by Sivapalan et at. (1983). Samples of a red-yellow podzolic soil (clay loam, pH
4.7) from Sri Lanka were treated, in vegetation pots, with the following plant materials
rich in polyphenols: 1. tea shoot tips; 2. mature tea leaf; 3. a mixture (1:1
weight/weight) of I and 2; 4. black tea; and 5. spent tea leaf, or with plant materials
poor in polyphenols: 1. dadap (Erythrina lithosperma) leaf; 2. mana grass
(Cymbopogam c01!{ertijlorus); 3. Guatemala grass (Tripsacum (axum); and 4. tobacco
(Nicotiana tahacum) leaf. The plant material was added at a rate of 5 g/kg soil and the
addition was repeated twice at 4-month intervals, i.e. the total amount was 15 glkg soil.
All soil samples were moistened and incubated in a greenhouse for 12 months, then
analyzed for urease activity, humic matter, and polyphenol contents.
Urease activity, expressed as Ilg NH/-N/g soil/5 hours, increased in the plant
material-treated soil samples as opposed to that of the untreated control soil. The
increase in urease activity was smaller in samples treated with polyphenol-rich plant
materials (from 14.9 to 32.3-51.0, on average, 44.7) than in samples treated with
polyphenol-poor materials (from 14.9 to 60.6-138.7, on average, 97.2). The hurnic
matter and polyphenol contents showed the reverse trend, with values of 12.68-13.80
mg humic-C/g soil and 26.6-69.2 Ilg polyphenols/g soil, respectively, in samples treated
with polyphenol-rich materials, and 11.56-12.19 mg humic-C/g soil and 4.0-6.0 Ilg
polyphenols/g soil, respectively, in samples treated with polyphenol-poor materials. The
smaller increase in urease activity of soil samples treated with polyphenol-rich materials
was attributed to the polyphenols. However, calculations indicated that there were no
significant correlations between urease activity and humic matter or polyphenol content.
This suggests that urease activity is influenced by the interaction of several factors,
including the inhibitory effect of the polyphenols and the stimulatory effect of the
hurnic substances.
The conclusion drawn is that the tea soils humified with tea leaf litter have a
partially inhibited urease activity and therefore are not susceptible to anunonia losses
following urea fertilizer application.
Under the experimental conditions described above, the polyphenol-rich and -poor
plant materials did not exhibit any inhibition of nitrification. This suggests that the soil
urease is more sensitive to polyphenols than are the nitrifying microorganisms
(Sivapalan et aT. 1985).
171
2.31.3.2. Extracts Containing Saponins

Sarsaponin is an extract from Yucca schidigera and consists of a mixture of steroid
saponins. According to the observations of Berg (1977), sarsaponion 40% added to the
feed of day-old turkey poults, at rates of 28.35 and 56,70 glt feed, reduced volatilization
of ammonia from poultry manure, indicating an inhibitory effect on urease activity in
manure and, possibly, on the soil urease activity as well.
But Broadbent et at. (1985) found, in a field experiment, that sarsaponin instead
exerted a negative effect on the test plant, maize (Zea mays) (see page 262).
2.31.3.3. Neem Cake. Oil. and Extracts

Neem or margosa (Azadirachta indica) is an Indian tree, from the seeds of which an
odorous oil is obtained. The neem oil is non-edible due to certain toxic substances it
contains. It is used in technics, mainly for manufacturing of soaps. The neem seeds are
also used for extracting a bioinsecticide. The extract contains a great number of
triterpenoids, including seven tetranortriterpenoid isomers called azadirachtins (AZ-A to
AZ-G), among which AZ-A (Figure 63) constitutes 85%, has the highest insecticidal
activity and is the strongest antifeedant for various insect pests (Sundaram, 1996).
Figure 63. Structure of azadirachtin A (AZ-A).
Persistence of azadirachtins A and B (active ingredients of the commercial

insecticide Margosan-O) in samples of a fertile soil from the Puyallup Research and
Extension Center of Washington State University was studied by Stark and Waiter
(\995). They found that both compounds were more persistent at 15°C than at 25°C and
were less persistent in nonsterile (microbially active) soil than in autoclaved soil. Thus,
the half-life times of AZ-A and AZ-B were 43.9 and 59.2 days at 15°C, and 19.8 and
20.8 days at 25°C in nonsterile soil; and 91.2 and 115.5 days at 15°C, and 31.5 and 42.3
days in autoclaved soil, respectively. The values of half-life times indicate that the
contribution of microbial activity to decomposition of both compounds was more
pronounced than that of the abiotic factors.
In Sundaram's (1996) experiments nonsterile and autoclaved samples of a Canadian
forest nursery soil were amended with 5 Ilg of pure AZ-A/g soil and incubated at 21°e.
The half-life time of AZ-A was 25.77 days in nonsterile soil and 37.65 days in
autoclaved soil. Sundaram (1996) also determined the half-life time of pure AZ-A in
172
buffer solutions and in a pond water: it was only 2 hours at pH 10, 12.9 days at pH 7,
and 19.2 days at pH 4 in buffer solutions, and 6.91 days in the pond water (PH 8.08). In
other words, AZ-A is very labile at alkaline pHs.
The inhibitory effect of neem cake (cake obtained after expulsion of oil from the
seeds) on nitrification has been known since the 1940's. The study of its effect on urea
hydrolysis in soil was initiated much later.
Reddy and Prasad (1975) added urea or a urea-neem cake mixture (containing by
weight 80% urea and 20% finely powdered neem cake) to 200-g air-dried samples of an
Indian sandy clay loam soil (PH 7.8). The rate ofurea-N addition was 100 ppm (on soil
basis). After thoroughly mixing the soil and fertilizer, enough water was added to bring
the moisture content of soil to field capacity. Incubation was carried out at a mean room
temperature of 29°C. At weekly intervals, the soil was analyzed for residual urea, NH/,
NO)-, and N0 2- contents. The residual urea in the urea-only treatment represented 9, 4,
3, and 0%, of the added urea after 1,2,3, and 4 weeks of incubation, respectively_ The
corresponding values in the urea-neem cake treatment were 15, 12, 8, and 0%,
respectively. In other words, neem cake had only a weak influence on urea hydrolysis.
Its inhibitory effect on nitrification was a little stronger during the first two weeks of
incubation_ Similar results were obtained when this experiment was repeated with
samples of different rice-growing soils (Thomas and Prasad, 1983).
The effects of neem cake on urea hydrolysis, urease activity, and ammonia
volatilization from urea in soil were the objectives of a series of studies performed in
India. First the pot and laboratory experiments, then the field experiments wiu be briefly
described.
Pot and Laboratory Experiments. Nair and Sharma (1979) used I-kg samples of a
silty clay loam (pH 6.9) placed in pots. Urea was applied at rates of 30 and 60 mg N/kg
soil, whereas neem cake had a proportion of 15% by weight of urea. Soil moisture was
brought to field capacity. Incubation took place at 30°C and lasted 4 weeks, during
which the residual urea, ~ +, N0 3 -, and N0 2 - contents were determined weekly. In the
urea-only treatments, 6 and 11 % of the added urea (30 and 60 mg N/kg soil,
respectively) remained unhydrolyzed after 1 week, and the hydrolysis became complete
in 2 weeks. In the urea-neem cake treatments, the corresponding values were 16 and
25% (after 1 week), 7 and 15% (after 2 weeks), 1 and 4% (after 3 weeks), and 0% (after
4 weeks). These values show that the neem cake reduced the rate of urea hydrolysis. It
has also been found that the neem cake was more inhibitory to nitrification than
ureolysis.
Reddy and Mishra (1983) studied the effect of neem cake on urease activity and
ammonia volatilization from urea in samples of an alkaline soil (sandy loam, pH 8.0).
Urea priUs were applied on the surface of moist soil, at a rate of 100 kg Nlha, without
cake or in mixture with 20% (weight/weight) powdered cake. Urease activity, expressed
in mg of hydrolyzed urea/kg soillhour at 30°C, was reduced from 11.8 (soil treated with
urea only) to 9.4 (soil treated with urea + cake). Cumulative NH3 losses in 16 days
decreased, under the influence of cake, by 31.4% as compared to losses registered in the
urea-only treatment. However, the neem cake was less effective than p-benzoquinone
used at a rate of 1% relative to weight of urea.
The inhibiting effect of neem cake on urea hydrolysis in flooded soils was also
reported by Hulagur and Shinde (1984).
173
In the pot experiments of Bopaiah and Biddappa (1987), 500-g samples of three
acid, cocoa palm (Cocos nucifera)-growing, soils (coastal sand, red sandy loam, and
laterite) were submitted to the following treatments: urea alone (250 ppm); urea mixed
with 30% neem cake powder; neem cake and coaltar mixed with urea in the presence of
kerosene (1 %); coir' dust coated with urea (at 9: 1 ratio). Soil moisture was kept at 50 %
of WHC during incubation (29°C). In each of these treatments, only trace amounts of
urea were detected after 10 days of incubation of the sandy loam and laterite soils. In
the coastal sand, unhydrolyzed urea was found, even after 10 days of incubation, in the
following amounts (in mg Nil 00 g soil): 0.68 (urea <IDly), 1.14 (urea + cake), 1.18 (urea
+ cake + tar), and only traces (urea + coir dust). In other words, neem cake did not
prove to be a potent inhibitor of urea hydrolysis in any of the three soils studied.
In the laboratory experiment performed by Sharma and Gupta (1989), 250-g air-
dried samples of an alkaline sandy soil (pH 8.2) were moistened up to and maintained at
field capacity. Urea prills at a rate of 100 kg Nlha were surface-applied. The other
treatments comprised: urea mixed with neem cake (1: 1 weight/weight); urea coated with
neem oil (urea dipped for half an hour in neem oil and then taken out and dried); urea
coated with shell-lac. During incubation (for 18 days at 35°C), the evolved ammonia
was assessed. Cumulative volatile NH3 losses in 18 days were the following: 20.4%
(urea only), 13.2% (urea + neem cake), 11.2% (urea + neem oil), and 9.1 % (urea +
shell-lac) relative to the added urea-No This means that the amendments reduced the
volatile NH3 losses from urea.
Mahajan and Tripathi (1991) collected samples from different depths (0-15, 15-30,
30-45 ,and 45-60 cm) of a silt loam soil (PH 5.9). The samples were ground and passed
through 2-mm sieve and packed in polyethylene columns (of 75-cm length and 8-cm
diameter) in the same order as they were obtained from the field. Five sources ofN (115
ppm): urea, urea supcrgranule (USG), lac-coated urea (LCU), sulfur-coated urea (SCU),
and neem cake-coated urea (NCCU) were mixed in the upper 0-10 em soil layer. Two
methods of watering were used: a) submergence (5-cm standing water was kept
constantly on the soil columns) and b) intermittent watering (the soil was irrigated to
maintain saturation). To determine leaching losses, the leachates were analyzed for urea,
NH/, and N0 3' at different intervals during 61 days of incubation and finally the
cumulative losses were calculated.
Total cumulative N leaching losses from the five N sources expressed as percentages
of the added N increased in the order:
SCU (l9.0%)<LCU (34.9%)<USG (S1.9%)<urea (62.S%)<NCCU (67.6%) under
submergence, and
USG (35J)%)<SCU (57.8%)<urea (65.5%)<LCU (76.6%)<NCCU (90.8%) under
intermittent watering.
These data show that leaching losses of N were lower under submergence (except
for USG) than under intermittent watering, and the losses were always highest from the
NCCU. Under submergence, the major form of the cumulative leaching losses ofN was
urea-N from each N source, but under intermittent watering, these losses were in form
of N0 3--N from LCU and SCU, in form of both urea-N and NH/-N from USG and
urea, and in the form ofurea-N (34.2%), NH/-N (21.5%), and N0 3 --N (35.1%) from
'Coir is a material for cordage, matting etc., consisting of prepared fibres ofthe outer husk of the coconut.
174
NCCU (which indicates that in this experiment the neem cake inhibited urea hydrolysis
and nitrification almost to the same extent).
Kubade and Mohite (1994) found that neem cake coating on urea retarded the
release of NH4 + from urea and nitrification of NH4 + to a lesser extent in samples of a
gypsum-amended saline sodic loamy soil (pH 8.8S) than in samples of a nonsaline clay
soil (pH 8.20).
In a pot experiment, Singh et af. (1996) studied the effect of various soil water
regimes on the volatilization of ammonia from prilled urea, urea supergranule (USG),
lac-coated urea (LCU), and neem oil-coated urea (NOCU). Samples (3.S kg/pot) of a
clay soil (pH 8.2) were used. Deionized water was added to the samples to bring the soil
to field capacity, SO% field capacity or saturation. Other samples were submerged under
a 3-cm water layer. The urea rate was 131.4 mg N/kg soil. The USGs of I-g weight
were placed at S or 10 cm below the surface. NOCU was obtained by mixing urea
granules and neem oil in a proportion of 1: 1 (weight/weight). The prilled urea and the
coated ureas were surface-applied. The volatile NH3 was measured during 4 weeks.
Depending on the water regimes, the cumulative NH3 losses from each N source
presented the order:
field capacity>SO% field capacity>saturated>submerged.
Depending on the nature of N sources, the following order of the cumulative NH3
losses was established:
prilled urea>USG placed at Scm >USG placed at 10 cm>LCU::::NOCO.
Consequently, in this experiment, urea granules coated with neem oil or lac were
more efficient than urea supergranules in reducing the volatile NH3 losses.
Field Experiments. To study the effect of neem cake (ID leaching losses of N from
urea, Singh and Singh (1986) used lysimeters installed in the field on a silty clay loanl.
The lysimeters had the dimensions of 120 cm in diameter and ISO cm in depth. The test
plant was wheat. The experiment was conducted over two years (1980/81 and 1981182).
The soil was fertilized with 0,60, and 120 kg urea-N/ha or with the same N rates of
urea to which neem cake was mixed in a proportion of 1: I by weight. The soil was also
fertilized with 36 kg of P (as single superphosphate) and 33 kg of Klha (as muriate of
potash). All fertilizers were mixed in the surface soil and then wheat seed~ were sown.
Soil was irrigated and the leachate was collected at 20-day intervals and analyzed for
NO]' and NH/. The soil was also analyzed for N0 3' and NH/. Total N content in the
plants was also detemlined.
In both years and at both urea-N rates, the neem cake significantly decreased the
leaching loss and downward movement of NO] -N, which was attributed to its inhibiting
effect on nitrification. At the same time, neem cake addition resulted in increased
leaching loss and downward movement ofNH/-N, which can be interpreted as lack of
its urease-inhibiting effect. However, the total (N03'-N + NH/-N) loss from the added
N was significantly reduced by the neem cake. The reduction was IS and 22% in the
first and second year, respectively, at the 60 kg urca-Nlha rate, and 33% in both years,
at the higher urea-N rate (120 kg/ha).
Agrawal et al. (1988) carried out a field experiment on microplots (l m 2), moistened
to field capacity. Urea granules (S g), urea + neem cake (I 6% weight/weigllt), urea +
neem oil (1'% weigllt/weight) or urea supergranules (S balls of I g each) were placed in
the center of microplots at IS cm depth. Soil samples were drawn from each microplot,
laterally designated as LJ (IS cm), L2 (30 cm), and L3 (4S cm) and vertically as DJ
175
(5 cm), D2 (15 cm), and D3 (25 cm) from the point offertilizer placement, after 7, 21,
and 28 days and analyzed for NH/ and N0 3-. Diminution of urea hydrolysis was
calculated for the LJD2 sample at day 7, obtaining the following values: 20.7% (urea +
neem cake), 27.9% (urea+neem oil), and 33.8% (supergranules), relative to the urea-
only treatment. It is evident that neem cake and oil inhibited urea hydrolysis, but this
process was best regulated through supergranule application. Neem cake and oil also
inhibited nitrification.
Santra et al. (1988) studied the effect of neem cake on volatilization of ammonia
from urea in a lowland rice field on an alluvial silty clay loam (PH 8.4). Urea was
applied in form ofprilled urea (90 kg Ntha) or neem cake-coated urea (90 kg Ntha) or
prilled urea (45 kg Ntha) in combination with a green manure [6-week-old dhaincba
(Sesballia aculeata) plants] incorporated into the soil at a rate of2.2 ttha for 45 kg Ntha.
After fertilization the evolved NH3 was measured. During 15 days, the cumulative
volatile NIl3 losses were highest (12.9-13.4%) in the urea-only treatment and lowest
(5.5-7.0%) in the urea + green manure treatment, and intermediary in the urea + neem
cake treatment. This means that the NIl3 losses were reduced under the influence of
neem cake.
A similar experiment was conducted by Mishra et at. (1990) on a rice field. The soil
of the plots was a silty clay loam (PH 7.6). Prilled urea, neem cake-coated urea, and
shell-lac coated-urea were applied at a rate of 90 kg urea-Ntha. In the dhaincha + urea
treatment, shoots of this plant as green manure supplied 45 kg Ntha and the rate of urea
was 45 kg Ntha. The fertilizers used as basal dressings were applied before
transplantation of rice plants. After transplanting the plots were kept flooded (5-7 cm)
until flowering stage. The prilled urea was also applied in three splits: 45 kg Ntha as
basal dressing + 22.5 kg Ntha as top dressing at tillering + 22.5 kg Ntha as top dressing
at panicle initiation. Ammonia volatilization was determined during 15 days after rice
transplanting. After harvest, the grains and straw were analyzed for total N.
The cumulative NIl3 losses, expressed in kg Ntha, presented the order:
prilled urea (19.22), green manure + urea (14.41), shell-lac-coated urea (13.47),
neem cake-coated urea (13.08).
It is evident that neem cake was most effective in reducing the volatile NIl3 losses
from urea. But the total N uptake by rice plants was highest in the treatment with split
application of urea.
2.31.3.4. Karanja Cake

The seed" of the karanja (Pollgamia g/abra) tree growing in India are used for
extracting a non-edible oil. The karanja cake has nitrification-inhibitory properties.
Sahrawat and Mukerjee (1977) proved that the principal crystalline chemical component
of the karanja seeds and oil, karanjin - which is a furanoflavone, 3-methoxy furano-
2' ,3',7 ,8-flavone (Figure 64) - causes the inhibition of nitrification and that the presence
176
Figure 64. Structure ofkaranjin.
of a furan ring in the karanjin molecule is the crucial structural factor for the inhibitory
activity.
Kamire and Sonar (1979) initiated a study for evaluating the effect of karanja cake
on both urea hydrolysis and nitrification in a calcareous soil (PH 8.1). Neem cake served
for comparison.
Soil samples (300 g) were uniformly mixed with 300 mg of N in the form of urea
alone and with urea replaced either by karanja or neem cake on N basis in proportions
of 25,50, 75 or 100%. All samples were moistened and incubated at 30D e for 60 days.
During incubation NH/ and N03 ' contents in soil were periodically determined. In the
presence of cakes, smaller amounts of NH4+ and N0 3' were produced than in their
absence. The rate of urea hydrolysis and nitrification decreased with an increase in the
proportion of cakes added to urea. Karanja cake was better than neem cake in delaying
the availability ofurea-N.
2.31-3.5. Mahua Cake

The seeds of mahua (Bassia lati/olia), like those of neem and karanja, are a source for
non-edible oil used in technics.
The slow-acting urea fertilizer, patented by Subramoney and Ramanathan (1979), is
prepared from urea (e.g.• 800 kg) coated with cashew (Anacardium occidentale)
nutshell oil resin (25 kg) and blended with neemlmahua cake powder (175 kg).
Results from the investigations of Mago and Totawat (1989) show that the rnahua
cake should be considered an inhibitor not only of nitrification, but also of urea
hydrolysis. In these investigations, urea granules or differently coated urea granules
were incorporated into samples of a sandy loam soil (2 kg/pot) at a rate of 40 mg N/kg
soil.
The granules were coated with an emulsion of coaltar and kerosene (1 :2), the
proportion of emulsion and urea being 1.5:300. A great number of fresh emulsion-
coated granules were also coated with finely ground mahua cake, neem cake, sulfur or
gypsum (at rates of 150 and 300 g/kg urea). Besides urea or coated urea, the soil also
received P and K fertilizers (8.8 mg of P and 16.6 mg of K/kg soil). The soil in each pot
was moistened with water of high salinity and medium sodicity. Incubation took place
at 30-32D C and lasted 21 days, during which the soil was analyzed several times for
determination of residual urea, NH/, and N0 3 ' contents.
The analytical data showed that both urea hydrolysis and nitrification were most
intense in the urea-only treatment and were inhibited in treatments with coated ureas in
the following order:
neem cake>mahua cake>sulfur>gypsum>coaltar.
177
This order means that the neem and mahua cakes were more inhibitory than the
other coating materials. The inhibitory effect of all coating materials was stronger at
their higher rather than lower rate.
2.31.3.6. Other Plant Materials

Sen and Bandyopadhyay (1986) tested the effects of fresh tamarind (Tamarindus
indica) leaves on volatilization of ammonia from floodwater of a rice field on a coastal
saline soil (silty clay, pH 7.5). Prilled urea (100 kg Nlha) was broadcast into floodwater.
The tamarind leaves (at a total amount of 5 tlha) were incorporated into the soil in two
splits, one day before and on the 4th day after fertilizer applications. Volatilization of
NH3 was measured over 16 days. Cumulative NH3 losses were reduced by 18.3% in the
treatment with urea and tamarind leaves as compared with the urea-only treatment. The
effect of the leaves was attributed to their acidity, due to which the pH of floodwater
decreased. Much lower NH3 losses were recorded when 3-g urea briquettes, without
tamarind leaves, were placed at 5-cm depth in the soil.
A flooded rice soil was studied by Xue and Li (1987). Mixtures were prepared from
5-g soil samples, 0, 0.05, 0.1, 0.2, 0.3, and 0.4 g of tobacco or tea leaves, rape or tea oil
seed cake, 0.4 g of castor-oil plant leaves, and 10 rnl of 1% urea solution. After 48 hours
of incubation at 31'C, inhibition of urease activity was highest at the rate of 0.4 g plant
material/5 g soil and had the following values: 41.12% (tobacco leaves), 35.51% (tea
leaves), 29.05% (rape oil seed cake), 22.02% (tea oil seed cake), and 44.05% (castor-oil
plant leaves). After prolonging incubation time up to 12 days, the inhibitory effect of
tobacco leaves was more marked than that of the castor-oil plant leaves (both plant
materials having been used at the rate of 0.4 g/5 g soil), but remained lower than that of
CuS04.5H 20 (0.4 g) or catechol, hydroquinone, p-benzoquinone, and quinhydrone (I 00
ppm). Castor-oil seed cake was also tested and found to be a better inhibitor of soil
urease activity than the other plant materials.
2.31.4. Microhial Products

Lin et at. (l997a,b) isolated and identified a urease inhibitor from Aspergillus
ochraceus. The inhibitor obtained in form of white, water-soluble, orthorombic crystals,
is a pyrane derivative (C9 H 12 0 4), namely 3-(1',2'-epoxypropyl)-5,6-dihydro-5-hydroxy-
6-methylpyran-2-one (Figure 65).
Figure 65. Structure of the urease inhibitor produced by Aspergillus ochraceus.
Initially, the inhibitory effect of this compound was demonstrated on jackbean

urease, then it was found to also be able to retard urea hydrolysis catalyzed by soil
urease. Its urease-inhibiting capacity was comparable to that of nBTPTA. The inhibition
is of non-competitive type.
178
2.32. A MISCELLANEOUS GROUP OF ORGANIC COMPOUNDS
Under conditions of the 5-hour test, Bremner and Douglas (1971) established that a
series of compounds (benzene, anisole, veratrol, benzoic acid, vanillic acid, maleic acid,
citraconic acid, phthalimide, iodoacetic acid, and iodocetarnide), used at a rate of 50
ppm (on soil basis), gave less than 4% inhibition of urease activity in the three Iowa
soils studied.
For inhibiting soil urease activity, Liesegang et al. (1976) patented the compounds
which have the general formula NC-CX 2-C(O)NR]R2 , where X=H, CI, Br or J; R] and
R2 = C]-C5 alkyl, cyclohexyl or part of a heterocycle with N. Evaluation of the
inhibitory effect was performed with 30-g soil samples mixed with 214.1 mg of urea
(100 mg N) and 4 mg of test compound, then moistened to 50% ofWHC and incubated
at 30°C for 24 hours, during which time the evolved ammonia was assessed.
Satisfactory inhibitions occurred even when the rate of test compound was reduced from
4 to 0.5 mg. It was also found that these compounds are more persistent in soil than p-
benzoquinone. They are recommended as additions to urea at preferred rates of 0.05-5%
relative to urea-N for fertilization of light- and medium-textured soils, meadows and
pastures.
Applying the short and long time tests (see page 51), Hartbrich et al. (1978) found
that bis(acetylvinyl) sulfide is a weaker soil urease inhibitor than mucochloric acid.
Sahrawat (1979) studied, using the 5-hour test, the effect of several chelating
compounds [nitrilotriacetic acid (NTA) trisodium salt, ethylenediarninetetraacetic acid
(EDTA) disodium salt, tartaric, citric, and oxalic acids], used at a rate of 50 ppm (on
soil basis), on the urease activity of an Indian sandy clay loam alluvial soil. The
inhibition degree was 4% (NTA), 1% (EDTA), and 0% (other compounds). It should be
mentionated that in the red-brown earth studied by Cai et al. (1989) EDTA gave more
marked inhibitions, ranging from 1.2 to 39.6% (see page 209).
According to Liao and Raines (1982), p-phenylenediamine, as an inhibitor of soil
urease activity, is comparable to organic mercury compounds, polyhydric phenols, and
quinones as well as phosphoryl and thiophosphoryl triarnides.
In the tests described by Kiss and Pintea (1987) (see page 35), the time required for
complete hydrolysis of urea was, under the influence of p-anisidine, prolonged from 6
to 10 days in an alluvial soil and reduced from 9 to 8 days in a leached chemozem. The
reaction mixtures were prepared from 5 g of air-dried soil, 10 rnl of aqueous phase
containing 6 mg of urea and 0.12 mg of p-anisidine; incubation took place at laboratory
temperature.
Caffeine, used at concentrations of 20,40, 60, 80, and 100 ppm (on soil basis) and
nicotine at concentrations of 60, 80, and 100 ppm, manifested an inhibitory effect on
urease activity in samples of a periodically water-logged paddy soil. The reaction
mixtures, consisting of 5 g of dry soil, 10 ml of 1% urea solution with or without
caffeine or nicotine, were incubated at 37°C for 48 hours. It should be noted that in the
case of caffeine the inhibition degrees were not significantly different (48.01-48.89%) at
the 20-60 ppm concentrations and were lower (43.61%) at the two higher
concentrations, but nicotine gave maximum inhibition (45.81 %) at its highest
concentration (Xue and Li, 1987).
179
Chapter 3. Combined Use of Inhibitors of Soil Urease Activity
3.1. COMBINED USE OF HEAVY METAL COMPOUNDS WITH OTHER

INHIBITORS
Geissler et al. (1970), of the Esso Research and Engineering Company worked, as
specified in their patented invention, with pellets prepared only from urea and CUS04,
i.e., without any hydrophobic material. Before pelletizing, the urea was melted at 130-
135°C, then CUS04 was added (in amounts of 0.01-1 0%, most preferably 0.2-4% based
on the weight of urea) to the urea melt and uniformly dispersed in the melt by stirring.
Some pellets were prepared from urea + borax or from urea + CUS04 + borax or from
urea + CUS04 + borax + sodium fluoride (NaF). Urea prills without inhibitor served for
comparison.
In an experiment conducted to study ammonia volatilization from urea, the pellets
(in an amount equivalent to rates at which urea is generally applied under field
conditions) were placed on the surface of moist soil samples. The cumulative NH3
losses measured during the incubation period (20 days) and expressed as percentages of
the added urea-N gave the following mean values (in the different treatments): 43.3%
(urea only); 26.2% (urea + 2% CUS04); 23.8% (urea + 4% borax); 20.3% (urea + 1%
CUS04 + 2% borax); 28.0% (urea + 0.6% CUS04 + 1.4% borax + 0.33% NaF). It is
evident that NH3 loss was lowest from the pellets that had been prepared by adding 1%
CUS04 + 2% borax to the urea melt. It should be mentioned that the NH3 losses did not
significantly diminish when the urea prills used had been merely coated with the same
amounts of the same inhibitors.
In another experiment, the same CUS04 amount was spread on soil samples at
intervals of 3, 10 or 17 days before the urea prills (without inhibitor) were applied to the
soil surface. After adding urea, the soil samples were submitted to incubation, during
which NH3 volatilization was assessed. The results showed that in this experiment
CUS04 had very little effect on reducing the rate of NH3 volatilization from urea.
However, this experiment was not repeated with borax or inhibitor mixtures (CUS04 +
borax and CUS04 + borax + NaF).
Huang et al. (1993) patented a long-acting urea fertilizer prepared by adding urease
inhibitors (Zn, Mn, Cu, Fe, B, and Ba) to the fertilizer composition.
3.2. COMBINED USE OF FORMALDEHYDE OR ANOTHER ALDEHYDE WITH

OTHER INHIBITORS
Sor et al. (1971), also of the Esso Research and Engineering Company, prepared three
kinds ofCH 20-containing urea pellets:
a) 2% CH20 + 2% borax + I % hydrophobic material (Marcol 72);
b) 1% CH 20 + 1% octadecylamine (ODA);
c) 1% CH 20 + 4% CUS04 without any hydrophobic material.
Volatilization of NH3 from these pellets applied to soil samples incubated for 3 days
was lower than from the control (no inhibitor) pellets. The NH3 losses were reduced
from 2.65-3.08 to 1.28% (pellets a), from 4.95 to 3.93% (pellets b) or to 3.07% (pellets
c). It should however be mentioned that application of urea-borax-ODA pellets led to a
more marked reduction ofNH3 loss from urea.
180
Sor et al. (1971) also prepared pellets from urea + 4% borax + 1% acetaldehyde or
1% p-to1ualdehyde or 4% a-naphtha1dehyde which proved to be more effective in
reduction of NH3 loss than pellets b and c, as - under similar conditions of incubation -
the NH3 loss was reduced from 4.95% (control pellets) to 1.13% (urea-borax-
acetaldehyde pellets), 0.54% (urea-borax-tolualdehyde pellets), and 0.48% (urea-borax-
naphthaldehyde pellets).
3.3. COMBINED USE OF HEXAMETHYLENETETRAMINE (HMTA) WITH

OTHER INHIBITORS
According to the invention patented by Neumann and Richter (1976), molten urea
(1 mole) was mixed with a) 0.02 moles ofHMTA; b) 0.0238 moles ofHMTA + 0.0145
moles of CuS04.5H20; c) 5.4 g of the dehydrated product of the reaction between
1 mole ofHMTA and 1.78 moles of boric acid; d) 0.025 moles of HMTA + 0.05 moles
of phosphoric acid (80%), then the mixtures were pelletized. Another method was also
used: urea and inhibitor(s) were melted together and the resulted melt was transformed
into pellets. Pellets containing urea only were the controls.
Volatilization of NH3 from the pellets was tested with 600-g samples of an arable
soil (moisture content:::: 50% of WHC). The pellets placed on the surface of each soil
sample contained 100 mg N. During incubation (at 30°C for 10 days), the volatilized
NH3 was measured. In comparison with NH3 volatilization from control pellets (100%),
the following NH3 volatilization values were recorded (in the different treatments): 15%
(urea + HMTA), 7% (urea + HMTA + CUS04), 5.6% (urea + HMTA + boric acid), and
9% (urea + HMTA + phosphoric acid). It should be added that HMTA also has a
physical effect: it reduces aggregation of urea pellets and, thus, makes it possible to
store the fertilizer for a longer period.
3.4. COMBINED USE OF mTHIOCARBAMATES WITH OTHER INHIBITORS
Tomlinson (1967) patented both dithiocarbamates and their combination with

halogenated alkanes as inhibitors of soil urease activity (see page 53).
3.5. COMBINED USE OF HYDROXAMIC ACIDS WITH OTHER INHIBITORS
Pugh and Waid (1969a,b) found that acetohydroxamic acid and sulfanilamide, when
applied together, exhibited a synergistic effect in inhibition of ammonia volatilization
from urea-treated samples of three English soils (see page 163).
3.6. COMBINED USE OF POLYHYDRIC PHENOLS AND QUINONES WITH

OTHER INHIBITORS
As already shown in Table 18, catechol and hydroquinone acted synergistically with
2,5-dimethyl-p-benzoquinone in inhibiting volatilization of ammonia from urea-treated
soils (Anderson, 1969, 1970).
Thieme et al. (1976) proved that this inhibitory effect of hydroquinone, p-benzo-
quinone, and quinhydrone was more intense and long-lasting when were used in
combination with tetramethylthiuram disulfide (see also page 88).
181
The data in Table 42 show that hydroquinone (HQ), 1:1 mixtures of HQ + maleic
hydrazide (MH), sodium or ammonium salt of MH prolonged complete hydrolysis of
urea in a light-textured (alluvial) soil by 4 days. In a heavier-textured soil (leached
chemozem) HQ acted more markedly than did the mixtures, complete hydrolysis of
urea having been prolonged by 7 and 5 days, respectively. In both soils, MH was a weak
inhibitor. The reaction mixtures, prepared from 5 g of air-dried soil + 10 ml of aqueous
phase containing 6 mg of urea + 0 or 0.12 mg of test compound or mixture of
compounds, were incubated at laboratory temperature (Kiss and Pintea, 1987).
TABEL 42. Effect of combined use of hydroquinone and maleic hydrazide on soil
urease activity"
Time necessary for complete
Co~ound or mixture of
hydrolysis of urea (days)
compounds
Alluvial soil Leached chemozem
Control 6 9
Hydroquinone (HQ) 10 16
Maleic hydrazide (MH) 9 12
HQ + MH (1:1) 10 14
HQ + MH sodium salt (1:1) 10 14
HQ + MH ammonium salt (1:1) 10 14
"Adapted from Kiss and Pintea (1987).
3.7. COMBINED USE OF PHENYLPHOSHORODIAMIDATE (PPDA) WITH

OTHER INHIBITORS
Based on laboratory experiments in which the short and long time tests (see page 51)
were applied, but the inhibitors were used at four rates relative to urea-N (0.5, 1,2, and
4% in the short time test, and 0.5, 1, 1.5, and 2% in the long time test), Hartbrich et al.
(1976b) patented not only PPDA, but also its mixtures with mucocbloric acid as
inhibitors of soil urease activity. The inhibitory effect of the two compounds in mixture
was strongly synergistic, especially when the molar ratio between them was 1: 1. Thus,
this PPDA-mucocbloric acid mixture at total rates of 0.5, 1, 1.5, and 2% inhibited
ammonia volatilization from urea for 10, 14,20, and 20 days, respectively. At 1.5 and
2% rates, the inhibitions were considerable even after 24 days of incubation (81 and
85%, respectively). Under the same conditions, total inhibitions by PPDA alone and
mucocbloric acid alone lasted only 4 and 6 days, respectively. Mixtures of PPDA-
mucochloric acid were recommended for practice at rates of 0.01-20%, preferably at
0.05-5% relative to urea-No
Lang et af. (1976) found that PPDA used with thiram (in proportions of 1:2.33 and
1:9, at a total concentration of 1% relative to urea-N) or with ferbam (in a proportion of
1:4, at 1% total concentration) or with dawmet (in a proportion of 1:1, at 2% total
concentration), 4-chloro-PPDA used with thiram (in proportions of 1: 1 and 1:9, at 1%
total concentration), and di-PPA used with thiram (in proportions of 1:4 and 1:9, at 1%
total concentration) manifested a synergistic effect with these compounds in inhibition
of soil urease activity, increasing the degree and prolonging the duration of inhibition.
182
The finding by Kurze (1981) that diammonium and Ca salts of imidodiphosphoric

acid diphenyl ester acted synergistically with PPDA has already been mentioned on
page 111.
In the pot experiment of Byrnes et al. (l989a), the soil urease-inhibiting effect of
PPDA was enhanced by H3 B03 (see page 248).
Luo et al. (1994) used PPDA in combination with nBTPTA. Samples were collected
from the 0-15-cm layer ofa vertisol (PH 7.6) and an alfisol (PH 6.7) in a rice-growing
area in New South Wales, Australia. The vertisol is more urease-active than the alfisol.
Reaction mixtures were prepared from 15 g of air-dried soil + 30 rnl of water + 14 mg
of urea with or without PPDA (1%) or nBTPTA (1%) or PPDA+nBTPTA (1+1, 2+2,
5+5, and 10+ 10%). The percentages are relative to weight of urea. The inhibitors were
added, followed immediately by the addition of urea. The mixtures were incubated at
25°C for 10 days, during which the residual urea was determined at 2-day intervals.
In both soils the 1% PPDA + 1% nBTPTA mixture was more inhibitory on urea
hydrolysis than 1% PPDA or 1% nBTPTA alone. For example, the rate of urea
hydrolysis was 100% (control, nBTPTA), 92% (PPDA), and 44% (PPDA+nBTPTA) in
the more urease-active vertisol after 6 days of incubation, and 100% (control), 60%
(nBTPTA), 36% (PPDA), and 18% (PPDA+nBTPTA) in the less urease-active alfisol
after 10 days of incubation.
Increasing the concentration of inhibitors in mixtures led to further increases in the
inhibitory effect. For example, in the presence of the 10% PPDA+ 10% nBTPTA
mixture, the rate of urea hydrolysis was only 6% in vertisol after 10 days of incubation.
For explaining the strong urease-inhibiting effect of the PPDA +nBTPTA mixtures,
the following mechanism was suggested: initially the soil urease was inhibited by
PPDA and during this time a part of the nBTPTA was converted to its oxygen analogue
(nBPTA); then, as the effectiveness ofPPDA declined, nBPTA blocked the subsequent
hydrol ysis of urea.
It was concluded that the combination of PPDA and nBTPTA has the potential to
improve the efficiency of urea fertilizer in flooded rice fields.
In the experiments conducted by Chaiwanakupt et at. (1996) and Phongpan et at.
(1995,1997) on a flooded rice field located on a clay soil (PH 5.1) in the Central Plain
region of Thailand, urease inhibitors were also used in combinations.
The experimental plots were 4 m x 4 m. Triple superphosphate (24 kg Plha) and KCl
(28 kg K/ha) were broadcast onto the drained soil and incorporated immediately before
transplanting 20-21-day-old rice seedlings. The water depth was adjusted to 5 cm and
maintained near that depth for the duration of the experiment.
In the first experiment carried out in the wet season of 1991 and described by
Chaiwanakupt et at. (1996) and Phongpan et at. (1997), the algicide terbutryn [2-(t-
butylamino)-4-(ethylamino)-6-(methylthio)-s-triazine], the nitrification inhibitor acetyl-
ene (provided by wax-coated calcium carbide, Cac2 ), and the urease inhibitors
nBTPTA, PPDA, N-(diaminophosphinyl)benzamide (DAPBA), and acetohydroxarnic
acid (AHA) were studied. The algicide (0.2 mg active ingredient/l floodwater) was
applied on days 0, 3, 6, and 9. Calcium carbide was applied at the rate of 40 kg active
ingredientlha. The rate of urease inhibitors was 0.5% relative to weight of urea. There
were two nBTPTA applications: single (0.5%) and multiple (0.5% on days 0, 3, 6, and
9), and a mixed urease inhibitor treatment: nBTPTA, PPDA, DAPBA, and AHA (each
183
at 0.5% rate) applied in succession at 3-day intervals. The rate of urea added was 75 kg
Niha.
There were seven treatments without algicide: 1. control (only urea); 2. nBTPTA; 3.
nBTPTA repeated; 4. mixed inhibitors; 5. Cac2; 6. nBTPTA + Cac 2; and 7. mixed
inhibitors + Cac 2 , and six treatments with algicide (algicide was not added to the
repeatedly applied nBTPTA). First the algicide, then the urease inhibitors and, finally,
the urea were broadcast into the floodwater. During 10 days after urea application,
floodwater pH, ammoniacal (NH3 + NH/)-N, temperature as well as wind speed were
measured and from these data the ammonia losses were calculated.
The cumulative NH3 losses measured in treatments with and without algicide and
expressed as percentages of the applied N were the following: 19.5 and 10.4 (control),
15.8 and 14.7 (nBTPTA+CaC 2 ), 15.2 and 12.4 (Cac2 ), 13.8 and 13.l (mixed
inhibitors+CaC 2 ), 9.8 (nBTPTA repeated), 9.3 and 7.6 (nBTPTA), and 9.1 and 7.0
(mixed inhibitors), respectively.
It is evident from these numerical data that the urease inhibitor mixture was most
efficient in reducing the volatile NH3 losses; the algicide had, in each treatment, a loss-
reducing effect, but Ca~ diminished this effect of urease inhibitors, although when
used without urease inhibitors CaC 2 also reduced the NH3 loss; nBTPTA in a single
application was more effective than in multiple applications and approached the
effectiveness of mixed inhibitors.
The second experiment, described by Phongpan et al. (1995, 1997), was carried out
during the dry season of 1992. The algicide terbutryn (again at 0.2 mg/l floodwater) was
applied at 4-day intervals in all treatments. The urease inhibitors studied were nBTPTA
and PPDA used alone or in combination. Their rates (on urea weight basis) were 0.5,
1.0, and 2% when used alone, and 0.5+0.5, 1.0+0.5, and 2+0.5% when used in
combination. Urea was applied at 60 kg Niha rate. The order of the additions to the
floodwater was, as in the first experiment, algicide, urease inhibitors, and urea. The
control received only algicide and urea. Ammonia volatilization was determined during
11 days after urea application.
It was found that the rate of inhibitors used alone or in combination had no
significant effect on NH3 loss, which means that the 0.5% inhibitor rate was sufficient to
inhibit soil urease activity and thus to reduce NH3 volatilization. The cumulative NH3
losses from the added urea-N had the following mean values: 15.0% (control), 5.4%
(nBTPTA), 7.3% (PPDA), and 3.0% (nBTPTA+PPDA), which proves the high
efficiency of the combined use of the two inhibitors.
To explain the strong urease-inhibiting effect of the nBTPTA+PPDA combination,
the suggestion made by Luo et at. (1994) (see page 182) was reiterated.
A shorter report on the second experiment was published by Freney et al. (1993).
3.8. COMBINATIONS CONTAINING HUMIC SUBSTANCES
Al-Kanani et at. (1990b) studied the effect of three inhibitors: humic substance (HS)
from leonardite, boric acid (BA), and ammonium thiosulfate (ATS) and two inhibitor
combinations: HS + ATS and BA + ATS on the transformation of urea-ammonium
nitrate fertilizer in a sandy clay loam soil (PH 6.5) and a sandy soil (PH 5.9) from
Quebec.
184
The urea-ammonium nitrate (VAN) fertilizer solution contained 10% N by weight (5

+ 5% urea-N and ~NOrN) and was surface-applied on air-dry soil samples (44-47 g)
at a rate equivalent to 147 kg Nlha, calculated on surface area basis. 1.8 and 3.6% (by
TABLE 43. Effect of humic substance (HS), boric acid (BA), ammonium thiosulfate (ATS), and combinations
ofHS+ATS and BA+ATS on transformation of urea-ammonium nitrate fertilizer in two Canadian soils·
____~~~~~--~~I~nh~ib~it.io~n~(~%~)----~~~~------
San~ c1a~ loam soil Sand~ soil
Inhibitor
Ammonia Urea Nitrifi- Ammonia Urea Nitrifi-
Volatilization hydrolysis cation vo latili zatio n hydrol~sis cation
HS (1.7%) 21.1 16 29.0 15.3 5 34.2
HS (3.4%) 20.2 22 25.5 17.4 11 34.2
BA (1.8%) 13.6 IS 20.0 3.2 5 15.5
BA (3.6%) 21.1 30 23.5 15.8 12 18.7
ATS(1.8%) 31.9 22 -2.6 33.7 7 9.3
ATS(3.6%) 37.1 25 -12.0 24.7 9 -4.7
HS (1.7%)+ATS (1.8%) 38.5 26 21.5 36.3 4 18.7
BA (1.8%)+ A TS (1.8%) 35.7 28 14.5 31.1 6 12.4
"Adapted from A1-Kanani et al. (1990b).
weight of VAN solution) of BA and ATS and 1.7 and 3.4% of the 0.1%
(weight/volume) HS were used. The samples were moistened and incubated at
laboratory temperature. Ammonia volatilization, urea hydrolysis, and nitrification were
evaluated after 10 days of incubation. The results are summarized in Table 43.
These results show that the combination HS+ATS was most inhibitory on NH3
volatilization, but the strongest inhibition of urea hydrolysis and nitrification was
caused by BA and HS, respectively. Surprisingly, the effect of ATS on nitrification was
stimulatory rather than inhibitory. The combination of HS+ATS was in general, more
effective than BA+ATS. The environmentally friendly HS as an inhibitor of soil urease
and/or nitrification should be considered superior to BA and ATS.
3.9. COMBINATIONS CONTAINING LIGNOSVLFONATES
The effect of ammonium lignosulfonate (ALS) used in combination with phenyl-

phosphorodiamidate (PPDA) or N-(n-butyl)thiophosphoric triarnide (nBTPT A) on
volatilization of ammonia from urea-treated samples of a Canadian clay soil (PH 5.2)
was studied by Al-Kanani et al. (1994).
Jars were filled with 150-g air-dried soil samples, which were then moistened and
preincubated at 23°C for 4 days. After preincubation, the samples were amended with
urea and urea + inhibitor(s) used in three forms: a) in solutions of urea and
urea+inhibitor(s); b) as dry urea prills and as dry blends - mixtures ofurea+inhibitor(s);
and c) as tablets prepared from urea+inhibitor(s). The solutions were surface-applied,
while the dry urea and dry blends as well as the tablets were both surface-applied and
banded 2 cm below the surface.
Rates of applications per jar were: 135.5 mg urea-N (equivalent to 387 kg Nlha on
surface area basis), 810 mg ALS, 1.54 mg PPDA or nBTPTA. The ratio of urea, ALS,
PPDA or nBTPTA was 1:2.7:0.005 on dry weight basis.
185
The NH3 volatilized was measured daily during l5-day incubation at 23.1°C. After
incubation the soil was analyzed for residual urea, ~+, and (N02- + N0 3}
Cumulative NH3 losses were highest from samples treated with urea only and
decreased with each inhibitor and inhibitor combination_ The extent of this effect
depended on the nature of inhibitors as well as on form and place of the urea
application.
Following surface application of solutions and dry blends and banded application of
dry blends and tablets, the decreasing effect on NH3 volatilization is as follows:
nBTPTA:::::: PPDA > ALS > ALS+PPDA:::::: ALS+nBTPTA.
In the case of surface-applied tablets, the order was:
nBTPTA:::::: PPDA:::::: ALS > ALS+nBTPTA:::::: ALS+PPDA.
These orders indicate that, in these experiments, PPDA was always as effective in
decreasing the volatile NH3 losses as nBTPTA, but ALS exhibited a comparable
effectiveness only when it was surface-applied in urea tablets. ALS used in
combinations limited the urease-inhibiting effectiveness of PPDA and nBTPTA. The
mechanism(s) by which ALS and PPDA or nBTPTA interact has not been delineated; it
was only supposed that the soil urease reacts with the negatively charged ALS and this
reaction makes the enzyme less susceptible to the inhibitors.
It follaws from the soil analyses that recovery ofN from the inhibitor-amended urea
solutions and dry blends increased in the same order as the decrease of volatile NH3
losses. However, recovery ofN from the tablets is partially different:
nBTPTA> ALS:::::: PPDA > ALS+nBTPTA:::::: ALS+PPDA from the surface-applied
tablets, and
nBTPTA:::::: PPDA > ALS+PPDA > ALS+ nBTPTA:::::: ALS from the banded
application of tablets.
187
Chapter 4. Comparative Studies on tbe Efficiency of Different Inbibitors of Soil

Urease Activity
These studies will be grouped according to the compound which proved to be the most
efficient inhibitor in the respective comparative study. But this criterion of grouping
will not be applied for those comparative studies in which a well-known urease inhibitor
was used as a reference compound.
4.1. COMPARISON OF HEAVY METAL COMPOUNDS WITH OTHER INHIBITORS
Laboratory Experiments. Xue and Li (1987) have found that CUS04 was a stronger
inhibitor of soil urease activity than were the plant materials (see Section 2.31.3 .3).
Sheudzhen et al. (1991) used surface (0-20 em) samples of a chernozemic soil
cultivated with rice in Russia. The samples were NPK-fertilized (N 180 kWba as urea, P
120 kWba as double superphosphate, K 60 kWba as KCI) and treated separately with Zn,
Mn, Cu, and Co (4, 4, 3, and 2 kWba, respectively, as sulfates), Mo (2 kglha as
ammonium molybdate), and B (2 kglha as boric acid). The control samples received
only NPK. Urease activity was assayed after 4 and 8 days. After both 4 and 8 days,
urease activity showed decreased values in the Zn- and Mn-treated samples and
increased values in the Mo-treated ones. Co did not influence, while Cu like B
decreased urease activity during the first 4 days and increased it later.
Sengik and Kiehl (1995b) have compared the effect of five inorganic salts (ferric
chloride, ferrous sulfate, simple and triple superphosphate, and monoammonium
phosphate) on volatilization of ammonia from peat- and urea-treated and moistened
samples of a red earth (PH 4.9) from Brazil. Peat and urea were added at rates
equivalent to 10 t and 72 kg Nlha, respectively. The ratios between the weight of salt
and that of the urea were 1:1,2:1, and 3:1. Soil samples treated only with peat and urea
were the controls. All samples were incubated for 22 days, during which the volatilized
NH3 was determined daily. Soil pH was measured periodically during 18 days.
The cumulative volatile NH3 losses were reduced by the salts at their 1:1, 2:1, and
3: 1 ratios to urea in the following proportions expressed as percentages of the NH3 loss
from the controls: 23, 52, and 70% by ferric chloride; 16, 46, and 49% by ferrous
sulfate; 14, 17, and 19% by simple superphosphate; 0, 0, and 25% by triple
superphosphate; and 0, 0, and 11 % by monoammonium phosphate.
Soil pH was also reduced by the salts in parallel with their reducing effect on NH3
volatilization.
It is evident from these data that the two iron salts were most effective in reducing
NH3 volatilization and soil pH. Their effect on soil urease activity was not evaluated.
Field Experiments. Sanikidze et al. (1987b) compared the effects of Zn and B on the
urease activity of a red earth under mandarin (Citrus nobilis) plantation in the humid
subtropical area of western Georgia (Gruzia). All experimental plots were NPK-
fertilized and limed in the 1974-1977 period. Some plots received, additionally, 4,8 or
12 kg Zn (as sulfate)lha or 3,6, and 9 kg B (as boric acid)lha, in the 1977-1982 period.
Urease activity in the 0-20-cm soil layer was measured systematically in the 1981-1983
period. For this period, the mean values of urease activity (expressed in mg NH3
produced by 1 g soil in 24 hours) were, in the different plots, the following:
188
0.3 (control) < 0.36 (12 kg Zn/ha) < 0.4 (8 kg Zn/ha) < 0.5 (4 kg Zn/ha, 3 and 9 kg
B/ha) < 0.6 (6 kg B/ha).
This order means that, at the applied concentrations, both Zn and B stimulated
urease activity, this effect of Zn being weaker than that ofB.
4.2. COMPARISON OF ALKALINE EARTH METAL SALTS WITH OTHER

INHIBITORS
Laboratory Experiments. Sloan and Anderson (1995) compared the effects of CaCh and
ammonium thiosulfate (ATS) to reduce ammonia volatilization from surface (0-4 cm)
samples of two soils possessing contrasting physical and chemical properties, namely an
acid Lufkin fine sandy loam soil (pH 4.9) and a calcareous Ships clay soil (PH 7.8),
both located in Brazos County, Texas.
The air-dried soil samples (30 g) wetted to 20% moisture (Lufkin soil) or to 30%
moisture (Ships soil) were incubated at 25°C for 24 hours in order to reestablish
microbial activity, then urea was surface-applied at a rate equivalent to 200 kg N/ha
based on surface area of soil samples. CaCl 2 was used at a Ca:N equivalent weight ratio
of 0.25, whereas ATS was applied in 10% amount relative to weight of urea. The
volatilized NH3 was measured for 192 hours (Lufkin soil) and for 408 hours (Ships
soil).
Inhibition of NH3 volatilization by CaCb and ATS at 24 hours after fertilizer
application was 84.9 and 8.1 % respectively (Lufkin soil), and 42.1 and 2.6%,
respectively (Ships soil). But the inhibitory effect of CaCl 2 decreased, and that of ATS
slightly increased with time. Thus, at 48 hours the inhibition of NH3 volatilization by
eaCb and ATS was 20.9 and 9.6%, respectively (Lufkin soil), and 32.5 and 5.1%,
respectively (Ships soil). The corresponding values registered at 192 hours in the Lufkin
soil were 1.2 and 11.1 %, and those registered at 408 hours in the Ships soil were 8.1 and
1.0%. The conclusion can be drawn that eaCh was always more effective than ATS in
the Ships soil, but only during the first 48 hours in the Lufkin soil.
In another experiment, in which the soil samples were submitted to rapid drying
after fertilizer application, CaCb significantly (p<0.05) reduced NH3 volatilization from
both soils, whereas the effect of ATS was not significant in the Lufkin soil, either.
4.3. COMPARISON OF INORGANIC BORON COMPOUNDS WITH OTHER

INHIBITORS
Laboratory Experiments. Sor et al. (1971) have compared ammonia volatilization from
urea granules containing an inhibitor (borax, CUS04, NH4F, formaldehyde, thiourea or
o-phosphoric acid) and octadecylamine or another hydrophobic material. The granules
were surface-applied on samples of a loamy sand soil (pH -6.4) from New Jersey. The
volatile NH3 was measured during 3-4 days. The results showed that the volatile NH3
loss was reduced most effectively by borax and least effectively by CUS04.
Bayrakli (1990) evaluated the effect of five compounds on the volatilization of
ammonia from urea-treated samples of a representative alluvial soil (PH in KCl 7.52) of
the Bafra Plain, Turkey. The effect of test compounds on the rate of urea hydrolysis was
also estimated.
189
The 40-g air-dried samples were treated with 40 mg N as urea and with nonidentical
amounts of the test compounds, namely with 2 mg of thiourea (TU), hydroquinone
(HQ) or 2,4-dinitrophenol (DNP), 20 mg of H3B03, and 100 mg of CaCI 2. The control
received only urea. All samples were moistened and incubated for 21 days (at 38 C for
Q
the first 5 days, then at 21 QC).

The cumulative NH3 losses expressed as percentages of the added urea-N had the
following values:
27.5 (control), 27.4 (HQ), 26.8 (DNP), 26.3 (CaCh), 24.7 (TU), and 21.7 (H3B03).
During the 21-day incubation period, the added urea was hydrolyzed in the
following percent proportions:
98 (CaCI 2), 94 (control), 89 (HQ), 88 (DNP), 87 (TU), and 65 (H 3B0 3).
In other words, boric acid was the most effective compound in both inhibiting urea
hydrolysis and reducing the volatile NH 3 loss.
In another experiment, phosphogypsum was tested. The mixture of 2 or 4 g of
powdered phosphogypsum + 870 mg of prilled urea was incorporated into 40-g soil
samples, then moistened and incubated for 25 days (again at 38°C for the first 5 days,
then at 21 QC). The control was treated only with urea.
The cumulative NH3 loss from the control was 32.6% of the added urea-N, whereas
the loss was only 5.1 % at both phosphogypsum rates.
4.4. COMPARISON OF INORGANIC SULFUR COMPOUNDS WITH OTHER

INHIBITORS
Laboratory Experiments. McCarty et al. (1991) prepared reaction mixtures from 5-g
air-dried samples of two Iowa soils (a silty loam and a sandy clay loam), 2 ml of water
containing 10 mg of urea or 10 mf of urea and different amounts of the compounds
compared. The mixtures were incubated at 20 Q C and, after 3 and 10 days, analyzed for
the residual urea. The compounds and their amounts were the following: sodium
thiosulfate and tetrathionate 1,000, 2,500, and 5,000 J.lglg soil; Fe 2+ (as FeS04) and
Mn2+ (as MnCl 2) 100, 250, 500, 1,000, and 2,500 J.lglg soil. Phenylphosphorodiamidate
(PPDA) was the reference compound used in an amount of 10 J.lglg soil.
Thiosulfate was found to be less inhibitory on soil urease activity than was
tetrathionate. The inhibiting effect of Fe2+ and Mn + was inexistent or very weak. As
expected, PPDA was the strongest inhibitor. Thus, at the 2,500 J.lglg soil addition, the
percent inhibitions of urease activity in the two soils after 3 and 10 days of incubation,
respectively, were: 18 and 3%, and 0 and 0% for thiosulfate; 48 and 22%, and 0 and
21 % for tetrathionate; 8 and 11 %, and 0 and 0% for Fe2+; and 10 and 10%, and 0 and
4% for Mn2+. The corresponding values for PPDA used, as mentioned above, only in the
amount of 10 J.lglg soil, were 89 and 93%, and 22 and 32%.
Using samples of three Kansas soils treated with Na2S203.5H20, Na2S406.2H20,
FeS04.7H20 or MnS04.H20 at rates of 0,0.05,0.25,2.50 or 25 mM S/kg soil, Sullivan
and Havlin (1992b) registered the following maximum urease inhibition levels: 30% for
S20/-, 29% for s40l-, 14% for Fe 2+, and 8% for Mn2+.
190
4.5. COMPARISON OF ORGANIC MERCURY COMPOUNDS WITH OTHER

INHIBITORS
Laboratory and Pot Experiments. According to the analytical data obtained by

Kozlovskaya et al. (1972), p-chloromercuribenzoate. at a concentration of 2 J.l.M in the
reaction mixtures with different peat bog soils, inhibited 1.15-1.50 times more strongly
the urease activity than did CUS04, the concentration of which in the reaction mixtures
was 84 mM.
Shih and Souza (1978), who used p-hydroxymercuribenzoate (PHMB) as a soil
urease inhibitor (see page 44), applied the same procedure for inhibiting urease with
thiourea. The reaction mixtures consisted of 0.2 g of soil + 0.1 ml of 1 or 2 M thiourea
in phosphate buffer (50 mM, pH 7.0) + 0.1 ml of 14C_urea (0.01 /lCi activity) in the
same buffer. Although the concentrations of thiourea on molar basis were about 100-
500 times higher then those of PHMB, its inhibitory effect was weaker in comparison
with that of PHMB. Thus, the residual 14C02 evolution from soil samples treated with
the higher thiourea rate was 6.69, 4.94, and 6.15% of the 14C02 evolved from samples
treated with urea only and incubated for 2, 4, and 6 hours, respectively.
Reddy and Chhonkar (1990a, 1991) have compared the effect of phenylmercuric
acetate (PMA), hydroquinone (HQ), and alcoholic extract of neern kemel (NKE) on
urea hydrolysis and urease activity in soil and floodwater. Samples of an Indian sandy
clay loam soil (pH 7.6) were studied.
The experiments comprised four principal variants: no flooding and cropping of rice
or no cropping, and flooding and cropping or no cropping of rice. For cropping pots
filled with 3 kg soil, and for no cropping beakers with 1 kg soil were used. Within each
variant there were pots and beakers in which the soil was not amended or was amended
with farmyard manure at a rate of 50 glkg soil. Rates of other additions per kg soil were:
urea 100 mg, PMA and HQ each 2 mg, and NKE 10 mg (i. e. 10% on urea weight basis).
In the cropping experiments, four 15-day-old rice seedlings were transplanted in each
pot. In the nonflooded soil, the moisture content was maintained at 50% ofWHC, while
the flooded soil was kept in submergence with a 5-cm standing water. On day 12 after
transplanting, urea without or with an inhibitor was broadcast on the surface and well
mixed in the soil.
Soil and floodwater were sampled on days 3, 10, and 20 after urea application for
determination of the rate of urea hydrolysis by analyzing the unhydrolyzed urea, and on
days 3, 10,20,30,40, and 60 after urea application for determination of urease activity
by using the 5-hour incubation test (see page 6).
In the urea hydrolysis experiments, on day 3 unhydrolyzed urea was detectable in
both nonflooded and flooded soil, noncropped and cropped soil, and in floodwater.
More urea remained unhydrolyzed in flooded than in nonflooded soil, in cropped than
in noncropped soil, and in soil than in floodwater, and always less urea remained
unhydrolyzed when the soil was amended with farmyard manure.
The amount of the unhydrolyzed urea increased always in the order:
control (no inhibitor) < NKE < HQ < PMA, which means that PMA was the
strongest and NKE was the weakest inhibitor. Farmyard manuring led to weakening of
the effect of each inhibitor.
191
On day 10, urea was not detectable in the soil. Small amounts of urea were present
in floodwater, in which the inhibitory effect in the order PMA> HQ > NKE remained
evident. By day 20, urea disappeared from the floodwater, too.
In the urease activity experiments, the activity was measurable in soil and unfiltered
floodwater during the whole 60-day incubation period (through filtering urease was
removed with the suspended particles and colloids).
The inhibitors decreased urease activity in the order PMA > HQ > NKE, which
confirms the results of the urea hydrolysis experiments. Although this order of
inhibitors remained unchanged during prolongation of the incubation time, urease
activity increased with time in the inhibitor-treated soil. For example, urease activity
(expressed in Ilg urea-N/g soil/hour) in the nonflooded and noncropped soil was 0.993
(control), 0.896 (NKE), 0.653 (HQ), and 0.623 (PMA) on day 3, and 0.730 (control),
0.747 (NKE), 0.760 (HQ), and 0.763 (PMA) on day 60. The activity-increasing effect of
inhibitors was attributed to their microbial decomposition, during which the
microorganisms, using the inhibitors or their degradation products as carbon and energy
sources, produced new urease molecules.
4.6. COMPARISON OF UREA DERIVATIVES WITH OTHER INHIBITORS
Field Experiments. Malhi and Nyborg (1979) compared the effect of thiourea on urea
hydrolysis in an Alberta silty clay loam soil with that of calcium sulfide (CaS) and
phosphorus pentasulfide (P 2S5; P4 SIO ) (see page 35) as well as with that of thio-
acetamide (CHr CS-NH 2). Urea was mixed (and not copelleted) with each sulfur
compound in a ratio of 2:1. The control plots were treated only with urea and the
experimental plots with urea + sulfur compound mixtures. Both urea and mixtures were
banded at a depth of 5 cm. Rate of N application was 112 kg/ha. Analysis of the NH/
and N0 3- contents in the 0-15-cm soil layer has shown that 5 weeks after fertilization,
urea hydrolysis was complete in the control plots and partial in the experimental plots:
68% (P2SS), 71 % (CaS), 42% (thiourea), and 43% (thioacetamide). That is, thioacet-
amide was as effective as thiourea. After 8 weeks, hydrolysis of urea became complete
in all plots.
4.7. COMPARISON OF POLYHYDRIC PHENOLS AND QUINONES WITH

OTHER INHIBITORS'
Laboratory Experiments. According to a short report (Anonymous, 1973), several

compounds were tested as inhibitors of urea hydrolysis in soil. Their inhibitory effect
decreased in the order:
catechol > hydroquinone > p-benzoquinone > Na p-chloromercuribenzoate >
acetohydroxamic acid.
Matzel et al. (1978) studied the effect of p-benzoquinone (BQ) with the same soils
and methods as those also used for studying the effect of acetohydroxamic acid (AHA)
(see page 64). Rate of BQ was 3% relative to weight of urea. The effect of BQ, as
compared to that of AHA, was more marked and long-lasting. For example, in samples
The comparative studies in which p-benzoquinone or hydroquinone was used as a reference compound have
already been referred to on pages 59,104. 160-162,164. and 181.
192
of a black soil (loess) which had been treated with urea+AHA and incubated for 19
days, no residual urea was detectable, whereas in the urea+BQ-treated samples the
residual urea represented 12.1 % of the initial urea amount. Cumulative ammonia losses
by volatilization from samples of an acid sandy soil treated with urea, urea + 3% of
AHA, and urea + 3% ofBQ during 21 days of incubation were 19.1, 18.1, and l3.2%,
respectively (see also Matzel and Heber, 1979).
Ashworth et af. (1979) mentioned that hydroquinone (HQ) at a rate of 50 mglkg soil
was a little more effective in inhibiting urease activity of a silty clay loam than was
potassium ethyl xanthate at a rate of 100 mglkg soil.
Lichko and Kiselev (1985) compared the inhibitory effect of HQ and Cu 2+ on urease
activity in samples of a grey forcst soil, a chemozem, and an alkaline light chestnut soil.
Both inhibitors were applied at rates of 50. 250. and 500 Ilg/g soil. The reaction
mixtures were incubated at 30-37°C for 1-3 hours. It was found that in each soil and at
each rate of inhibitors HQ was more effective than was Cu2+. Thus, at the 250 Ilg/g soil
rate, HQ caused 89-96% inhibitions, whereas under the influence of Cu 2+ only 45-56%
inhibitions occurred.
In the experiments of Xue and Li (1987), dihydric phenols and quinones were
stronger inhibitors of soil urease activity than the plant materials (see Section 2.31.3.6).
Studying the alkaline alluvial soils in Pakistan, Hamid and Ahmad (1987) incubated
samples of a calcareous soil (pH 8) with urea, with or without addition of 5%
hydroquinone or phenol (on urea weight basis), and determined the ammonia evolved
during incubation. Cumulative NH3 losses during 112 days were reduced by l3% in
hydroquinone-treated and by 5% in phenol-treated samples as compared to the control
soil.
Among 28 substances tested at a rate of 20 ppm (on soil basis), quinhydrone,
hydroquinone, p-benzoquinone, and caffeine were the most effective inhibitors of soil
urease activity, causing -61, 59, 39, and 48% inhibitions, respectively (Li and Xue,
19(1).
Thorrnahlen and du Preez (1991) compared the effect of seven inhibitors, namely
hydroquinone (HQ), p-benzoquinone (BQ), phenylphosphorodiamidate (PPDA),
catechol (CT), phenylmercuric acetate (PMA), thiourea (TU), and ammonium
thiosulfate (ATS) on urease activity in four widely different soils from the central
irrigation areas in South Africa.
Air-dried soil samples (500 g) were treated with 25 rnl aqueous solution containing
25 mg of urea and 0 or 25 mg inhibitor. The samples moistened to field capacity were
incubated at 30°C. During incubation, the urea-N was measured at regular time intervals
until hydrolysis of urea was completed. These data were used to calculate the urease
activity and its inhibition by each inhibitor in each soil.
On average, the inhibitions caused by the seven inhibitors studied in the urease
activity of the four soils were the following:
HQ 94%; BQ 93; PPDA 85%; CT 75%; PMA 70%; TV 30%; and ATS 19%.
The minimum time necessary for complete hydrolysis of urea was registered in the
control (non-inhibited) sample of each soil, and the prolongation of this time was
maximal in the HQ-treated samples. The minimum and maximum times (in hours) for
the four soils are specified below: 42 (control) and 374 (HQ and PMA) in soil 1; 24
(control and ATS) and 156 (HQ) in soil 2; 60 (control) and 540 (HQ) in soil 3; and 8
(control) and 192 (HQ and BQ) in soil 4.
193
4.8. COMPARISON OF PHENYLPHOSPHORODIAMIDATE (PPDA) WITH

OTHER INHIBITORS·
Laboratory Experiments. Martens and Bremner (1982, 1984b) have described

experiments for comparing the urease-inhibiting effectiveness of PPDA and phosphoro-
diamidic acid (PDA) with that of 11 phosphoric triamide (PTA) compounds, namely
with compounds No. 1-9 (in 1982 and 1984) as well as with compound No. 10 (in 1982)
and compound No. 11 (in 1984) (see Table 44).
TABLE 44. PTA compounds compared with PPDA and PDA in experiments described by Martens
and Bremner (1982, 1984b)
No. Compound Structural formula
I Phosphoryl triamide H2N-P(O)(NH2)2
2 N-Phenylphosphoric triamide C.H,-HN-P(O)(NH2)2
3 N-(4-Nitrophenyl)phosphoric triamide 4-02N-C.H.-HN-P(O)(NH2h
4 N-(Diaminophosphinyl)benzamide C6H,-CO-NH-P(O)(NH2)2
5 4-Chloro-N-(diaminophosphinyl)benzamide 4-Cl-C.H.-CO-NH-P(OXNH212
6 4-Fluoro-N -( diaminophosphinyl)benzamide 4-F-C6H.-CO-NH-P(O)(NH2h
7 4-Cyano-N-(diaminophosphinyl)benzamide 4-NC-C6H.-CO-NH-P(OXNH2)2
8 N-(Diaminophospinyl)benzeneacetamide C.H,-CH 2-CO-NH -P(O)(NH2)2
9 N-(Diaminophosphinyl) 3-pyridinecarlJoxamide 3-C,H.N-CO-NH-P(O)(NH212
10 3-Trifluoromethyl-N-(diaminophosphinyl)benzamide 3-F,C-C6 H.-CO-NH-P(O)(NH2)2
II N-(3-Trifluoromethylphenyl)phosphoric triamide 3-F,C-C.H.-HN-P(O)(NH2)2
Experiments performed with the aim to compare PPDA and PDA with phenyl-
mercuric acetate, catechol, hydroquinone, p-benzoquinone, and 2,5-dimethyl-p-benzo-
quinone were also described by Martens and Bremner (I 984b).
The seven Iowa soils selected for the experiments were very varied in terms of their
pH (5.0-8.0), texture (5-56% sand, 13-32% clay), organic C content (0.30-4.23%),
CaC03 equivalent (0-20.8%), urease activity (14.2-80.2 /lg of hydrolyzed urea/g
soillhour at 37°C), and their other properties. The reaction mixtures (5 g of air-dried soil
+ 2 ml of solution containing 10 mg of urea without or with 50 /lg of inhibitor) were
incubated at 20°C and analyzed for residual urea after 3, 7, and 14 days of incubation.
The analytical data indicated that PPDA was the most effective urease inhibitor in
each soil, whereas PDA was a weak inhibitor. Of the PTA compounds, 4-cyano-N-
(diaminophosphinyl)benzamide was the least effective inhibitor. It is noteworthy that
phenylation of PDA (weak inhibitor) leads to the formation of the strongest inhibitor
(PPDA). Conversely, the effect of phosphoryl triarnide was a little greater than that of
its phenyl derivative, N-phenylphosphoric triamide.
The inhibitory effectiveness of all compounds decreased with prolongation of the
incubation time .
• The comparative studies in which PPDA was used as a reference compound have already been referred to on
pages 101, 103, 144, and 189.
194
In another experiment performed with samples of five soils, the effectiveness of

three concentrations of PPDA and hydroquinone (HQ) (5, 25, and 50 11g15 g soil) was
compared through 10-day incubations at 20°e. Each concentration of PPDA was more
effective than the corresponding concentration of HQ in each soil. Mean values of the
percent inhibitions brought about by the three inhibitor concentrations in the five soils
studied were as follows: 47, 61, and 75%, respectively (PPDA) and 9, 22, and 28%,
respectively (HQ).
Results of the investigations performed by Liao and Raines (1982, 1985) make
possible a comparison of the inhibitory effectiveness of PPDA with that of phosphoryl
triarnide (PTA), thiophosphoryl triarnide (TPTA) , potassium phosphoroamidate, and
sodium thiophosphorodiamidate.
The reaction mixtures, prepared from 5 g of air-dried soil, 2.5 ml of a solution
containing 5 mg of urea-N and 250 I1g of test compound or only 5 mg of urea-N, were
incubated at 30°C for 24 hours. Then, the NH4 + released from urea was determined and
percent inhibition calculated.
One can see from Table 45 that PPDA proved to be the most potent inhibitor of
urease activity in each of the four soils studied. Inhibitory effect of PTA and TPTA was
TABLE 45. EtTect ofehosehoroamides on soil urease activitt

Inhibition (%2
Silty clay Fine sandy Fine sandy Silty
Compound Struc tural formula
loam loam loam loam
pH 7.3 e H7 .8 e H7 .7 e H7 .O
Phosphoryllriamide H2N-P(O)(NH,h 75 73 89 69
Thiophosphoryl triamide H,N-P(S)(NH,h 68 63 67 83
K phosphoroamidate KO-P(O)(OH)NH, 19 7 14 10
Na thiophosphorodiamidate NaO-P(S)(NH2h 53 44 44 46
Phenylphosphorodiamidate C6 H,O-P(O)(NH2h 94 96 98 95
"FromLiao and Raines (1985), by permission ofKluwer Academic Publishers,
also marked; PTA was more effective than TPTA in three soils. In each soil, potassium
phosphoroamidate was the weakest inhibitor.
The effect of different concentrations of PTA, TPTA, and PPDA (2.5,5,10,25,50,
and 100 ppm relative to soil weight) on urease activity in four soils was also studied.
The inhibition increased with increasing inhibitor concentration up to 25 ppm and
tended to level off at higher concentrations. The maximum inhibition brought about by
PTA and TPTA at 100 ppm was 93% (in fine sandy loam, pH 7.7) and 86% (in silty
loam), respectively, whereas PPDA produced nearly complete inhibition at
concentrations of 10-50 ppm.
Persistence of the inhibitory effect was followed in the silty clay loam. The reaction
mixtures were incubated for 3 and 7 days, then the residual urea was assessed. After 3
days, PTA, TPTA, and PPDA caused similar inhibitions (of about 64%). Inhibition by
PTA and PPDA persisted at a level of about 15-20% even after 7 days of incubation.
The observations made by Liao and Raines (1985) that PTA was a more effective
and persistent inhibitor than was TPTA should be emphasized since in other
195
experiments (Anonymous, 1985a; Radel et al.. 1987) TPTA proved to be superior to

PTA and even to PPDA in inhibition of urea hydrolysis in soil (see page 144).
Under conditions identical to those described on page 93, Gorelik et al. (1983) have
also studied PPDA added to urea in a proportion of 1% relative to weight of urea.
During the first 7 days of incubation, PPDA acted more inhibitorily on urea hydrolysis
than did hydroquinone (HQ) also used at a rate of 1% relative to weight of urea, but
after 14 days urea hydrolysis became complete in both HQ- and PPDA-treated samples
of each soil. Similarly, PPDA was more effective than HQ in diminution of ammonia
volatilization from urea during the first 7 days, but the cumulative NH310sses in 14 days
were practically the same in untreated and inhibitor-treated samples of each soil.
Rodgers (1984b), who has established that of five aminocresols tested 4-amino-o-
cresol had the most evident inhibitory effect on urease activity in the three English soils
studied (silty clay loam, sandy clay loam, and loamy sand) (see page 80), compared the
effect of this aminocresol with that of PPDA. The reaction mixtures had the following
composition: 8 g of air-dried soil + 1.2 ml of 0.4% aqueous urea solution (=280 ~g N/g
soil) + 0.8 ml of 60% methanol or 0.8 ml of solution of 4-amino-o-cresol or PPDA in
60% methanol (50 ~g of test compound/g soil). Periodic analysis of the residual urea
and ~ + during incubation (90 hours at 30°C) showed that in the silty clay loam the
inhibitory effect of the two compounds was practically identical, but in the other two
soils PPDA was more effective. In the sandy clay loam, the inhibitory effect of both
compounds decreased sharply during incubation and disappeared at the end of
incubation period. During incubation of the silty clay loam, only a little decrease
occurred in the inhibitory capacity of both compounds. In the loamy sand, the inhibitory
effect of PPDA remained essentially unchanged during incubation, in contrast with that
of 4-arnino-o-cresol which markedly decreased with incubation time.
The comparative study devoted by Hendrickson and O'Connor (1987) to PPDA and
phenol has already been referred to on page 136.
In the experiments of Germann-Bauer (1987), 100-g samples of a loess brown earth
(PH 6.5) were amended with urea (120 mg N), 0.1 or 0.4 mg of PPDA or 2 mg of
guanylthiourea (GTU) and moistened to 60% of WHC. No inhibitor was added to the
control. All mixtures were incubated at 8°C and analyzed for residual urea at days 1, 3,
6, and 8 of incubation.
No unhydrolyzed urea was found at day 6 in the control and at day 8 in the urea +
GTU treatment, whereas at day 8 urea was still detectable in amounts of 0.6 and 1.9 mg
in the treatments with urea + 0.1 or 0.4 mg PPDA, respectively.
But there was no significant difference between the inhibitory effects of PPDA and
GTU on volatilization of ammonia from 500-g samples of a sandy brown earth (PH 5.9)
amended with urea (50 or 100 mg N) and PPDA (0.25 or 0.5 mg) or GTU (5 or 10 mg)
and incubated at 25°C for 6.6 and 13 days.
Studying a light soil (PH 7) and a heavy soil (PH 5) and PPDA 1%, dazomet 2%,
and thione 2% (on urea weight basis), Winiarski (1990) has established that the
inhibitory effect of these compounds on hydrolysis of urea in samples of both soils
presented the order: PPDA > dazomet> thione.
In the experiments of Luo et al. (1994), the effects of nBTPTA, PPDA, N-( diamino-
phosphinyl)benzarnide (DAPBA), and acetohydroxamic acid (AHA) to inhibit urease
activity in two Australian rice soils (vertisol, pH 7.6 and alfisol, pH 6.7) were
compared. The vertisol is more urease-active than the alfisol. Air-dried soil samples
196
(15 g) were flooded with 30 ml of water, then amended with urea (14 mg) without or
with inhibitor (1, 2.5, 5, and 10% relative to weight of urea) and incubated at 25°C for 8
days, during which, at 2-day intervals, the residual urea was determined.
At 1% inhibitor concentration, rate of urea hydrolysis was 99% (control), 96%
(AHA), 91 % (DAPBA), 89% (nBTPTA), and 30% (PPDA) in the more urease-active
vertisol after 2 days of incubation, and 100% (control), 99% (AHA), -82% (DAPBA)
and -40% (PPDA, nBTPTA) in the less urease-active alfisol after 8 days of incubation.
Increasing concentration of AHA and DAPBA from I to 10% did little to reduce the
rate of hydrolysis of urea in either the vertisol or the alfisol. Essentially, all the urea was
hydrolyzed by day 4 in the vertisol and day 8 in the alfisol, irrespective of the
concentration of these compounds.
In contrast, increasing the concentration of PPDA and nBTPTA from I to 2.5, 5 and
10% led, at least at the 10% concentration, to a decrease in the rate of urea hydrolysis.
Thus, the rate of urea hydrolysis in the vertisol after 8 days of incubation was 100, -90,
70, and 58%, respectively, in the PPDA treatments, and 100, 100, 100, and 75%,
respectively, in the nBTPTA treatments. The corresponding values registered in the
alfisol after 8 days of incubation were: 40, 25, 17, and 9%, respectively, in the PPDA
treatments, and 28, 25, 21, and 20%, respectively, in the nBTPTA treatments.
All results indicated that in the two flooded rice soils studied PPDA was the most
effective urease inhibitor.
Wang and Douglas (1996) used surface (0-15 cm) samples of two sandy soils,
namely a solonized brown soil (PH 7.7) and a podzol (PH 5.5), both from Victoria,
Australia, for comparing the inhibitory effects of PPDA, cyc1ohexyl-PTA (CHPTA),
and nBTPTA on urea hydrolysis. The reaction mixtures, prepared from 6 g air-dried soil
and 3 ml of aqueous solution containing 0, 5, 10 or 25 J.lg inhibitor and 1 mg of urealg
soil, were incubated at 37°C for 4 hours, then analyzed for NH4 +.
The analytical data showed that each compound was more inhibitory in the brown
soil than in the podzol.
At the 5,10, and 25 J.lglg soil rates ofPPDA. CHPTA, and nBTPTA, the following
inhibitions were registered in the brown soil:
100, 100, and 100% (PPDA); 87, 100, and 100% (CHPTA); and 68, 73, and 83%
(nBTPTA).
The corresponding values for the podzol were:
79, 79, and 71% (PPDA); 78, 79, and 71% (CHPTA); and 65, 66, and 71%
(nBTPTA).
Thus, in these soils, PPDA and CHPTA were stronger inhibitors of urea hydrolysis
than was nBTPTA.
Field Experiments. Beyrouty et al. (l988a,b) conducted two field trials on a silt loam
soil (pH 5.7) (located at the Purdue University Agronomy Fann, West Lafayette,
Indiana) with the aim of studying the effect of six phosphoroamides (N,N-dimethyl-,
N,N-diethyl-, N-cyc1ohexyl- and N-benzyl-N-methyl-PTA, trichloroethyl-PDA, and
PPDA)" on urea hydrolysis and ammonia volatilization from urea-No The trials were
carried out on conventional till (CT) and no-till (NT) microplots. In the previous year,
"It should be emphasized that in these field trials N-(n-butyl)thiophosphoric triamide (nBTPTA) was not
evaluated.
197
the experimental field was cropped to maize; the maize residue covered about 60% of
the surface of the NT microplots. All microplots remained unsown and in all of them
weed control was accomplished with application of herbicides followed by hand
cultivation as needed. Urea prills (200 kg N/ha) with or without an inhibitor (4 kg/ha)
were uniformly broadcast over the soil of microplots. Each inhibitor used was coated
onto urea prills with paraffin oil. The control microplots received no fertilizer.
Trial 1 was initiated on 7 June 1983, and trial 2 on 5 July 1983. At fertilizer
application, the soil surface was moist in trial 1, and air-dry in trial 2. After fertilization,
analyses were made at 2-5-day intervals for determining residual urea and volatilized
ammonia during 24- and 20-day periods, respectively, Air and soil (5-cm depth)
temperatures had average maxima of 29 and 35°C, respectively, in trial 1, and 32 and
40°C, respectively, in trial 2. The urea prills had completely dissolved within 12 hours
(trial 1) or within 6 days (trial 2) in both CT and NT microplots. In the absence of
inhibitors, the rate of urea hydrolysis was more than twice as great in trial 1 than in trial
2, due to slower dissolution of urea prills in trial 2.
In trial 1, in the CT microplots, the most effective inhibitors were PPDA, trichloro-
ethyl-PDA and N-benzyl-N-methyl-PTA, decreasing the rate of urea hydrolysis by 68,
66, and 60%, respectively. The other compounds had limited effects on the urea
hydrolysis rate. The inhibitory effect always decreased with time and was evident for 19
days in the case of PPDA and trichloroethyl-PDA, but no inhibition was present at day
9 in the case of the other compounds. In the NT microplots, only PPDA had significant
(>70%) inhibitory effect on urea hydrolysis for 4 days, but its effectiveness declined
rapidly by day 9. However, some effect of PPDA on urea hydrolysis was observed for
as long as 19 days. Inhibition disappeared in each treatment at day 24 after fertilization.
In trial 2, in the CT microplots, none of the compounds tested significantly affected
the rate of urea hydrolysis. In the NT microplots, PPDA was the single effective
inhibitor (reduction of urea hydrolysis rate was higher than 63%).
In both trials, the weakest urease inhibitors were N,N-dimethyl- and N,N-diethyl-
PTA.
Cumulative NH3 losses in 12 days (trial 1) or in 20 days (trial 2) were smaller in the
CT microplots than in the NT ones, in both absence and presence of inhibitors. Thus,
cumulative NH3 losses from the urea-N applied without inhibitors in CT and NT
microplots were 30 and 31 %, respectively, in trial 1, and 7 and 35%, respectively, in
trial 2. In trial 1, NH3 volatilization was significantly reduced only by PPDA in both CT
microplots (degree of inhibition: 90%) and NT microplots (degree of inhibition: 61 %).
In trial 2, PPDA was again the only inhibitor which significantly reduced NH3
volatilization, but only in the NT microplots (degree of inhibition: 46%) (see also
Nelson et al., 1986).
Referring to the results obtained with two acid soils and an alkaline soil under
aerobic and water-logged conditions, Van Cleemput and Wang (1991) have drawn the
conclusion that the inhibitory effect of PPDA on urea hydrolysis and NH4 +
accumulation was more marked than that of nBTPTA in the acid soils under both
aerobic and water-logged conditions as well as in the alkaline soil under water-logged
conditions, while in the alkaline soil under aerobic conditions nBTPTA was a stronger
inhibitor than PPDA (see also Wang and Van Cleemput, 1992).
198
4.9 COMPARISON OF PHOSPHORIC TRIAMIDE (PTA) AND THIOPHOSPHORIC

TRIAMIDE (TPTA) COMPOUNDS WITH OTHER INHIBITORS
4.9.1. Comparative Studies on the Efff!Ct of PTA and TPTA Compounds and Other
Inhibitors on Soil Urease Activity. Urea Hydrolysis. and Ammonia Volatilization
4.9.1.1. Comparison ofnBTPTA with Ammonium Thiosu(f{lle (ATS)

Laboratory Experiments. In a short report, Bremner et af. (1990) emphasized that ATS
significantly retarded urea hydrolysis only when applied at rates as high as 2,500 and
5,000 ~glg soil, whereas nBTPTA caused substantial retardation of urea hydrolysis
when applied at rates as low as I ~g/g soil.
These investigations were described in detail by McCarty et af. (1990). Four soils
possessing markedly different physical and chemical properties were studied. Reaction
mixtures were prepared from 5-g air-dried soil + 2 ml of water containing 10 mg of urea
with or without ATS (500, LOOO, 2.500 or 5,000 ~g/g soil) or nBTPTA (1 or 10 ~lg/g
soil), then incubated at 20°C for 3 and 10 days, followed by analysis of residual urea.
The results emphasized by Bremner et al. (1990) can be reiterated. For
exemplification, we specify only the percent inhibitions of urea hydrolysis registered in
the four soils at the highest ATS rate vs. the lower nBTPTA rate (5,000 VS. 1 ~g/g soil)
after 3 and 10 days of incubation: 63 and 30 vs. 62 and 30%; 7 and 0 vs. 83 and 45%; 29
and 0 vs. 33 and 12%; and 20 and 0 VS. 60 and 54%.
Field Experiments. Grant et al. (1996a,b) conducted two microplot studies in 1995,
under no-till conditions on a chernozemic soil (fine sandy loam, pH 7.3) located on
Canadian prairie. The first study was initiated on May 20, 3 days after seeding of wheat,
but before emergence. The second study began on August 14, after the wheat had
headed; wheat was mowed at a height of 4 cm and the fresh residue removed. The
treatments included: urea-ammonium nitrate (VAN) solution (100 kg Nlha) , VAN +
0.25% nBTPTA (relative to fertilizer N); VAN + 10% ATS (again relative to fertilizer
N). The fertilizer was placed in a 2-cm circle on the soil surface in the center of the
microplot. After fertilization the volatile ammonia was measured for 7 days.
The cumulative NH3 losses were higher in the second study than in the first,
presumably due to higher soil and air temperatures and lower initial soil moisture levels
in August as compared to May.
nBTPTA significantly (p=0.05) reduced total NH3 losses in both studies, whereas
ATS was ineffective in the first study and less effective than nBTPTA in the second
study.
4.9.1.2. Comparison ofnBTPTA with Pyrite. Phosphogypsum. and KCl

Field Experiments. For this comparison, two experiments were carried out on a
calcareous clay soil (PH 8.44) at the Regional Soil Research Institute in Konya, Turkey.
The test plant was winter wheat under dry-land conditions in the first experiment
(Gezgin and Bayrakli, 1995) and sugarbeet under irrigated conditions in the second
experiment (Bayrakli and Gezgin, 1996).
In both experiments plot size was 2.5 by 4 m. The rates of additions were: fertilizer
200 kg Nlha; nBTPTA 0.25% relative to fertilizer N (nBTPTA1) and 0.5%
(nBTPTA2); pyrite (pH 3.5) 1 and 2 tlha (PRI and PR2); phosphogypsum (PH 2.5) 1
199
and 2 t/ha (PG 1 and PG2). KCl was applied only in the sugarbeet experiment, at rates of
540 and 1,080 kg/ha (KCn and KCI2). The N fertilizers were urea, ammonium sulfate,
and ammonium nitrate in the wheat experiment, while only urea was applied in the
sugarbeet experiment. The N fertilizer mixed with the test substance was placed on the
surface of plots in May 1993 (wheat experiment) or in July 1993 (sugarbeet
experiment). Only N fertilizer was added to the control plots.
Volatilization of ammonia was assessed up to 57 days in the wheat experiment and
up to 78 days in the sugarbeet experiment.
Total NH3 loss from ammonium sulfate and ammonium nitrate was significantly
(p<0.05) increased by the substances tested, including nBTPTA.
Loss from urea in the wheat experiment was significantly increased by PG2 from
10.6% of the added N (control) to 12.0%, whereas the loss in the other treatments was
significantly lower than in the control, with values of 9.2% (PGl), 8.6% (PRl), 8.2%
(PR2), 5.0% (nBTPTAI), and 3.9% (nBTPTA2).
In the sugarbeet experiment total NH3 loss from the added urea-N (12.6% in the
control) was significantly increased by PRI (17.2%), PGl (17.7%), and KCn (23.6%),
not affected significantly by nBTPTAI (12.6%) and PG2 (11.5%), and significantly
reduced by PR2 (11.1 %), KCl2 (10.5%), and nBTPTA2 (7.0%).
All results show that nBTPTA at the 0.5% rate was most effective in reducing the
volatile NH3 losses from urea-treated plots.
4.9.1.3. Comparison ofnBTPTA and/or TPTA. PTAs and Alkyl-PDAs with PPDA
Laboratory Experiments. Swerdloff et al. (1984, 1985d) and Van Der Puy et al. (1985b)
treated samples of a sandy loam soil with urea with or without a urease inhibitor and
incubated them at 35°C for 12 days. The rates of additions per 20 g soil were: 42.8 mg
of urea and 0.2 mg of inhibitor. The unhydrolyzed urea was determined at 2-day
intervals. Urea hydrolysis was complete in 2 days in the urea-only treatment, in 6 days
when the inhibitor was PPDA, whereas about 35 and 65% of the added urea remained
unhydrolyzed after 12 days in samples treated with N-methyl-N-(4-nitrophenyl)-PTA
and N-(2-chloroethyl)-PTA, respectively.
In a similar experiment described by Kolc et aT. (1984, 1985d), 20-g samples of a
silt loam soil were amended with 42.8 mg of urea with or without 0.2 mg of inhibitor,
then incubated at 25 or 35°C for 22 days. The residual urea content was determined at
1-2-day intervals. No urea was detected after 4-day incubation at 25°C and after 2 days
at 35°C in the soil to which only urea was added. Urea disappeared from the PPDA-
treated soil in -9 days at 25°C and -6 days at 35°C. In the N,N-diethyl-PTA-treated soil
-50 and nearly 20% of the added urea remained unhydrolyzed at 25 and 35°C,
respectively, although the incubation time was 22 days.
In another experiment performed by Kolc et al. (1984, 1985d), nBTPTA, five PTAs,
namely N-(n-butyl)-PTA, N-(sec-butyl)-PTA, N-ethyl-PTA, N-(n-octyl)-PTA, and N-
cyclohexyl-PTA, as well as PPDA at a rate of 0.2 mg were preincubated with 20-g
samples of a silt loam soil, at 25 and 35°C or in the open, for 11 days. The next
operations were: addition of urea (42.8 mg), incubation (at 25°C for 3 days), and
analysis of residual urea.
The following results were registered: nBTPTA was the most effective urease
inhibitor; the inhibitory effectiveness of each compound (except nBTPTA) decreased
during the preincubation; the decrease was more marked at 35 than at 25°C and less
200
marked in samples kept in the open; PPDA as a urease inhibitor was inferior not only to
nBTPTA, but also to the five PTAs.
Beyrouty et al. (1988a) compared the inhibitory effect of two PTAs (N-cyclohexyl-
PTA and N-benzyl N-methyl-PTA), nBTPTA, and PPDA on urea hydrolysis in a silty
clay loam soil from Indiana. The surface (0-15-cm) soil layer was sampled in two areas.
Soil pH was 5.6 in one area and 7.4 in the other area which was under a long-term
liming experiment.
The soil samples (20 g) with contrasting initial pH values of 5.6 and 7.4 were treated
with 4.8 ml of a solution containing 20 mg of urea-N with or without 0.4 mg of
inhibitor, then the reaction mixtures were incubated at 25 D e for 0, 2,4, 7, and 14 days,
after which they were analyzed for residual urea. Table 46 shows that at pH 5.6, PPDA
and nBTPTA were the most effective inhibitors of urea hydrolysis. At pH 7.4, PPDA
TABLE 46. Effect of initial soil pH on inhibition of urea hydrolysis by several

EhosEhoroamides"
Inhibition (%2
Incubation time Initial
N-cyclo- N-benzyl-N-
(days) soil pH nBTPTA PPDA
hexz:I-PTA methz:I-PTA
2 5.6 55 b* 24 a* 76 c 96 d*
7.4 89b 57 a 83 b 82 b
4 5.6 46b* 15 a* 81 d* 72 c*
7.4 85 c 55 b 93 d 47 a
7 5.6 21 b* o a* 60 d* 41 c*
7.4 77 c 33 b 86 d 5a
14 5.6 o a* o a* 2 b* Oa
7.4 8c 2b 14 d Oa
"From Beyrouty et al. (l988a), by permission of Williams and Wilkins Publishers.
Values within rows followed by the same letter are not significantly different (p=O.05).
Paired values for different pH values within a compound and an incubation time are
significantly different (p=O.05) iffollowed by an asterisk.
had practically no inhibitory effect after 4 days of incubation, whereas N-cyclohexyl-

PTA and nBTPTA were effective inhibitors for 7 days. Taking into account both pH
values, one can state that nBTPTA was the most effective inhibitor. The least effective
inhibitor at both pHs was N-benzyl-N-methyl-PTA. The weak effectiveness ofPPDA at
pH 7.4 is attributed to its more rapid hydrolysis than at pH 5.6, which is in accord with
the results described on page 137, showing that PPDA effectively inhibited
volatilization ofNH3 from urea at pH 5.6, less effectively at pH 6.5, and not at all at pH
7.2.
Bremner et al. (1991) compared nBTPTA, TPTA (thiophosphoryl triamide), and
PPDA. Surface (O-IS-cm) samples of four Iowa soils, markedly different in pH, texture,
and organic matter content were used. Field-moist samples (equivalent to 5 g of oven-
dry soil) were preincubated at 15 and 300 e for 0, 3, 7, 14 or 28 days after treatment with
I ml water containing 0 or 25 Jlg of test compound. The ability of the pre incubated soil
samples to hydrolyze urea was then assayed by treating them with 1 ml water
containing 10 mg of urea and incubating them at 20D e for 3 days, after which the
residual urea was determined.
201
Persistence of the urease-inhibiting effect of each test compound decreased with an

increase in preincubation temperature from 15 to 30°C, and whereas the effect ofPPDA
decreased with an increase in preincubation time, the effects of nBTPTA and TPTA
sometimes increased before decreasing with increased preincubation time.
In each soil, the inhibitory effect of nBTPTA was higher at zero time of
preincubation and considerably more persistent than that of TPTA and PPDA. This was
significant even in those soil samples which, after having been treated with nBTPTA,
were preincubated at 15 or 30°C for 28 days.
At zero time of preincubation, PPDA was a little more inhibitory on urea hydrolysis
than was TPTA in three of the four soils studied, although a reverse trend was evident in
each soil during the preincubation at both temperatures.
Field Experiments. The maize experiments conducted by Bundy and Oberle (1988)
were mentioned on page 28. Under similar conditions, nBTPTA, N,N-diethyl-PTA, and
PPDA added to urea prills and nBTPTA and PPDA added to urea-ammonium nitrate
(UAN, 28-0-0) solution were also studied. The urease inhibitors were applied at a rate
of 2% on fertilizer N basis (56 and 112 kg N/ha). Urea prills and UAN without
inhibitors were used for comparison. In these experiments also, field measurements of
ammonia volatilization were made at the higher fertilizer N rate only and lasted 8-10
days.
nBTPTA, added to urea and UAN and applied only at Arlington in 1984,
significantly reduced NH3 volatilization from urea (18%) and UAN (5%) to 1 and 2%,
respectively. Urea and urea-PPDA, which were used at both locations in both years,
gave rise to the following N losses as NH3: 8 and 1% (Arlington, 1983), 18 and 3%
(Arlington, 1984),24 and 16% (Lancaster, 1983), and 19 and 14% (Lancaster, 1984),
respectively. It is evident that PPDA was more effective at Arlington than at Lancaster.
UAN-PPDA, used only in 1984, was effective at both locations; the NH3 losses were
reduced from 5 to 1% (at Arlington) and from 16 to 7% (at Lancaster), respectively.
N,N-Diethyl-PTA, added to urea and applied at both locati(ms in 1983, was ineffective
at Arlington (NH3 loss in both presence and absence of N,N-diethyl-PTA was 8%) and
effective at Lancaster (NH3 loss was reduced from 24 to 8%).
Fertilization experiments on a silt loam under an established orchardgrass (Dactylis
glomera/a) pasture at Lancaster were also performed in 1983 and 1984. Urea prills
(67.2 kg N/ha) with or without 2% (on urea-N basis) nBTPTA (in 1984), PPDA (in both
years), N,N-diethyl-PTA (in 1983), and trichloroethyl-PDA (in 1983) were surface-
applied in mid-June following the first harvest of the orchardgrass forage. Field
measurements ofNH 3 volatilization revealed significant reductions in NH3 losses due to
the urease inhibitors. Thus, in 1983, the NH3 losses were of 19% from urea, 11 % from
urea-PPDA, 8% from urea-N,N-diethyl-PTA, and 10% from urea-trichloroethyl-PDA.
In 1984, 9% of the N was lost as NH3 from urea, 1% from urea-nBTPTA, and 6% from
urea-PPDA, which means that nBTPTA was more effective than PPDA in controlling
NH 3 10sses.
4.9.1.4. Comparison ofnBTPTA. PTAs. and Alkyl-PDAs with PPDA and Hydroquinone
(HQ)
Laboratory Experiments. Chai and Bremner (1985, 1986) and Bremner (1986)
presented experimental data on the comparison of six compounds, namely nBTPTA,
202
cyclohexyl-PTA (CHPTA), N,N-dimethyl-PTA, N,N-diethyl-PTA, N-benzyl-N-methyl-

PTA, and trichloroethyl-PDA with PPDA and HQ for evaluation of their effect on urea
hydrolysis, ammonia volatilization, and nitrite accumulation in urea-treated samples of
different Iowa soils. Of the eight compounds tested, the most effective inhibitors were
nBTPTA> CHPTA > PPDA, whereas HQ was the least effective inhibitor.
Detailed description of these investigations is the subject of the papers by Chai and
Bremner (1987) and Bremner and Chai (1989).
Chai and Bremner (1987) used six Iowa soils. The reaction mixtures were composed
of 5 g of air -dried soil + 2 ml of a solution containing 10 mg of urea or 10 mg of urea
and 50 /1g of test compound. Incubation took place at 10. 20, 30. and 40°C and lasted 3,
7, and 14 days, then the unhydrolyzed urea was determined and percent inhibition
calculated. The results point out that nBTPTA and CHPTA were more effective than
PPDA at 20, 30, and 40°C, but not at 10°C. HQ was the weakest inhibitor at each
temperature. The effectiveness of all test compounds for inhibiting urea hydrolysis
decreased with an increase in temperature, and increased with an increase in
concentration from 1 to 5 and 10 /1g inhibitor/g soil.
Bremner and Chai (1989) used five Iowa soils. The effect of four inhibitors
(nBTPTA, CHPTA, PPDA, and HQ) on urea hydrolysis, ammonia volatilization, and
nitrite accumulation in urea-treated soil samples was studied. The reaction mixtures
contained 109 of air-dried soil, 1 ml of a solution containing 10 mg of urea-N, 3 ml of
water or 3 ml of an aqueous solution containing 0.1 mg of inhibitor and were incubated
at 30°C. The NH3 evolved during 2, 7, and 14 days of incubation was determined and
the incubated soils were analyzed for urea, exchangeable NH/, N0 2-, and N0 3-. The
results showed again that urea hydrolysis as well as NH3 volatilization and nitrite
accumulation in urea-treated soils were reduced by the four inhibitors in the order:
nBTPTA> CHPTA» PPDA > HQ.
The inhibitory effect was more marked with light- than with heavier-textured soils.
On average, volatile NH3 loss and nitrite accumulation were decreased from 52 to 5%
and from 11 to 1%, respectively, by the most effective inhibitor, nBTPTA.
Wang et al. (1991a) studied the effect of nBTPTA, PPDA, and HQ on urea
hydrolysis and ammonia volatilization in aerobically and anaerobically incubated
samples of a Belgian alkaline loam soil (PH 8.2). In studying urea hydrolysis, 10-g soil
samples, to which 2 ml of solution containing 2 mg ofurea-N + 1% inhibitor (on urea
weight basis) were added, and incubated at 25°C, at two-thirds of field capacity or
under water-logged conditions (H20:soil ratio = 2:1). Periodically the mixtures were
analyzed for urea, ~ +, N02-, and N0 3-. Figure 66 shows that the best inhibitor of urea
hydrolysis was nBTPTA under aerobic conditions and PPDA under anaerobic
conditions. There were no large differences between PPDA and HQ under aerobic
conditions and between nBTPTA and HQ under anaerobic conditions concerning their
effect on urea hydrolysis. Accumulation of NJ4 + in the mixtures was retarded by the
inhibitors in the same order as was urea hydrolysis. Lower N0 2- but higher N0 3 -
accumulations were observed when nBTPTA and PPDA were added to the soil under
203
100
A -e- Urea
.. -- ....
.•. HQ
e:. '11
-Ir PPDA
.nBTPTA
~- ,
.,
~
::>
30
0
0 2 3 7
Incubation lime (davs)
100
11
~Urea
. • HQ
~ -tr PPDA
·il 10 "nBTPTA
~'
..
.E' ~
1l
:5 ""
20 '-.
"'.
.....•. ~.::-
Q
0
Incubalion lime (days)
Figure 66. Hydrolysis of urea as a percentage of the urea applied, under aerobic (A) and
anaerobic (B) conditions./From Wang et al. (199\a), by permission of Springer-Verlag.I
aerobic conditions. This was mainly due to the effect of these inhibitors on retarding
urea hydrolysis, resulting in a lower accumulation of NH4 + which provided a more
favorable environment for oxidation of N0 2- to N0 3- by Nitrobacter. In the control and
HQ-treated soil samples, more N0 2- accumulated, probably because the higher NH/
concentration hindered oxidation of N0 2- to N0 3-. Little N0 2- was detected under
anaerobic conditions due to lack of O2 •
Ammonia volatilization was studied with mixtures prepared from 600 g of soil +
120 mg of urea-N with or without 1% inhibitor and water to two-thirds of field capacity
or from 350 g of soil + 70 mg of urea-N with or without 1% inhibitor and sufficient
water to form a 2-cm layer on the soil surface. During incubation (at 25°C), the NH3
evolved was assessed daily for 16 days. Less NH3 volatilized under aerobic than
anaerobic conditions. In concordance with inhibition of urea hydrolysis, cumulative
NH3 loss was lowest in the nBTPTA-treated soil (3% vs. 20% in the control soil) under
aerobic conditions, and in the PPDA-treated soil (15% vs. 40% in the control soil) under
anaerobic conditions.
In another experiment, Wang et al. (1996) compared the effects of three urease
inhibitors on movement of urea and its transformation products at two soil moisture
204
levels (10 and 20%, on a soil dry weight basis). The same inhibitors and the same soil
were used as in the investigations described in the preceding paragraphs, i.e. nBTPTA,
PPDA, and HQ and a Belgian alkaline soil, respectively.
The soil sampled from the 0-20-cm depth was preincubated at 25°C at 10 and 20%
moisture contents for 5 days to restore microbial activity. Afterwasds, the soil was
packed into plastic cylinders 10-cm high and 9.5-cm in diameter. Urea (300 mg N/kg
soil) alone or together with an inhibitor (1 % relative to weight of urea) was applied 3-4
cm below the surface of soil columns which were then incubated at 25°C. After 2, 4,7,
12, and 17 days, the soil columns were sliced into 10 sections, each 1 cm thick. The
soils were analyzed to determine moisture, urea, NH4 +, and (N0 3·+ N0 2-) contents.
The results clearly showed that the effect of each inhibitor in retarding urea
hydrolysis was stronger at 10% than at 20% soil moisture level. nBTPTA, in
comparison to PPDA and HQ, was the strongest inhibitor at both moisture levels,
whereas PPDA, compared to HQ, was a weaker inhibitor at 10% moisture and stronger
at 20% moisture.
By inhibiting urea hydrolysis, the inhibitors affected movement of urea, formation
and movement ofNH/ and (N03- and NO z-) in the soil columns during incubation. It
was demonstrated that distribution of urea and its transformation products after 7 days
of incubation at 20% moisture was comparable with that observed after 17 days at 10%
moisture.
4.9.1.5. Comparison ofnBTPTA with PPDA and Ammonium Thiosulfate (ATS)

Laboratory Experiments. Bremner et al. (l986a) compared nBTPTA with PPDA and
ATS in samples of various soils treated with urea and incubated at 20, 25, and 30°C,
and concluded that both nBTPTA and PPDA applied even at rates as low as I I1g/g soil
caused a substantial retardation of urea hydrolysis, whereas ATS did not have any
inhibitory effect even when it was added to the reaction mixtures at rates as high as
1. 000 11g!g soil.
Field Experiments. Joo et al. (1989) used a Kentucky bluegrass (Poa pratensis) turf
established on a fine loamy soil (PH 7.5) to compare the effectiveness of nBTPTA,
PPDA, ATS, as well as K+ and Mg2+ in reduction of the volatilization of ammonia from
urea-treated plots. Rates of addition were: 10% N urea solution 49 kg Nlha; nBTPTA
0.5, 1, and 2%; PPDA 1,2, and 3%; ATS, K+ (as KCI), and Mg2+ (as MgCl z) 5, 15, and
25%. All percentages represent weights relative to weight ofurea-N. The control plots
were treated only with urea. The volatile NH3 was measured for 4 days.
The cumulative NH3 losses expressed as percentages of the added urea-N gave the
following values for the control plots and for those treated with the three rates of the test
compounds:
18.5 (control); lOA, 7.9, 7.2 (nBTPTA); 7.0, 6.9, 5.6 (PPDA); 14.5, 14.0, 14.7
(ATS); 17.2, 17.3, 17.6 (K+); and 15.3, 15.2, 15.9 (Mgz+).
It is evident that the effectiveness of the compounds tested for reduction of NH3
losses after fertilization with urea decreased in the order:
nBTPTA > PPDA » ATS > Mgz+ > K+.
205
4.9.1.6. Comparison oInBTPTA with PPDA

Laboratory and Pot Experiments. Bremner and Chai (1986) prepared reaction mixtures
from 5-g air-dried soil samples + 2 ml of solution containing 10 mg of urea with or
without 5. 25. and 50 Ilg of nBTPTA or PPDA. The incubation temperature and time
TABLE 47. Effect ofdiflerent amounts ofnBTPTA and PPDA on urea hydrolysis in soil"
Inhibition of urea h:t:drol:t:sis (%2
Incubation
Amount added (/-1g!g soil)
Soil temperature
(DC) nBTPTA PPDA
5 10 5 10
Sandy loam 20 49 60 73 49 66 71
I2H 8.0 30 44 58 69 0 17 28
Sandy loam 20 80 86 87 19 26 40
I2H 8.3 30 68 SO 84 0 0 0
Loam 20 58 75 79 49 67 73
I2H 6.4 30 4 39 55 0 6 13
Silty clay loam 20 26 64 72 3 27 30
I2H6 .O 30 0 0 8 0 0 0
Average 20 53 71 78 30 47 55
30 29 45 54 0 6 10
"From Bremner and Chai (1986). by courtesy of Marcel Dekker. Inc.
were 20 and 30°C and 10 days, respectively. Four soils were used. Results of the
comparison (Table 47) prove that nBTPTA was considerably more effective than PPDA
for inhibition of urea hydrolysis at 20°C and was much more effective than PPDA at
30°C.
Beyrouty et al. (1988b) used samples of a silty loam soil (PH 6.5) from Indiana to
compare the effects of nBTPTA and PPDA on volatilization of ammonia from urea. The
experiment comprised three variants: bare soil samples (20 g); 20-g soil samples to
which 0.3 g of finely ground maize residue was added and partially incorporated below
the soil surface; and 0.3 g of maize residue alone. All samples were treated with a urea
solution (20 mg N) with or without 0.4 mg of nBTPTA or PPDA. The next step was
incubation. Ammonia volatilized during 2, 4, 8, 14, 21, and 28 days was determined.
The NH3 volatilized in 28 days was practically identical in soil only and soil + residue,
with 26-27% of the applied urea-N volatilized in the absence of inhibitors and 1-3% in
their presence. This means that in the soil studied (pH 6.5) both inhibitors were very
effective. In the variant with residue only, the NH3 volatilization loss in 28 days was
92% in the absence of inhibitors, and it was reduced by nBTPTA and PPDA to 39 and
43% respectively. The weaker effectiveness of inhibitors in this variant is explained by
the finding that urease activity in the residue was approximately 47 times greater than in
the soil on the same weight basis. It should be added that in the absence of inhibitors,
volatilization of NH3 during the first 2 days of incubation was 5 times higher from the
soil + residue variant than from the residue alone, but, as already mentioned, during 28
days the same amounts of NH3 were lost by volatilization from both variants.
Influence of redox potential on the inhibitory effectiveness of nBTPT A and PPDA
in retarding urea hydrolysis was studied by Lu et al. (1989). Samples of four rice soils
were used: a silt loam, pH 5.8 {soil 1) and a clay, pH 5.3 {soil 2) from the U.S.A., a silty
206
clay, pH 6.9 (soil 3) from China, and a silty loam, pH 8.3 (soil 4) from India. Air-dried
ground soil equivalent to 400 g of oven-dry weight and 1,600 rn1 of deionized water
were introduced into flasks. One-half of the flasks were continuously stirred and purged
with air (oxidized treatment) and the remaining set stirred and purged with argon gas
(reduced trcatment). Then the soil suspensions were preincubated in the dark at 30°C.
During preincubation, the redox potential (Eh) and pH were measured. They reached
steady state values after 19-23 days of preincubation. Afterwards, soil suspension
aliquots equivalent to 4 g of oven-dry weight were removed from the flasks and
centrifuged. The water layer was discharged while maintaining aerobic and anaerobic
conditions. respectively. The sedimented soil was treated with urea (400 /lg N/g soil)
with or without 2% (on urea weight basis) nBTPTA or PPDA. Untreated soil served for
comparison. The reaction mixtures prepared in this way were incubated at 30 D C and
after 1,3,5, 7, and 15 days they were analyzed for residual urea.
The oxidized treatment aimed at simulating the thin oxidized surface layer in
flooded soils and the oxidized zone around the roots of rice plants, whereas the reduced
zone under the soil surface was simulated by the reduced treatment.
At steady state, Eh values in suspensions of the four soils studied ranged from +630
to +730 mY in the oxidized treatment, and from -290 to -340 mY in the reduced
treatment; pH decreased in the oxidized treatment (except soil 1 in which the pH
slightly increased) and increased in the reduced treatment (except soil 4 in which the pH
remained unchanged).
The results obtained in analysis of the residual urea are presented in Table 48. In
soils treated only with urea, urea hydrolysis during 1 and 3 days was more marked in
reduced san1ples than in the oxidized ones of soils 1 and 4, whereas the opposite was
true for soils 2 and 3. Urea hydrolysis was complete in 5 days in all samples of each
soil. Both inhibitors retarded urea hydrolysis in each soil. nBTPTA was more effective
in oxidized samples and PPDA in the reduced ones. Thus, at day 7 after incubation, the
residual urea (expressed as average of the values recorded in the four soils studied) was
49 and 17% of the added urea in oxidized samples treated with nBTPTA and PPDA,
respectively. The corresponding average values registered in the reduced samples were
4% (nBTPTA) and 26% (PPDA), respectively. At day 15, urea remained unhydrolyzed
in considerable amounts only in three samples: the PPDA-treated reduced sample of soil
1, nBTPTA-treated oxidized samples of soils 3 and 4, containing -20, 44, and -54%
residual urea, respectively.
In the experiment by Bronson et al. (1989) to compare the effectiveness of nBTPTA
and PPDA on a111l110nia volatilization from urea, samples of a loamy sand soil (PH
6.9)were used. Some samples were acidified with sulfuric acid to adjust the pH to 6.5
and 6.0. The reaction mixtures contained lO g of air-dried soil + 10 mg ofurea-N + 0 or
50 ~lg of nBTPTA or PPDA. During incubation (at 25 D C for 27 days), amounts of
volatile NH3 were assessed. The results (Table 49) show that the inhibitory effect of
both compounds increased slightly when soil pH was adjusted from 6.9 to 6.5 and 6.0.
nBTPT A was more effective than PPDA. Thus, NH3 volatilization was completely
inhibited at each pH for 9 days by nBTPTA and only for 4 days by PPDA. At day 27,
the degree of inhibition by nBTPTA was between 34 and 43 at the three pHs, whereas
the inhibition by PPDA became equal to zero at each pH.
TABLE 48. Effectiveness ofnBTPTA and PPDA in retarding urea hydrolysis in four rice soils under oxidized and reduced conditions·
Soil 1 Soil 2 Soil 3 Soil 4
Residual urea (~N/g soil}
Iyi' Inhibitor'
Oxidized Reduced Oxidized Reduced Oxidized Reduced Oxidized Reduced
conditions conditions conditions conditions
nBTPTA 292 a 185 b 324 a 332 b 376 a 143 b 250 a 170 b
PPDA 292 a 271 a 349 a 391 a 307b 288 a 273 a 299 a
No 216 b 186 b 239b 304c 121 c 168 b 183 b 117 c
nBTPTA 294 a 26b 247 a 191 b 295 a 32 b 218 a 5b
3 PPDA 294 b 255 a 183 b 327 a 193 b 213 a 202 a 254a
No 30 c Ob 66c 82 c Oc 5b 13b Ob
nBTPTA 274 a 20b 170 a 32b 246 a 17 b 218 a Ob
5 PPDA 96b 238 a 133 a 182 a 120 b 101 a 142 b 254 a
No Oc Ob Ob Oc Oc Ob Oc Ob
nBTPTA 252 a 20b 52 a 18 b 260 a 18 a 217 a Ob
7 PPDA 36 b 144 a 28b 44a 80b 23 a 118 b 199 a
No Ob Oc Oc Oc Ob Ob Oc Ob
nBTPTA 25 a Oc 5a 2b 176 a Ob 214 a Oa
15 PPDA 12 b 78 a Ob 4a 4b 7a Ob 2a
No Oc Ob Ob Oc Ob Ob Ob Oa
"Adapted from Lu et aI. (1989), by courtesy of Marcel Dekker, Inc.
bIncubation time (days).
'Inhibitor added to urea (400 Ilg urea-N/g soil) at a rate of2% relative to weight ofurea-N.
No - Only 400 Ilg urea-N/g soil.
Values with the same letter are not significantly different (p=0.05).
N
o
-...J
208
TABLE 49. Inhibition of ammonia volatilization by nBTPTA and PPDA in urea-treated samples of a
loa~ sand at three initial EH values at 25°C·
Inhibition ~%l
Initial
Treatment Incubation time ~da:z:sl
pH
2 4 6 9 12 15 18 21 24 27
6.9 100 100 100 100 95 84 71 57 46 34
Urea + nBTPTA 6.5 100 100 100 100 97 87 75 64 53 42
6.0 100 100 100 100 97 89 77 66 54 43
6.9 100 100 88 53 19 8 4 2 1 0
Urea + PPDA 6.5 100 100 93 64 27 9 5 2 0 0
6.0 100 100 100 73 30 6 0 0 0 0
"From Bronson et aI. (1989), by courtesy of Marcel Dekker, Inc.
Complex investigations related to nBTPTA and PPDA were also conducted in New
South Wales, Australia, by Cai et al. (1989).
In the first experiment, air-dried samples (1 kg) of three soils were placed in pots
and flooded to a depth of 5 cm. Then nBTPTA and PPDA (at rates of 0.005,0.01,0.1,
1, and 5% of the weight of urea) were applied into the floodwater followed by the
addition of 366 mg of urea (equivalent to 80 kg N/ha). The pots were kept in a
glasshouse, in which the temperature ranged from 22 to 32°C. At zero time and after 2,
4, and 6 days, the soils were analyzed for urea.
The analytical data show that effectiveness of nBTPTA and PPDA as urease
inhibitors increased with the rate of addition. The low rates (0.005-0.1 %) only slightly
retarded the hydrolysis of urea. At higher rates (1 and 5%), both inhibitors were
effective, but the extent and duration of inhibition varied with soil type (Figure 67).
A B a
,~ \ \:
~---,~U
2 • 8
\::
'-----'-_-'-.
2
-_~.u
"Ii
~,
I
2
~~I
4 a
Timeldaysl Time (daysl Time (deysl
Figure 67. Effect of rate of addition ofnBTPTA and PPDA on urea hydrolysis in flooded soils in a
pot experiment.
A - Yellow podzolic soil, pH 6.2. B - Grey soil, pH 8.4. C - Red-brown earth, pH 5.6. U - Urea. B I -
Urea + 1% nBTPTA. B5 - Urea + 5% nBTPTA. PI - Urea + 1% PPDA. P5 - Urea + 5% PPDA.
IFrom Cai et al. (1989). by permission of Pergamon Press PLC'/
Thus, in two soils (yellow podzolic and grey soil), nBTPTA was more effective than
PPDA, whereas in the third soil (red-brown earth), only PPDA was effective. The
209
higher inhibitor rate acted always more markedly than the lower rate. During
incubation, the degree of inhibition decreased only slowly in the yellow podzolic soil
and rapidly in the two other soils.
In other experiments, carried out in bottles, the influence of a) soil moisture content
and b) growth of algae on the inhibitory effectiveness of nBTPTA as well as the
influence of c) the chelating compound ethylenediaminetetraacetic acid (EDTA) on
inhibitory effectiveness ofnBTPTA were studied.
a) Air-dried samples (15 g) of five soils received water up to field capacity (moist
samples). Thirty ml of water was added to other samples (flooded samples). The added
water contained 14 mgofurea with or without 5% nBTPTA or PPDA relative to weight
of urea. Incubation took place in the dark at 25°C. After 2, 4, and 6 days, the
unhydrolyzed urea was assayed. In the moist samples of each soil, nBTPTA inhibited
the urea hydrolysis more markedly than PPDA. In flooded samples the inhibition was
nearly complete in 6 days for 4 and 3 out of the five soils treated with PPDA and
nBTPTA, respectively. Urea hydrolysis in the flooded red-brown earth proved to be
resistant to inhibition by nBTPTA in this experiment, as well.
b) Samples (15 g) of three soils were flooded with 30 ml of water containing urea
with or without nBTPTA or PPDA in the same amounts as in experiment a. Three
treatments were applied. Some samples were incubated in the dark (at 25°C) for
preventing growth of algae (treatment D). Other samples were exposed to light under
normal daylight conditions in a glasshouse to promote growth of algae (treatment L). In
order to enhance additional algal growth, some flooded samples were exposed to light
in the glasshouse for 10 days, before urea was added with or without inhibitor and new
exposure to light as in treatment L (treatment LL). Determination of urea after 2, 4, and
6 days of incubation showed that in each soil urea hydrolysis was inhibited by PPDA
more effectively under dark than light conditions, whereas the inhibition by nBTPTA
was practically unaffected by exposure to light. This means that in the presence of light
and algae, PPDA was less effective than nBTPTA. The effectiveness of PPDA was
much lower in treatment LL than in treatment L. To explain these findings, it is
considered possible that the algae in the soils studied contain other enzymes, such as
urea carboxylase (hydrolyzing)', which catalyze decomposition of urea and that these
enzymes are not inhibited by PPDA, but are by nBTPTA
c) EDTA was studied with the red-brown earth, in which nBTPTA did not inhibit
urea hydrolysis. The reaction mixtures had the following composition: 15 g of soil +
0.67 or 2 mM of EDTA + 30 mg of urea + 0 or 1 or 5% nBTPTA (relative to weight of
urea) in 30 m1 of water. After 2 and 4 days of incubation in the dark at 25°C, the
mixtures were analyzed for residual urea. The analyses indicated that EDTA manifested
an inhibitory effect on urea hydrolysis. At 0.67 mM of EDTA, the degree of inhibition
after 2 and 4 days of incubation was 26.8 and 1.2%, respectively, whereas at 2 mM of
EDTA the corresponding values were 39.6 and 4.2%, respectively. At the same time,
EDTA improved the inhibitory capacity of nBTPTA and this effect became stronger
with the increase in concentration of the two compounds. Moreover, after a 2-day
incubation the combined effect of EDTA and nBTPTA (at each concentration) was
This enzyme catalyzes hydrolytic decomposition of urea according to the reaction:

urea + A TP + CO2 -> 2 NH3 + 2 CO2 + ADP + a-phosphate.
210
significantly greater than the sum of their individual effects (synergism), but after 4
days, only the combined effect of 2 mM EDTA and 5% nBTPT A (degree of inhibition:
21.6%) was significantly greater than their additive individual effects (degree of
inhibition: 4.2 + 3.2%).
These investigations were also referred to by Freney et al. (1989).
Byrnes and Amberger (1989) compared the urease inhibition by nBTPTA and
PPDA in flooded, unplanted samples of a silty clay loam soil (pH in H20 5.9). The 300-
g samples were placed in plastic containers 10.8 cm in diameter and 6 cm deep, and
were flooded and puddled to provide a soil depth of about 3 cm and 2.5 cm of
floodwater. The rates of additions to floodwater were: 75 mg of urea, 0, 0.5, I, 2, and
5% nBTPT A or PPDA relative to weight of urea. The samples were incubated in a
greenhouse under normal daylight conditions.
Daily analysis of urea in floodwater showed that urea disappeared in 5 days from the
control sample and in 5-8 days from the samples treated with 0.5-5% PPDA. Urea was
still detectable in all nBTPTA-treated samples even at day l3; the amount of
unhydrolyzed urea was proportionate to the rate of nBTPT A. Even at its highest rate
(5%), PPDA was less inhibitory on urea hydrolysis than was nBTPTA at its lowest rate
(0.5%).
In a pot experiment with flooded rice under normal daylight conditions, it was also
found (Byrnes et aI., 1989b) that nBTPTA was a stronger inhibitor of urea hydrolysis in
soil than PPDA (see page 285).
Bhupinderpal-Singh et al. (1992) used surface (0-15 cm) samples of two
representative soils from semiarid regions of northwestern India (a silty loam, pH 8.3
and a sandy loam, pH 8.4). Soil samples (200 g) were treated with urea (40 mg N) and 1
mg of nBTPT A or PPDA. The control received only urea. The mixtures moistened to
field capacity were incubated at 35°C. The unhydrolyzed urea was estimated after 1,2,
5, and 9 days of incubation.
Urea hydrolysis was complete in 2 days (control), 5 days (PPDA), and 9 days
(nBTPTA). After 1 day, rates of urea hydrolysis in the silty loam and sandy loam were
97 and 55% (control), 49 and 41% (PPDA), and 43 and 15% (nBTPTA), respectively.
Thus, nBTPTA was superior to PPDA in retarding urea hydrolysis in the soils studied.
Joo et af. (1992) transplanted sod from an established Kentucky bluegrass turf (on a
fine loamy soil, pH 7.5) into plastic pots (21 cm diameter, 16 em height). The
transplanted sod was allowed to grow in the greenhouse for -3 months before receiving
10% N urea solution (49 kg N/ha) with nBTPTA (0.125, 0.25, 0.5, and 1% of the
weight of urea-N) or PPDA (0.5, 1, and 2% of the weight of urea-N)o Only urea was
added to the control pots. The volatilized ammonia was determined daily for 7 days.
The cumulative NH3 loss from the control pots (49.9% of the added urea-N) was
reduced by the four rates of nBTPTA to 29.0, 24.6, 22.8, and 20.4%, and by the three
rates ofPPDA to 32.8, 26.7, and 24.2%.
These numerical data prove that nBTPT A was a more effective inhibitor than PPDA
in reducing the volatile NH3 loss from urea, and the effectiveness of both inhibitors
increased with their rates.
Field Experiments. Schlegel et af. (1986) determined volatilization of ammonia from

conventional till (CT) and no-till (NT) microplots on a silty clay loam, cropped to maize
and fertilized with surface-applied urea priUs at a rate or 180 kg N/ha. The urea prills
211
were or were not coated with paraffin oil (0.8% relative to weight of urea) and with
powdered nBTPTA (0.5 or 2 kg/lOO kg N) or PPDA (2 kg/lOO kg N). After fertilizer
application, the NH3 volatilized during 18 days (CT microplots) <r 15 days (NT
microplots) was assessed. Cumulative NH3 losses from urea applied on CT microplots
were the following (in kg Nlha): 6.7 (no inhibitor), 5.7 (PPDA), and 2.0 (0.5 and 2%
nBTPTA). The corresponding values for the NT plots were 13.3 (no inhibitor), 5.7
(PPDA), and 4.0 (both nBTPTA rates). Thus, less NH3 was lost from the CT microplots
than from the NT microplots. nBTPTA at each rate was more effective than PPDA.
Buresh et al. (1988a) described experiments in the 1986 dry season for comparing
the urease-inhibitory effectiveness of nBTPTA and PPDA on two flooded rice fields in
the Philippines. One field on a clay soil is located at Pila, the other on a silty clay soil at
Munoz. Urea was applied at three rates (35, 70, and 140 kg N/ha) with or without 0.9%
nBTPTA or 2% PPDA relative to weight of urea. Two-thirds of each rate of urea with
or without inhibitor was broadcast into the 5-cm deep floodwater at day 18 after
transplanting rice seedlings and one-third at day 5-10 after panicle initiation. The
control plots were not fertilized. Following each urea application, the floodwater was
analyzed daily for 8-10 days for residual urea and ~ +; vapor pressure of NH3 (pNH3)
was also assessed.
Analysis of urea has shown that after administration of the first urea amount
nBTPTA was a more effective inhibitor of urea hydrolysis than PPDA, especially at the
Munoz rice field. For example, at day 7 after application of the first urea amount (23,
47, and 93 kg Nlha, respectively), residual urea represented 21, 31, and 31 % of the urea
applied with nBTPTA, and 7, 13, and 0% from the urea applied with PPDA at Munoz,
whereas at Pila the corresponding values were 2, 2, and 9% (nBTPTA) and 1, 1, and 0%
(PPDA), respectively. In contrast, after the second application of urea, PPDA proved to
be more effective than nBTPTA at both Pila and Munoz.
Determination of the ~ + content gave results concordant with those obtained by
analysis of residual urea. Thus, after the first administration of urea, nBTPTA prevented
the accumulation ofNH4+ in floodwater for 10 days, whereas PPDA tended to delay and
not to eliminate accumulation ofNH/. These findings were valid for each urea rate and
for both rice fields. After the second application of urea, nBTPTA reduced less
markedly the accumulation of~+ in floodwater in comparison with PPDA.
Variation of pNH3 followed trends similar to those of~+.
4.9.2. Comparative Studies on the Effect of nBTPTA and Other Inhibitors on the
Immobilization of Urea-N in Soils
In a variant of the pot experiment performed by Cai et al. (1989) and referred to on page
208, the effect of nBTPTA on distribution of urea-N in upper layers of a flooded grey
soil and the effect of both nBTPTA and PPDA on immobilization of urea-N into
organic matter of two flooded soils (grey soil and yellow podzolic soil) were studied.
nBTPTA or PPDA at a rate of 5% of the urea weight was added to the floodwater of the
I-kg soil samples and followed immediately by 366 mg of J5N-labeled urea (5 atom%
excess 15N). The next step was incubation in a glasshouse. After 0, 1,2,4,6, 8, and 35
days, total N and 15N were determined in the 0-1-, 1-2-, and 2-4.5-cm layers for
establishing the depth of penetration of urea into the soil. For assessing 15N immobilized
into organic matter, total N and 15N were determined after extraction with a eaCh
solution to remove mineral N.
212
The bulk of the added N was recovered in the O-l-cm layer, although appreciable N
had diffused to the 2-4.5-cm layer. Considerably more N was retained in the soil treated
with nBTPTA than in the soil receiving urea only. Thus, after 35 days of incubation the
0---4.5-cm soil layer contained 47.4% of the added 15N in the urea-only treatment and
74.7% in the urea + 5% nBTPTA treatment.
Both inhibitors increased immobilization of added urea_ 15 N into organic matter of
both soils studied. After 35 days of incubation, the following percentages of the added
urea- 15N were immobilized in the urea, urea + 5% nBTPTA, and urea + 5% PPDA
treatments: 37.5, 46.3, and 47.5%. respectively (in the grey soil), and 43.8, 47.2, and
49.8%, respectively (in the yellow podzolic soil). Increased immobilization should
reduce losses of applied N and may increase the efficiency of fertilizer N.
Wang et al. (1991b) studied the immobilization of urea-N in samples of a Belgian
loam soil treated or not treated with air-dried, ground barley straw and nBTPTA, PPDA,
and hydroquinone (HQ). The reaction mixtures consisted of 20 g of soil + 0, 1 or 2%
straw + 4 mg ofurea-N, either unlabeled or labeled with 15N (5.297 atom% excess 15N;
i.e .•. 10.594 mg 15Nlkg soil) and 1% of inhibitor (based on weight of urea). The soil
moisture was brought up to two-thirds of field capacity. Then, the reaction mixtures
were incubated at 25°C for 15 days (mixtures with unlabeled urea) or for 20 days
(mixtures with labeled urea). Urea and inorganic N, including NH/, N0 3 -, and N0 2 -
were periodically extracted with 1 M KCl solution and determined. The reaction
mixtures with labeled urea were washed twice with 2 M KCl solution to remove the
mineral N. Then the mineral 15N and the 15N remaining in the soil organic fraction were
determined.
Analysis of residual urea showed that addition of straw to the soil caused an
extremely rapid urea hydrolysis and decreased the inhibitory effect of each test
compound. HQ and PPDA were more affected by straw than was nBTPTA. The order
of their inhibitory capacity, HQ < PPDA < nBTPTA, was the same in both the absence
and presence of straw. The inhibitors, especially nBTPTA reduced the formation of
NH4 +. Much less NH4 + was formed in the soil treated with 2% straw, due to
immobilization in microbial biomass and chemical fixation by humus. From the 9th day
on, a little more N0 3- + N0 2- was found in mixtures with 1% straw + inhibitor. This
means that the inhibitors slightly decreased the N immobilization. However, this effect
was less clear when 2% straw was used, since all reaction mixtures showed a low level
ofN0 3 - + N0 2-.
In contrast to the mixtures amended with 1% straw in which the inhibitors slightly
decreased immobilization ofurea-N, immobilization of 15N into soil organic matter was
enhanced and the amount of mineral 15N decreased in the unamended mixtures (Table
50). The immobilizing effect of the inhibitors increased in the order: PPDA < HQ <
nBTPTA. The order was the same for the recovery of 15N under the influence of
inhibitors.
These investigations were also referred to in a communication presented in 1990 at
an International Symposium held in Vienna (Van Cleemput et al.• 1991).
213
TABLE 50. Effect of urease inhibitors on immobilization ofurea_ 15 N in soil samples incubated at 25°C
for 20 days·
Distribution of 15Nb
Treatment ISN, Il Nm IlN/'N f x 100 IlNm/IlNfX 100 Recovery
(m!ikg soil) (m!ikg soil) (%) (%) (%)
Urea' 4.2 5.1 39.4 48.0 87.4
Urea + nBTPTA 7.4 3.2 69.4 30.2 99.7
Urea + PPDA 4.8 4.9 44.9 46.3 91.1
Urea + HQ' 5.2 4.6 49.4 43.5 93.0
Q Adapted from Wang et al. (l99Jb), by permission of Springer-Verlag.
b 15N, _ I5 N in the soil organic fraction. 15 Nm _ 15 N in the soil extract (mineral 15N). 15 Nr _ 15N initially
added (=10.594 mg/kg soil).

, These numerical data, obtained in the urea and urea + HQ treatments, were also published in another
paper, whose co-author is Wang (Zhou et al.. 1992).
4.9.3. Comparative Studies on the Stability of PTA and TPTA Compounds and Other
Inhibitors
4.9.3.1. Comparative Studies on the Stability of Phosphoryl Triamide (PTA),

Thiophosphoryl Triamide (TPTA), nBTPTA, and Other Inhibitors in Solutions
(Including Urea Melt) and in Solid State
It was demonstrated in laboratory studies (Anonymous, 1985a, 1987; Radel et al., 1987;
Gautney, 1987) that phosphoryl triamide (PTA) is a very labile compound; it decom-
poses rapidly in urea melts at 140°C (e.g., a quarter of the PTA decomposed in 4
minutes). This means that PTA can not be cogranulated with urea. It is also not stable in
solid state.
Thiophosphoryl triamide (TPTA) dissolves relatively rapidly in urea melts, in which
it remains intact for about 4 minutes; i.e., it is more stable than PTA and even than
PPDA. Consequently, TPTA can be cogranulated with urea. However, its instability in
solid mixtures with urea, proven by kinetic i'>tudies OIl its solid-state decomposition,
precludes its use with solid urea.
In these studies the decomposition of pure TPTA served for comparison. To
simulate ambient air moisture levels, the decomposition studies were conducted at a
constant water partial pressure of 12.7 mm Hg. Zero-order reaction half-lives for the
solid-state decomposition of pure TPTA and TPTA in urea mixture (l0% TPTA + 90%
urea) at 42, 50, and 60°C were estimated experimentally. The reaction half-lives at 25°C
were calculated from Arrhenius plots of the higher temperature data, and they were
found to be 212 hours for pure TPTA and 38 hours for TPTA in urea mixture. Thus,
TPTA is not stable enough for storage at ambient air moisture conditions either alone or
in mixtures with urea. As shown on page 134, the calculated first -order reaction half-life
at 25°C is longer for pure solid PPDA (254 years) and zero-order kinetics indicated
56% decomposition per year at 25°C for PPDA in mixtures with urea.
Pure solid TPTA is stable in sealed Nalgene containers under ambient air and when
stored in glass under dry N2 , but decomposes when stored in plastic bag or in glass
under dry air.
Solid-state decomposition ofTPTA results in a number of different compounds, two
of which were identified as ammonium thiophosphorodiamidate (ATPDA) and
dian1ITIonium thiophosphoroamidate (DATPA), respectively. This means that at least a
214
part of the TPTA is decomposed via two partial hydrolysis reactions (Figure 68). The
other decomposition products and reactions were not identified.
H~-P
s
II ......NH 2
'NH2
Thiophosphoryl triamide
(TPTA)
+ H:P
- S
II ......NH 2
NH40-P,
NH2
Ammonium thiophosphorodiamidate
(ATPDA)
ATPDA
s
II.,....NH2
NH40-P
' NH2
+ H:P
- NH4
s
NH4o-....11
i fP - NH2
Diammonium thiophosphoroamidate
(DATPA)
Figure 68. Solid-state decoJ1llosition of thiophosphoryl triarnide via partial hydrolysis reactions.
(Adapted from Radel et al. (1987). by permission ofTennessee Valley Authority.!
The possibility for using TPTA in fluid fertilizers was also studied. This possibility
was substantiated by observations that TPTA is very soluble and, additionally, very
stable for up to 30 minutes in 75 and 87% urea solutions at 90 and 99°C, respectively.
The fluid fertilizers studied comprised: urea-ammonium nitrate suspension (UAN,
250
~ur..
--
aolut1OD
pH 8.54
191
DB
31-0-0
H2O pH S.lO
ua pH 7.24 !l3
)6-0-0
pH 7.)6 68
I 43
t
Figure 69. Stability ofTPTA in water and fluid fertilizers at 25°C. !From Anonymous (1987), Radel
et al. (1987), Gautney (1987), by permission of Tennessee Valley Authority.!
36-0-0) and solution (UAN, 31-0-0), and 40% urea solution. TPTA in water served for
comparison. In each case concentration of TPTA was 1% (weight/weight). Figure 69
shows that the first-order reaction half-life for decomposition of TPTA increases with
215
the pH of fluid fertilizers. As these half-life values are high (43-191 days), there is
sufficient time for adding TPT A to fluid fertilizers and for their application on the soil.
Decomposition of TPTA in solution phase is the result of a single hydrolytic
reaction at slightly alkaline pHs, the only decomposition product being the ammonium
thiophosphorodiamidate (ATPDA).
Stability of nBTPTA in aqueous solutions was studied by Bremner and Chai (1986).
nBTPTA was not directly analyzed. An indirect method was applied: diminution of the
inhibitory effectiveness of nBTPTA solution for urease activity in soils was evaluated
after its storage, because diminution of effectiveness indicates decomposition of
nBTPTA. The aqueous solution, containing 50 fig of nBTPTAlml, was stored at 5, lO,
20, and 30°C for 0, 3, 7, 21, and 28 days, and then 1 ml of stored solution and 1 ml of a
urea solution (10 mg of urea) were added to 5-g air-dried samples of three Iowa soils.
After incubation of the reaction mixtures (20°C for 2 days), the residual urea was
assessed and percent inhibition calculated.
The results showed that storage of the nBTPTA solution at 5°C for 3 days did not
significantly affect its ability to inhibit urease activity in soils, i.e., nBTPTA did not
decompose under these conditions. With increasing temperature and prolongation of
storage time, the inhibitory effectiveness of nBTPTA gradually decreased, i.e., more
and more nBTPTA molecules were decomposed. For example, the inhibitions, that the
fresh nBTPTA solution and the solution stored at 30°C for 28 days exhibited in the
urease activity of the three soils studied, were: 98 and 71 %, 95 and 61 %, and 69 and
47%, respectively.
Douglass and Hendrickson (1991), who developed a sensitive high-performance
liquid chromatography method for direct analysis of nBTPTA and its oxygen analogue
(nBPTA), studied stability of these compounds in solutions. lO,5 M nBTPTA solutions
were prepared in deionized water or 20% Hoagland's nutrient solution (adjusted to
either pH 5.5 or 7.5 with H2S04 or KOH, respectively). Similar solutions were prepared
to a final concentration of 40% (volume/volume) methanol. The solutions were stored at
25,4, and -20°C for 3, 7, and 14 days prior to analyses. The results obtained after 14-
day storage are summarized in Table 51.
TABLE 51. Stability of nBTPT A in 10-5 M solutions following storage for 14 days
at various temperatures
Q
Methanol nBTPTA remaining ('Yo)

Solution
('Yo)
Water 0 100 100 96
Nutrient solution, pH 5.5 0 97 101 58
Water 40 84 96 102
"Adapted from Douglass and Hendrickson (1991).
One can see from Table 51 that solutions of nBTPT A in water and nutrient solutions
can be stored without decomposition for at least 2 weeks at room temperature or when
refrigerated. Considerable nBTPTA decomposition occurred, however, when the
solutions were stored at -20°C (i.e., in frozen state), especially if they were acidic (pH
216
5.5). The decomposition at _20DC was completely eliminated by adding methanol to the
solutions to prevent freezing. However, for storage at 25 and 4 DC, addition of methanol
to the solutions is not recommended, since at these temperatures methanol enhances
decomposition of nBTPTA in non-alkaline solutions.
Similar results were obtained with solutions of nBTPTA.
4.9.3.2. Comparative Studies on the Stability Qf PTA. TPTA, nBTPTA, and Other
Inhibitors in Soils
In the field experiment conducted by Beyrouty et al. (1988a, b) and referred to on page
196, N-benzyl-N-methylphosphoric triamide (in contrast to PPDA which retarded urea
hydrolysis for 19 days in rnicroplots covered in a proportion of about 60% by maize
residue) was not able to retard urea hydrolysis under similar conditions. This finding
was interpreted as evidence of a deactivating action of plant residue on this
phosphorotriarnide.
The mechanism through which nBTPTA is decomposed in soil was studied by Chai
et al. (1988), Byrnes and Christianson (1988), and McCarty et al. (1989). These
investigators found that a) nBTPTA is a strong inhibitor of soil urease activity, but a
very poor inhibitor of ureases of plant (jackbean) and bacterial (Bacillus pasteurii)
origin; b) aqueous extracts from soils previously treated and incubated aerobically with
nBTPTA inhibited activity of jackbean urease; c) nBTPTA is rapidly converted in soil
(largely abiotically) to a compound that is much more effective than the parent
compound for inhibition ofjackbean urease activity; d) data from 31p nuclear magnetic
resonance and infrared spectroscopy of unfractionated soil extracts indicated that the
decomposition product of nBTPTA is its oxygen analogue, i.e.. N-(n-butyl)phosphoric
triamide (nBPTA). This means that the mechanism through which nBTPTA is
converted in soil to nBPTA is oxidative' desulfuration:
Creason et al. (1990) extracted this compound with water from samples of a silt
loam soil (PH 6.9) previously treated with nBTPTA (5 mglg soil) and incubated in
loosely capped bottles (for access of air) at 25 DC for 24 hours. Then, the compound was
purified by high-performance liquid chromatography fractionation and identified by
mass spectroscopy as nBPTA. The purified nBPTA was shown to be highly active,
giving 50% inhibition of jackbean urease at concentrations between 10 and 100 mM.
This urease-inhibiting ability was almost equivalent to that of PPDA. The further fate of
nBPTA in soil was not studied.
'The oxydative mechanism is also supported by the results of an experiment of Luo et al. (1994). Air-dried
samples (15 g) of an Australian vertisol (pH 7.6), placed in 120-ml glass bottles, were flooded with 30 ml of
water, then nBTPTA (1% relative to weight of urea) and urea (14 rng) were added and the mixtures were
oxygenated by bubbling O2 into the floodwater for 3 hours or by adding 1 ml of 30% H20 2 • Mixtures
without nBTPTA, O2, and H20 z were the controls. All mixtures were incubated at 25°C for 8 days, during
which at 2-day intervals the residual urea was determined.
It was found that rate of urea hydrolysis in the nBTPTA-treated mixtures was significantly (p<0.05)
slowed by O2 and very significantly (p<O.OO 1) by HzO z, indicating that at least a part of the nBTPTA was
oxidized to nBPTA, which was more inhibitory than the parent compound.
217
Studying the decomposition of thiophosphoryl triamide (fPTA) in soil, Bremner

and McCarty (1989) and McCarty and Bremner (1989b) found that TPTA behaved
similarly to nBTPTA.
In one experiment the effect of TPTA and its oxygen analogue, phosphoryl triarnide
(PTA) on activity of jackbean urease was studied. The reaction mixtures, containing
0.02 nmol of jackbean urease, 0 or 0.1 to 500 nmol ofTPTA or PTA and 0.2 mmol of
urea in 0.1 M tris buffer (pH 7.0) in a total volume of 10 rnl, were incubated at 30°C for
15 minutes, then analyzed for ~ + released from urea. The analytical data clearly
showed that TPTA had very little (only 6%) inhibitory effect on jackbean urease
activity even when its concentration was as high as 50 nmollrnl, whereas PTA caused
significant inhibition of this activity even when its concentration was as low as 1
nmol/rnl (97% inhibition) or 0.05 nmol/rnl (24% inhibition).
In another experiment, 0.5-g samples of three Iowa soils and 1 rnl of a solution
containing 0.1 Ilmol of TPTAlml were incubated at 30°C for 0-24 hours, then treated
with 99 ml of water and filtered (membrane filter, 0.45 J.1ffi). Afterwards, 1 ml of each
filtrate was tested for its ability to inhibit jackbean urease using the same method as in
the first experiment. The data obtained indicated that the filtrates inhibited activity of
jackbean urease. The inhibitory capacity of filtrates increased with the time of
incubation of the TPTA solution + soil mixtures and varied with the nature of soil.
Thus, after 24 hours of incubation, the filtrates gave the following inhibitions of the
jackbean urease activity:80% (filtrate from silty clay loam), 73% (filtrate from sandy
clay loam), and 71 % (filtrate from sandy loam).
To identify the inhibitor formed from TPTA in soil, 3-g samples of the silty clay
loam were incubated with 5 rnl ofTPTA solution (25 mM) at 30°C for 0,2,4 or 6 days,
then the filtrates were analyzed by 31p nuclear magnetic resonance spectroscopy. The
analysis proved the presence of PTA in the filtrates. This means that TPTA like
nBTPTA was decomposed in soil by oxydative desulfuration:
This mechanism greatly differs from the decomposition mechanism of TPTA in

solid state and in liquid fertilizers (see page 214).
In contrast with fmdings by Chai and Bremner (1985, 1986, 1987) and Bremner and
Chai (1989), according to which nBTPTA was a more effective soil urease inhibitor
than N-cyclohexylphosphoric triamide (CHPTA), Chien et al. (1988) found, in
preliminary soil studies under submerged conditions without rice plants, that CHPTA
and its thio analogue, N-cyclohexylthiophosphoric triarnide (CHTPTA) seemed to be
more effective sustained-action soil urease inhibitors in comparison with nBTPTA, its
oxygen analogue (nBPTA), and PPDA. However, according to a short report by
Carmona et al. (1988), who studied the effect of nBTPTA on ammonia volatilization
and urea hydrolysis in a variety of soils, the effectiveness of nBTPTA was comparable
to that of its oxygen analogue (nBPTA), CHTPTA, and CHPTA over a similar range of
urea:inhibitor ratios.
To study the stability of nBTPTA and its oxygen analogue (nBPTA) in soils,
Douglass and Hendrickson (1989) carried out analyses by means of high-performance
liquid chromatography (HPLC), and found that nBTPTA was more stable, especially in
acidic soils, than nBPTA.
218
The aim of other laboratory studies was to compare the effects of nBTPTA and
nBPTA on hydrolysis of urea in samples of 24 soils obtained from different sites
throughout the U.S.A. The reaction mixtures, containing 1 mg ofurea-N and 0 or 10 flg
of inhibitor/g soil, were incubated at 25°C for periods up to 10 days. Both compounds
significantly extended the persistence of urea in all soils, but inhibition of urea
hydrolysis with nBTPTA was more persistent than that with nBPTA in nearly every
soil. The average half-life of urea was more than 8 days for nBTPTA, 6 days for
nBPTA, and less than 2 days without inhibitors. The superior effectiveness of nBTPTA
can be explained by its greater stability. Although nBPTA is the active inhibitor,
application of nBTPTA - due to the subsequent generation of nBPTA - is more
effective in most soils than direct application of nBPTA.
This conclusion gained further support in a laboratory experiment described by
Hendrickson and Douglass (1993). nBTPTA and nBPTA were applied to an acid (PH
4.9) silt loam soil (soil A) and to same soil that had been neutralized (PH 7.1) by long-
term liming (soil B) or by recent application of Ca(OHh (soil C). In addition to the
effect of the two inhibitors on urea hydrolysis, their stability in soil was also studied.
The reaction mixtures contained 2 g of soil A, B or C, 0 or 2 mg ofurea-N/g soil and
o or 10 flg of nBTPTA or nBPTAIg soil. Additionally, reaction mixtures were prepared
from 2 g of soil B, 0 or 2 mg of urea-N/g soil and O. 1 or 100 flg of nBTPTA or
nBPTAIg soil. All reaction mixtures were incubated at 25°C for 14 days and analyzed
for remaining urea several times during the incubation. The results showed that the
inhibition of urea hydrolysis was weak in soil A and strong in soils B and C. nBTPTA
was more inhibitory than nBPTA in each soil and also in the soil B amended with 1 or
100 flg nBTPTA or nBPTAIg soil.
The reaction mixtures were also analyzed also for quantitative determination (by
HPLC) of the inhibitors, including nBPTA formed from nBTPTA during incubation of
the reaction mixtures.
In soil A, B, and C, to which nBTPTA or nBPTA was applied at the rate of 10 flglg
soil, nBTPTA, nBPTA formed from nBTPTA during incubation and nBPTA applied
disappeared in 8, 8, and 2 days, respectively (soil A), in 14, 14, and 8 days, respectively
(soil B), and in 10, 10, and 8 days, respectively (soil C).
In soil B, depending on the rate of nBTPTA or nBPTA application (1, 10, and 100
flglg soil), nBTPTA disappeared in 3, 14, and more than 14 days, respectively, nBPTA
formed from nBTPTA disappeared in 8, 14, and more than 14 days, respectively, while
the disappearance of the applied nBPTA occurred in 2,8, and 8 days, respectively.
It is evident from these data that both nBTPTA and nBPTA were less persistent
(and, thus, less urease-inhibitory) in the acid soil A than in the neutral soils B and C.
Therefore, a primary role can be attributed to soil pH or pH-associated characteristics in
controlling persistence of both inhibitors. It is also evident that the nBPTA formed from
nBTPTA during incubation was more persistent (and consequently nBTPTA was more
urease-inhibitory) than the nBPTA applied directly to soil.
Christianson et af. (1990) compared the effects of the two thio and two oxygen
analogues, nBTPTA and nBPTA, CHTPTA and CHPTA on volatilization of ammonia
from urea and hydrolysis of urea in soil samples. The same Guthrie soil and methods
were used as those used in the laboratory experiments of Carmona et al. (1988, 1990)
(see page 146). The rate of urea addition was again 100 kg N/ha on a surface area basis,
219
but rates of inhibitors were only 0.01 and 0.1% relative to weight of urea. Ammonia
volatilization and urea hydrolysis were assessed several times during the incubation of
TABLE 52. Cumulative ammonia volatilization from urea during 14 days and urea
hydrolysis during 10 days as influenced by urease inhibitors"
N volatilized (%) Urea hydrolysis (%)
Inhibitor Inhibitor rate (% relative to weight of urea)
0.1 0.01 0.1 0.01
No (urea alone) 47 47 87.0 a 87.0 a
nBTPTA 7 28 65.4 be 84.7 a
nBPTA 7 15 71.3 b 77.5 ab
CHTPTA 16 30 56.6c 78.0 ab
CHPTA 10 14 60.0c 60.5 b
"Adapted from Christianson et al. (1990).
Numbers followed by the same letter are not significantly different at p=0.05.
14 and 10 days, respectively. The results obtained at the end of the incubation periods
are summarized in Table 52.
It is evident from this table that the inhibitory effect of the four triarnide compounds
studied was higher at their 0.1 % than at their 0.01% rate.
At the 0.1 % rate, NH3 volatilization was inhibited to the same extent by nBTPTA
and its oxygen analogue, and to a smaller extent by CHTPTA than by its oxygen
analogue. At the 0.01 % rate, the thio analogues were less inhibitory than the oxygen
analogues.
The inhibition of urea hydrolysis was stronger by the thio analogues at the 0.1 % rate
and weaker at the 0.01% rate in comparison with the oxygen analogues, but the
differences in inhibition degrees were not significant at p=0.05. The inhibiting effect of
the four compounds on urea hydrolysis increased, although insignificantly, in the
following orders: nBPTA < nBTPTA < CHPTA < CHTPTA (at 0.1 %), and nBTPTA <
CHTPTA < nBPTA < CHPTA (at 0.01 %).
A comparative study on the mobility of these four triarnides and urea was also done
(Christianson and Howard, 1994), by means of soil thin-layer chromatography (TLC).
Five soils were used, from which TLC plates were prepared. Each triamide (0.5 mg
dissolved in methanol) and urea were applied as separate spots on each plate, then the
plates were placed in a developing tank containing water and wetting front was allowed
to advance 10 cm. The plates were then removed, dried at 60°C, cooled and placed in an
iodine developing chamber overnight. The triamides and urea were readily discerned as
golden-brown spots.
nBTPTA moved on plates of all soils at a rate comparable with that of urea and
although no nBTPTA remained at the starting point, it exhibited significant trailing.
Contrarily, the other thio analogue, CHTPTA was very immobile and most of the
compound remained at the starting point. On some plates however a narrow, faint streak
was visible which emanated from the CHTPTA spot and migrated distances similar to
those of urea. The two oxygen analogues, nBPTA and CHPTA moved as compact spots
at rates similar to that of urea.
220
The Rr values of urea and triamides on the plates prepared from the five soils varied
between 0.68 and 0.86 (urea), 0.71 and 0.88 (nBTPTA), 0.67 and 0.79 (nBPTA), 0.64
and 0.75 (CHPTA) or were invariably 0 (CHTPTA).
The trailing spot of nBTPTA eluted from the plates produced a significant inhibition
of jackbean urease, while the inhibitory effect of the eluted faint streak above the
CHTPTA spot was very weak on jackbean urease. These findings indicate that in soil a
partial, but rapid conversion of nBTPTA to its more urease-inhibiting oxygen analogue,
nBPTA took place, while conversion of CHTPTA to a more active inhibitor was very
limited. Immobility and limited conversion of CHTPTA in soil are disadvantageous
properties of this inhibitor.
The investigations performed by Keerthisinghe and Freney (1994) make it possible
to compare the urease-inhibiting effectiveness of nBTPTA and TPTA with that of their
oxygen analogues, nBPTA and PTA, respectively. Flooded samples of two Australian
soils were amended with urea and inhibitors (at a rate of 1% relative to weight of urea),
then incubated for 12 days and analyzed for remaining urea every 2 days.
The conclusion which may be drawn from the results concerning comparative
efficiency of the two pairs ofthio and oxygen analogues is that nBTPTA was a stronger
inhibitor than nBPTA and that TPTA was superior to PTA.
These investigations, in which the effect of an algal inhibitor on the effectiveness of
urease inhibitors was also studied, are referred to in Chapter 6, on page 249.
4.10. COMPARISON OF CYCLOTRIPHOSPHAZATRIENE (CTPAT)

DERIVATIVES WITH OTHER INHIBITORS
By using the procedure a (see page 156), Medina and Sullivan (1986, 1987) found that
the inhibitory effect of 2,2,4,4,6,6-hexaarnino-CTPAT on soil urease activity was more
marked not only than that of phosphoryl triarnide, but also than that of acetohydroxamic
acid, hydroxyurea, thiourea, and ammonium thiocyanate.
221
Chapter 5. Compounds Tested for Evaluation of Their Inhibiting Effect on Both

Soil Urease Activity and Nitrification
First, the effect of nitrification inhibitors on soil urease activity, then the urease
inhibitors also possessing nitrification-inhibiting capacity will be dealt with.
5.1. EFFECT OF NITRIFICATION INHIBITORS ON SOIL UREASE ACTIVITY
5.1.1. Effect of Nitrapyrin and Other Nitrification Inhibitors, Except Azide and
Dicyandiamide, on Soil Urease Activity
Aiming at evaluating the effect of nitrification inhibitors on urease activity in three Iowa
soils, by applying the 5-hour test, Bremner and Douglas (1971) studied 17 compounds
(Figure 70) that were patented as nitrification inhibitors by different companies. Each
compound was used at a rate of 50 ppm (on soil basis). Inhibition of soil urease activity
was very weak: 1-2% with CL-1580 and less than 1% with the other compounds. The
finding that nitrapyrin did not inhibit soil urease activity is in concordance with the
observation of Goring (1962), according to which this nitrification inhibitor, added at
rates of 0.05-10 ppm to 50-g samples of a California sandy loam soil (pH 7.3) treated
with urea (200 ppm N), did not prevent the conversion of urea-N to Nl4+ during
incubation (4,8 or 12 weeks at 21°C).
Bundy and Bremner (l974b) studied three nitrification inhibitors: nitrapyrin, CL-
1580 (see Figure 70), and 4-amino-l,2,4-triazole (Figure 71) in form of hydrochloride
(ATC), determining their effects on urea hydrolysis, ammonia volatilization, and
nitrification. Ten-g samples of three Iowa soils (two clay loams and a sandy clay loam)
were treated with 1 ml of urea solution containing 4 mg of N and with 2 ml of water or
2 ml of aqueous solution containing 100 f.lg of inhibitor. Water was then added to bring
the soil moisture content to 60% of WHC. During incubation (at 30°C), the NH3
volatilized was determined. After various times, the unhydrolyzed urea, NH/, N02',
and N0 3', were analyzed.
Nitrapyrin did not have any effect on urea hydrolysis. CL-1580 and ATC reduced
the amount of hydrolyzed urea by 2-8% during the first 12 and 24 hours, but in 3 days
hydrolysis of urea became complete in both absence and presence of inhibitors.
Volatilization ofNH3 has intensified under the influence of inhibitors in the order:
nitrapyrin> ATC > CL-1580,
as they inhibited nitrification of NH/ (and accumulation of N0 2' and N0 3") in this
order:
Due to low water solubility ofnitrapyrin and CL-1580, the effect of higher inhibitor
rates could be studied only in the case of ATC. It was established that application of 250
and 500 f.lg of ATC/1O g soil did not have any significant effect on urea hydrolysis and
volatilization of urea-N as NH3 but appreciably increased the percentage inhibition of
nitrification.
In the laboratory and field experiments conducted by Ashworth et al. (1977) and
already mentioned on page 36, the effect of nitrapyrin (1.25 kglha) on urea hydrolysis
·Other investigators (e.g.. Knop, 1982) also found that nitrapyrin increased the volatile NH3 losses from
different urea-treated soils which indicates that nitrapyrin did not inhibit urease activity of these soils.
N
N
N
NH2
IN N.J..,.N
CI,c-O-CI H,c~.,)LNH2
N N
H,N~N..Jl- CCls
2-Chloro-6-(trichlorometl1yl)pyridine 2-Amino-4-chloro-6-metl1ylpyrimidine 2,4-Diamino-6-trichloromethyl os-triazine
(nitrapyrin; N-Serve) (AM) (CL-1580)
H,N-o-~-NJl~ q-NH2 NH 0 , N - Q -NH 2

P- 2
NO. NO.
Sulfatl1iazole (ST)
o-Nitroaniline m-Nitroaniline p-Nitroaniline
q-NH2 NH C I - Q -NH 2
P- 2
CI CI
o-Chloroaniline m-Chloroaniline p-Chloroaniline
?-NH-OC-C H 3 ?-NH-OC-CH3
~N)LN 2
0H ~.,)l-CI
o N
CI NO.
2-Aminopyridine 2-Chloropyridine 3-Chloroacetanilide m-Nitroacetanilide
OH
HSC ~ N"CH3
I
HC=C-C-CH3
'N-NO
H,c"
o-
6 H3
- 'NO
N-Nitrosodimetl1ylamine N-Metl1yl-N-nitrosoaniline 2-Metl1yl-3-butyn-2-o1
Figure 70. Structure of nitrification inhibitors tested by Bremner and Douglas (1971) for evaluation of their effect on soil urease activity.
223
was also studied, and it was found that nitrapyrin, like Na 2 CS 3 , (NH4hCS 3 , and CS 2 , had
no effect on urea hydrolysis.
A H:tJ-N--CH B
I II
HC~/N
Figure 71. Structure of 4-amino-l,2,4-triawle (A) and 3-amino-I,2,4-triazole (B).
In an experiment performed in India, Sahrawat (1977) treated 200-g samples of an

alluvial soil (sandy loam, pH 7.7) with a solution containing 20 mg ofurea-N and 0, 1,
2.5, 5 or 10% of biuret (on urea weight basis). During incubation (8 weeks at a mean
temperature of 28°C). the samples were moistened periodically to maintain them at 60%
of WHC and analyzed to determine their urea, NH/, N0 2 -, and NO)- contents. Urea
hydrolysis was complete in 7 days in all the samples, which indicates the lack of the
inhibitory effect of biuret on soil urease activity. Contrarily, biuret inhibited both phases
of nitrification (oxidation of both NH4 + and N0 2}
Abdel Hadi et al. (I980) studied the transformation of urea in two Egyptian soils
(1oanlY clay, pH 8.3 and calcareous sand, pH 7.9). Soil samples (25 g) were treated with
5 mg of urea in a solution also containing 0, 004, 2,4 or 8% biuret (on total N basis),
then incubated at constant moisture content (60% of WHC) at 30°C, and analyzed for
urea, NH/, N0 2-, and NO)- after 3,7,14,21, and 28 days of incubation. The analyses
showed that biuret did not affect urea hydrolysis as this was complete, in both absence
and presence of biuret, after 3 days (loamy clay) and 7 days (calcareous sand). Biuret
inhibited nitrification in loamy clay during the first 7 days and exhibited a weak
retarding effect on nitrification in the calcareous sand.
Guthrie and Bomke (1981) dealt with the effect of nitrapyrin and ATC on urea
hydrolysis in a Omadian silt soil and incubated the soil samples not at 30°C, as did
Bundy and Bremner (1974b), but at 2 and l20e, temperatures that are commonly found
in the soil during late autumn and early spring. The reaction mixtures were prepared
from 5 g of soil + I ml of aqueous solution containing 2 rug of urea + 0 or 10 or 100 Ilg
of nitrapyrin or ATe. The final water content was 40% by weight. These nitrification
inhibitor additions were equivalent to 2.5 and 25 kg inhibitorlha. During incubation that
lasted 21 days, the amount of residual urea was determined periodically. Urea
hydrolysis was almost complete within 21 days at 2°C and complete within 7 days at
l2°C in both absence and presence of nitrapyrin or ATC. Consequently, these
nitrification inhibitors did not affect soil urease activity.
The effect of nitrapyrin on urease activity in other Canadian soils was also studied
(Table 53). This effect was stimulatory rather than inhibitory on soil urease activity.
224
TABLE 53. Effect ofnitrapyrin on urease activity in Canadian soils

Significant effect (P<0.05) on
Amount ofnitrapyrin urease activity' Reference
Soil pH
(,",gig soil) Incubation time (days)
2 7 14
Sandy loam 7.6 10 N.D. o o Tu,1990
30 o N.D. o Tu, 1993a
40 N.D. I N.D. Tu, 1992b
100 S S S Tu, 1980
200 S S S Tu, 1980
Clay loam 7.2 30 S S N.D. Tu, 1981a
60 S S N.D. Tu, 1981a
Loamy sand 7.6 30 0 N.D. 0 Tu, 1993b
Organic soil 7.2 30 N.D. I S Tu, 1981b
60 N.D. S Tu, 1981b
Organic soil 6.8 10 N.D. 0 S Tu, 1990
's -Stimulatory effect. 1- Inhibitory effect. 0- No effect. N.D. -Not determined.
Khadzhiev (1985) reported that ATC added to irrigated grey soils under cotton
plantations in Uzbekistan effectively inhibited nitrification and, contrarily, increased
urease activity.
Raguotis and Shleinys (1986) introduced urea (90 or 180 kglba) with or without
nitrapyrin (1 or 2% relative to weight of urea) under the canopy in a pine forest.
Following fertilization, the volatilized ammonia was measured during 41 days.
Nitrapyrin at 1% rate did not reduce the volatile NH3 losses, but when it was applied
at 2% rate 14 and 25% reductions occurred in the cumulative NH3 losses from 90 and
180 kg urealha, respectively. This experiment carried out in 1980 was repeated in 1981,
when nitrapyrin had, even at its 2% rate, no significant reducing effect on the volatile
NH310sses.
Simon and Bergerova (1986) determined urease activity in samples of two soils
(chernozern, pH 7.4 and pararendzina, pH 7.8) from Slovakia. The samples were
previously incubated with or without urea (or ammonium sulfate) and with or without
nitrapyrin (2% relative to fertilizer N) at 28°C for 14 days. The results showed that
urease activity was slightly inhibited in samples previously treated with nitrapyrin
without fertilizers, but it was practically not affected in samples previously treated with
nitrapyrin + urea or nitrapyrin + (N~)2S04.
In a pot experiment, Mekhtiev et al. (1988) fertilized samples of a Moldavian soil
with P and K (each at a rate of 60 kglha) and urea (180 kg NIha) with or without
nitrapyrin (0.5 or 1% relative to urea-N). The control soil was fertilized only with P and
K. There were variants unsown and sown with a mixture of vetch + oats. Urease activity
assayed after three incubation times' gave the following mean values (mg NH3/g soil):
2.4 (control), 0.7 (urea), and 1.0 (urea + nitrapyrin) in the unsown variants, and 2.7
(control), 3.0 (urea), and 4.1 (urea + nitrapyrin) in the sown variants. This means that
nitrapyrin attenuated the negative effect of the high urea rate on urease activity in the
unsown soil and intensified the positive effect of urea on urease activity in the sown
"Length (in days) ofint;ubation times is not specified in the paper.

225
soil. In other words, nitrapyrin administered together with urea did not act inhibitorily
on soil urease activity.
All investigations mentioned above indicate that nitrapyrin, used at concentrations
efficient for inhibition of nitrification, does not inhibit urease activity in soil. This
conclusion should be compared with the result described by Reddy and Prasad (1975).
These investigators treated 200-g air-dried samples of a sandy clay loam soil (PH 7.8)
from India with urea (100 ppm N) with or without sulfathiazole (ST) or nitrapyrin (1 %
relative to urea-N). Then the samples were moistened to field capacity and incubated at
room temperature of 29°C for 4 weeks. At weekly intervals, the samples were analyzed
for residual urea, NH/, N02-, and N03-. ST had slight influence on urea hydrolysis.
Concerning nitrapyrin (N-Serve), the authors write: ''N-Serve showed considerable
retarding influence on the hydrolysis of urea in the present investigation but this needs
confirmation by other workers". But we should reiterate that this result was not
confirmed in other investigations.
The effect of ST on hydrolysis and nitrification of urea was also studied by
Muthuswamy et al. (1975) in pot experiments with red, black, and alluvial soils from
India. When urea granules coated with tar (from different coals) and ST were used
instead of uncoated urea, formation of both NH4 + and N03- was reduced and,
consequently, the N loss was decreased.
We mention here that the isomer of ATC, 3-arnino-l,2,4-triazole (the herbicide
amitrole; Figure 71) was also evaluated as an inhibitor of soil urease activity.
Gauthier et of. (1976) used three clay soils from Quebec. The soil samples were
arranged in pots (45 cm high x 23 cm diameter), each preserving the natural layering.
Soybeans (Glycine max) were planted in each pot and, after 3 weeks, the soil received
10 ml of 2.10-3 , 2.10-4, 2.10-5 or 2.10-6 M amitrole solution. The control soil was not
treated with arnitrole. After 1 week, samples were collected from the pots, 2.5 cm below
the surface (S samples) and 5 cm from the bottom, i.e. 40 cm below surface (deep, D,
samples), and their urease activity was assayed. The results showed that amitrole
inhibited soil urease activity. There was a linear relationship between percentage
inhibition and arnitrole concentration. The inhibition was stronger in the S than in the D
samples. Thus, arnitrole applied at 2.10-3 and 2.10-4 M concentrations broUght about
complete inhibition of urease activity in Sand D samples of two soils and inhibitions of
82 and 74% in S samples and of 68 and 63% in D samples of the third soil. Amitrole at
2.10-5 and 2.10-6 M concentrations reduced urease activity by 56-38% in S samples and
by 63-17% in D samples of the three soils studied. The only exception was sample D of
the third soil in which urease activity was not inhibited by the lowest amitrole
concentration. Further observations were that amitrole also manifested nematocidal
effect and, unfortunately, inhibited nodulation on soybean roots.
The effect of amitrole on urea hydrolysis in soil was also studied by Vlek et al.
(1980). Prilled urea alone (at a rate of 100 kg N/ha) or in a mixture with arnitrole (2%
on urea weight basis) was incorporated into a silt loam soil placed in pots and then
flooded (depth of floodwater = 5 cm). During incubation (10 days at 35°C day and 25°C
night), the floodwater was analyzed daily for residual urea. After 0 and 1-6 days, the
following values were registered: 140, 71, 29, 5, 1, 0, and 0 mg of urea-Nil,
reo;pectively, in the control soil (not treated with amitrole), and 137, 67, 22, 5, 2, 1, and
omg of urea-Nil, respectively, in the amitrole-treated soil. It is evident that in these pot
226
experiments, contrary to those of Gauthier et al. (1976), amitrole had practically no

effect on urea hydrolysis.
Rodgers ef al. (1984) injected a urea solution (at a rate of 375 kg N/ha) to 10-cm
depth of a clay loam soil covered by ryegrass ley. In other treatments, a nitrification
inhibitor, namely nitrapyrin or 5-ethoxy-3-trichloromethyl-l,2,4-thiadiazole
(etridiazole, terradiazole, Dwell; Figure 72) in an amount of 1.5 kglha was also added to
the urea solution. The treatments were carried out in either winter or spring of 1981,
N--C-CCla
II II
CHa-CH2-o--C, ..,.....N
S
Figure 72. Structure of 5·ethoxy·3-trichlorom:thyl·I,2.4-thiadiazole (etridiazole,

terradiazole, Dwell).
1982, and 1983. In all treatments, urea was completely hydrolyzed in 7-14 days after
injection; the two nitrification inhibitors did not slow the rate of urea hydrolysis.
This finding is in concordance with the results obtained by Abdullatif and Stroehlein
(1990) in a study with three Arizona soils (sandy loam, pH 7.2, loam, pH 7.8, and
loamy sand, pH 7.7). Soil samples (50 g), treated with urea only (at a rate equivalent to
224 kg N/ha) or with urea and etridiazole (1.27 kglha), were incubated at 33°C and their
pH was measured at intervals of 0, 1, 2, 3, 4, 6, 8, and 16 days. Changes in pH were
similar in untreated and etridiazole-treated samples which was interpreted as evidence
proving that etridiazole did not affect urea hydrolysis, did not reduce urease activity. In
another experiment with the same three soils, etridiazole was found to inhibit
nitrification of urea and ammonium sulfate for 10-15 days.
Bremner (1986) mentioned that 2-ethynylpyridine and phenylacetylene (Figure 73),
which proved to be the most effective nitrification inhibitors among 15 monosubstituted
(RC=CH) and 6 disubstituted (RC=CR) acetylenes tested in different <NH4hS04-
treated Iowa soils, had no significant effect on urea hydrolysis in soil. In another study
O-C5CH
N
B
o-C=CH
Figure 73. Structure of2-ethynylpyridine (A) and phenylacetylene (B).
(McCarty and Bremner, 1990a), it is specified that in reaction mixtures (5 g of air-dried

soil + 2 ml of solution containing 10 mg of urea + 0, 50 or 500 J.lg of 2-ethyny1pyridine)
incubated at 20°C for 7 days, urea hydrolysis was not inhibited by the lower 2-
ethynylpyridine rate in any of the three soils studied, whereas at the higher 2-
ethynylpyridine rate the inhibitions were 0, 5, and 6%, respectively.
227
De Boer et al. (1989) observed that nitrapyrin completely inhibited nitrification in

suspensions of fertilized, acid (PH 4.1) heath soil sampled in the central area of the
Netherlands. As the ammonium-oxidizing autotrophic bacteria living in this acid soil
are able to hydrolyze urea at pH<5.0, elimination of these bacteria by nitrapyrin results
in the loss of both nitrification and urea-hydrolyzing capacities.
In laboratory experiments described by Pisareva and Muravin (1988) and Pisareva
(1989), 3-methylpyrazole-l-carboxamide (MPC; l-carbamoyl-3-methylpyrazole; Figure
r T CH3
"'-N. . . N
ICONH2
Figure 74. Structure of3-methylpyrazole-l-carboxamide.
74) added to 100-g samples of three soils (soddy podzol, common chernozem, and
calcareous chemozem) had no effect on urea hydrolysis but inhibited nitrification of
NH4 + released from urea. The samples received 20 mg of urea-N with or without 2%
MPC (on urea-N basis) and water to 60% of WHC and were incubated at 28°C. Their
residual urea content was determined after 6-72 hours of incubation. For NH4 + and
N0 3 -, the samples were analyzed after 15,30, and 45 days of incubation.
Similar results were registered by McCarty and Bremner (1990b), who treated
samples of three Iowa soils with MPC at a rate of 10 or 50 ~glg soil, and also by Popov
et al. (1990), who carried out a pot experiment using a grey forest soil, fertilized with
NPK, treated with MPC (1.6 ~glg soil) and sown with maize.
Shcherbakov and Stakhurlova (1990) studied the effect of MPC on nitrification and
urease activity in microplots on a leached chernozem. Urea was applied with or without
MPC in bands at 10-cm depth. Urea rate was equivalent to 90 kg of Nlha, whereas that
of MPC was 3% relative to urea-No The soil was seeded to cucumber. MPC inhibited
nitrification for approximatively 4 weeks. Contrarily, urease activity in the period of 8-
32 days after fertilization showed higher values in the urea + MPC treatment than in the
urea-only treatment. The increased urease activity, which should be ascribed to
microbial synthesis of new urease molecules, indicates that MPC did not inhibit activity
and microbial synthesis of urease.
In Kucharski's (1991) experiment, 50-g air-dry samples of a loamy sand soil (PH
6.7) from Poland were amended with urea (10 mg N) and nitrapyrin, A TC, and MPC at
rates of 0,0.35,0.70, 1.05,2.10, and 3.15% relative to urea-No The soil moisture was
kept at 60% ofWHC during the incubation (120 days) at 20°C. Analyses ofNH4 +-N and
N0 3--N at days 10,20,30,60,90, and 120 showed that none of the three nitrification
inhibitors at any rate inhibited hydrolysis of urea.
In the pot experiments of Khabirov et al. (1992) (see page 58), besides thiram, five
pyridine derivatives, all being nitrification inhibitors (Figure 75), were also tested for
evaluation of their effect on urease activity in a leached chernozem. The experimental
conditions were identical to those under which thiram was tested.
228
H3C---H2C~ H3C- (H~)~(CH~~CH3

~.,)J-cH3
N ~.,)L(CH~:r;H3
N
2-Methyl-5-ethylpyridine 2-n-Buty~3,5-di- n-pro pylpyrid ne

(A) (B)
CH3
'''~'" N
2,4,6-Trimethylpyridine 2-n-Propyl-3,5-c1ethylpyridine
(C) (0)
HaC- H
2-Pheny~3,5-diethylpyridine
(E)
Figure 75. Structure of the nitrification inhibitors tested by Khabirov et al. (1992) for evaluation of their effect
on soil urease activity.
Compound A, 2-methyl-5-ethylpyridine, which is the most effective nitrification

inhibitor among the five pyridine derivatives tested as well as compound B, had a weak:
inhibitory effect on urease activity, whereas compound C did not affect and compounds
D and E increased urease activity.
5.1.2 . .t;nect of Sodium and Potassium Azide on Soil Urease Activity

Sodium azide (NaN 3 ) exhibited only negligible «1 %) inhibitions in urease activity of
two soils, under the conditions of the 5-hour test. Rate of NaN 3 was 50 ppm on soil
weight basis (Bremner and Douglas, 1971).
Bremner and Bundy (1976) proved that potassium azide (KN3) has a weak and
short-lasting inhibitory effect on urea hydrolysis in soiL They used four soils having pH
between 6.8 and 7.9. Two of these soils, following their treatment with urea without
KN 3, did not produce appreciable amounts of N0 2- during incubation. Under similar
conditions in the other two soils, accumulation of a higher amount ofN0 2- occurred.
Soil samples (lO g) were treated with 1 rnl of urea solution (4 mg ofN) and 2 rnl of
water or 2 ml of aqueous solution containing 100,250 or 500 J.1g ofKN3 • The samples,
whose moisture content was brought to 50% of WHC, were incubated at 30°C.
Determination of residual urea showed that KN3 , depending on its rate, inhibited urea
hydrolysis in the four soil studied in the following proportions: 5-9, 10-19, 8-25, and 3-
11 %, respectively, during the first 12 hours of incubation. The inhibitions became
smaller or remained practically unchanged after 24 hours, namely: 0, 11-20,0-7, and 2-
12, respectively. At 3 days, the inhibitory effect of KN3 disappeared as urea hydrolysis
was complete in each soiL After 14 days, it was established that in those two soils,
229
which in absence ofKN 3 did not produce N0 2- from urea in appreciable amounts, KN3
strongly inhibited nitrification, whereas in the other two soils, which in absence of KN 3
accumulated more N0 2-, KN3 did not have any significant inhibitory effect on
nitrification ofurea-N as it did not decrease accumulation and maximum concentrations
of N0 2-. It is assumed that the lack of inhibitory effect on nitrification is due to
decomposition ofKN3 by its reaction with N0 2-:
In addition, in acidic soils KN3 may hydrolyze with formation of volatile hydrazoic
acid':
KN3 + H;P - KOH + HN3.
In comparison with nitrapyrin, KN3 is a weaker nitrification inhibitor.
5.1. 3. F;fJect qfDicyandiamide on Soil Urease Activity

Calcium cyanamide ("lime nitrogen") is an N fertilizer, from which cyanamide is
released in soil. Cyanamide is then transformed to urea. Cyanamide can also give rise in
soil to other products: dicyandiamide (dimeric form of cyanamide, cyanoguanidine,
DCD)*', guanylurea, and guanidine which all are decomposed to urea (Figure 76).
NH
II
Ca=N-C:=N H2N-C-NH -C=N
Calcium cyanamide Cyanamide Dicyandiamide (DCD)
NH a NH
II II II
H2N-C-NH -C-NH2 H2N- C- NH 2
Guanylurea Guanidine
Figure 76. Structure of calcium cyanamide and of products resulting from its transformations in soils.
Finally, urea is hydrolyzed and the released ~ + can be nitrified. DCD, produced
by chemical synthesis (didin) and added to soil, acts, at the beginning, as a nitrification
inhibitor. Then it is slowly decomposed and mineralized with formation of plant-
'Hydrazoic acid is not only volatile, but it is also poisonous and spontaneously explosive. Its explosion hazard
increases in the presence ofHg and Ag salts. Thus, mercuric azide formed in soil is detonated even under the
action ofa fiiction (Rozicky and Bartha, 1981).
•• Guanylthiourea can also give rise in soil to dicyandiamide (see page 75).
230
available N and concomitant loss of inhibitory capacity. Consequently, DCD,

containing 67% N by weight, is both a nitrification inhibitor and slow-release N
fertilizer (e.g.. Amberger and Gutser, 1978; Amberger and Vilsmeier, 1979; Rodgers et
01., 1985; Amberger, 1989).
Using four Polish soils (two active and two less active as concerns nitrification
capacity), Ostromecka (1986) treated air-dried soil samples (100 g) with urea (10 mg of
N) with or without DCD (5% relative to urea-N) and water to 50% of WHC. During
incubation (22°C/45 days), the ~ + and NO)- contents were determined periodically.
The results showed that DCD had no effect on urea hydrolysis; it inhibited nitrification
to a large extent in the active soils and nearly completely in the less active soils.
Amberger and Vilsmeier (1979) tested the effect of DCD, guanylurea, and guanidine
on urease activity in four German soils. Nitrite served for comparison. The reaction
mixtures had the following composition: 100 g of air-dry soil + 20 mg of N as urea + 10
mg of N in form of the test compound + water to 40% of WHC. Incubation was carried
out at 5°C for 5 days, during which the residual urea was determined daily. The results
indicated that in each soil DCD and guanidine did not inhibit urea hydrolysis but, quite
contrarily, increased it by 5-15%. Guanylurea brought about a weak inhibition, whereas
nitrite decreased considerably the urea hydrolysis in each soil; the decrease was greatest
(50%) in a loamy sand and smallest (25%) in a silt loam.
In another experiment, it was established that N in cyanamide and urea was
completely transformed to NH4 + in 5 days, at temperatures of 5, 10, 20, and 30°C. This
finding proves that the cyanamide molecules not transformed yet to urea molecules did
not inhibit hydrolysis of those urea molecules that already appeared as a result of
transformation of some cyanamide molecules.
These results do not confirm the assumption of Sommer and Rossig (1978),
according to which the inhibitory effect of DCD on soil urease activity would explain
depressions in spring wheat yields in plots on a sandy loam that had been treated with
urea + DCD, in comparison to yields obtained in other plots of the same soil that had
been fertilized with aqueous ammonia with addition ofnitrapyrin.
Rodgers (1983) studied the effect of DCD on urea hydrolysis in three English soils
(silty clay loam, clay, and loamy sand). Two experiments were performed. In the first
experiment, 50-g air-dried soil samples were moistened with 10 ml of distilled water;
then on their surface urea prills or DCD-containing urea prills were placed. Rate of urea
application: 50 mg ofN, and that of DCD: 7.2% relative to urea weight. Incubation was
carried out at 30°C and lasted 4 weeks. At weekly intervals, the amounts of volatilized
NH), residual urea, NH/, N0 2-, and NO)- were recorded.
In the second experiment, urea or urea-DCD prills were covered with 50 g of air-
dried soil forming a 2-cm layer, then moistened with 10 ml of water and incubated at
30°C for 4 weeks. Following incubation, the same analyses were made as in the first
experiment.
The analytical data indicated that hydrolysis of urea was complete in 7 days in each
soil in both absence and presence of DCD. This means that DCD did not inhibit soil
urease activity. Of course, DCD inhibited nitrification ofN~ +. At the same time, DCD
enhanced volatilization of ammonia from two of the three soils studied, the NH3 lost by
volatilization having been higher by 20-68% compared to treatment with urea alone.
More NH.1 volatilized from the loamy sand than from the clay, and in the first
231
experiment (surface application of urea or urea-DCD) than in the second experiment

(incorporation of urea or urea-DCD into soil).
The conclusion was drawn that in some soils the positive effect of DCD by
inhibition of nitrification may be counteracted by its increasing effect on NH3
volatilization.
In other experiments, conducted by Rodgers et al. (1984), under field conditions on
a clay loam and a silty clay loam covered by ryegrass leys at Rothamsted, DCD added
to urea prills inhibited nitrification but did not reduce the rate of urea hydrolysis and did
not increase ammonia volatilization. DCD and urea were applied at a rate of 37 kg
DCD-N and 338 kg urea-Nlha as a single dressing or as three equally divided dressings.
Rodgers et at. (1986) conducted similar field experiments on two silty clay loams
cultivated with winter oil-seed rape. DCD did not delay urea hydrolysis irrespective of
the mode of urea and urea-DCD application (single dressing or two equally divided
dressings). Ammonia volatilization was reduced or not affected by DCD when urea-
DCD, like urea, was applied as a single dressing, but it was always increased by DCD
following divided application of fertilizer.
In the 2-year field experiment conducted by Rodgers et al. (1987) on a light sandy
soil under a grass ley at Woburn Experimental Farm, Bedfordshire (see page 125), there
were also plots fertilized by broadcast application of prilled urea (375 kg Nlha) or
prilled urea (338 kg Nlha) plus DCD (37 kg Nlha). Eleven days after broadcasting the
fertilizers, no urea-N but more N0 3--N were present in the urea-only treatment and no
urea-N but less N0 3--N were recovered in the urea+DCD treatment. These findings
indicate that DCD did not inhibit urease activity but inhibited nitrification.
According to Khadzhiev (1985), DCD like ATC inhibited nitrification but increased
urease activity in cotton soils (see also page 224).
The stimulating effect of DCD on volatilization of ammonia from urea (which
indicates absence of the inhibitory effect of DCD on soil urease activity) was also
observed by Prakasa Rao and Puttanna (1987), when in field experiments urea (187 kg
NIha) was applied together with DCD (15 and 20% relative to ureac N) on the surface of
an Indian sandy loam soil (PH 7.3) cultivated with Java citronella (Cymbopogon
winterianus), a perennial, aromatic grass. When the urea + DCD mixture was
introduced to soil at 5-cm depth, the nitrogen losses by NH3 volatilization became very
small. In pots filled with the same soil, DCD (20 ppm on soil weight basis), applied
with urea (100 ppm), increased and prolonged volatilization ofNH3 from urea up to 105
days, whereas in absence of DeD the NH3 volatilization loss was negligible even after
15 days. At the same time, effectiveness of DCD (15 and 20 ppm) in inhibiting
nitrification was confirmed.
The effect of DCD on urea hydrolysis was also studied with a loamy soil (PH 5.3)
under a banana plantation in Costa Rica (Vilsmeier et al., 1987). Air-dry soil samples
(50 g), treated with urea (10 or 30 mg of N) with or without DCD (1 and 3 mg of N,
respectively), were moistened and incubated at 20 and 30D e for periods of up to 8
weeks. It was established that urea was completely hydrolyzed in two weeks, at both 30
and 20D C and in both absence and presence of DCD. This means that DeD did not
inhibit urea hydrolysis.
In Georgian (Gruzian) red soils, fertilized with PK and urea and cultivated with
subtropical plants, DCD enhanced urease activity. This is the conclusion drawn by
232
Sanikidze et al. (1987a) based on an experiment carried out in lysimeters during a 3-

year period. DCD waf> applied at rates of 10 and 15% relative to weight of urea.
McCarty and Bremner (1989a) found that DCD had very little, if any, effect on urea
hydrolysis. Air-dried samples (5 g) of three Iowa soils, treated with 2 ml of solution
containing 10 mg of urea and 0, 5, 50, 125, 250 or 500 Ilg of DCD, were incubated at
20°C for 7 days and then analyzed for residual urea. None of the DCD amounts affected
urea hydrolysis in one soil, and the highest DCD rate caused only 1-2% inhibitions of
urea hydrolysis in the other two soils.
In a pot experiment described by Popov et al. (1990), samples (10 kg) of a grey
forest soil were fertilized with PK and urea (8 mg of Nil 00 g of soil) with or without
10% of DCD (relative to urea-N) and sown with maize. Urease activity determined
periodically during the growing season showed only slight differences between the
values recorded in the urea-treated and urea+DCD-treated pots.
According to Clay et at. (1990), DCD had no effect on volatilization of ammonia
from urea-treated samples of a Minnesota sandy loam soil and when the soil in the field
was covered with maize residue, but it had an increasing effect on NH3 volatilization
when the urea was applied on bare soil (see also page 244).
In the experiment of Blaise et al. (1997), 100-g samples of a loess brown earth were
amended with 10 mg of N as urea or with 9 mg of N as urea + 1 mg of N as DCD and
incubated at 20°C for 30 days. The volatile ammonia losses were higher from
urea+DCD-treated samples than from the urea-treated ones. This finding indicates, as
did similar findings already cited in this section, that DCD does not inhibit urease
activity.
Surprisingly, in a periodically water-logged paddy soil, calcium cyanamide and
DCD exhibited inhibitory effect on urease activity (Xue and Li, 1987). Soil samples (5
g) were incubated with 10 rn1 of 1% urea solution, containing or not containing 0.1 or
0.2 g of Ca cyanamide, or 0.05 or 0.1 g of DCD, at 37°C for 48 hours. Inhibition of
urease activity was 30.84 and 75.70%, respectively, under the influence of Ca
cyanamide, and 27.10 and 33.64%, respectively, under the influence of DCD. It was
also established that the inhibition was evident even after 12 days.
In a 2-year experiment (1983-1984) on a flooded rice field (on silty clay loam, pH
7.6), Chauhan and Mishra (1989) studied the prilled urea (PU) in comparison with neem
cake-coated urea (NCCU), shell-lac-coated urea (LCU), dicyandiamide-treated urea
(DTU), and urea supergranule (USG) applied at rates of 40, 80, and 120 kg Nlha.
NCCU was prepared by mixing prilled urea with finely powdered neem seed cake,
coaltar, and kerosene in a proportion of 100:30:3:1, respectively. Dicyandiamide was
mixed with prilled urea in 1:9 ratio on N basis. Fifty per cent of fertilizer N except USG
was applied as basal dressing through incorporation in the soil during puddling. The
remaining 50% of NCCU, LCD and DTU, and 25% ofPU were applied as top dressing
at tillering stage. The remaining 25% of PU was topdressed at panicle initiation stage.
The total amount of USG was placed in the soil at 10-cm depth 7 days after
transplantation of rice seedlings.
Ammonia volatilization was measured during 3 weeks after both basal and top
dressings. The cumulative NHJ losses, expressed as percentages of the applied N, varied
depending on fertilizer rate and experimental year between the following limits: 19.0-
23.6 (PU), 12.0-15.3 (DTU), 7.6-9.4 (LCU), 6.1-8.0 (NCCU), and 0.5-1.4 (USG). These
values indicate that dicyandiamide reduced the volatile NH3 losses, but to a lesser extent
233
than did the neem cake- and lac-coatings. The loss was lowest from the urea
supergranule.
Some of these investigations are also referred to by Sahrawat (1989) and Prasad and
Power (1995).
5.2. UREASE INHIBITORS ALSO POSSESSING NITRIFICATION-INHIBITING

CAPACITY
5.2.1. Inorganic Compounds
5.2.1.1. Heavy Metal Compounds

Daif and van Beusichem (1981) studied the effect of three heavy metallic ions on urease
activity of eight soils (see page 10). One of these soils, namely a sandy loam (PH 5.9),
was selected for studying the effect of Fe2+, Cu 2+, and Zn2 + on nitrification. Rate of
metallic ion addition was 20 J.lg/g soil. After 14 days of incubation at 20°e. the (N0 2' +
N0 3') content was determined. It was found that Cu2+, which was most inhibitory on
urease activity, was also the only metallic ion possessing nitrification-inhibiting
capacity.
In the experiment of Skujins et at. (1986), CrCI 3 , in comparison with CUC\Z, was a
stronger inhibitor of urease activity in samples of a forest soil (see page 14). The two
heavy metal salts acted similarly on the nitrification, except in the two lowest rates of
Cu2+ (50 and 200 J.lg/g soil) which had a slight stimulatory effect on nitrification.
In the pot experiment of Badura et al. (1986), urease activity in samples of a forest
soil was not affected by Zn2 + and Cd2+ used separately at a rate of 5,000 ppm or in
mixture at 2,500 ppm + 2,500 ppm (see page 14). But when used at these rates, they
inhibited both phases of nitrification: phase I was inhibited in the order Zn 2 + > Cd2+ ~
Zn2+ + Cd2+ and phase II in the order Cd2+ ~ Zn2+ + Cd2+ > Zn2+.
The pot experiment performed by Kandeler et al. (1990) (see page 15) made it
possible to establish that the inhibiting effect of heavy metal salts was markedly
stronger on urease activity in a sandy loam and a little weaker on urease activity in a
clay loam than was their inhibiting effect on nitrification.
Gupta and Chaudhry (1994) studied the effect of four heavy metals on both urea
hydrolysis (see page 16) and nitrification in a Hisar soil. Nitrification was inhibited by
the heavy metals in the same order as was urea hydrolysis. For example, when the soil
samples were amended with 100 mg of urea-N and 400 mg of heavy meta1lkg soil and
incubated at 30°C for 14 days, the following amounts of N0 3'-N (mg/kg soil) were
found in the treated and untreated (control) samples: 55.7 (Hg), 67.7 (Zn), 72.0 (Ni),
84.2 (Pb), and 84.2 (control).
Agricultural grade pyrite, which in experiments of Blaise et al. (1996, 1997) reduced
the volatile N losses as ammonia and nitrous oxide from urea-treated soil samples (see
page 18), was also found to inhibit nitrification ofurea-N (Blaise et al., 1997). Samples
(100 g) of a loess brown earth (pH 6.8) were amended with 10 mg of N as urea and 0 or
0.01 g up to 0.1 g of pyrite. DCD was also used as a reference compound in mixtures of
100 g soil + 9 mg N as urea + 1 mg N as DCD. All mixtures, moistened to 60% of
WHC, were incubated at 20°C and analyzed for N0 2'-N and N0 3'-N after 3, 6, 9, 15,
and 30 days.
234
Percent inhibition of nitrification increased with the rate of pyrite and decreased
with incubation time. After 30 days of incubation, inhibition of nitrification was 40.3%
at the highest pyrite rate and 55.9% in the DCD treatment.
5.2.1.2. Salls ofAlkali Metals and Alkaline Earth Metals

Vostal et af. (1975,1976) amended 100-g samples of two sandy loam soils (PH 4.4 and
4.7) and two sandy soils (pH 5.0 and 7.3) with urea (30 mg N) + P salt (13.2 mg) + K
salt (33 mg). Superphosphate, Kl HP04, C<t(H1P04 )2, KCl, K2S04 were applied in
different combinations. No salts were added to the control. The samples were moistened
to 60% ofWHC and incubated at 25°C. After 2,7,14,21, and 28 days of incubation the
samples were analyzed for ammonium, nitrite, and nitrate contents.
Superphosphate in combination with KCl or K1 S04 inhibited both hydrolysis and
ni trification of urea in the acid soils and only the formation of nitrite in the alkaline soil.
Similar results were obtained when the acid soils were amended with Ca(H2P04h
instead of superphosphate.
For comparing the effects of 12 salts, namely Na, K, C<t, and Mg chlorides, sulfates,
and carbonates on hydrolysis and nitrification of urea, Elkholei et af. (1982) treated
samples of an Egyptian alluvial soil with urea and solution of these salts having
concentrations of 40, 60, 80, and 100 milliequivalents/l. Following incubation and
analysis of the samples it was found that nitrification of urea was more sensitive than its
hydrolysis to high salt concentrations. The inhibitory effect increased in the order: Na+
> K+ > Mg2+ > Cal +, and cr > CO/' > sol'.
El-Shahawy and Mashbady (1984) studied the effect of NaCI and CaClz on urease
activity and nitrification in an Egyptian soil (sandy clay loam, pH 7.4). The air-dried
soil samples (300 g) were treated with equivalent mixtures of NaCI and CaCl l , obtained
four total salt concentrations: 0.114, 0.228, 0.456, and 0.912% (on soil weight basis).
Addition of urea (200 ppm N) and distilled water to 60% of WHC was followed by
incubation at 30°C for 12 weeks, during which the contents of NH/, NO)', and NO l '
were detem1ined at weekly intervals. The results showed that hydrolysis of urea was
complete after the first week in the untreated samples and in those treated with low salt
concentrations (0.l14 and 0.228%) and required 3 and 4 weeks, respectively, in samples
treated with the two higher salt concentrations. At the same time, the two lower salt
concentrations resulted in delaying nitrification for a period of 1-5 weeks over the
untreated soil, whereas the two higher salt concentrations caused complete inhibition of
nitrification for 5 and, at least 12, weeks, respectively.
Agrawal et al. (1985) amended samples of three Indian soils (an acidic, a neutral,
and a calcareous soil) with urea (70 mg N/kg soil) alone or together with KCl (40 mg
Klkg soil). Inhibitions of nitrification in the three soils were 22, 20, and 18%,
respectively, after 1 week of incubation at 27°C, and 14, 12, and 8%, respectively, after
4 weeks.
5.2.1.3. Fluorides
Sodium fluoride, which was patented as an inhibitor of soil urease actlVlty (see
Subchapter 1.5), is also able to inhibit nitrification in soil (Baumgartner and Otlow,
1985).
235
5.2.1.4. Sulfur Compounds

As shown in Subchapter 1.7, the inorganic sulfur compounds tested for evaluation of
their effect on soil urease activity inhibit nitrification, and they were found to be
stronger nitrification than urease inhibitors. It should be mentioned here that ATS,
inhibiting the second phase of nitrification, may have an undesirable effect: nitrite
accumulation (Janzen and Bettany, 1986).
5.2.2. Organic Compounds
5.2.2.1. Organic Mercury Compounds

In Moe's (1967) laboratory experiment (see page 43), p-chloromercuribenzoate
(PCMB) reduced not only the rate of urea hydrolysis but also inhibited nitrification.
Bundy and Bremner (1973b) prepared reaction mixtures from 10-g air -dried samples
of two Iowa soils (clay loam, pH 7.6 and clay loam, pH 7.3), 1 ml of solution containing
2 mg of nitrifiable N as (N"lL)zS04 or urea, 2 rnl of water or 2 rnl of solution containing
100 /1g of phenylmercuric acetate (PMA) and water to bring the soil moisture content to
60% of WHC. Following incubation at 30°C for 14 days, the reaction mixtures were
analyzed for their (NOz' + N0 3')-N contents. Inhibition of nitrification of ammonium-N
and urea-N was 1 and 2%, respectively, in the pH 7.6 clay loam and 31 and 38%,
respectively, in the other clay loam.
In another experiment. Bundy and Bremner (l974a) added 100 and 500 /1g PMA or
sodium PCMB to lO-g air-dried samples of the pH 7.6 and 7.3 clay loams and a sandy
clay loam (PH 7.2). The nitrifiable N was ammonium sulfate. The other experimental
conditions were the same as those mentioned in the preceding paragraph. Inhibition of
nitrification by the higher rate of PMA and PCMB was 88 and 5% (PH 7.6 clay loam),
86 and 32% (pH 7.3 clay loam), and 95 and 70% (sandy clay loam), respectively. Thus,
PMA was more effective than PCMB.
5.2.2.2. Urea Derivatives

Thiourea (TU) is an inhibitor of both soil urease activity and nitrification. Its effect on
nitrification is more long-lasting than that on urease activity (Malhi and Nyborg, 1979,
1984).
Moawad e1 at. (1984), who proved the urease-inhibiting effect ofTU in a Nile Delta
soil (see page 51), also found that TU inhibited nitrification as well.
Invcstigators from the National Fertilizer Development Center (Tennessee Valley
Authority, Muscle Shoals, Alabama) (Anonymous, 1983; Gautney et al., 1983, 1984,
1985) emphasize that, besides the capacity to inhibit urease activity and nitrification,
TU has other advantageous properties, nanlely: it can be cogranulated with urea; it is
compatible with anhydrous and aqueous an1illonia and a wide variety of fluid fertilizers;
the relative cost of adding 2 % TU to urea (on N basis) is only 18% more than the cost of
urea alone. It should be mentioned that technology for cogranulation of TU with urea
was elaborated by Gautney et af. (1983, 1984).
Bock and Williams (1984), collaborators of the Tennessee Valley Authority,
evaluated the effects of TU on urea hydrolysis, ammonia volatilization, and nitrification
in a laboratory experiment with samples of four soils. Urea alone and urea amended
with 2.5-7.5% or 11-33% TU-N were surface-applied on soil samples which were then
incubated at 25°C and moisture contents near field capacity. It was found that amending
236
urea with 2.5-7.5% TU-N either increased or had no effect on NH3 volatilization,
because nitrification was inhibited but rate of urea hydrolysis was not reduced.
Amending urea with 11-33% TU-N reduced NH3 volatilization between 11 and 94%,
because urea hydrolysis rate was reduced.
Germann-Bauer (1987), who proved that guanyithiourea (GTU) inhibits soil urease
activity (see page 52), also established that GTU is a stronger inhibitor of nitrification
than of the soil urease activity. One of the abiotically catalyzed transformations of GTU
in soil is dicyandiamide, a potent nitrification inhibitor (see also Amberger, 1989).
5.2.2.3. Thiuram Disu(fides

In the pot experiments of Khabirov et at. (1992), tetramethyithiuram disulfide (thiram)
inhibited urease activity in samples of a leached chernozem amended with 500 mg of
urea-N/kg soil and 0 or 10% thiram (on urea-N basis) and analyzed after 1, 2, and 4
weeks of incubation at ambient temperature (see page 58). Now we add that in these
experiments thiram also inhibited nitrification. Percent inhibition of nitrification after I,
2, and 4 weeks was 64, 68, and 63%, respectively.
5.2.2.4. Xanthates
Ashworth et of. (1979) found that potassium ethyl xanthate and cellulose xanthate
inhibited nitrification more markedly than ureolysis (see page 58). The data published
by Ashworth et 01. (1980) confirm the inhibitory effect of some xanthates on both
urease activity and nitrification.
5.2.2.5. Heterocyclic Sulfur Compounds

As shown on page 71, Held et at. (1974) patented tetrahydro-l,3,5-thiadiazine-2-
thiones, including dazomet, as inhibitors of soil urease activity. Now we add that these
compounds were patented by Held et at. (1974) also as nitrification inhibitors.
According to the example given in the patent description, dazomet induced a 43%
inhibition in nitrification ofurea-N when soil samples moistened to 50% ofWHC were
treated with urea amended with 2% dazomet (on urea weight basis) and incubated at
30°C for 1() days.
In the field experiment of Tu et of. (1995), the soil was treated with 0 or 56 kg
dazometlha on May 10, 1978, and sampled for analyses on May 23 and June 27 (see
page 72). The effect of dazomet was weaker on urease activity than on nitrification: due
to the dazomet treatment, decrease of urease activity was insignificant, while that of the
nitrification was significant on May 23, and the insignificant increase of urease activity
was accompanied by a significant increase in nitrification on June 27.
5.2.2.6. Monohvdric Phenols

Inhibitory eff~t of nitrophenols on soil urease activity was weak (Bremner and
Douglas, 1971) (see page 79) or relatively strong (Shen et 01., 1997) (see page 82).
Putt anna et of. (1999) found that 2-nitrophenol, added to samples of an Indian sandy
loam soil (pH 8.3) at a rate of 10 Ilg/g soil, effectively inhibited nitrification of urea-N
(l00 Ilg/g soil) during incubation of samples at 30°C for 30 days.
237
5.2.2.7. Polyhydric Phenols and Quinones

Bundy and Bremner (1974a) tested the effect of two polyhydric phenols and six
quinones on nitrification in three Iowa soils. For comparison three nitrification
inhibitors were used. The reaction mixtures (lOg of air-dried soil + I ml of a solution
containing 2 mg of NH4 +-N as ~)2S04 + 2 ml of water or aqueous solution
containing 100 or 500 Ilg of test compound + water up to 60% of WHC) were incubated
aerobically at 30°C for 14 days, then analyzed for NH/, NO}-, and N0 2-. Based on the
(NO}- + N0 2-)-N contents, percent inhibition of nitrification was calculated.
TABLE 54. Effect of eight urease inhibitors on nitrification in soils, as compared to that of three nitrification
inhibitors"
Inhibition of nitrification (%)
Amount of
Clay Clay Sandy
Compound compound
loam loam clay loam Average
(flglg soil)
~H 7.6 EH7.3 EH7.2
Urease inhibitors:
Catechol !O 3 0 5 4
50 4 8 20 II
Hydroquinone 10 2 7 11 7
50 4 54 71 43
50 3 56 77 45
2,3-Dimethyl-BQ to 2 8 4
50 3 43 72 39
2,5-Dimethyl-BQ 10 2 10 17 10
50 30 83 96 70
2,6-Dimethyl-BQ 10 I 6 13 7
50 27 87 94 69
2,S-Dichloro-BQ 10 2 4 7 4
50 3 32 52 29
2,6-Dichloro-BQ !O 2 3 16 7
50 3 38 46 29
Nitrification inhibitors:
Nitrapyrin to 69 85 96 83
2-Amino-4-chloro-6-methylpyrimidine (AM) 10 1 25 68 31
Sultathiazole (ST) 10 1 24 40 22
"Adapted from Bundy and Bremner (1974a), by permission of Pergamon Press PLC.
Table 54 shows that the eight urease inhibitors tested had little effect on nitrification
when applied at the rate of 10 Ilg/g soil. but at the rate of 50 Ilg/g soil some of them,
especially 2,5-dimethyl-p-benzoquinone and 2,6-dimethyl-p-benzoquinone, markedly
inhibited nitrification. However, none of them inhibited nitrification as effectively as
nitrapyrin. Degree of inhibition was higher with the light-textured soil than with the
clay loams.
Using samples of four Indian soils (alluvial, black, red, and laterite), Sachdev et al.
(1977) prepared reaction mixtures from 10 g of air-dried soil + 10 g of quartz sand + 1
ml of a solution containing 2 mg of N as (NH4hS04 or urea with or without 500 Ilg of
2,5-dimethyl-p-benzoquinone + water to 60% of WHC. The mixtures were incubated
aerobically at 30°C for 14 days, after which the NH/-N and N0 3--N contents were
determined. It was established that 2,5-dimethyl-p-benzoquinone strongly inhibited
238
nitrification of both (NH 4hS04 and urea; degree of inhibition was 91.7-98.0% and 92.2-
97.3%, respectively.
Studying a great number of polyhydric phenols and quinones, Mishra and Flaig
(1979) and Mishra et al. (1980) found that some of these compounds inhibit both soil
urease activity and nitrification (see pages 91-92).
Rodgers (1984a) mentioned that the 2,6-dimethyl-p-benzoquinone concentrations
needed to inhibit nitrification in soil are 5-10 Jlg/g soil.
Hera et al. (1986) compared the effects of hydroquinone (HQ) and two nitrification
inhibitors, nitrapyrin and ATC, on the formation of NO)- in urea-treated samples of a
chemozernic soil. The three compounds were applied at rates of I, 2, and 10 mglkg soil.
The experiment lasted 200 days, during which the NO)- content was determined
periodically.
By inhibiting urea hydrolysis, HQ reduced the formation of NO)-. But the effects of
nitrapyrin and ATC were stronger. With each compound, the relation between dose and
effect was directly proportionate. In comparison with the soil treated with urea only, HQ
reduced the formation of NO)- for 14 days when applied at rates of 1 and 2 mglkg soil
and for 28 days when its rate was 10 mglkg soil. After this period, nitrification showed
a trend for recovery_ But, even after 200 days, the amount of NO)- remained smaller in
the soil treated with urea and an inhibitor than in the soil treated with urea only.
Ten-g air-dried samples of a Belgian loam soil were used by Wang et al. (1990) for
evaluating the effect of HQ, PPDA, and nBTPTA on nitrification of NH/ and N0 2 -.
One ml ofa solution containing 2 mg ofNH/-N as (NH4hS04 or 0.1 mg of N0 2 --N as
NaN0 2 and I ml of a solution containing 0.04 g of urease inhibitor were added to the
soil samples; the resulting moisture content was two-thirds of field capacity, assuring
aerobic conditions. During incubation at 25 D C, the samples were analyzed periodically
for NH/, NO)-, and N0 2-.
HQ brought about a 2-day delay in oxidation of the added NH4 +. HQ delayed the
oxidation of soil-derived + added N0 2- similarly by 2 days. Thesefmdings suggest that
HQ inhibited oxidation of NH/ to N0 2 (including oxidation of soil NH/ to soil-
°
derived N0 2 -) and had little or no effect on the oxidation of N0 2 - to NO)-. PPDA and
nBTPTA had no effect on nitrification since in both absence and presence of these
urease inhibitors complete oxidation of added NH4 + and that of the added N0 2 - occurred
in 5 and 9 days, respectively.
A part of these investigations, namely those referring to HQ are also described in
another paper whose co-author is Wang (Zhou et aI., 1992).
Xu et al. (1994) presented evidence that HQ is an inhibitor of both soil urease
activity (see page 98) and nitrification. In one of their experiments, samples of an acid
soil (pH 5.58) were treated with (NH4hS04 or urea (50 mg N/lOO g soil) with or
without 2 mg HQ/IOO g soil. HQ inhibited nitrification of both ammonium sulfate and
urea_ The inhibition was 92.5 and 92.1 % respectively, after I-week incubation and 78.5
and 84.1 %, respectively, after 2 weeks.
5.2.2_8_ N-Halamine Compounds

As shown in Subchapter 2_18, Gautney ef al. (1990) patented N-halamine compounds as
dual purpose, urease and nitrification inhibitors. The four N-halarnine compounds tested
for evaluation of their urease-inhibiting ability in a silt loam soil were specified in
239
Figure 33. The same compounds, but another silt loam soil, were used for testing their
nitrification-inhibiting ability.
Soil samples (384 g) wetted to field capacity were amended with 410 mg of urea and
o or 41 mg of N-halamine compound. Dicyandiamide (DCD) served as a reference
compound. The samples were then incubated at 25°C for 5 weeks and analyzed weekly
for N0 3--N, N02--N, and urea-No
Compounds ABC, AB, and IE were stronger nitrification inhibitors, and compound I
was a weaker one than DCD. The inhibition caused by compounds ABC, AB, and IE
was 100% during 4 weeks and 99.6, 98.3, and 99.6%, respectively, after 5 weeks. The
corresponding values recorded after week I to week 5 were: 78.2, 88.6, 87.4,87.4, and
90.0%, respectively, for DCD, and 74.4, 68.9, 4.9, -2.8, and -1.5%, respectively, for
compound I.
Compounds ABC and IE were identically potent inhibitors of nitrification, but
compound ABC was a stronger urease inhibitor than IB. Therefore, compound ABC
(l-bromo-3-cWoro-4,4,5,5-tetramethyl-2-imidazolidinone) may be considered the best
dual purpose inhibitor among the N-halamine compounds studied.
5.2.2.9. Phosphorodiamides
One of the phosphoroamides patented and tested as urease inhibitors by Swerdloff et al.
(1985a), namely 3-(1 ',1 ,-dimethylethyl)-4-hydroxy-PPDA (see page 113 and Figure 40)
was also tested for evaluating its effect on nitrification. Air-dried samples (20 g) of a
New York soil (Cazenovia silt loam, pH 7.3) were treated with 0 or 0.8 mg of test
compound in 5 ml of water and, immediately or after 14 days of preincubation, with 6
mg of diammonium phosphate in I ml of water. Then all samples were incubated at
25°C for 14 days, followed by their analysis for nitrate content.
Inhibition of nitrification was found to be 39% in non-preincubated soil samples and
0% in the preincubated ones. This finding indicates that the test compound was
degraded in soil during the 14 days of preincubation.
Bremner et al. (l986b) studied the effect of nine phosphoroamides, including
trichloroethyl-PDA and nBTPTA, on nitrification ofNH/. Three Iowa soils were used
(see Table 54). Nitrapyrin and etridiazole served as reference compounds. The reaction
mixtures (lOg of air-dried soil + 1 ml of a solution containing 2 mg of N as (NH4)2S04
+ 3 ml of water or aqueous solution containing 0.05, 0.10, 0.50 or 1.0 mg of
phosphoroamide or 0.05 mg ofnitrapyrin or etridiazole) were incubated at 20°C for 21
days or at 30°C for 14 days and analyzed for (N03- + N02-)-N.
Nitrification was inhibited only by trichloroethyl-PDA at each of its rates and by
nBTPT A at rates of 50 and 100 Ilg/g soil. The inhibitory effect increased with
increasing inhibitor rate and was more marked in the light-textured soil than in the
heavier-textured ones but less marked at 30°C than at 20°C (see also Bremner, 1986).
It should be emphasized that trichloroethyl-PDA and nBTPTA, even at their highest
rate, were much weaker nitrification inhibitors than were nitrapyrin and etridiazole used
at a rate of only 5 Ilg/g soil.
5.2.2. JO. Phosphoric Triamides

For studying inhibition of nitrification, Kolc et al. (1985a) used the same soil and
methods as those applied by Swerdloff et al. (1985a) and described briefly in the
preceding section.
240
Three of the four N-acyl phosphoric triamide compounds evaluated as urease

inhibitors, namely N-(diaminophosphinyl)-2-chloro-, 2,2-dichloro-, 2,2,2-trichloroacet-
amide (see Table 41) as well as N-(diaminophosphinyl)-4-methoxybenzamide [4-CH30-
C6Rt-CO-NH-P(O)(NH2)2], were tested for evaluation of their effect on nitrification.
Percent inhibitions of nitrification in the non-preincubated and preincubated soil
samples were 47 and 3 (trichloro compound), 18 and 9 (dichloro compound), 2 and 0
(chloro compound), 4 and 13 (methoxy compound), respectively. Thus, preincubation
led to great diminution or disappearance of the nitrification-inhibiting ability of the
three chloro compounds and to an increase in the weak inhibitory ability of the methoxy
compound.
In the non-preincubated soil samples, inhibition of nitrification caused by the three
chloro compounds increased in the order:
trichloro » dichloro » chloro,
which is the opposite of the order of their urease-inhibiting capacity (see Table 41).
Nevertheless, N-(diaminophosphinyl)-2,2,2-trichloroacetamide causing 64% inhibition
in urease activity and 47% inhibition in nitrification may be considered the best dual
purpose inhibitor among the N-acyl phosphoric triamides studied.
The inhibitory effect of nBTPTA on nitrification, as studied by Bremner et al.
(1986b), is mentioned in the preceding section.
As shown on page 144, thiophosphoryl triamide (TPTA) was patented as a urease
inhibitor by Gautney (1987) and as a dual purpose, both urease and nitrification
inhibitor by Radel (1990). The descriptions in Radel's patent are dedicated nearly
exclusively to experiments on inhibition of nitrification.
Air-dried samples (384 g) of a silt loam soil wetted to field capacity were amended
with 887 mg of ammonium sulfate (the N supplied is equal to that of 410 mg of urea)
and with 4.1 or 41 mg of test compound. The control samples were amended with urea
only. Besides TPTA, nBTPTA was also tested, and dicyandiarnide (DCD) was used as a
reference compmmd. All samples were incubated at 25°C and analyzed weekly for
N0 2--N, N0 3--N, and NRt +-N for 5 weeks.
No significant nitrite accumulations were found in any of the five analyses.
Therefore, calculation of percent inhibitions of nitrification was based only on the N03--
N contents.
After 5 weeks of incubation, the percent inhibitions registered in the TPTA-,
nBTPTA-, and DCD-treated samples were 2.3, 4.3, and 68.3, respectively, at the lower
(4.1-mg) inhibitor rate, and 53.2, 4.7, and 97.2, respectively, at the higher (41-mg) rate.
Thus, DCD was a stronger inhibitor of nitrification of NH4 + from ammonium sulfate
than was TPTA, whereas nBTPTA had practically no effect on nitrification.
In another experiment, the conditions were identical to those of the first experiment,
with the following exceptions: urea was used (410 mg!384-g soil sample) as a nitrifiable
substrate; the inhibitors were applied only at the 41-mg rate; after each incubation time,
the residual urea was also analyzed.
The nitrification inhibitions registered after 5 weeks were 97.3% (TPTA) , 84.6%
(nBTPTA), and 90.0% (DCD). In other words, TPTA was a stronger inhibitor of
nitrification ofurea-N than was DCD, and nBTPTA was also inllibitory on nitrification.
The analyses for urea showed that urea was not detectable after 1 week in the
control samples and after 2 and 4 weeks in the DCD- and TPTA-treated samples,
respectively, whereas a considerable amount of urea was present in the nBTPTA-treated
241
samples even after 5 weeks. Thus, the order of urease inhibition was: nBTPTA »
TPTA» DCD.
The NH/-NIN0 3--N ratio in the urea+inhibitor-treated samples had, after 1-,4-, and
5-week incubation, the following values: 7.0, 106.1, and 28.2 (TPTA); 3.4, 0.24, and
0.15 (nBTPTA); and 78.2, 5.7, and 7.2 (DCD). It was emphasized that the time of
maximum ratio can be varied by varying the amount of TPTA for maintaining proper
ammonium/nitrate nutrition of agricultural plants. Therefore, TPTA can be designated
as an ammonium/nitrate ratio control agent (Radel et al., 1992).
In the experiment performed by Bronson and Mosier (1994), 20-g fresh samples
taken from the 0-15-cm layer of two native shortgrass prairie soils from northeastern
Colorado (a fine sandy loam, pH 6.1 and a sandy clay loam, pH 5.7) were treated with
NH4 Cl at a rate of 25 flg N/g soil and with 0, 5, and 25 flg nBTPTA or PPDAIg soil.
Percent inhibitions of nitrification registered after 6 days of incubation at 28°C at the 5
and 25 flglg inhibitor rates were 0 and 13 (nBTPTA) and 3 and 12 (PPDA) in the fine
sandy loam and 6 and 17 (nBTPTA) and 7 and 12 (PPDA) in the sandy clay loam. The
oxygen analogue of nBTPTA was also tested in the fine sandy loam at a single rate (25
flglg soil), and it was found that its nitrification-inhibiting effect was 13% of that of the
nBTPTA.
In the laboratory experiments of Vittori Antisari et al. (1996), concentration of
nitrite was decreased and that of nitrate was increased by nBTPTA during incubation of
urea-treated samples of three Italian soils (see also page 153).
5.2.2.11. Humic Substances and Lignosulfonates

Sachdev et af. (1977) studied the effect of N-lignin (an oxidatively ammoniated
lignosulfonate) on nitrification of urea and ammonium sulfate in four Indian soils
during 14-day incubations and found that N-lignin inhibited more markedly nitrification
of urea (14.1-44.5% inhibitions) than that of ammonium sulfate (1.6-26.2% inhibitions).
Based on these findings, the authors assume that N-lignin and its degradation products
also inhibited soil urease activity. However, no experimental data are presented in favor
of the assumption.
The inhibiting effect ofhurnic substances and lignosulfonates on soil urease activity
and/or nitrification is also dealt with in Section 2.31.2 and Subchapters 3.8 and 3.9.
5.2.2.12. Plant Residues

According to Krishnapillai (1979), tea waste (tea fluff) inhibits nitrification. Contrarily,
Sivapalan et al. (1985) found that polyphenol-rich plant materials, including tea
residues, inhibit soil urease activity but had no inhibitory effect on nitrification (see
page 170).
Cakes and oils from neem, karanja, and mahua are, in general, stronger inhibitors of
nitrification than of urease activity (see Sections 2.31.3.3-2.31.3.5). In a laboratory
experiment of Blaise and Prasad (1997), neem cake used for coating urea prills was a
little stronger inhibitor of nitrification than was pyrite blended with urea. But in the pot
experiment ofBiau et al. (2000), neem cake had a less evident and less stable inhibiting
effect on nitrification ofurea-N in comparison with that of DCD and 3-methylpyrazole
(rates of additions were: 150 mg urea-N/kg soil, neem cake 50%, DCD 10%, and 3-
methylpyrazole 2% relative to weight of added N).
242
5.2.3. Mixtures ofInorganic and Organic Compounds

Ashworth et al. (1979) found that the inhibitory effect of a 1: 1 (weight/weight) mixture
of sodium trithiocarbonate (Na2CS3) and potassium ethyl xanthate (KEtX) 00 both
urease activity and nitrificatioo was synergistic (see also page 59).
In a short report, Ziyamukbamedov et al. (1986) mentioned that in laboratory,
greenhouse, and field experiments a CuS0 4 + KEtX mixture added to urea which was
then applied as fertilizer to cotton soils inhibited urease activity for 15 days and
nitrification for 1.5 months. By using 15N-Iabeled urea they established that, under the
influence of CUS04 + KEtX, the N losses decreased by 12-20%.
243
Chapter 6. Soil Urease Inhibitors Used in Combination with Nitrification and/or

Algal Inhibitors
6.1. COMBINED USE OF UREASE AND NITRIFICAnON INHIBITORS
For improving the storage and/or fertilizer properties of urea, Richter et aT. (1978)
patented a technology for coating the urea prills with distillation residue of synthetic
fatty acids manufactured by oxidation of paraffins and for amending the coated prills
with biologically active compounds, including urease and nitrification inhibitors. But
application of the technology for urease and nitrification inhibitors is not exemplified, in
the patent description, by any nominalized urease or nitrification inhibitor.
Amberger (1989) described an experiment in which ammonium thiosulfate (ATS)
(nitrification and urease inhibitor) used with dicyandiamide (DCD) prevented the rapid
degradation of DCD and, thus, prolonged the nitrification-inhibiting effect. Amberger
(1989) also recommended the combined use of guanylthiourea (nitrification and urease
inhibitor) and DCD with the same aim to prolong inhibition of nitrification. According
to the data published by Amberger and Gutser (1984), thiourea (TO) (nitrification and
urease inhibitor) markedly reduced decomposition of DCD. Soil samples (50 g each)
were amended with 1.5 mg DCD and 0 or 0.5 or 1.0 mg TU and incubated at 15°C for 3
weeks. The residual DCD content was 0.66 mg (no TO), 0.80 mg (0.5 mg TU), and 0.87
mg (l.OmgTO).
Goos et al. (1990) and Goos and Johnson (1992) conducted laboratory, microplot,
and field experiments to compare the effects of ATS, DCD, and ATS+DCD mixtures on
nitrification of urea and urea-ammonium nitrate (UAN) in North Dakota soils.
In the laboratory experiment, a silt clay (PH 6.6) and a loam (PH 6.9) were studied.
Urea with or without inhibitor or inhibitor mixture was applied in solutions containing
186 g Nil. Rate of ATS was 8.7% (volume/volume) and that of DCD was 0.5, 1, 2 or
5% ofN as DCD-N. A droplet (0.1 ml) of fertilizer solution was placed on the surface
of 50-g soil samples. Incubation took place at 25°C (silty clay soil) or at 20°C (loam
soil) and lasted 17 days. The ammonium-N and (nitrite+nitrate)-N contents determined
after incubation served for calculation of the percent inhibition of nitrification. Similar
results were obtained in the two soils studied. The following average inhibitions were
registered: 68% (ATS alone), 48-92% (DCD alone at rates of 0.5-5%), and 76-94%
(ATS+DCD 0.5-5%). Thus, the inhibition was greater with ATS + DCD than with DCD
alone. It should also be mentioned that some nitrite accumulated when ATS was added,
but little or no nitrite accumulated when both ATS and DCD were present in the
fertilizer solution.
The microplots were installed, in the spring or autumn of 1988 or in the spring of
1989, in the field at 11 sites on different soils commonly used for wheat production. The
microplots were "buried bags". Top soil samples (300 g each) packed in nylon mesh
bags were amended with a droplet (0.25 ml) of fertilizer solution; then other 300-g soil
samples were added, and the bags were buried in holes having 10 cm in diameter. The
fertilizer solution prepared from urea Of from urea + ATS or DCD or ATS+DCD
contained 186 g N/l. Rate of ATS was 10% (volume/volume) and that ofDCD was 2%
of N as DCD-N. Soils were taken from the bags 4 or 8 or more weeks after fertilizer
application for determination of ammonium-N contents. The results showed that ATS
added to urea solution significantly (p:SO.lO) increased residual ammonium contents
244
over urea alone at six of II sites. ATS was usually a less effective nitrification inhibitor
than was DCD, and ATS+DCD outperformed DCD at only one of 11 sites.
Three field experiments were carried out in the spring of 1990. Size of the plots was
2.5 x 9 m. The fertilizer, UAN solution, was applied at a rate of 75 kg N/ha. Rate of
ATS was 10% (volume/volume) and that of DCD was 2% of N as DCD-N. The
fertilizer solutions were banded 10 cm deep on 35-cm centers of the plots. The soils
were sampled 1 day, 2, 4, and 8 weeks after fertilizer application. The I-day, 2- and 4-
week samples were analyzed for (urea+ammonium)-N and the 8-week samples for
ammonium-No In all three field experiments, ATS added to UAN increased residual
ammonium contents. Again, ATS was less effective than DCD, and no ATS+DCD
synergism was observed.
Sutton et al. (1991) patented a homogeneous granular fertilizer composed of urea
(90-98% by weight), DCD (1.4-3.0%), and ATS (0.4-1.0%), and, optionally, of a
phosphate, most preferably, of an ammonium polyphosphate (APP) (0.3-1.0%). Besides
the technology for manufacturing of urea+DCD+ATS and urea+DCD+ATS+APP,
testings of urea+DCD+ ATS are also described in the patent. The testings were carried
out on maize fields (see page 262) and on turf (see page 313).
In the field experiment conducted by Clay et al. (1990), urea alone or urea with
nBTPT A or DCD or nBTPTA + DCD were surface-applied on soil covered by dried
maize leaves (2-4 cm in diameter) and on bare soil (a sandy loam from Minnesota).
Rate of additions per m2 was: 160 g of urea-N, 0.8 g of nBTPTA and/or 2 g of DCD.
Ammonia volatilization was measured during the first 4 days after fertilization.
Volatile NH3 losses in the same treatment were lower from the residue-covered than
from the bare soil. nBTPT A reduced NH3 volatilization by 100 times over urea only,
and the effect was even stronger when urea was applied with nBTPTA + DCD. DCD
alone had no effect on NH3 volatilization from the residue-covered soil but increased
the volatile NH3 loss from the bare soil.
In a laboratory experiment with the same soil and treatments as in the field
experiment, ammonia volatilization was measured during 8 days of incubation at 35°C.
Volatilization of NH3 presented the order: control (urea only);::: DCD > nBTPTA >
nBTPTA+DCD.
Weston et af. (1994) patented a liquid fertilizer, namely an aqueous solution of urea,
ammonium nitrate, nBTPTA, and DCD, and, optionally, a clay as a suspending agent.
This fertilizer contains 24-32% (preferably 26-32%) of urea, 34-42% (preferably 36-
42%) of ammonium nitrate, 0.01-0.4% (preferably 0.02-0.3%) of nBTPTA , and 0.01-
2% (preferably 0.03-1.5%) of DCD. The nBTPTA to DCD ratio should exceed 0.01,
should preferably be between 0.02 and 8.0 and most preferably between 0.05 and 1.0.
nBTPTA as a concentrated solution in an N-alkyl-2-pyrrolidone, preferably N-methyl-
2-pyrrolidone, is incorporated into a urea-ammonium nitrate (UAN) solution or
suspension, whereas DCD is added to UAN as a solid, a suspension or in dissolved form
along with the nBTPTA.
In the laboratory experiments of Chen et aZ. (1995) and in the pot experiments of
Chen et al. (1998), hydroquinone (HQ) was used in combination with either DCD or
acetylene provided by encapsulated calcium carbide (CaC 2). Besides hydrolysis and
nitrification of urea, N 20 emission was also studied.
In the laboratory experiments, 250-g samples of a Chinese soil (pH 6.66) were
treated with: 1. urea only (1.2 g); 2. urea + 0.3% HQ (on urea weight basis); 3. urea +
245
0.3% HQ + 3% DCD; and 4. urea + 0.3% HQ + 20% CaC z• The samples were wetted to
22% soil moisture content or were water-logged. Incubation took place at 30°C.
The ammonium-N content in the wet and water-logged soils was determined
periodically during 90 days, and this content was also measured periodically in the
floodwater during 60 days. The following orders were established in the different
treatments: control (urea only) ~ HQ+CaC 2 < HQ < HQ+DCD in the wet soil and
control < HQ < HQ+CaCz < HQ+DCD in the water-logged soil. The floodwater
ammonium-N contents were not significantly different between the treatments.
The nitrate-N content in the wet soil and floodwater was determined periodically
during 90 and 21 days, respectively. In both cases, the following order was established:
HQ+DCD < HQ+CaCz < HQ < control.
The ammonia volatilized from the wet and water-logged soils was measured during
90 and 28 days, respectively. The NH3 loss from the wet soil presented the order:
HQ+DCD < HQ < HQ+CaC z ~ controL and an almost reverse order was established for
the water-logged soil: control < HQ < HQ+DCD < HQ+CaC z.
NzO emission from the wet and water-logged soils was detectable during the first 14
and 28 days, respectively, in the same order: HQ+DCD < HQ < control < HQ+CaC z•
In the pot experiments, 2.5-kg samples of a sandy loam soil (PH 7.2) were submitted
to the same four treatments as were the soil samples in the laboratory experiments.
Rates of additions were: urea 0.58 g N/kg soil, HQ 0.3%, DCD 8%, and CaCz 20%.
There were also pots to which no urea and no inhibitor were added. The soil moisture
content was maintained at 20% (about field capacity). Thirteen spring wheat plants were
grown in each pot. The experiments were carried out at 22°C.
The amounts of ammonium-N, nitrate-N, and NzO were measured periodically
during 56 days. The ammonium-N content showed the order: control ~ HQ ~ HQ+CaC 2
«HQ+DCD, whereas the reverse of this order was valid for the nitrate-N content. NzO
was emitted in the order: HQ+DCD < HQ+CaC z < HQ < control.
The conclusion drawn from both laboratory and pot experiments was that HQ+DCD
was an effective combination for inhibition of hydrolysis and nitrification of urea and
reducing ofNzO emission.
This conclusion was confirmed by results of another pot experiment in which
labeled urea (urea_ 15 N) was applied and the test plant remained spring wheat (Xu et al.,
2(00).
Grego et al. (1995a,b) conducted a field experiment on a loam soil (PH 7.0) at the
Experiment Station of the Pisa University. Urea (60 and 120 kg Nlha) unamended or
amended with 0.25% nBTPTA or with 0.25% nBTPTA + 4.5% DCD (on urea weight
basis) was surface-applied on plots (2 by 5 m) installed in a wheat field. Soil was
sampled from the O-15-cm layer for analysis of residual urea, exchangeable NH/-N,
and N03--N contents at days 5,12, and 19 after fertilizer application.
The amount of residual urea presented the orders: nBTPTA+DCD < control (urea
only) < nBTPTA (at the urea rate of 60 kg Nlha), and control < nBTPTA+DCD <
nBTPTA (at the urea rate of 120 kg Nlha) at day 5 and was very low and negligibly low
at days 12 and 19, respectively (at both urea rates). These findings indicate that a)
nBTPTA efficiently inhibited urease activity only for 5 days and b) DCD enhanced
hydrolysis of urea. Contrarily, the effect of DCD in inhibiting nitrification was evident
even at day 19, at which the exchangeable ammonium-N content was highest and the
nitrate-N content was lowest in the nBTPTA+DCD treatment at both urea rates.
246
Grego et al. (1995b) described a similar wheat field experiment on a sandy loam soil
(PH 6.3) at the Experiment Station of the Bologna University. Rates of additions were:
urea-N 120 kgtha, nBTPTA 0.25%, and nBTPTA 0.25% + DCD 4.5% relative to
weight of urea. As in the Pisa experiment, the fertilizer was surface-applied on plots (2
by 5 m). Soil was sampled from the 0-5-cm layer at days 2, 5, and 12 after fertilizer
application. The samples were analyzed as were the Pisa soil samples.
The results were not different from those registered in the Pisa experiment.
Palazzo et al. (1996) conducted experiments in wheat fields on a silty clay soil (PH
8.11) and a sandy clay soil (PH 8.12) at Metaponto (southern Italy). Ammonia
volatilization was studied in the following treatments: control (urea only, 120-150 kg
Nlha), urea + 0.25% nBTPTA, and urea + 0.25% nBTPTA + 4.5% DCD. The volatile
NH3 was measured during 6 weeks after fertilization.
The cumulative NH3 losses from the urea-N added to the silty clay soil were: 7.6%
(control), 3.3% (nBTPTA), and 8.1% (nBTPTA+DCD), whereas the losses from the
sandy clay soil were: 16.1% (control), 5.0% (nBTPTA), and 10.0% (nBTPTA+DCD).
Thus, nBTPTA applied without DCD markedly reduced the NH3 losses, but DCD
enhanced volatilization ofNH 3 from the silty clay and hindered the effect of nBTPTA in
the sandy clay. Nevertheless, the grain yield (tlha) increased from 4.49 (control) to 4.50
(nBTPTA) and 4.67 (nBTPTA+DCD).
Based on the finding that nBTPTA decreased the concentration of nitrite but
increased that of nitrate during incubation of urea-treated soil samples, Vittori Antisari
et al. (1996) also suggested the combined use of urease and nitrification inhibitors.
Hong and Chen (1997) patented a slow-release granular urea fertilizer to which both
urease and nitrification inhibitors are also added. Heavy metal salts [FeS04, ZnS04,
MnS0 4, ~hMo04]' boric acid, sodium borate, hydroquinone are among the applied
urease inhibitors, whereas thiourea and DCD are the preferred nitrification inhibitors.
The average amount of urease + nitrification inhibitors is about 0.5% relative to weight
of urea.
Montemurro et al. (1998) studied nBTPTA, DCD, and nBTPTA+DCD applied with
urea on a clay soil (PH 7.8). The experiment was carried out in a cold greenhouse at
Metaponto. The best plant was lettuce. Before planting, the soil was fertilized with 20
kg Plha as superphosphate and 40 kg Klha as K2 S0 4 • On 14 December 1995, 3-week-
old seedlings were transplanted and the soil was irrigated. On 15 January 1996, at the
stage of four leaves, urea (60 kg Nlha) was incorporated into top 20 cm of soil in four
treatments: urea only (control), urea amended with 0.25% nBTPTA or 4.0% DCD or
0.25% nBTPTA+4.0% DCD (percentages mean weight relative to weight of urea). The
volatile ammonia was measured up to the harvest time (2 April 1996). Ammonium-N
and nitratc-N were determined in the 0-20-cm soil layer at days 8, 19,33, and 59 after N
fertilization and at harvest.
The cumulative NH3 losses expressed in kg Nlha were the following: 11.3 (control),
2.7 (nBTPTA), 8.3 (DCD), and 7.4 (nBTPTA+DCD). Thus, nBTPTA alone was more
effective than in combination with DCD. During the whole experimental period, the
ammonium-N content was higher and the nitrate-N content was lower in the DCD and
nBTPTA+DCD treatments than in the nBTPTA treatment and control, which means
that DCD inhibited nitrification for more than 2 months.
247
6.2. COMBINED USE OF UREASE AND ALGAL INHIBITORS
In the pot experiments conducted by Vlek et al (1980) and referred to on page 60, the
effect of phenylphosphorodiamidate (PPDA) used alone and in combination with the
herbicide (algicide) simazine was also studied. Urea prills (100 kg of N/ha) with and
without 1 or 2% PPDA (relative to urea weight) were puddled into soil covered by 5-cm
deep floodwater. In other pots, only urea was incorporated into the soil, whereas PPDA
with or without simazine was administered directly into the floodwater (rate of PPDA
was 1% relative to weight of urea and that of simazine was 3 mg/l floodwater). During
incubation at 35°C day and at 25°C night for 10 days, the daily analyses for residual
urea and NH/ + N0 3- from floodwater showed that complete hydrolysis of urea
occurred in 3 days in the absence of PPDA, in 6 days when PPDA had been added to
floodwater and in 6-8 days when PPDA with urea had been incorporated into the soil.
Inhibition of urease activity was strong only during the first 3 days of incubation,
and the degree of inhibition was higher with PPDA incorporated into soil than with
PPDA added to floodwater. This is explained by the finding that urease activity in
floodwater was negligibly low in comparison to that of soil. PPDA was more effective
at 2 than at 1% amount. Inhibition of urease activity was accompanied by diminution of
NH4+ concentration in floodwater; thus, susceptibility of NH3 to volatilization was
reduced. Simazine administered in combination with PPDA had little additional effect.
Moreover, simazine may even have a negative effect by preventing development ofN2-
fixing blue-green algae (cyanobacteria) (see also Vlek and Craswell, 1981).
But in other pot experiments, the algicide added to floodwater in combination with
PPDA delayed the decomposition of PPDA, prolonged its urease-inhibiting effect,
which means that, from this viewpoint, the algicide had a positive effect. According to
the first data published on these investigations, CUS04, acting as an algicide, decreased
the decomposition of PPDA and maintained urea in the floodwater for a longer time
(Anonymous, 1986). These investigations were developed and published in detail by
Byrnes et al. (1989a, b).
Byrnes et al. (1989a) used two soil systems: 300-g samples of a silt loam (PH in
H20 5.2) and mixtures of 60 g silt loam + 260 g coarse washed sand. Urease activity is
high in the soil and low in the soil+sand mixture due to the sand. The samples of soil
and soil+sand mixture were flooded and kept flooded at 3-cm depth for 3 weeks. Then
an acidifying agent or sodium acetate or algicide + urea + PPDA were added to the
floodwater in the following amounts per pot: 600 mg of Ab(S04}3.16H20; 126 mg of
Ca~(P04bH20; 62 mg ofH300 3; 136 mg of CH3COONa; 16 mg of CuS04.5H20 (as
algicide); 35 mg ofurea-N (-200 ppm urea-N); 9% PPDA (on the weight of urea) (-40
ppm PPDA). The control soil and soil+sand mixture received only urea. Daytime pH,
concentrations of PPDA, urea, and ammonium-N were determined daily for 26 days
after addition of urea.
The average daytime (afternoon) pH of the floodwater, which was -9 and 8 in the
control soil and soil+sand mixture, respectively, was reduced by the acidifying agents,
namely to 4.3 and 4.2 by Alz(S04)3, to 8.2 and 5.1 by Ca~(P04)2' and to 8.2 and 7.1 by
H3B03, respectively. The average daytime pH remained 9 and increased to 9.1 in the
floodwater of the CH3 COONa-treated soil and soil+sand mixture, respectively.
The pH below 5 caused rapid decomposition of PPDA, and PPDA was essentially
undetectable in 3 or 4 days. In the less urease-active soil+sand mixtures, the acidifying
248
agents, except Alz(S04)3, did not diminish the floodwater pH below 5. Thus, PPDA
remained in the floodwater for 9-12 days.
At the very alkaline floodwater pH resulting from addition of CH3COONa, the
decomposition of PPDA was, as at pH below 5, very rapid in both soil and soil+sand
mixture.
The algae are responsible for the very high daytime pH (9-10) of paddy floodwaters
because of algal uptake of HC0 3' during photosynthesis. By inhibiting the growth of
algae, CUS04 reduced the average daytime floodwater pH to -8 in soil and to -6 in
soil+sand mixture. Thus, effectiveness of PPDA was prolonged for about 3 days more
than without the algicide.
The conclusion concerning fate of PPDA in floodwater was that at daytime pHs of
approximately 9 the half-life of PPDA was only 10 hours; at neutral or slightly acidic
pHs the half-life could be extended to about 25 hours with the soil and up to about 90
hours with the soil+sand mixture.
Inasmuch as the longevity of PPDA in floodwater of samples submitted to different
treatments was different, disappearance of urea took place during different incubation
times: in 5 days (urea-only treatment), 6 days (PPDA), 7 days (PPDA + CH3COONa),
11 days (PPDA + CUS04), and 12 days (PPDA + H3B03)'
The relatively long-lasting effect of PPDA + H3B03 is attributed not only to the
acidifying effect of H3B03 but also to its urease-inhibiting capacity in addition to that of
PPDA.
In the pot experiment carried out with flooded rice, Byrnes et al. (1989b) studied the
influence of Cu-chelate (commercial algicide; tradename: Cutrine Plus) on the
inhibitory effectiveness of nBTPTA and PPDA. In the case of PPDA, the influence of
an acidifying agent, Alz(S04)3, was also studied.
A silt loam (brown earth from loess; pH 6.5) was used. The soil (7.22 kg dry
weight/pot) was flooded for 3 weeks, then fertilized with 0.43 g of P [Ca(H2P04)2.H20]
and 1.00 g of K and 0.22 g of S (K2S04)/pot, incorporated in the top 8 cm of soil. Two
25-day-old IR36 rice plants were transplanted in each pot. The treatments comprised:
control (no N added), urea alone, urea + nBTPTA with or withoutCu-chelate, urea +
PPDA with or withoutCu-chelate or Alz(S04)3 or Cu-chelate + Al 2(S04)3, added to
floodwater. Rates of additions were: 300 mg of N/pot as 1~-labeled urea (4.67 atom%
excess 15N) at 15 days after transplanting (DAT) and 100 mg ofN/pot as unlabeled urea
at 42 DAT; 13 mg of urease inhibitor/pot (20 glkg urea); 1 mg of Cu/kg soil as Cu-
chelate; 0.1 g of Al2(S04)3.16H20/pot. Each treatment was performed either with water
percolation of 5 mm/day through the soil or without percolation. Floodwater was
maintained at 3 em depth. Urea and NH4 + in floodwater were analyzed daily for 20 days
after fertilization, and pH was measured every afternoon for 10 days after fertilization.
Plants were harvested at 42 DAT and at maturity (119 DAT) to determine their total N
and 15N contents and yields. Total N and 15N contents in soil + roots were also
measured.
Complete hydrolysis of urea occurred in 2 days in absence of inhibitors, in 14 and 4
days in the urea+nBTPTA and urea+PPDA treatments, respectively. This means that
nBTPTA was more effective than PPDA. There was no difference in urea disappearance
when nBTPTA was used with Cu-chelate compared with nBTPTA itself. In the urea
+PPDA+Cu-chelate and urea+PPDA+Ah(S04)3 treatments, urea disappeared in 7 and 5
days, respectively, which shows that Cu-chelate improved more markedly the urease
249
inhibition by PPDA than did Ah(S04h. There was no significant difference in the rate
of urea disappearance between Cu-chelate with PPDA and Cu-chelate+AI2 (S04h with
PPDA.
In the urea+nBTPTA treatment, NH/ concentration in floodwater was very low
throughout the 20 days following fertilization; urea disappeared from the floodwater
mainly by movement into the soil, and urea hydrolysis was so slow that adsorption and
immobilization by algae allowed no accumulation of NH4 + in the floodwater. Addition
of PPDA decreased the peak concentration of NH4 + in floodwater to about one-half that
of the urea-only treatment, and Cu-chelate and Alz(S04)3 reduced it further.
Addition ofCu-chelate to nBTPTA resulted in reduction of floodwater pH by about
one-half pH unit. In the urea+PPDA treatment, the lower daytime pH was maintained
more effectively by Cu-chelate than by Alz(S04)3. Even though the pH reductions were
not great, they decreased decomposition of PPDA by basic hydrolysis and, thus,
extended its inhibitory effectiveness.
In the laboratory experiments of Keerthisinghe and Freney (1994), the urease
inhibitors tested were thiophosphoryl triarnide (TPTA), N-(n-butyl)thiophosphoric
triarnide (nBTPTA) and their oxygen analogues PTA and nBPTA, as well as
cyclohexyl-PTA (CHPTA). The algicide was terbutryn [2-(t-butylarnino)-4-(ethyl-
arnino)-6-(methylthio)-s-triazine]. Two Australian clay soils (PH 5.8 and 6.7,
respectively) cultivated with rice were studied. Air-dried soil samples (15 g) were
placed in 120-ml glass bottles, and the soils were flooded by addition of 30 ml distilled
water. Terbutryn (0.2 mg active ingredientll) was added with the water to half of the
bottles, which were kept in the dark, and water only to the other half, which were kept
in the light. All samples were maintained at 25°C. After 3 weeks, a profuse growth of
algae was noted in the samples kept under light, whereas no algae were visible in the
samples kept in the dark. At this stage, the urease inhibitors were added to the samples
at a rate of 1% of the weight of urea (14 mg urea per bottle). The samples were then
incubated for 12 days and analyzed every 2 days.
Results of the analysis of residual urea showed that elimination of the algal growth
led to a) diminution of the urease-inhibiting effect of TPTA and nBTPTA (due,
probably, to lack of O2 produced by the algae, the thio analogues were not oxidized into
the more urease-inhibitory oxygen analogues) and b) to increased urease-inhibiting
capacity of the oxygen analogues and CHPTA in both soils. In the absence of algae, the
effectiveness of the inhibitors was CHPTA> nBPTA>PTA>nBTPTA>TPTA.
The conclusion was drawn that CHPTA and nBPTA in conjunction with an algicide
have the potential to reduce considerably ammonia loss from flooded rice soils.
6.3. COMBINED USE OF UREASE, NITRIFICATION, AND ALGAL INHIBITORS
In the rice field experiment described by Chaiwanakupt et af. (1996) and Phongpan et
af. (1997), the algicide terbutryn reduced the volatile ammonia losses to a greater extent
than did the nitrification inhibitor acetylene provided by calcium carbide. In addition,
terbutryn enhanced, while calcium carbide diminished, the effect of urease inhibitors to
reduce the NH3 losses (see page 182).
Freney et af. (1995) and Phongpan et af. (1997) described an experiment which was
carried out in a rice field located on a clay soil (PH 5.1) in the Central Plain region of
Thailand during the dry season of 1993. The urease inhibitors tested were N-(n-
250
butyl)phosphoric triamide (nBPTA) and cyclohexyl-PTA (CHPTA). Phenylacetylene

(C6HS-C=CH) (PA) was the nitrification inhibitor. and copper sulfate and terbutryn were
the algicides.
The drained plots (4 by 4 m) were PK-fertilized (24 kg Plha as superphosphate and
28 kg Klha as KCI) immediately before transplanting 2l-day-old rice seedlings on IS
March. The floodwater depth was adjusted to 5 cm. In the next steps, the inhibitors and
urea were applied on the surface of floodwater. Rates and dates of additions were the
following: nBPTA and CHPTA (1% of the weight of urea), PA (2.5 kgiha), copper
sulfate (5 kg Culha) on 16 March; urea (60 kg Nlha) on 17 March; terbutryn (0.2 mg
active ingredient/l floodwater) on 19 March. The addition of copper sulfate and
terbutryn was repeated on 23 March and 25 March, respectively.
There were six treatments with and six treatments without algicides: 1. urea; 2. urea
+ PA; 3. urea + nBPTA; 4. urea + nBPTA + PA; 5. urea + CHPTA; and 6. urea +
CHPTA + PA.
Urea content in floodwater and ammonia volatilization were assessed during the first
11 days after urea application. The urease inhibitors slowed the rate of disappearance of
the urea. The urease-inhibiting effect of CHPTA was stronger than that of nBPTA. The
addition of algicides alone and with urease inhibitors increased the amount of urea
remaining in the floodwater, whereas PA alone had no effect on the disappearance of
urea. The cumulative NH3 loss from the urea-only treatment was 14.5% of the added
urea-No The losses ranged from 1.1 to 11.8% from the other treatments, i.e., all
inhibitors reduced the volatile NH3 losses. The reduction was insignificant with PA
alone and algicides alone, whereas the losses were significantly (p<0.05) reduced to 1.2-
2.4% in the four treatments with CHPTA, but the difference between the 1.1 % and
2.4% losses was not significant.
For studying recovery of applied urea-N, microplots were installed within the plots.
The microplots were treated as the plots, but the urea applied (60 kg Nlha) was labeled
with lsN at 5.27 atom% excess. Total N and 15N in soil and plants were determined at
harvest (on 5 July). It was found that the urease inhibitors increased the recovery of
applied N from an average of 37.6% for the four treatments without urease inhibitors to
averages of51.5 and 61.6% for the four nBPTA and CHPTA treatments, respectively.
251
Chapter 7. Effect of Soil Urease Inhibitors on Germination, Growth, and Yield of

Plants
7.1. EFFECT OF UREASE INHIBITORS ON MAIZE (Zea mays)
7.1.1. ~Uect qfAlkali Metal and Alkaline Earth Metal Salts

Based on the results of a laboratory experiment in which KCl addition to urea reduced
the volatile N losses as ammonia [for example combination of 1 part of urea with 1 part
of KCl (by weight) applied in solution on samples of a loamy sand soil reduced the NH3
losses from 24.0 to 4.5%], Rappaport and Axley (1984) initiated field experiments on 3
by 15 m plots installed on a silt loam soil in Maryland. The solution containing only
urca (8.4 g N/m2) and that containing both urea and KCI (at urealKCl weight ratio of
1:1) were applied 2 days after planting of maize. KCI added with urea significantly
(p=0.05) increased maize yield over urea alone from 768 to 881 glm 2 • But when KCl
was added 10 days after urea addition, no yield increase was recorded.
Fox et af. (1986) conducted I-year (1983) field experiments on a silt loam soil (pH
6.1-6.5) in central Pennsylvania for evaluating the effect of urea-KCl on no-till maize
yields and N uptakes. The solid urea-KCl used contained 300 g Nand 166 g K/kg. Urea
alone and urea-KCl were surface-applied, at planting, at three rates of N: 67, 134, and
202 kglha. The grain yields and N uptakes by plants were not significantly different
between the urea-only and the urea-KCl treatments at any fertilizer rate.
The field experiments conducted by Bundy and Oberle (1988) for studying several
N fertilizers, including urea-CaCh and urea-KCl solution and urea prills surface-applied
at rates of 56 and 112 kg Nlha on silty loam soils cultivated with maize at Arlington and
Lancaster (Wisconsin), are referred to on page 28. While the volatilized ammonia was
measured at the higher N rate only, the grain yield and N uptake by plants were
evaluated at both N rates. At the higher N rate, significant differences among the three
N fertilizers that can be attributed to NH3 volatilization were observed only at
Lancaster. At this location, also at the lower N rate, grain yield and N uptake were
significantly higher in the urea-CaCh than in the urea treatment. In contrast, grain yield
and N uptake were not significantly different in the urea-KCl and urea treatments.
MacKenzie et al. (1988) studied the effect of urea-KCl on the silage maize yield in
4-year (1981-1984) field experiments on an eastern Canadian clay loam soil (pH 6).
Urea was applied at rates of 0, 90, and 180 kg Nlha for each year. Rates of added KCl
were 0, 60, and 120 kg K2 0lha for the first 2 years, and 0, 120, and 240 kg K2 0lha for
the last 2 years. Both urea and KCl were surface-applied at planting. Except in 1981,
when the highest dry matter yield of maize plants (11.3 tlha) was registered in the 180
kg N + 60 kg K2 0lha rates, in the next 3 years the yields were highest (15.6, 11.3, and
17.0 tlha, respectively) at the highest N + K20 rates. The mean values of yields were
lower in the urea-only than in the urea-KCl treatments in each year.
7.1.2. Effect qfFluorides

In the pot experiments of Gaponyuk and Kuznetsova (1984), urease activity was not
inhibited in soil samples treated with NaF at rates of 0.1-3 g F/kg soil (see page 34), but
root growth of maize seedlings was strongly reduced at rates ~ 0.5 g F/kg soil.
252
7.1.3. Effect ofInorganic Sulfur Compounds

Hoeft (1986) described field experiments carried out on two Illinois soils in 1984 and
1985 for studying the effect of ammonium thiosulfate (ATS) on maize. Urea-
ammonium nitrate (VAN) solution was the fertilizer at rates of 134 and 202 kg Nlha.
ATS solution was used at a rate supplying 5% of the total N. The two solutions were
mixed and surface-applied on soil. The results showed that ATS had no increasing
effect on grain yield on any soil and at any fertilizer N rate.
For studying the effect of ATS on no-till maize, Fox et al. (1986) carried out I-year
(1984) field experiments on a silt loam soil in central Pennsylvania. In the ATS solution
used, the content of N was 120 g and that of S was 260 glkg. Fifty ml of the ATS
solution and 950 ml of a UAN solution were mixed. Rates ofN addition to soil were 67,
134, and 202 kglha. UAN alone and UAN-ATS were surface-applied at planting. The
grain yields and N uptakes by plants were not significantly different in the UAN-ATS
and UAN treatments at any fertilizer rate.
But in 2-year field experiments carried out by Fox and Piekielek (1987) on the silt
loam soil in central Pennsylvania, better results were obtained with UAN-ATS than
with UAN alone. As the soil was not tilled, approximately 50-80% of the soil surface
was covered by plant residues at the time fertilizers were applied (more precisely, after
maize was sown but before it emerged). UAN and UAN + ATS were applied as a fine
spray. In other treatments in which ATS was not used, UAN was dribbled and
ammonium nitrate surface-broadcast. N rates were 50, 100, 150, and 200 kglha in all
treatments, except in the UAN + ATS treatment, in which N rate was 100 kglha. ATS
represented 5% of the volume of UAN. In both years, grain yield in the UAN + ATS
treatment was higher than that in the UAN treatment and only insignificantly lower than
that registered in treatments with dribbled UAN and broadcast mN0 3 • Similar results
were found in respect of N uptake by maize plants. Therefore, it appeared evident that
ATS increased the efficiency of spray-applied UAN. Contrarily to this conclusion, Fox
and Piekielek (1993), based on the results of new experiments, conducted over 3 years
(1989-1991) on three no-till maize fields on silt loam soils in central Pennsylvania,
concluded that amending UAN with ATS had no significant effect on N fertilizer use
efficiency of sprayed UAN and only a slight effect on banded UAN.
In the field study of Zadak et al. (1987), UAN with and without ATS was surface
band-applied (105 kglha) to a clay loam soil cultivated with maize in a ridge-tillage
system in Minnesota. To determine the importance of rainfall on N loss, water was
applied at various times and rates in the 10-day period following N application. The
conclusion was drawn that no significant reductions in urea hydrolysis and no
significant increases in maize yields occurred when ATS was used.
Leis et al. (1989) and Varsa et al. (1989) conducted four field experiments in 1987
and 1988 on silt loam soils at two locations (Belleville and Carbondale) in southern
Illinois to evaluate the effect of ATS on no-till maize. ATS was added to solution of
urea and UAN at rates of 2.5 and 5% of the applied N as A TS-N such that a total
application of 134 kg Nlha was made. ATS addition at the 5% rate to urea solution
which was then dribble-applied led to a significant (12.4%) grain yield increase only in
one of the four experiments. ATS was less effective when broadcast and added to UAN
solution.
Eckert et al. (1990), Lamond et al. (1991b), Maddux and Bames (1991), and McVay
et al. (1991) also found, in field experiments carried out in Ohio, Kansas, and Missouri,
253
that addition of ATS to UAN had little effect on the fertilizer efficiency of UAN
applications to no-till maize.
Using samples of four Iowa soils and applying ATS concentrations ranging from
100 to 5.000 J.lglg soil. Bremner et af. (1990) and McCarty et af. (1990) found that ATS
had an adverse effect on germination of maize and wheat seeds when applied at the rate
of 2.500 or 5,000 J.lglg soil and caused a dramatic reduction of early growth of maize
and wheat seedlings when applied at rates:::: 1,000 J.lglg soil.
McCarty et at. (1991) studied the effect of sodium thiosulfate, tetrathionate, sulfite,
and sulfate on growth of maize and wheat seedlings in 7 days. Samples of two Iowa
soils were used. Rates of salt additions were: 1,000. 2,500, and 5,000 J.lg anion/g soil.
Sodium thiosulfate was more toxic than sodium tetrathionate, whereas sodium sulfite
and sulfate had no adverse effect on seedling growth.
Graziano (1990) conducted field experiments in 1988 in the Po Valley near
Mantova, Italy, on a clay loam soil (pH 7.85) for evaluating the efficiency of ATS as an
additive to UAN fertilizer for maize. Before seeding, the ploughed soil of plots (6 by 6
m) was sprayed with a mix of UAN + ammonium polyphosphate + KCI ± ATS. Rates
of additions per hectare were: 300, 200, and 150 kg N, 65 kg P, 166 kg K, 100 kg of a
60% ATS solution. Nand ATS were also applied as top dressings at rates of 100 or 50
kg Nlha and 50 kg ATS solutionlha. There were five pairs of treatments, each pair
comprising plots treated with UAN and with UAN+ATS. Grain yields were always
higher in the UAN+ATS-treated plots than in the corresponding plots treated with UAN
without ATS, but the yield differences were not significant at p < 0.05.
But in other field experiments, Graziano and Parente (1996) recorded significant
maize grain increases by using ATS. These experiments were carried out during the
growth seasons of 1992, 1993, and 1994, at Spilimbergo, on. a stony sandy loam soil
(pH 7.8) typical of the northeastern Italian plain. In each year, all plots (75 m 2) were
fertilized with 35 kg Plha, 125 kg K/ha, 250 kg N as UAN or 239.7 kg N as UAN +
10.3 kg N as ATSlha (10% by weight ATS to UAN) and irrigated.
Grain yields (tlha) in the plots treated with UAN and with UAN + ATS were: 6.2
and 8.1 (in 1992), 10.5 and 12.2 (in 1993), and 9.5 and 11.1 (in 1994), respectively. N
uptake by plants was also higher in the UAN+ATS-treated plots than in those treated
with UAN without ATS. The increases in grain yields and N uptakes were significant
(p<O.OI or p<O.05) in 1992 and 1993 but not in 1994. The conclusion was drawn that
adding 10% by weight ATS to UAN seems to be a beneficial practice for maize
production in the northeastern plain of Italy.
7.1.4. Effect o/Organic Mercury Compounds

See Table 61.
7.1.5. E;f!ect o/Polyhydric Phenols and Quinones

Arziani et al. (1985) added 14C-labeled hydroquinone (HQ) to axenic culture of maize
seedlings and found that the exogenous HQ taken up by the seedlings was inactivated
(detoxified) in the tissues mainly by glycosylation and, to a lesser extent (about 8%), by
conjugation with peptides.
Zhou et af. (1988) studied the effect of HQ on maize plants in field experiments on
brown soils in the areas of Changtu and Tieling counties (Liaoning province, China), in
1984 and 1985. Urea with or without HQ was applied before sowing. During the
254
growing season, no fertilizers were applied. Grain yields were significantly higher in the
urea+HQ treatment than in that with urea alone. For example, the yield increase in 1985
was 6.5-8.5% in Changtu county and 6.9% in Tieling county.
In the field experiments of Zhao et af. (1993b), 10-m2 plots on a silty loam soil (PH
6.6) were submitted to the following treatments: control (no urea and no HQ); urea
alone (138 kg N/ha); HQ alone ( 3 kg/ha); urea (138 kg Nlha) + 0.9, 1.5, 3.0 or 9.0 kg
HQlha. All plots were treated with 22.2 kg/ha of P as ~H2P04 and then sown with
maize.
As expected, the maize yield was lowest in the control plots. HQ alone increased
insignificantly, whereas urea alone increased significantly the yield. Addition of HQ to
urea led to further yield increases. The yield was highest (7.8 tlha) in plots treated with
urea + 1.5 kg HQlha. The N content in grains also increased in the urea + HQ
treatments. The highest N content in grains (110.88 kg/ha) was recorded in the
treatment with urea + 3 kg HQlha.
See also Table 61.
7.1.6. F;tfect ojPhosphorodiamides

In the field experiment briefly described by Omholt and Hendrickson (1983),
phenylphosphorodiamidate (PPDA) (2.2 kg/ha) applied with a 50% urea solution (95 kg
Nlha) on plots cropped to maize, at the beginning of June, increased grain yield by as
much as 1 tlha relative to the urea-only treatment, but statistically the increase was not
significant.
Broadbent et af. (1985), whose field experiment with maize fertilized with urea (180
kg Nlha) with or without PPDA (2.5 kg/ha) is referred to on page 117, also determined
the grain yield. In plots fertilized with urea and urea-PPDA, the grain yields were 13.7
and 14.9 tlha, respectively. Thus, PPDA caused a yield increase, but the difference
between the two values was not significant (p=0.05).
Similarly, PPDA had no significant effect on dry matter yield of silage maize in the
field experiment of Tomar et af. (1985). It should be added to the description of this
experiment (see page 121) that, besides plots untreated or treated with timothy straw
and unfertilized or fertilized with urea with or without PPDA, plots untreated or treated
with straw and fertilized with ~N03 (used, like urea, at rate of 75 kg Nlha) were also
installed. Dry matter of silage maize was determined at day 114 after sowing.
One can see from the results (Table 55) that the dry matter yield presented values
oscillating depending on the rate of PPDA, and, at the same time, the differences
betwee11 these values are not significant. Neither did PPDA lead to yield increase in
straw-treated plots. The effect of straw on dry matter yield was also insignificant in
plots fertilized with urea without PPDA. Significant yield increases were registered only
in some fertilized plots compared to the control that had not been fertilized and treated
with straw.
The effect of two urease inhibitors, trichloroethylphosphorodiarnidate (TPDA) and
diethylphosphoric triarnide (DPTA) on maize yield was studied by Hoeft (1986) in field
experiments on three Illinois soils in 1983. Solid urea and urea-an1ffionium nitrate
(UAN) solution were the fertilizers. Urea was surface-applied, whereas UAN was
255
TABLE 55. Effects of nitrogen fertilizers, timothy straw, and PPDA on dIY matter yield
of silage maize"
Timothy straw PPDA DIY matter yield
Fertilizer
(kg/ha) (kg/ha) (tlha)
Unfertilized control 0 0 8.49d
4600 0 8.94 bed
NH4N03 0 0 9.46 abed
4600 0 10.30 a
Urea 0 0 9.72 abed
4600 0 9.99 ab
0 0.25 9.17 abed
4600 0.25 10.01 ab
0 0.50 9.75 abed
4600 0.50 10.09 ab
0 1.00 10.44 a
4600 1.00 10.17 ab
0 2.00 9.19 abed
4600 2.00 9.78 abc
0 3.74 8.68 cd
4600 3.74 9.63 abed
Fvalue 2.35"
Coefficient of variation 7.87
"From Tomaretal. (1985).
Values followed by the same letter or no letter are not significantly different (p=O.OS).
"Significant (p=O.OS).
surface- and dribble-applied. Rates of additions per hectare were: 134 kg N as urea +
2.7 kg inhibitor and 202 kg N as urea + 4 kg inhibitor; 134 kg N as VAN + 2.24 kg
inhibitor and 202 kg N as VAN + 2.24 kg inhibitor.
TPDA significantly increased grain yield on one soil which had been fertilized with
surface-applied urea and dribble-applied VAN at the 202 kg N/ha rate. DPTA had a
significant increasing effect on grain yield on the second soil fertilized with dribble-
applied VAN at the 202 kg N/ha rate. The inhibitors had yield-increasing effect on the
third soil.
The maize experiments conducted by Bundy and Oberle (1988) are referred to on
page 28. In these experiments, the effect of PPDA on grain yield and N uptake was also
evaluated. PPDA was added to urea prills and VAN solution at a rate of 2% on fertilizer
N basis (56 and 112 kg N/ha). The PPDA-containing fertilizers were surface-applied on
silt loarns at Arlington and Lancaster in 1983 and 1984. PPDA had no significant effects
on grain yield and N uptake, except in a single case: at the lower fertilizer N rate, urea-
PPDA VS. urea significantly increased N uptake (without increasing grain yield) at
Lancaster in 1984.
In pot experiments of Winiarski (1990), 1% PPDA added to urea fertilizer brought
about the increase of dry matter yield of maize by about 6%; the N uptake by plants also
increased.
The experiments in which Bremner and Krogrneier (1989) found that PPDA added
to soil samples completely eliminated the adverse effect of urea on germination of seeds
of four plant species, including maize, are dealt with on page 276. See also Table 61.
256
7.1.7. Effect of Phosphoric Triamide (PTA) and Thiophosphoric Triamide (TPTA)

Compounds
In most studies, the effect of PTA and TPTA compounds on maize plants was compared
with that of other inhibitors.
Broadbent et al. (1985) conducted a field experiment on a Califomia silt loam soil.
Main plots were clean-cultivated vs. zero-tillage, with residues from the previous year's
maize crop remaining on the zero-tillage area. All plots were planted to maize at a
population density of 86,000 plantslha. After emergence of the seedlings, urea was
applied with or without a phosphoroamide: N,N-diethylphosphoric triamide (DPTA),
trichloroethylphosphorodiamidate (TPDA) or PPDA. The rate of urea was 160 kg Nlha
and that of the phosphoroamides - 3.2 kgiha. In some treatments, urea labeled with an
excess of l'N was applied in form of solution with or without DPTA, TPDA or PPDA in
bands on the soil surface, whereas in other treatments unlabeled urea in form of
granules coated or not with DPTA or TPDA was broadcast. Leaf samples were
collected at days 29, 60, and 84 after planting. At harvest (116 days after planting),
yields of grain, cobs, and stover were measured. Total N was determined in each plant
material, and 15N in plant materials from the urea solution treatments was also analyzed.
The results showed that total dry matter, grain yield, and total N uptake were all
higher in the cultivated plots than in the zero-tillage plots. The phosphoroamides did not
significantly affect total N and 15N contents in leaves, except in a single case: total N
content in leaves at day 84 after planting significantly decreased in the urea + PPDA
treatment as compared to urea alone.
The effect of phosphoroamides at plant maturity was in general also insignificant. In
the urea solution treatments, TPDA increased, though insignificantly, the total dry
matter, grain yield, and total N content, and decreased, also insignificantly, the 15N
content. Each of these four parameters was decreased insignificantly by DPTA. Under
the influence of PPDA, the total dry matter, grain yield, and 15N content decreased
insignificantly, but total N content showed a significant decrease. When urea had been
applied in form of granules, both TPDA and DPTA acted positively: they increased,
though insignificantly, the total dry matter, grain yield, and total N content. These
findings suggest that it is more advantageous to apply granules coated with TPDA or
DPTA than urea solutions with addition of these phosphoroamides.
Hoeft (1986), who studied the effect of a phosphorodiamidate and a phosphoric
triamide on maize (see page 254), also dealt with the effect of a thiophosphoric triamide
[N-(n-butyl)thiophosphoric triamide; nBTPTA] on maize. Three Illinois soils were
studied in 1984 and 1985. The N fertilizers: solid urea was surface-applied and UAN
solution was surface- and dribble-applied. Rates of additions per hectare were: 134 kg N
as urea + 0.67 and 1.0 kg nBTPTA; 202 kg N as urea + 2.7 and 4.0 kg nBTPTA; 134 kg
N as UAN + 0.56 and 2.24 kg nBTPTA; 202 kg N as UAN + 0.56 and 2.24 kg
nBTPTA.
nBTPTA significantly increased the grain yield only in two cases: in 1984 on one
soil which had been fertilized with 134 kg Nlha as dribble-applied UAN amended with
2.24 kg nBTPTAlha, and in 1985 on another soil fertilized with 134 kg Nlha as urea
amended with 0.67 and 1.0 kg nBTPTAlha.
Schlegel et al. (1986, 1987) evaluated, under field conditions in Indiana, the effect
of seven phosphoroamides, including nBTPTA and PPDA, on N uptake and grain yield
of maize in 1983 and 1984.
257
In the 1983 experiments, plots sited on a silty clay loam (pH 5.9) and a silt loam (PH
5.7) were installed. The sites were cropped to maize in 1982. The plots on silty clay
loam were tilled (conventionally tilled, CT plots), whereas those on silt loam were left
untilled except for chopping the stalks (no-tilled, NT plots). At day 10 (first soil) or at
day 21 (second soil) after sowing (at a population density of 64,000 seedslha), urea
(prills and solution) and UAN solution, at a rate of 84 kg Nlha, with or without a
phosphoroamide, were broadcast on the plots. UAN solution was also applied in bands
on the soil surface. Each of the six phosphoroamides tested (N,N-dimethyl-, N,N-
diethyl-, N-cyclohexyl- or N-benzyl-N-methylphosphoric triamide or trichloroethyl-
phosphorodiamidate, TPDA, or PPDA) was adhered to urea prills by first coating the
prills with paraffin oil (0.8% relative to urea weight) by mixing, then adding powdered
phosphoroamide at 2 kg/100 kg N and mixing. In the case of urea and UAN solution,
the phosphoroamides were dissolved and applied at a rate of 2.24 kglha.
The 1984 experiments were also carried out on two soils. The silty clay loam (PH
5.9) had been cropped to maize the previous year and was divided into two areas, CT
and NT plots. The NT plots, also in these experiences, were left untilled except for
chopping the stalks. The plots on the other soil (silt loam, pH 5.3), on which the
previous crop was winter wheat, were left untilled (NT plots). At day 15 (first soil) and
at day 12 (second soil) after sowing (60,000 seeds/ha), urea prills and UAN solution
with or without an inhibitor were broadcast on plots, at rates of 60, 120, and 180 kg
Nlha. Two inhibitors were tested: nBTPTA and PPDA. The rate of nBTPTA was 2 or
0.5 kg/IOO kg N (as coating on urea priUs) or 2.24 and 0.56 kglha (dissolved in UAN
solution). Rates of PPDA were 2 kg/100 kg N (on urea prills) and 2.24 kglha (in UAN
solution).
Fertilizer application was followed by a rainy period in 1983 but by a dry one in
1984.
In 1983 grain yield and N content in grain did not significantly increase under the
influence of inhibitors. In 1984 the inhibitors applied as coatings on urea prills led to
significant grain yield increases in the NT plots on both soils. Thus, average of grain
yields recorded at the three N rates was 7.46 tlha in urea-only treatment and 7.95 tlha in
urea + PPDA treatment on the silty clay loam soil; on the other soil, nBTPTA, at the
higher rate, increased the mean grain yield from 5.14 to 5.99 tlha. In addition, grain N
levels in the NT plots on silty loam were higher from urea with the higher nBTPTA rate
than those from urea alone. Urease inhibitors did not increase grain yield when applied
on CT maize or when added to UAN solution.
The conclusion was drawn that urease inhibitors will be most beneficial under the
following conditions: urea is applied on soil surface with large amounts of plant residue
cover; there is adequate moisture on soil surface to promote urea hydrolysis and
ammonia volatilization; the precipitations in the first 10 days after urea application are
insufficient to leach urea into the soil (see also Nelson et al.. 1986).
Leis et al. (1989) and Varsa et al. (1989) conducted seven experiments on no-till
maize fields on silt loam soils, at two locations (Belleville and Carbondale) in southern
Illinois in 1985, 1987, and 1988. nBTPTA was used in form of coating on urea
granules, copelleted with urea, and in urea and UAN solutions.
In 1985 urea coated with 0.92% nBTPTA was used at rates of 84 and 168 kg Nlha.
In 1987 and 1988, the urea-nBTPTA pellets containing 0.25 or 0.50% nBTPTA and
urea-nBTPTA and UAN-nBTPTA solutions containing 2.24 kg nBTPTAlha were used
258
at a single rate of urea, namely 134 kg N/ha. The controls received urea and UAN
without nBTPT A. In all years, the N fertilizers were broadcast or dribbled.
In six of the seven experiments, nBTPTA significantly increased the grain yield.
Lack of a yield response from nBTPT A was associated with a rain event of major
proportions soon after fertilizer application. nBTPT A was more effective with dribbled
than with broadcast urea; at 0.5% than at 0.25% nBTPT A content in urea pellets; and in
urea solution than in UAN solution.
The investigations were continued at both Belleville and Carbondale, and the results
obtained in 1990 were reported by Varsa (1991).
Granular urea and urea cogranulated with 0.33% nBTPTA, UAN solution with and
without 1.12 kg nBTPTAlha as well as granular ammonium nitrate (AN) were used at
rates of90, 134, and 180 kg N/ha. The fertilizers were all broadcast-applied.
At both locations, use urea cogranulated with nBTPT A resulted in significant grain
yield increases as compared with urea alone; addition of nBTPTA to UAN had no
significant increasing effect on grain yields; the significantly highest yields were
obtained with AN.
Varsa and Ebelhar (1992) reported on investigations conducted at Belleville and
Dixon Springs in 1991. The studied fertilizers were: urea, urea+0.3% nBTPTA. UAN,
AN as well as Super Urea and Super N. At Dixon Springs UAN with 1.12 kg
nBTPT A/ha was also used. All were broadcast and dribbled at rates of 90 and 180 kg
N/ha.
At Belleville the grain yields presented the orders: AN > urea-nBTPTA:::; urea>
UAN:::; Super N :::; Super Urea (broadcast fertilizers), and AN > urea-nBTPTA > urea:::;
Super N:::; UAN :::; Super Urea (dribbled fertilizers). The dribbled urea- nBTPTA
performed better than the broadcast urea-nBTPTA.
At Dixon Springs the orders were: AN > UAN-nBTPT A:::; Super Urea:::; urea :::;
Super N:::; urea-nBTPTA:::; UAN (broadcast fertilizers), and urea-nBTPTA > AN >
UAN-nBTPTA > UAN:::; Super N :::; urea:::; Super Urea (dribbled fertilizers). Thus,
urea-nBTPTA was more effective than AN. Urea-nBTPTA and UAN-nBTPTA gave
better results when dribbled than when broadcast. The active ingredients in Super Urea
and Super N were not as effective as nBTPTA in increasing grain yields.
Goodroad and Wilson (1989) conducted field experiments in the Piedmont region of
Georgia using urea fertilizer with or without nBTPTA. Grain yields of irrigated maize
were higher when urea+nBTPTA was used.
Bronson et al. (1990) also found that nBTPT A can increase the N use efficiency of
urea surface-applied to no-till maize.
In a short report on 2-year (1989-1990) field experiments carried out in North
Carolina, Baird and Ngueguim (1991) pointed out that in 1989 urea, as compared with
nBTPTA-amended urea, produced more grain yield, total dry matter, and total N
uptake, but in 1990 the nBTPT A-amended urea performed more efficiently than urea
alone with respect to grain yield, grain N uptake, and total above-ground dry matter
uptake ofN.
Schlegel (l99Ia,b) conducted field experiments on silt loam soils in west-central
Kansas, over 3 years (1988-1990), for evaluating the effect of nBTPTA to reduce the
damage caused to maize by ammonia released from urea under the action of soil urease.
Size of plots was 3 by 9 m. UAN solution (0, 12.5, 25, 50, and 100 kg N/ha) and
nBTPT A (0 and 1.12 kglha) were applied in dribble band with seed at planting. The
259
addition of nBTPTA allowed the rate ofN to be doubled without reducing emergence or
early plant growth and reduced delays in emergence by 50%. Thus, nBTPT A was
effective in reducing the phytotoxic effects of urea fertilizer of maize.
In field studies by Mullen et al. (1991) and Howard et af. (1992), 168 kg N/ha as
solid urea or liquid UAN with or without 1.12 kg nBTPTA/ha was broadcast on plots
planted to no-till maize on a silt loam soil with maize stubble or wheat residue cover.
Plots with injected UAN were the control.
Grain yields with urea+nBTPTA were not different from injected UAN yields for
both plant residues (approximately 5,895 kglha). The yields with urea alone were
significantly lower at approximately 4,421 kglha. For UAN broadcast in maize stubble,
the yields were 5,330 and 4,640 kglha with and without nBTPTA, respectively. For
UAN broadcast in wheat residue, the yields dropped to 4,703 and 3,198 kglha with and
without nBTPTA, respectively. Comparison of yield values makes it evident that
nBTPTA was more efficient in the urea than in the broadcast UAN treatments.
Fox and Piekielek (1993) studied the effect of urea-nBTPTA and UAN-nBTPTA on
no-till maize in three field experiments carried out on silt loam soils in central
Pennsylvania in 1989, 1990, and 1991. Rates offertilizer additions were: 56, 112, and
168 kg N/ha. The urea-nBTPTA contained 0.5% nBTPTA by weight. The nBTPTA
concentration in UAN was adjusted so that 1.12 kg nBTPTA/ha was applied at all N
rates. Urea treatments were broadcast at planting and applied as surface band at
sidedress. UAN was sprayed at planting, broadcast at planting, banded at sidedress, and
injected at sidedress.
All average grain yield was higher in the urea-nBTPTA than in the urea-only
treatment. The increase was significant at p=0.01 when urea and urea-nBTPTA were
broadcast at planting and at p=0.05 when they were surface-banded at sidedress. UAN-
nBTPTA compared with UAN increased significantly (p-0.1) the grain yield only when
these fertilizers were applied by spraying at planting.
Murphy and Ferguson (1992, 1997) evaluated, under field conditions, the effect of
nBTPTA-amended and unamended urea and UAN solution (28% N) on ridge-till,
irrigated maize. The investigations were carried out on silt loam soil at the University of
Nebraska South Central Research and Extension Center located near Clay Center over 3
years (1990-1992). Rate of N application was 112 or 224 kg/ha and that of nBTPTA
1.12 kglha. Methods of application for urea and UAN were broadcast and surface band.
Additionally, the subsurface band (knifed) method was applied for UAN.
It was found that the climatic conditions are the primary factors influencing the
effectiveness of nBTPTA to reduce rate of urea hydrolysis and ammonia volatilization
from urea and to increase the maize yield.
Precipitation soon (during the first 14 days) after fertilization in 1990 and 1991
resulted in little or no benefit from the use of nBTPTA, as precipitation moved urea into
the soil, reducing the potential for NH3 volatilization. Limited precipitation and low
humidity for an extended period following fertilization in 1992 resulted in an
approximately 3,600 kglha increase in grain yield when nBTPTA was applied with urea
(averaged over rates and application methods), but there was no yield increase when
nBTPTA was applied with UAN. No differences in yield were observed between
broadcast and surface band methods. In 1991 (but not in 1990 and 1992), nBTPTA in
the UAN applied by the knifed method led to a yield decrease of 2,156 kglha, perhaps
260
indicating that nBTPTA slowed the hydrolysis rate of urea too much than when VAN
was applied by the other methods.
The conclusion was drawn that in some years in south-central Nebraska the use of a
urease inhibitor such as nBTPTA will help protect surface-applied urea from volatile
NH 3 10ss.
Lamond et al. (1993, 1994) reported on field experiments conducted in 1993 and
1994 on no-till maize sites in Kansas. N fertilizers were surface-broadcast just prior to
or shortly after planting. In 1993 two continuous maize sites and two maize sites after
soybeans were studied. In 1994 four other maize sites were established. In both years,
ammonium nitrate and urea with nBTPTA performed better than urea and VAN. Thus,
in 1993, at three sites, urea + nBTPTA produced significantly higher grain yields than
urea and VAN.
In the 3-year field experiments of Palazzo et al. (1995, 1996) (see page 153),
nBTPTA addition to urea resulted in maize grain yield increases ranging from 11.0 to
30.6%. nBTPTA at a concentration of 0.1 % (on urea weight basis) was as efficient as at
its 0.25% concentration.
See also Table 61.
* *
*
Results of 78 field experiments conducted from 1984 to 1989 across the V.S.A. for
evaluation of the effect of nBTPTA on maize were summarized by Hendrickson (1992).
nBTPTA was applied with urea at rates of 0.25 to 1.0% (weight/weight) and with VAN
solution at rates of 0.56 to 2.24 kglha. On overall average, nBTPTA increased grain
yields by 4.3 bu/acre when applied with urea and by 1.6 bulacre when applied with
VAN. Average responses to nBTPTA were greater on sites responsive to N (6.6 bulacre
for urea and 2.7 bulacre for VAN). Results from 21 experiments employing multiple N
rates showed that maximum grain yields could be obtained using an average of 83 kglha
less N when nBTPTA was included with surface-applied urea.
Varsa et al. (1993) summarized the results from 7 years of experiments on no-till
fields at two locations in southern Illinois (Belleville and Carbondale). nBTPTA
addition to broadcast urea, when evaluated across N rates and locations, gave grain
yield increases averaging 8.4 bulacre in 13 experiments. For dribbled urea the response
to nBTPTA was 12.0 bu/acre for 9 experiments across the two locations. Grain yield
responses to nBTPTA added to VAN solutions were much smaller. In 8 experiments
broadcast VAN-nBTPT A resulted in an average grain yield increase of 2.3 bulacre
across the two locations. In 13 experiments, in which dribbled VAN-nBTPTA was
evaluated, yield increases of 3.3 bu/acre were obtained.
The manufacturer of the commercial nBTPTA under the registered trade product
name of Agrotain has elaborated a collection of 38 summaries of reports presenting
results obtained with Agrotain in experiments carried out on maize fields at a great
number oflocations in the U.S.A., namely in 19 states (IMC, 1995).
7.1.8. Effect ofCyc!otriphosphazatriene (CTPAT) Derivatives

Zehruilek et al. (1990) dealt with the effect ofhexaamino-CTPAT on plants in a pot and
a field experiment. The test plant was maize. The Mitscherlich pots contained 6 kg of
261
soil-sand mixture (1:1). After sowing, 100 ml of urea solution (1.2 g N) with or without
2% hexaamino-CTPAT (relative to weight of urea) was applied on the surface of soil. In
the field experiment, the fertilizer was DAM 390 at rates of 100 and 200 kg Nlha,
whereas the rate of hexaamino-CTP AT was I % relative to fertilizer N. In both
experiments, the over-ground parts of 43-day-old plants were analyzed for fresh and dry
weights and N, P, Ca, and Mg contents.
In the pot experiment, fresh and dry weights and N contents were higher, and P, K,
Ca, and Mg contents were lower, in the urea-hexaamino-CTPAT treatment than in the
urea-only treatment. The weight increase was about 10%. Similar results were
registered in the field experiment, but Ca and Mg contents were higher in plants
growing in plots received DAM 390 + hexaamino-CTPAT.
See also Table 61.
7.1. 9. Effect ofAntimetabolites

Of the five antimetabolites patented by Peterson and Walter (1970) for inhibition of soil
urease activity (see page 163), three were selected (pyridine-3-sulfonic acid,
oxythiamine chloride, and o-chloro-p-aminobenzoic acid) for studying their effect on
growth of maize plants. The antimetabolites were added in amounts ranging from about
2.2 to 17.8 kglha to different areas of soil fertilized with urea and planted to maize. No
phytotoxic effects were noted.
7.1.10. Effect ofLignosulfonates

In the field experiments conducted by Alkanani and MacKenzie (1996), two soils,
representing the major maize-growing areas in eastern Quebec, were chosen (a silty
clay, pH 6.2 and a clay, pH 5.9).
The plots (3 by 5 m) received banded solutions of ammonium lignosulfonate (ALS)
(8 kg Nlha), urea (30 and 90 kg Nlha) + diammonium phosphate (DAP) (14 kg Nlha),
urea + DAP + ALS, applied at planting 5 cm to the side and 3 cm below the seeds.
Immediately after planting and banding, the plots received broadcast KCl (I80 kg Nlha)
and three rates (59, 119, and 163 kg Nlha) of broadcast granular urea which were
immediately incorporated into the soil. There were also plots which were treated with
banded ALS (8 kg Nlha) and broadcast urea (163 kg Nlha). Total N applied was 163
kglha (in plots with no ALS addition) and 163+8=171 kglha (in plots with added ALS).
The control plots received no urea, DAP, and ALS.
In comparison with the control, ALS alone had no effect on grain yield on the silty
clay and had an insignificant (3.6%) increasing effect on the clay soil. When applied
with broadcast urea, ALS induced a 21.8 and 19.6% increase in grain yield on the silty
clay and clay, respectively.
The yield increase in the treatment with 30 kg Nlha of banded urea + DAP was 30.9
and 23.2%, respectively, and it was higher in the 30 kg Nlha + DAP + ALS treatment
(34.5 and 33.9%, respectively). In the 90 kg Nlha banded urea + DAP treatment, the
yield increase was only 10.9 and 10.7%, respectively, and became 27.3 and 25.0%,
respectively, when ALS was also applied. These observations suggest that the banded
urea at its higher rate had an adverse effect on the grain yield, and ALS mitigated this
adverse effect.
ALS increased the uptake of N from urea by the maize plants and, thus, led to
increased crude protein content in grain on both soils.
262
In separate field experiments, 15N-labeled urea (at 33 atom% excess of 15N) was
used, and it was found that ALS resulted in significantly less 15N immobilized in soils.
7.1.11. Effect a/Plant Materials

Sarsaponin, which is, as shown on page 171, a Yucca schidigera extract containing a
mixture of steroid saponins, was tested by Broadbent et af. (1985) in a field experiment
in which three phosphoroarnides were also tested (see page 256). Sarsaponin at a rate of
3.2 kg/ha was added to solution of 15N-labeled urea (160 kg Nlha). The test plant was
maize. The analyses showed that in the urea + sarsaponin treatment an insignificant or
significant reduction occurred in total N and 15N contents in the leaves of 29-, 60-, and
84-day-old plants as compared to urea-only treatment. At plant maturity, total dry
matter and grain yield decreased insignificantly, whereas total N and 15N contents
decreased significantly under the influence of sarsaponin.
Vyas et al. (1991) established that the maize plants took up more N from urea
granules coated with neem extract (nitrification and urease inhibitor) than from
uncoated urea. The fertilizer N use efficiency was 56.4% from coated urea-N and 35.5-
44.4% from uncoated urea-No
In a 2-year field experiment on a sandy loam soil (PH 8.0) at the Indian Agricultural
Research Institute, New Delhi, Sharma and Prasad (1996) compared the effects ofneem
cake (considered as an inhibitor of nitrification only) and dicyandiarnide (DCD), both
applied with urea, on the grain yields in two crop years 199011991 and 199111992 in a
maize (June to October)-wheat (November to April) rotation. Prilled urea (100 kg) was
coated with neem cake (20 kg) (NCCU). DCD (7 kg) was blended with urea prills (93
kg) (DCDU). Prilled urea (PU) alone served for comparison. Rates of N addition were
0, 60, and 120 kg/ha. The control received no N fertilizer.
In 1990 maize responded well to 60 kg Nlha. At this N rate, PU increased maize
yield by 1.03 tlha, whereas NCCU and DCDU increased maize yield by 1.55 and 1.18
tlha over the control. Thus, NCCU was more efficient than DCDU. The yield increases
produced by NCCU and DCDU were equivalent to 127 and 94 kg Niha as PU,
respectively. Furthermore, when the results were averaged over the two years of
experiment, residual N from NCCU and DCDU at 60 kg/ha left after maize cropping
increased the grain yield of the succeeding wheat grown with 60 Nlha as PU by 1.97
and 1.68 tlha, respectively, over the control or 60 kg Nlha as PU applied to the maize.
This was equal to 96 and 82 kg Nlha as PU to wheat.
7.1.12. Effect a/Combined Urease and Nitrification Inhibitors

The maize fields on which the granular urea+DCD+ATS fertilizer patented by Sutton et
at. (1991) (see page 244) was tested are located in Kentucky, Illinois, and Indiana. Urea,
urea+DCD or ATS, ammonium polyphosphate (APP), ATS+APP served for
comparison. Rate of addition was 156 kg Nlha. Results of three experiments, each
repeated nine times, are described in the patent. Average values of grain yields are
presented. In two experiments, the patented fertilizer was most effective: the yield
increase was 8 and 17%, respectively, in comparison with the urea treatment. In the
third experiment, the yield obtained with the patented fertilizer was 97% of the yield
registered in the urea treatment.
In an experiment conducted by Lamond et al. (1994) in 1994, plots on an irrigated
no-till maize field at Sandyland, Kansas, were fertilized with 1. urea, 2. urea +
263
nBTPTA. 3. urea + DCD, and 4. urea + nBTPTA + DCD at rates of 0, 67, l34, and 202
kg N/ha. The following grain yields (bu/acre) were regictered with the four fertilizers:
143 (no N added), 174, 188, 174, and 176 (at 67 kg Nlha), 190, 192, 183, and 202 (at
l34 kg Nlha), and 196, 198, 183, and 186 (at 202 kg Nlha), respectively. Thus,
urea+nBTPTA was most efficient at the lowest and highest N rates and
urea+nBTPTA+DCD at the medium N rate, but the differences between the yields were
not significant at p=0.05.
Varsa and Jan (1997) conducted experiments in 1992 and 1994 to 1996 at
Carbondale, Kansas, to evaluate the effect of nBTPTA and DCD alone and in
combination on no-till maize. The urea rates were 90, 135, and 180 kg Nlha. nBTPTA
and DCD were incubated alone with urea at concentrations of 0.14 and 2.2%,
respectively. Combination included nBTPTA at 0.14% and DCD at 0.55 and 1.1%.
When a response to inhibitors was obtained, usually nBTPTA alone was the highest or
equal to urea treatments that included a combination of nBTPT A and DCD. Urea plus
DCD alone never increased the maize yield over urea. Inhibitor response failures were
usually associated with seasons of severe drought or when high N levels remained in the
soil from the previous crop.
7.2. EFFECT OF UREASE INHIBITORS ON WHEAT (Triticum aestivum)
7.2.1. Effect of Fluorides

root growth of wheat plantlets was strongly reduced at rates 2: 0.5 g F/kg soil.
7.2.2. Effect of Inorganic Sulfur Compound~'

Ammonium thiosulfate (ATS) gave good results in experiments carried out on wheat
fields in Idaho. Thus, in Kissel's (1984) experiments, ATS improved winter wheat
production when banded below the seeds. Mahler and Lutcher (1989) applied ATS and
urea-ammonium nitrate-ATS (UAN-ATS) solutions and solid AN and ammonium
sulfate (AS) mixture (AN+AS) at rates of 0, 45, 90, and 135 kg Nlha and 0 or 22.5 kg
S/ha to N- and S-deficient silt loam soils. No N and S fertilizers were added to the
control. Three experiments were carried out in 198211983 and three more experiments
in 1983/1984. The plot size was 3 by 9 m (in 1982/1983) and 2.75 by 9 m (in
1983/1984). The fertilizers were applied to the soil surface in the autunm prior to
seeding winter wheat and topdress-applied in the spring at Zadok's growth stage 21, 33
or 37.
ATS applied alone, in the autumn or in the spring before the plants reached growth
stage 22, improved winter wheat yields over the unfertilized control and was as
effective as AS in supplying S, and UAN-ATS and AN+AS were equally effective as N
sources.
But ATS, topdress-applied after the plants reached growth stage 21, resulted in
tissue damage which translated into yield loss. Consequently, UAN-ATS which
provides both N and S for wheat plants should be applied in the autUnID or in the early
spring up to the time plants reach growth stage 22.
In the field experiments conducted by Scharf and Alley (1987) over two growing
seasons at two Virginia locations each season, ATS and ATS added to urea or UAN
264
solution were spring topdress on winter wheat. None of the treatments significantly
increased yield in any experiment, or consistently increased mid-season N uptake.
Bremner et af. (1990) and McCarty et af. (1990,1991) found that the adverse effect
of ATS, sodium thiosulfate, and tetrathionate on germination of maize and wheat seeds
and growth of maize and wheat seedlings is similar (see page 253).
7.2.3. Ej{ect o/Organic Mercury Compounds

May and Douglas (1975, 1978) used two techniques to study the effect of
phenylmercuric acetate (PMA) on germination of wheat seeds. In the first technique,
paper towelling was placed in sterile plastic Petri dishes, moistened and then treated
with 0, 0.003, 0.015, and 0.03 g of powdered PMA/dish. Twenty-five wheat seeds were
placed on the moistened, treated paper. The next step was incubation (at 20°C for 8 days
in the dark); thereafter, the number of seeds that had germinated was recorded. It was
found that PMA, at each concentration, significantly inhibited the germination.
In the second technique, 50 g air-dried soil (grey clay, pH 8.9 or solonized brown
soil, pH 7.9, both from Victoria, Australia) replaced the paper towelling in the Petri
dishes. The wheat seeds (in the same quantity as in the first technique) were placed at a
depth of 5 mm in soil. PMA was applied in solution at rates of 0, lO, 50, and lOO ppm
(on soil basis). Incubation proceeded and germination was recorded as in the first
technique. In contrast with the results registered in the first germination test, PMA
applied to soil did not significantly inhibit germination, except in a single case, namely:
PMA at the 100 ppm rate significantly inhibited germination in the brown soil.
Rao and Ghai (1986a) studied the effect ofPMA on germination and yield of wheat
and also on N uptake by plants in vegetation pots filled with an Indian alkali soil (sandy
loam, pH 9.3). Soil (20 kg/pot) at field capacity, 20 mg of P (as single
superphosphate)/kg soil and 10 mg of ZnSOJkg soil were mixed and treated, in two
splits, with urea or urea + PMA mixture at a total rate of 40, 80, or 120 mg N/kg soil;
PMA was applied at a rate of 10% by weight of urea. One half of the urea or urea +
PMA mixture (basal dose) was mixed as a solid in the surface 5 em of soil. Then wheat
was sown (20 seeds/pot). After 5 weeks of growth, the other half of urea or urea + PMA
mixture was topdressed. Germination counts were made 15 days after sowing. At day
21, the plants were thinned and 8 plants were maintained per pot. A total of 9 irrigations
(45 cm water) were applied during the 125 days of growing season. Total dry matter
and N uptake were recorded at harvest. The results showed that PMA had no adverse
effect on germination. Under its influence, dry matter in straw decreased significantly,
but grain yield and N content in straw and grain remained similar to those recorded in
the control (treated with urea only).
See also Table 61.
7.2.4. Ejfect ojDithiocarbamates

Tomlinson (1967), patenting dithiocarbamates as inhibitors of soil urease activity,
applied two testing methods. The first method is described in Subchapter 2.6 (see page
53). The second method is described below. This method is based on the following
principle: brairding of (wheat) seedlings is hindered by the alkalinity resulted from the
rapid hydrol ysis of urea; thus, the action of the urease inhibitor will manifest itself by an
increased number of the emerged shoots. About 1.5-kg samples of five soils were
placed in vegetation pots and watered. Circular drills were made in each pot and urea
265
not treated or treated with inhibitor(s) was distributed along the bottom of the drills.
Then wheat seeds were placed on the bottom of drills and covered with the soil. The
treatment of urea consisted of mixing urea granules or prills with powdered inhibitor(s).
Urea was applied at rates of 16.8 and 33.6 kg N/ha (in one soil), 44.8 or 56 kg N/ha
(four soils). The inhibitors tested were: zineb 1% (on urea weight basis) + Br3C-CBr3
1% (one soil); zineb 0.5% and zineb 0.5% + ClzBrC-CBrC12 1% (four soils). The
control soils in pots were not treated with either urea or inhibitor(s). After sowing, all
pots were stored in the dark at lO°C. The shoots emerging above the soil surface were
counted every second or third day for 35 days.
One can deduce from the results tabulated in the patent that urea at low rate (16.8 kg
N/ha) with or without zineb + Br3C-CBr3 retarded emerging of shoots, but at day 35 the
number of shoots emerged from these treated soil samples did not differ from that
registered in the control soil. In the soil samples treated with 33.6 kg urea-N/ha, the
number of emerged seedlings was lower than in the control soil during the whole 35-
day period. This effect of urea was attenuated. to some extent, by the zineb + Br3C-
CBr3 mixture. At the rates of 44.8 and 56 kg N/ha, urea strongly inhibited emerging of
shoots. Thus, at day 35 the number of emerged seedlings in the four soils treated with
urea at these rates varied between 5 and 25%, whereas 84-92% of the seeds produced
shoots in the control soils. Zineb (0.5%) and zineb (O.5%) + C1 2BrC-CBrClz {l %}
attenuated somewhat the effect of urea, increasing the number of emerged shoots to 13-
26%. The action of zineb did not differ from that of the zineb + ChBrC-CBrCh
mixture. This means that CI 2BrC-CBrClz, which, when tested with the first method
(page 54), was more effective than zineb in reducing volatile ammonia losses, had no
inhibitory effect on soil urease activity in the brairding experiments.
7.2.5. Effect ofHydroxamic Acids

May and Douglas (1975, 1978) studied the effect of benzohydroxamic acid (BHA) on
germination of wheat seeds. They used the same two techniques, the same two
Australian soils and the same inhibitor concentrations already used for studying the
effect of PMA on germination of wheat seeds (see page 264). The results showed that
germination of wheat on paper towelling was strongly inhibited by the higher
concentrations of BHA. Contrarily, none of the BHA concentrations affected
germination of wheat in any of the soils.
7.2.6. Effect ofBromo-nitro Compounds

In the pot experiments performed by Norden et al. (1985, 1986), urea and urea-
ammonium nitrate (UAN) solutions with or without a bromo-nitro compound (BNC),
namely 2-bromo-2-nitropropyl-N-methylcarbamate as urease inhibitor, were applied for
foliar fertilization of spring wheat plants. Total amount of N applied in urea and UAN
solutions was equal, approximately 0.15 g/pot, and in the UAN solution content of urea-
N was equal with the N content of ammonium nitrate. The rates of inhibitor addition per
weight of urea-N were 2 and 4% (in urea solution) and 4 and 8% (in UAN solution).
Optimal crop yield, N uptake, and raw protein content were achieved with 4% inhibitor
concentration in both urea and UAN solutions.
266
7.2.7. EfJect ofPolyhydric Phenols and Quinones

In the field experiments conducted by Tomlinson (1970) in England, winter wheat plots
were fertilized with urea (50 or 100 kg Nlha) with or without I % (relative to urea-N) of
hydroquinone or 2,5-dimethyl-p-benzoquinone. No significant differences were
observed in crop yields between the treatments with urea only and with urea + inhibitor.
For evaluation of the effect of catechol, hydroquinone, p-benzoquinone, 2,5-
dimethyl-p-benzoquinone, and o-naphthoquinone on the germination of wheat seeds,
May and Douglas (1975, 1978) used the same two techniques and the same two
Australian soils they also used for studying this effect of PMA (see page 264). Rates of
inhibitors were also the same: 0.003,0.015, and 0.03 gldish, and 10, 50, and 100 ppm
relative to soil weight, respectively.
At rates of 0.003 and 0.015 gldish, inhibition of germination was not always
significant, but at the rate of 0.03 gldish all these compound significantly inhibited the
germination. All rates of 2,5-dimethyl-p-benzoquinone were strongly inhibitory.
Germination in soil was affected to a lesser extent. Thus, at 10 ppm none of these
compounds inhibited significantly the germination in either of the two soils studied. The
germination was significantly inhibited in both soils by 2,5-dimethyl-p-benzoquinone at
50 and 100 ppm and by a-naphthoquinone at 100 ppm.
In another experiment performed in vegetation pots, May and Douglas (1978) used
an acid (pH 4.6) sandy soil (2.6 kglpot). A basal dressing containing P, K, Cu, Mn, Zn,
Mo, B, Fe, and lime (to raise pH to 6.5) was given to soil. Six wheat seeds were sown in
a 2.5-cm deep furrow in the soil. Urea at a rate of 13.4 or 53.6 ppm ofN (on soil basis)
with or without an inhibitor was placed in this furrow. Catechol, p-benzoquinone, and
2,5-dimethyl-p-benzoquinone were added as aqueous solutions at a rate of 2.9 ppm.
Catechol was also added at this rate as a solid. Application of inhibitors at the rate of 2.9
ppm was estimated to result in a concentration of approximately 50 ppm in close
proximity to seeds. Three weeks after germination, plants were thinned to two per pot.
At maturity, grain yields and N contents in plants were determined.
Data of Table 56 show that urea at the lower rate (13.4 ppm N) led to an increase in
grain yield which was not affected significantly by the inhibitors. At the higher rate
(53.6 ppm N), urea caused the death of all plants which is explained by the phytotoxic
concentration of ammonium released from urea. In the presence of inhibitors, except
TABLE 56. Effect of urea and urease inhibitors on wheat grain yield"
Yield (g dry matter/potl
Treatment Urea-N awlication rate (Wm of soil)
13.4 53.6
Control (urea with no inhibitor) 3.00 ab 0 c
Catechol (solid) 2.87 ab 3.14 ab
Catechol (solution) 2.32 b 0 c
p-Benzoquinone 2.89 ab 4.16 a
2,5-Dimethyl-p-benzoquinone 2.86 ab 3.84 a
"From May and Douglas (1978).
"when no urea or inhibitor was applied, a yield of 1.29 g/pot was obtained.
Yields not followed by the same letter are significantly different (p=O.05).
267
catechol in solution, soil urease was inhibited, NH4 + in phytotoxic concentration was not
produced and, thus, the plants could grow. Inefficiency of catechol in solution is
attributed to its migration from the soil zone near the germinating seeds.
At the 13.4 ppm urea-N rate, little increase in N uptake by plants occurred when
urea was applied with an inhibitor as compared to urea applied alone. In treatments
where 53.6 ppm urea-N + either catechol (solid), p-benzoquinone or 2,5-dimethyl-p-
benzoquinone was added to soil, N recoveries by plants were 60, 57, and 72%,
respectively.
Mishra et al. (1980) also used wheat as a test plant. Samples (300 g) of a black earth
from Germany were mixed with sand (100 g), then treated with 5 ml of solutions of the
compounds specified in Table 57 (to obtain concentrations of 20 and 50 ppm in soil),
moistened to 60% of WHC and sown with 100 wheat seeds. Germination and, then,
growth took place at 20°C. The number of germinated seeds was determined after 4
days, the height of plantlets was measured after 8 days, and the dry weight of roots and
TABLE 57. Effect of urease inhibitors on germination and growth of wheat"

Concentration Number of Height of Dry weight (g)
Compound of compound germinated plantlets Root Shoot
(ppm of soil) seeds (em)
No (control) 0 92 7.4 0.75 2.06
p-Naphthoquinone 20 93 7.6 0.75 2.10
50 90 8.2 0.76 2.10
2-Methyl-p-naphthoquinone 20 97 7.1 0.80 2.08
50 90 5.9 0.80 2.04
2.3-Dichlorohydroquinone 20 96 8.3 0.73 2.05
50 89 9.0 0.72 2.08
4-t-Butylcatechol 20 94 7.8 0.74 2.06
50 92 7.5 0.70 2.02
4,6-Di-t-butylcatechol 20 91 7.5 0.78 2.00
50 91 6.3 0.72 2.01
"From Mishra et al. (1980), by permission of Kluwer Academic Publishers.
shoots was recorded aftcr 21 days. One can deduce from this table that none of the
compounds affected germination of wheat seeds. After 8 days, the plantlets were shorter
with the addition of 2-methyl-p-naphthoquinone and 4,6-di-t-butylcatechol at 50 ppm,
but this adverse effect disappeared later and, thus, after 21 days of growth dry weight of
plants was practically the same in the control and in all treatments.
In a short report, Edwards (1982) described a field experiment, in which winter
wheat was fertilized with prilled JSN-labeled urea (l00 kg N/ha) with or without
addition of p-benzoquinone (BQ) at a rate of 2.5% relative to urea-No Neither urea nor
BQ was added to the control. Under these conditions, BQ increased the uptake of urea-
N by an average of 7 kg/ha reduced plant uptake ofN from soil by 34% compared to the
unfertilized control and by 57% compared to the treatment with urea alone. Grain yields
were increased by \,280 kglha from the urea + BQ treatment and by 2,260 kg/ha from
the application of urea alone. It is assumed that the response difference represents the
summation of the negative effect of BQ on the mineralization of soil organic N and the
positive effect of urea on the release of complexed soil organic N.
268
Under conditions identical to those described on page 264, Rao and Ghai (l986a)
also studied the effect of catechol (CT) and hydroquinone (HQ), used at a rate of 10%
relative to weight of urea, and established that percentage of germinated wheat seeds,
estimated at day 15 after sowing, was not affected by CT but was reduced by HQ. A
similar situation was observed concerning dry matter of 21-day-old plants. Contrarily, at
day 35 after sowing, dry matter of plants in both CT and HQ treatments did not
significantly differ from that measured in the control plants (treated with urea alone),
whereas at the end of the growing season (125 days), grain yield was highest in the urea
+ HQ treatment, followed by the urea + CT treatment and exceeding by 20.0 and 14.8%,
respectively, the yield of the control plants. N content in grain also increased in the
order:
control (urea) < urea + CT < urea + HQ.
Dry weight of straw showed the order:
urea + CT < control (urea) < urea + HQ.
Accumulation of N in straw was not influenced by CT and HQ.
The explanation for the more favorable effect of HQ than of the CT is that HQ
moves together with urea, which means that it moves more rapidly as compared to the
movement of CT.
Hera et al. (1986) compared the effects of hydroquinone (HQ), nitrapyrin, and 4-
arnino-1,2,4-triazole hydrochloride (ATC) on the yield of wheat cultivated on a
chemozemic soil in vegetation pots. Urea (0.5 glpot) was applied only in autumn at
sowing time or in both autumn and spring (0.5 + 0.5 glpot). The inhibitors were used
together with urea at a rate of 5 mglpot. The controls received no urea and/or no
inhibitor. On 1st of June, some pots were submitted to simulated rainfalls (50 mm) for
creating excessive moisture, i.e.. conditions favorable for leaching of nitrates. The soil
in the other pots was maintained at optimum moisture content. Periodical analysis of
nitrates formed in soil showed that the inhibitors delayed the appearance of the
maximum nitrate amount from the time of stalk shooting (control) to times of
inspication (HQ), flowering (nitrapyrin), and maturation (ATC). The wheat yield
increased due to inhibitors in the following order: HQ>nitrapyrin>ATC. These
differences between the three compounds were more marked under conditions of
excessive moisture, although under these conditions, HQ, nitrapyrin, and ATC reduced
the losses through leaching of nitrates by 42.4, 52.9, and 61.2%, respectively, i.e., in an
order opposite that of their yield-enhancing effect.
The results obtained in pot experiments carried out by Zhou et al. (1988) for
studying the effect of HQ on spring wheat plants sown in a brown soil from China are
presented in Table 58. They show that the optimum amount of HQ for increasing plant
uptake ofurea-N was 10 mg with 0.9 g ofurea-N, whereas HQ at the rate of 40 mglpot
reduced uptake of N from urea. Grain yield was also highest at 10 mg of HQ/pot which
led to a significant (l 0.23 %) increase as compared to the control treated with urea
alone.
In another laboratory experiment, in which urea labeled with 15N was applied
together with HQ at rates of 0, 10, 20 or 40 ppm (on soil basis), the spring wheat plants
growing in soil treated with 0, 10, and 20 ppm of HQ took up more N from urea than
from soil (the ratio of N taken up from soil to N taken up from urea < 1) during the
whole period of experiment (90 days). At 40 ppm of HQ, this ratio was also smaller
269
TABLE 58. Effect of hydroquinone, applied at various rates, on uptake ofurea-N by spring wheat plants"
Rate of Rate of urea-N Urea-N taken up by Gaseous loss of Residual urea-N
hydroquinone plants urea-N in soil
(mg/pot) (g/po!) (g/pot) (%) (g/pot) (%) (g/pot) (%)
0 0.9 0.4403 48.92 0.4078 45.31 0.0519 5.77
10 0.9 0.4733 52.59 0.3655 40.61 0.0612 6.80
20 0.9 0.4678 51.98 0.3199 35.54 0.1123 12.48
40 0.9 0.3167 35.19 0.4106 45.62 0.1727 19.19
"From Zhou el al. (1988).
than 1 during the first 60 days; thereafter, it became greater than 1. But at all HQ rates,
utilization ofN from soil increased linearly with time.
It was also established that HQ applied at rates of 5, 10, 20, and 40 mg/pot did not
accumulate in soil, stems and grains of wheat (sec also Zhao et al.. 1991, 1992a; Zhou
et aI., 1992).
Xue et al. (1991) amended wheat fields with 15 kg of urea + 10 kg of KCl + 0 or 50,
100 or 150 g of hydroquinonc (HQ) or quinhydrone (QH). All rotes refer to an area of 1
mu' (1/15 ha). The groin yicld increased with increasing inhibitor rate. The increases
were 6.8, 14.1, and 19.8% (HQ), and 11.8, 16.9, and 22.8% (QH). The yield increases
were accompanied by increases in coefficient of plant utilization of N from urea, from
2.8 to 5.3% in the HQ treatments, and from 2.4 to 6.1 % in the QH treatments.
See also Table 61.
7.2.8. Effect of Phosphorodiamides

Matzel et at. (1979a,b) described experiments performed, in the 1976-1977 period, in
Mitscherlich pots, each containing 6 kg of soil (loamy sand, pH 5.4). The basal
fertilization with 1 g of K and 0.4 g of P/pot (as K2HP0 4) and with 0.4 g of N/pot (as
unlabeled NH 4N0 3) was followed by sowing of spring wheat (20 plants/pot). As soon as
the plants had reached a height of 25 or 40 cm, a second amount of 0.4 g of N/pot in
form of 15NH415N03 (12.0 atom% excess 15N)** or 15N-labeled urea (12.0 or 11.2 atom%
15N)*' or labeled urea mixed with 1% (on urea weight basis) of phenylphosphoro-
diamidate (PPDA) was applied on the soil surface. In 1976, at both 25- and 40-cm high
plants, the fertilizers were immediately washed into the soil with 200 ml of water. In
1977 the soil surface was moist when the plants were 25-cm high and dry'" when the
plants were 40-cm high; fertilizers were washed into the soil (with 50 ml of water) only
at day 14 after their application. Following the second fertilization, at different intervals
(including maturity stage of plants), the plants and soil were analyzed for total Nand
15N; crop yields (groin and straw) were also determined.
'Mu is a traditional Chinese unit of land surface area which in modem China is reckoned to be 1/15 ha.
"In the balance studies, the labeled NH 4 N0 3 and urea contained 52.2 and 50.6 atom% excess lIN,
respectively.
"'Water to the soil mass was supplied through a strip of glass fibre cloth, which was embedded in the soil
and. passing through the bottom of the pot, immersed in a subjacent water container.
270
One can see from Table 59 that, when the fertilizers were immediately washed into
the soil, the grain yield increased, though insignificantly, in order:
urea < urea+ I % PPDA < NH4N0 3,
whereas the straw yield was highest in the urea treatment; more N was taken up by
plants from NH4 N0 3 than from urea, and PPDA did not enhance plant uptake of N.
TABLE 59. Wheat yield and nitrogen uptake from fertilizers under different conditions of their application"
Wheat yield N uptake (% of the
(g/pot) applied N with the
Fertilizer Conditions of application
second dose)
Grain Straw Grain Straw Total
Experiments performed in 1976
NH 4NO, Application at a plant growth 41.6a 57.2 ab 49 15* Ma
Urea height or 25 em; immediately 40.7 a 60.6 b 43 a 15 58 b
Urea + 1% PPDA washed into the soil 41.3 a 57.5 ab 44 a 14 58 b
NH4NO, Application at a plant growth 43.6 a 53.1 a 57 14 71
Urea height of 40 cm; immediately 40.2 a 53.6 a 52 b 13 65 a
Urea + 1% PPDA washed into the soil 41.1 a 44.9 53 b 12 65 a
Experimel1lS performed in 1977
NH 4 NO J Application to a moist soil surface 32.5 a 39.3 a 58 a 15 73
Urea at a plant growth height of 25 cm; 30.1 a 38.2 a 40 II 51
Urea + 1% PPDA washed into the soil after 14 days 34.4 41.6a 54 be 13 67 a
NH 4 NO J Application to a dry soil surface 21.6 b 28.3 b 56 ab II 67 a
Urea at a plant growth height of 40 cm; 22.7b 29.3 b 47 d 9 56
Urea + 1% PPDA washed into the soil after 14 days 24.5 b 32.5 51 cd II 62
"Adapted from Matzel et al. (1979a,b).
The same letter indicates insignificant differences (p=O.05).
"No statistical evaluation.
Grain yield was not influenced significantly by timing offertilizer application (at 25- or
40-cm high plants), but total N uptake was higher when the fertilizers were
administered for 25-cm high plants than for 40-cm high ones. When the fertilizers were
not washed into the soil immediately but only at day 14 after their surface application,
the grain and straw yields decrcased, especially following fertilizer application on the
dry soil surface, at 40-cm high plants. The plants took up less N from urea than from
NH4N0 3 or urea+ I % PPDA; the uptake was minimal when urea was applied on moist
soil surface. PPDA increased both crop yield and plant uptake of N from urea; this
effect was more consistent with urea application on moist soil surface.
Balance offertilizer N at maturity stage of wheat in 1977 (Table 60) shows that the
plants took up by 11 and 5% more N from NH4N0 3 than from urea and urea+ I %
PPDA. respectively. This means that PPDA increased the uptake of N from urea. At the
same time, PPDA enhanced immobilization of urea-N in soil and reduced the N deficit
from 32 to 22%, i.e., to the same level as that recorded in the NH4N0 3 treatment. In
other words, application of urea alone led to a 10% loss by NH3 volatilization, but this
loss could be prevented by adding PPDA to urea.
271
TABLE 60. Balance of fertilizer nitrogen at maturity stage of wheat in an experiment

performed in 1977"
Fertilizer N at maturity stage of wheat (at day 69 after fertilization)
(% of the applied N)
Fertilizer"
In plants
In soil Deficit
(including roots)
NH4 NO, 69 9a 22a
Urea 58 10 a 32
Urea + I%PPDA 64 14 22 a
"Adapted from Matzel et al. (1 979a.b).
"Conditions of fertilizer application: at a plant growth height of 40 cm; washed into the soil
after 14 days.
The same letter indicates un significant ditferences (p=O.05).
Kampfe et al. (l982b) reviewed the results of 136 winter cereal (wheat and rye)
experiments carried out under field conditions, on mostly sandy soils as well as on
loamy sands, in the 1974-1979 period. In the review, the wheat and rye are evaluated
together as winter cereals.
Conditioned urea with or without 0.5 or 1% PPDA (on urea-N basis) and lime
ammonium nitrate were the fertilizers tested. They were surface-applied and not worked
into the soil.
Grain and straw yields and N uptake by plants on 9-17 _m2 plots were determined.
In 1974 and 1975, at the beginning of the growing season, 50 kg of N/ha were
applied. In the 1976-1979 period, the same amount of 50 kg of Niha was administered
at the beginning of the growing season, but later a second amount (40 kg ofNIha) was
added. The plots were also fertilized with 30 kg of P and 100 kg of Klha at the
beginning of growing season in each year.
For evaluation of the effect of N fertilizers on crop yield and N uptake by plants,
only the results of 21 winter wheat and 80 winter rye experiments conducted in the
1976-1979 period were taken into consideration, but for evaluation of the relationship
between crop yield and the amount of rainfall during the first 5 days after fertilization
the results registered in 1974 and 1975 were also taken into account.
It was established that the grain yield was significantly smaller in the treatment with
conditioned urea than in that with lime ammonium nitrate, when fertilization was
followed by dry weather. In this case, PPDA reduced the grain yield losses. Reduction
of these losses under the influence of 1% PPDA was total in loamy sands and about
50% in sands; at the 0.5% PPDA rate the effect of 1% PPDA was attained only in the
sands. PPDA also improved N uptake by plants. Straw yields were nearly the same in
all treatments.
When it rained during the first 5 days after fertilizer application, urea with or
without PPDA acted more efficiently on grain yield and N uptake than did lime
ammonium nitrate. The fertilizers did not exhibit significantly different effects on straw
yield.
One can deduce from Figure 77, which presents the relationship between grain yield
and amount of rainfall during the first 5 days after fertilization, that the yield losses
272
2,0
'iii'
i":~i
7'
'[i! -1.0
CJ
- 2.0 ""r--T'" i I I , i ,
o 5 10 15 20 25 )J 35
Rainfalls (mm)
Figure 77. Influence of rainfalls on the difference between cereal grain yield recorded in the
treatment with lime ammoniwn nitrate and grain yields obtained in the treatment with urea with or
without 0.5 or 1% PPDA addition.
a - Conditioned urea. b - Conditioned urea + 0.5% PPDA. c - Conditioned urea + 1% PPDA. /From
Kiimpfe e/ al. (l982b)./
were great whert it did not rain, especially in the urea-only treatrnertt. With increasing
amounts of rainfall, both urea+0.5% PPDA and urea+ 1% PPDA became more efficient
than lime ammonium nitrate, but at rainfalls ~ 30 mm only urea+ 1% PPDA preserved
the efficiency.
The results obtained in the winter wheat experimertts concerning the grain yield-
increasing effect of 1% PPDA addition to urea fertilizer were also referred to by Linke
and Kampfe (1987) in their communication presented at the International Fair
Symposium held in Leipzig.
The first results obtained in experiments carried out within an international
collaboration in Bulgaria, the former Czechoslovakia, (East) Germany, Hungary,
Poland, Romania, and the former U.S.S.R. for studying the efficiency of urea+PPDA
were published by Koren'kov et al. (1980). The experiments started in 1976. Soils in
Mitscherlich pots were surface-treated with urea (0.5 g N/pot) with or without 1%
PPDA relative to urea-No The test plants were spring wheat (on differertt soils, in all
countries, except in Romania) and oats (on a chestnut chernozem in Romania).
Generalization of the results of all experiments led to the conclusion that PPDA did not
significantly increase the crop yield but enhanced, to some extent, plant uptake of N
from urea.
The investigations were continued in 1977 and 1978, and their results were
published by Kampfe et al. (1982c). In these years, rate ofurea-N was increased to 1
glpot, whereas that of PPDA remained at 1% on urea-N basis. If in 1976 wheat (grain
and straw) yield and plant utilization offertilizer N in urea treatmertt (100%) increased
to only 102% in the urea+PPDA treatment, in 1977 and 1978 PPDA increased crop
yield by 8-12% and plant utilization ofurea-N by 9-18%.
Rao and Ghai (1986a) studied, under conditions identical to those shown on pages
264 and 268, the effect of PPDA used in a proportion of 10% relative to urea weight
(40, 80, and 120 mg N/kg soil). The principal results are specified below.
Germination of wheat seeds, evaluated at day 15 after sowing, was not affected by
PPDA. Dry matter of plants after 21 and 35 days of growth was higher in treatments
with urea + PPDA at three rates than in the control plants treated with urea alone (on
273
average, by 13.1 and 38.7%, respectively). PPDA increased grain yield by 25.1% and
decreased straw yield, and, in such a way, the total yield remained at a level similar to
that of the control plants. Under the influence of PPDA, N content in grain also
increased (by 25.8%), but that in straw remained unchanged; therefore, the total N
content (grain + straw) exceeded the value recorded in control plants only by 13.8%.
In the field experiments described by Schlegel et af. (1987), PPDA had no
significant effect on crop yield. Urea prills, urea solution, and urea-ammonium nitrate
solution with or without PPDA were applied on surface of two Indiana soils (silt loam
and loam), at a rate of 34 or 67 kg NIha, and 0.56 or 2.24 kg PPDAlha, then
immediately sown with winter wheat. Grain yield and N content in grains were not
significantly different in soils N-fertilized with and without PPDA.
In field experiments (Winiarski, 1990), PPDA added at a rate of 1% to urea fertilizer
had no significant effect on spring wheat yields. However, PPDA influenced positively
the N uptake by plants.
In a 3-year field trial on 16-m2 plots installed on a brown soil of loamy sand texture
(PH 6.3-6.4), Kucharski (1992) compared the effects of urea and urea+PPDA on winter
wheat. The plots were fertilized yearly with 35 kg Plha as triple superphosphate and 83
kg Klha as KCI in a single application, and with 120 kg Nlha as urea with or without
1% PPDA in single and divided applications (120 kg Nlha and 60 + 60 kg Nlha,
respectively). The results indicated that PPDA in both single and divided applications of
urea+PPDA exerted no significant effect on grain and straw yield, on uptake and
utilization ofN by the wheat plants.
It is mentioned in a short report by Yeomans and Cerrato (1993) that in growth
chamber experiments the wheat plants treated with urea and urea + PPDA had lower
yields than those treated with nitrate and nitrate + PPDA.
Czapla and Hurniecki (1998a,b) studied the effect of PPDA on spring wheat in 3-
year field experiments (1987-1989) on a light-textured soil (pH 5.7-5.9). Before sowing,
all plots (20 m2 each) were fertilized with 32 kg Plha as triple superphosphate and 100
kg Klha as KCl. For soil application, urea was used at rates of 0, 50, 100, and 150 kg
Nlha, whereas for foliar application rates of urea were 0, 25, 50, and 75 kg Nlha. PPDA
was used at a rate of 1% relative to weight of urea.
Grain and straw yield and N content data allowed the researchers to draw the
conclusion that PPDA in both soil and foliar applications with urea had no significant
effect of wheat yield. When applied with urea to soil, PPDA caused a several percent
increase in plant utilization of N from urea. In foliar application, PPDA increased the N
content in grain and decreased it in straw.
See also Table 61.
* *
*
Triticum durum was the test plant in a field experiment in which Katyal et af. (1987)
studied the effect of timing of urea, urea-PPDA, and urea-DCD applications relative to
irrigation on the fertilizer urea-N use efficiency. The experiment was carried out on a
coarse-textured alkaline (PH 7.9) loamy sand soil in the 198211983 growing season, at
the research farm of the Punjab Agricultural University, Ludhiana, India. The fertilizers
were 15N-Iabeled: urea alone C5N excess% 4.836), urea-PPDA (10 g PPDA/kg urea; 15N
excess% 4.733), urea-DCD (100 g DCD-N/kg N in the final product; 15N excess%
274
4.273). KNO) C5N excess% 4.085) was the reference fertilizer. The labeled fertilizer
were applied on microplots (1.2 by 1.2 m) installed within the large plots (5.5 by 2 m).
No N fertilizers was added to the control plots.
Rate of N fertilizers was 120 kg N/ha, applied in two equal splits - half basal and
half topdressed. For basal application, the fertilizers were broadcast and incorporated
into the moist seedbed before sowing; for topdressing, the fertilizers were surface-
applied on the dry soil within a half hour before the first irrigation or on the wet soil 20
hours after the first irrigation, which took place about 3 weeks after sowing.
Before the basal N fertilization, superphosphate (22 kg P/ha) and ZnS04 (10 kg
Zn/ha) were broadcast on all plots. The K that was added through KN0 3 was balanced
across all plots by adding equivalent amounts of K as KCl.
At maturity, besides estimation of grain yield, the plants and soil were analyzed for
total Nand 15N.
KN0 3 gave the highest grain yield irrespective of the timing of topdressing (before
or after irrigation). In the urea, urea-PPDA, and urea-DCD treatments, the grain yields
were significantly higher (p=0.05) when they were topdressed before irrigation than
when they were applied after irrigation. In the case of topdressings before irrigation, the
grain yield in the urea-PPDA treatment was significantly higher than that in the urea-
DCD treatment and not significantly different from that registered in the urea-only
treatment. The grain yield obtained with urea-PPDA topdressed after irrigation was
significantly higher than that with urea or urea-DCD also topdressed after irrigation;
following topdressings after irrigation, urea-PPDA yielded 400 kg/ha more than urea
alone.
15N loss was lowest in the KN0 3 treatment and had the following values for the
other fertilizers topdressed before and after irrigation: 15.8 and 42.2% (urea), 32.9 and
33.2% (urea-PPDA), and l3.7 and 46.2% (urea-DCD), respectively.
The highest grain yield and the lowest N loss in the KNO) treatment indicate that
leaching and denitrification were not significant loss mechanisms. The higher grain
yield and the lower N loss with urea-PPDA compared to yield and N loss with urea and
urea-DeD suggest that volatilization of ammonia released from urea was the major N
loss mechanism.
7.2.9. Effect of PhosphoriC Triamide (PTA) and Thiophosphoric Triamide (I'PTA)

Compounds
In most studies, the effect of PTA and TPTA compounds on wheat plants was compared
Schlegel et al. (1986, 1987) studied, under field conditions in Indiana, the effect of
several phosphoroamides, including PPDA, on N uptake and grain yield of winter wheat
in the 1982/1983 and 1983/1984 crop years.
The 1982/1983 experiments were conducted on a silt loam (PH 6.0) and a loam (pH
6.3). The plots, into which wheat was seeded in the autumn of 1982, were fertilized in
the spring of 1983. Urea prills and urea solution were broadcast and urea-ammonium
nitrate (VAN) solution was either broadcast or applied in bands on soil surface. Rate of
each fertilizer was 50 kg N/ha. The same six phosphoroamides were tested as in the
1983 maize experiments and at the same rates (see pages 256-257). They were coated
on urea prills or dissolved in urea and UAN solutions.
275
In the 1983/1984 experiments, plots on a silt loam (PH 6.3) and a loam (PH 6.3)
were fertilized by broadcast urea prills or UAN solution either in autumn (after sowing)
or in spring at rates of 34, 50, and 67 kg N/ha. The inhibitors tested were cyclohexyl-
PTA, trichloroethyl-PDA, and PPDA at rates of2 kg/I 00 kg N (in prills) and 2.24 kg/ha
(in UAN solution).
In all experiments, the inhibitors did not significantly affect grain yield and N
content. There were only two exceptions, both in the 1983/1984 experiments. The first
instance was in the autumn experiment on silt loam where grain N content was higher
with urea (50 kg N/ha) plus PPDA than urea alone (21.0 and 19.0 g/kg, respectively).
The second instance was in the spring experiment on loam where grain yield was higher
with UAN (34 kg N/ha) plus PPDA than UAN alone (4.09 and 3.46 t/ha, respectively).
The general inefficiency of inhibitors in the wheat experiments was attributed to
precipitation that fell on the first days after fertilizer application; the rainfall leached the
~urface-applied urea into the soil, preventing volatilization of ammonia.
The effect of nBTPTA on wheat was also studied in a greenhouse experiment
conducted at the Headquarters of the International Fertilizer Development Center
(Muscle Shoals, Alabama). Urea with 0.1 % nBTPTA, surface-applied on a wheat soil,
led to a 65% increase in urea-N uptake by the plants and to a 16% increase in grain
yield over those achieved with urea alone (Carmona et al.. 1988; Christianson and Vlek,
1991 ).
Bremner and Krogrneier (1988) studied the ability of 10 urease inhibitors to prevent
or reduce the adverse effects of urea on seed germination, seedling growth, and early
plant growth, these effects being caused by ammonia released from urea under the
action of soil urease. The inhibitors studied were: nBTPTA, PPDA, four phosphoric
triamides [phosphoryl triamide, phenylphosphoric triamide, N-(diaminophosphinyl)
benzamide, 4-fluoro-N-(diaminophosphinyl)benzamide], phenylmercuric acetate,
catechol, hydroquinone, and p-benzoquinone. Wheat, barley, oats, rye, sorghum, and
alfalfa served as test plants. Three Iowa soils were used.
In the germination test, air-dried soil samples (40 g) placed in Petri dishes (1.5 x 10
cm) were moistened with 10 ml of water or 10 ml of a solution containing 100 mg of
urea with or without 1-100 Ilg of inhibitor, then sown with 100 seeds and incubated in
the dark, at 20 0 e for 7 days, after which time the germinated seeds were counted.
In the soils not treated with urea, the seeds germinated in high proportions. In the
urea-treated soils, no seed germinated which proves toxicity of urea. In the presence of
both urea and inhibitors, the seeds germinated in the same number as in the untreated
soils or in a smaller number. In other words, the urease inhibitors eliminated or reduced
the adverse effect of urea on germination. The most effective inhibitor was nBTPTA,
followed by PPDA. Thus, the adverse effect of urea on germination of seeds (wheat,
barley, oats, and sorghum) was eliminated by nBTPT A used in a concentration as low
as 0.001 % relative to weight of urea (in two soils) or at 0.005% concentration (in one
soil).
The procedure used for studying seedling growth was similar to that used in the
germination test, but only 15 seeds were sown instead of 100 seeds. Shoot length was
measured after 7 days. No seedling growth occurred in soils treated with urea only.
Again, nBTPTA proved to be the most effective compound, eliminating even at 0.005-
0.01 % concentrations the adverse effect of urea.
276
For studying early plant growth, the following procedure was applied. Air-dried soil
samples (200 g) placed in small pots were moistened with 10 ml of a solution
containing 10 mg ofK2S04, 10 mg of NaH 2P0 4, and 50 mg of (NH4hS04, then 3 seeds
(only wheat or sorghum seeds) as well as 3 granules, each containing 25 mg of urea
with or without nBTPTA or PPDA (0.001, 0.01 or 0.1% on urca weight basis) were
placed 2 cm below the soil surface. Finally, 20 ml of water were added and, then, the
pots were introduced to a growth chamber (22°C). After 21 days of growth, dry matter
of plants was determined. It was established that nBTPT A, even at 0.0 I %
concentration, markedly reduced the adverse effect of urea on early plant growth. PPDA
acted less markedly.
However, PPDA was found very effective in a similar experiment in which Bremner
and Krogmeier (1989) studied the adverse effect of purified urea and fertilizer urea on
seed germination and elimination of this effect by PPDA. Samples of four Iowa soils
and seeds of wheat, maize, barley, and rye were used. Rates of addition per 40 g of air-
dried soil were 100 mg of urea and 1 mg of PPDA. As the adverse effect of fertilizer
urea was not significantly different from that of purified urea, it was deduced that the
effect of fertilizer urea was not due to impurities (biuret, cyanuric acid) but to ammonia
formed through hydrolysis of urea by soil urease. This deduction was supported by the
findings that biuret and cyanuric acid alone had little, if any, effect on germination in
soil when compared with urea, and the adverse effect of urea was completely eliminated
by PPDA. The results were similar with the four soils and seeds of the four plant species
studied.
In continuation of these investigations, Krogmeier et al. (l989a) performed other
experiments to give answer on the following question: does inhibition of soil urease
activity by nBTPT A or PPDA lead to accumulation of urea in plants grown in soil
fertilized with urea because, as known, urea may cause leaf tip necrosis?
For these experiments wheat and sorghum were grown on two Iowa soils.
In the first experiment, air-dried soil samples (500 g) in pots were moistened with 25
ml of a solution containing 20 mg of K2S04 and 20 mg of NaH2 P0 4, then sown with 15
seeds at 2-cm depth and treated with 50 ml of a solution containing 0.5 g of urea with or
without 0.05, 0.5 or 5 mg of nBTPT A or PPDA. The pots were then placed in a growth
chamber (22°C). After 21 days of growth, dry weight of plants was determined. Leaf tip
necrosis was assessed by separating the necrotic portions of the plant shoots from the
nonnecrotic portions and determining their dry weight. Urea content in both necrotic
and nonnecrotic portions was also assayed.
Dry weight of plants of both species was smallest, i.e., the growth was weakest, in
soils treated with urea without inhibitors. These plants did not manifest any symptoms
of leaf tip necrosis and contained only negligible amounts of urea. The weak growth
was due to ammonia which could be released from urea since soil urease had not been
inhibited. The plants grew incomparably better when urea and inhibitor were applied
together, but leaf tip necrosis occurred and urea content increased in necrotic portions
and remained negligibly low in nonnecrotic portions. Severity of necrosis and urea
content increased with increasing inhibitor conccntration and were much more marked
with the stronger inhibitor, i.e., with nBTPTA than with PPDA. Dry weight of necrotic
portion highly correlated (r=0.99) with their urea content. It results from these
observations that urea, which could not be hydrolyzed because of the presence of the
277
soil urease inhibitors, had accumulated in toxic concentrations in leaf tips and induced
necrosis.
In the second experiment, non-autoclaved and autoclaved soil samples were used
and the test plant was wheat. Autoclaving (at 120D e for 2 hours) led to disappearance of
urease activity in soil. The procedure was the same as in the first experiment, but urea
was applied at several rates (0, 0.0625, 0.125, 0.25, and 0.5 glpot) with or without
nBTPTA or PPDA at a single rate (2.5 mg). Analyses of plants after 21 days of growth
showed that in the absence of inhibitors the necrosis appeared and urea accumulated
only in plants grown in the autoclaved soil treated with 0.25 and 0.5 g of urea/pot. In the
presence of inhibitors, the necrosis occurred and accumulation of urea in necrotic
portions of leaves also took place at lower urea rates. These effects were more marked
in autoclaved soil than in the non-autoclaved soil and with nBTPTA than with PPDA.
In the third experiment, the effect of nBTPTA and PPDA on urease activity in wheat
and sorghum shoots was studied. The procedure of experiment 1 was applied, but the
soil samples were not treated with urea, and, after 21 days of growth, the shoots were
immediately weighed and analyzed for urease activity. The results indicated that
nBTPTA and PPDA did not significantly decrease, at any of their rates, urease activity
in shoots.
The conclusion is that nBTPTA and PPDA applied together with urea led, through
inhibition of urease activity, of urea hydrolysis in soil, to manifestation of the
phytotoxic effects of urea. However, these phenomena were observed only when the
soil concentrations of nBTPTA and PPDA markedly exceeded those likely to exist
when these compounds are used as fertilizer amendments to reduce problems
encountered in use of urea as a fertilizer.
The potential of nBTPTA and PPDA for inducing phytotoxicity should not preclude
their use to eliminate the adverse effects of urea fertilizer on seed germination and
seedling growth because the ammonia produced through hydrolysis of urea fertilizer by
soil urease is much more detrimental to plant growth than is urea accumulation induced
by urease inhibitors.
In the field experiments conducted by Goodroad and Wilson (1989) in Georgia, late
winter topdress application ofurea+nBTPTA to winter wheat resulted in increased plant
N content and also in increased grain yield when N was limiting.
Bremner and Krogmeier (1990) evaluated the effect of 23 urease inhibitors on the
germination of wheat, maize, barley, sorghum, and alfalfa seeds in soil samples to
which no urea was added. Three Iowa soils (loam, pH 6.9, clay loam 7.5, and silty clay,
pH 7.7) were studied. Rates of inhibitor addition were: 0, 50, 250, 500, and 2,500 J-lglg
soil. The urease inhibitors are nominalized. and their effect on germination is specified
in Table 61.
One can see from this table that 2,5-dimethyl-l,4-benzoquinone exhibited the
strongest adverse effect on germination and that wheat and sorghum seeds were most
sensitive to this urease inhibitor.
In the wheat field experiment of Gezgin and Bayrakli (1995), nBTPTA added to
urea at rates of 0.25 and 0.50% relative to weight ofurea-N (200 kg urea-N/ha) greatly
reduced the volatile ammonia losses (see page 198) and increased grain yield (t/ha)
from 3.763 (urea only) to 4.443 (urea+0.25% nBTPTA) and 4.313 (urea+0.50%
nBTPTA), but had no effect on the protein content in grain.
278
TABLE 61. Effect of23 urease inhibitors on germination of seeds in soils"

Inhibitor class Inhibitor
Urease inhibitors manifesting significant (p<O.Ol) adverse effect on germination
Polyhydric phenols and quinones Hydroquinone. at the 2.500 )!glg soil rate. inhibited germination of
maize seeds in the clay loam soil and germination of wheat seeds
in all soils.
1,4-Benzoquinone. at the 2,500 )!glg soil rate. inhibited germination
of wheat and sorghum seeds in all soils.
2,5-Dimethyl- L4-benzoquinone, at the 500 )!glg soil rate, inhibited
germination of wheat and sorghum seeds in all soils and. at the
2.500 J.lglg soil rate, inhibited germination of all seeds in all soils.
At lower rates. they had no inhibitory effect on germination of any
seed in any soil.
Urease inhibitors manifesting no significant adverse effect, at their any rate, on germination of any seed
in any soil
Organic mercury compounds Phenylmercuric acetate
Polyhydric phenols and quinones Catechol
2,5-Dichloro-l ,4-benzoquinone
Phosphorodiarnides Phosphorodiarnidic acid
Pheny Iphosphorodiamidate
4-Chlorophenylphosphorodiamidate
Phosphorotriamides and Phosphoryl triamide
thi oph osp horotriami des N-Phenylphosphoric triamide
N-(4-Nitrophenyl)phosphoric triamide
N-(3-Trifluoromethylphenyl)phosphoric triamide
N-(Diaminophosphinyl)benzamide
4-Chloro-N -(diaminophosphinyl)benzamide
4-F1uoro-N -( diaminophosphinyl)benzamide
4-Cyano-N -( diaminophosphiny I)benzarnide
N-(Diaminophosphinyl)benzeneacetamide
N-(Diaminophosphinyl)-3-pyridinecarboxamide
Thiophosphoryl triamide
N-(n-Butyl)thiophosphoric triamide
Cyclotriphosphazatriene (CTPA T) 2-Phenoxy-2,4,4,6.6-pentaamino-CTPA T
derivatives 2,4-Diphenoxy-2,4,6.6-tetraamino-CTPA T
a Adapted from Bremner and Krogrneier (1990).
Wang et at. (1995) studied the effect of nBTPTA on spring wheat under growth
chamber conditions. Five-kg samples of a black chernozemic soil from Manitoba were
placed in plastic pots (20 cm high with a 20-cm diameter). Urea was applied at rates
equivalent to 0,40, 80, and 120 kg Nlha, with or without 0.15 or 0.25% nBTPTA by
weight of urea, and was seed-placed or surface dribble-banded. Fifteen wheat seeds
were sown in each pot. The soil moisture was cycled between 70 and 100% of field
capacity by watering.
Seedling emergence was significantly decreased by seed-placed urea at its rates of
80 and 120 kg Nlha but not by surface-dribbled urea. The damage caused by seed-
placed urea was reduced by nBTPTA. There was no significant difference between the
rates of 0.15 and 0.25% nBTPTA. Vegetative yield measured at heading increased with
application of urea, but no difference in vegetative yield occurred due to urea placement
or the use of nBTPTA. N accumulation in plants increased with increasing urea rate,
and the increase was higher when urea was seed-placed than surface-dribbled. nBTPTA
279
consistently increased N accumulation, particularly with the surface dribble application.

There was no significant effect of urea placement or use of nBTPTA on residual soil N.
Wang and Douglas (1996), who found that PPDA, cyclohexyl-PTA, and nBTPTA
inhibited hydrolysis of urea in samples ofa solonized brown soil (PH 7.7) and a podzol
(PH 5.0), both from Victoria, Australia (see page 196), also studied the effect of these
inhibitors on wheat in a pot experiment using samples ofthe soils mentioned above.
Air-dried soil samples were treated with a micronutrient solution before potting.
Four wheat seeds were planted in all plastic pots containing the equivalent of 500 g
oven-dried soil. Urea-N (50 or 200 mg N, having enrichment of 5 atom% 15N excess)
and inhibitors (at rates of 0.5 or 1% of urea by weight) were added in solution to the
centers of the pots. Soil moisture was maintained at field capacity during the
experiment. Plants were grown in a glasshouse in which the average day and night
temperatures were 23 and lOoC, respectively, and harvested after 6 weeks of growth.
Then their dry weight and total N and 15N contents were determined.
Contrasting the results obtained in determination of urea hydrolysis, application of
the inhibitors with urea to soil did not result in significant increases in dry matter and N
uptake by plants. The lack of effect was attributed to the glasshouse conditions
(moderate temperature and no wind) that minimized ammonia loss from the soil surface.
7. 2.1 O. Effect ofLignosulfonates

In a pot experiment, Builov et al. (1979), treated 600-g samples of a solonetz soil from
the Rostov-on-Don area, Russia, with ammonium lignosulfonate (ALS) at rates of 0,
1.3, and 1.7% relative to soil weight. Then pregerminated wheat seeds were sown in the
soil samples.
Early growth of seedlings was retarded in the untreated soil, but it was vigorous in
the treated soil, at both ALS rates. But after 18 days of growth, there was no visible
difference between the plants in untreated and treated soil. Moreover, in a similar
experiment in which 600-g samples of a compact meadow soil were treated with 0, 0.3,
and 1% ALS, growth of wheat seedlings was not affected at all by the addition of ALS.
7.2.11. Effect ofPlant Materials

In the lysimeter experiment conducted by Singh and Singh (1986) (see page 174), neem
cake mixed with urea fertilizer in a proportion of 1:1 (weight/weight) led to
significantly increased yields and N contents of wheat grain and straw. The grain yield
increase was -24 and 25% at 60 and 120 kg urea-N/ha, respectively.
In the 1988/1989 and 198911990 growing seasons (December-April), Joseph and
Prasad (1995) conducted experiments on spring wheat fields on a sandy loam soil (PH
8.2) at the Indian Agricultural Research Institute, New Delhi. The N fertilizers used
were prillcd urea (PU), PU blended with 10 or 20% DCD-N and with 20 or 40% neem
cake (NC) (considered as a nitrification inhibitor only). Rates of N applied were 0, 60,
and 120 kgiha.
The following average grain yields (t/ha) were obtained in the different treatments:
3.0 (no N applied), 4.3 (PU), 4.7 (PU + 10% DCD-N), 4.3 (PU + 20% DCD-N), 4.5 (PU
+ 20% NC), and 4.6 (PU + 40% NC). It was calculated from response equations that for
a target grain yield of 4.5 t/ha 10% DCD-N and 40% NC reduced fertilizer N
requirement by 40 and 30%, respectively, as compared to PU alone.
280
The results obtained with neem cake-coated urea applied by Sharma and Prasad
(1996) in a 2-year maize-wheat rotation are referred to on page 262.
7.2.i2. Effect of Combined Urease and Nitrification inhibitors

As shown on page 246, in the field experiments of Palazzo et al. (1996), the combined
use of nBTPTA and DCD led to a marked increase in the wheat grain yield.
In the pot experiments of Chen et al. (1998) (see page 244), 11 spring wheat plants
were grown in each pot. The grain yields (glpot) were, in the different treatments, the
following: 2.13 (no urea and no inhibitor), 7.61 (urea alone), 7.98 (urea + HQ), 8.37
(urea + HQ+CaCz), and 10.01 (urea + HQ+DCD). The efficiency of plant uptake ofN
from fertilizer urea in the HQ+DCD treatment was 10 and 14% greater than were the
efficiencies in the HQ+CaC z and HQ treatments, respectively. Thus, the best results
were obtained with the combination of HQ and DCD.
Similar results were obtained in another experiment in which labeled urea (urea- 15N)
was applied and the test plant remained spring wheat (Xu et al.. 2000).
7.3. EFFECT OF UREASE INHIBITORS ON RICE (Oryza sativa)
7.3.1. Effict ofinorganic Boron Compounds

Application of urea cogranulated with up to 4% borax (Zhan et al., 1993) to rice field"
resulted in 5-20% yield increases in comparison with the yields obtained with urea
alone.
7.3.2. Effect of Organic Mercury Compounds

Reddy and Chhonkar (1990b) studied the effect of phenylmercuric acetate (PMA) on
rice in a pot experiment. Four 15-day-old seedlings were transplanted in pots containing
3-kg samples of a sandy loam soil (PH 7.6). The soil was not flooded (and kept at about
50% of WHC) or was flooded with 5-cm standing water. At day 12 after transplanting,
urea (100 mg N/kg soil) ± farmyard manure (FYM) at a rate of 5% relative to soil
weight ± PMA (2 mgikg soil) were broadcast on the surface.
Grain yield increased in the PMA treatments over the no PMA treatments to the
following extent: 12% (no flooding, no FYM addition), 10% (no flooding, FYM
addition), 33% (flooding, no FYM addition), and 61 % (flooding, FYM addition). Thus,
PMA was more effective in the flooded than in the nonflooded soil.
7.3.3. E.tfect ofPolyhydric Phenols and Quinones

The pot experiment of Reddy and Chhonkar (1990b) (briefly described above) was also
carried out with hydroquinone (HQ) (2 mgikg soil). The rice grain yield increased in the
HQ treatments to the following extent: 15% (no flooding, no FYM addition), 13% (no
flooding, FYM addition), 35% (flooding, no FYM addition), and 63% (flooding, FYM
addition). It is evident that HQ was more effective in flooded than nonflooded soil.
Xue et al. (1991) treated rice fields with 17.5 kg of urea + 0, 100 or 150 g of HQ or
quinhydrone (QH) per mu (1/15 ha). The grain yield increased under the influence of
inhibitors; the increases were 8,4 and 11.3% due to HQ, and 8.2 and 13.3% due to QH,
and were related to a better utilization of urea-N by the plants: the increases in the
coefficient of plant utilization of N from urea were 2.9 and 4.8% in the HQ treatment
and 4.7 and 5.8% in the QH treatments.
281
Zhao et al. (1993a) performed a pot trial for studying the fate of HQ and urea in a
soil-rice system. Samples of a brown soil of silty loam texture (PH 6.4) were amended
with 15N_urea (atom excess, 5.56%) at a rate of 61.33 !lg N/g soil with or without 14C_
HQ (having a specific activity of 235.727 x 104 Bq/mg) at a rate of 533.2 ng/g soil.
Thus, the initial ratio (weight/weight) of 14C to 15N was 1: 115. Then rice seedlings were
transplanted into the soil samples. During the growth period and after harvesting, the
aboveground parts of rice plants and the soil were analyzed for 14C_HQ and other 14C_
materials as well as for total N and 15N.
The results showed that the rice plants absorbed only a minor part (0.25%) of the
amount of 14C_HQ added to soil. The 14C_HQ content was 37 nglg refined grain and 47
nglg straw. Most of the absorbed 14C_HQ was degraded or converted by the plant tissues
into other forms of 14C-materials, the content of which was 396 nglg refined grain and
116 nglg straw. In soil, 14C_HQ was gradually decomposed. Thus, after 3 months of rice
growth, 1 g of soil at different depths in the 0-20-cm layer contained, on average, 220
ng 14C-materials and 71 ng 14C_HQ which is lower than the permissible ecological value
(2,000 nglg soil) and the permissible healthy value (200 nglg soil) of the 14C_HQ.
The initial 14C:15N ratio of 1:115 changed to 1:72 in soil, 1:1,160 in straw and
1:3299 in grain, i.e., the rice plants absorbed relatively more 15N and there was more
14C-material left in the soil. Nevertheless, in this experiment the aboveground plant
biomass was not significantly different in the urea and urea + HQ treatments.
The conclusion was drawn that applying appropriate concentration of HQ with urea
as a urease inhibitor will not produce harmful effects on human health and the
environment. This conclusion is consistent with the opinion of Zhao et at. (1988, 1989),
based on results obtained with unlabeled urea and HQ: the mipor amount ofHQ applied
with urea fertilizer will not accumulate in soil and plant and will not enter the
underground water and atmosphere and, therefore, will not induce pollution of
environment and poisoning of food chain.
In the experiment of Fan and Ye (1995), HQ applied with urea enhanced the
efficiency of utilization of fertilizer N by rice plants, as it prevented the evaporation loss
of N in the paddy field.
7.3.4. F/ject ofPh osphorodiamides

It is mentioned in a report presented by the International Fertilizer Development Center
(IFDC, 1981) that, in a pot experiment, rice plants took up more 15N from urea-PPDA
than from urea. Both urea and urea-PPDA were more efficient when the plants were
grown under continuously flooded conditions than when the soil was flooded only after
a 39-day dry period.
Another pot experiment mentioned in the report (lFDC, 1981) was described in
detail by Byrnes et al. (1983). Two 20-day-old rice seedlings (variety IR36) were
planted in 7 kg (oven-dry weight) soil which had been thoroughly puddled by hand and
kept flooded for 2 weeks before transplanting. The soil used was a silty loam (PH 6.2)
which had been fertilized with 700 mg of P as Ca(H2 P04)z.H 2 0, 700 mg of K and 287
mg of S as K 2 S04 by incorporating the salts into the top 8 cm of soil.
Urea was applied at a rate of 460 mg N/pot (115 kg N/ha on an area basis) with and
without 5% PPDA relative to weight of urea. Both urea and urea-PPDA were applied
basally, i.e.. incorporated into 5 cm of soil before transplanting or used in equal split
applications of 230 mg N each; the first was broadcast and incorporated before
282
transplanting and the second broadcast without incorporation at maximum tillering (30
days after transplanting). The basal urea was 15N-labeled. Two sets of split-applied
treatment were made, one with only the basal-incorporated portion 15N-labeled and the
other with only the broadcast or delayed application 15N-labeled. The labeled urea
contained 4.2 atom% excess 15N and was tableted with or without PPDA in a press to
produce pellets 2.4 mm in diameter and 10-15 mg in weight.
The soil in each treatment was kept flooded with 3 cm of water throughout the 77-
day experiment. The rice crop was harvested prematurely because poor light conditions
caused erratic heading and grain formation. Dry weight of the entire aerial portions of
the plants was determined. Total N and 15N contents in aerial parts as well as in soil and
roots were also assessed.
The results obtained (Table 62) are surprising: under the influence of PPDA, the dry
matter production decreased significantly for both basal and split applications in
contrast with the significantly increased plant recovery of applied 15N. The increase was
10-15%. The highest increase was registered in the treatment in which the urea applied
at maximum tillering stage was labeled (41.0% of 15N recovered by plants without
PPDA and 66.7% with PPDA); in this treatment, the soil and roots recovered 36.4 and
34.0% of 15N. Thus, in the presence of PPDA, the recovery of 15N in the plant-soil
system was 100%. The great effectiveness of PPDA at maximum tillering stage was
probably caused by the large capacity of the root system at this growth stage to take up
N, together with the poor competitive position of the algae which were shaded by the
rice plants.
TABLE 62. Effect ofPPDA on dry matter production by rice plants and on recovery of 15N in
a ~t e:seeriment"
I~N recove!1 (%J
Dry matter
Treatment Plants Soil and
(glpot) Total
(aerial ~rts) roots
Control 35.3 d
Basal urea 44.4 ab 13.4 d 33.1 d 46.5 c
Basal urea + PPDA 36.8 bc 23.8 c 42.3 c 66.1 d
Split application
First split 15N-labeled'
Urea 46.0 a 15.8 d 51.2 b 67.0 d
Urea + PPDA 40.8 bc 24.5 c 61.7 a 86.1 b
Second split liN-labeled
Urea 46.0 a 41.0 b 36.4 d 77.4c
Urea + PPDA 40.8 bc 66.7 a 34.0 d 100.7 a
"From Byrnes et al. (1983). by permission of the Soil Science Society of America, Inc.
Data in the same column not followed by the same letters are significantly different
(p=0.05).
'Dry matter yields for the split applications with 15N-labeling at different times were
averaged.
De Datta et al. (1983) mentioned that in field experiments conducted at the

International Rice Research Institute (the Philippines), PPDA, broadcast together with
prilled urea (2 or 5% PPDA relative to 67 kg of urea-Niha) into the floodwater at day 20
after transplanting of rice seedlings retarded urea hydrolysis and delayed appearance of
ammonium in floodwater, but did not increase grain yield and total N uptake when
compared with control without PPDA.
283
In the experiment of Simpson et al. (1985), the microplots cropped with flooded rice
were fertilized with urea (80 kg Nlha) labeled with 15N (at 1.48 atom% excess 15N) with
or without 1% PPDA (relative to weight of urea) addition. Urea and urea-PPDA were
applied at the stage when the plants reached 20 cm above the floodwater (see page 119).
At early heading stage (at day 47 after fertilization), the plants (shoots and roots)
contained significantly more 15N in the urea-PPDA treatment (47.6% ofthe 15N applied)
than in the urea-only treatment (28.2%). This effect persisted at maturity. Thus, at this
stage, 15N contents in grain and straw in the urea-PPDA treatment exceeded those
recorded in the urea-only treatment by 53 and 56%, respectively. However, there was
no increase in grain yield. This is explained by the property of the rice variety used
(Inga) not to respond well to N fertilizers applied at early tillering. Consequently, for
studying the effect of PPDA on grain yield, it is necessary to use more responsive rice
varieties.
The rice variety IR36, used by Fillery et al. (1986b) in the investigations mentioned
on page 122, responded differently to PPDA depending on locality and season. At Los
Banos, in both dry and wet seasons of 1982, PPDA had no increasing effect on plant N
uptake and grain yield. Neither did PPDA enhance the grain yield at Munoz. Contrarily,
application ofPPDA significantly increased the grain yield (from 4.7 to 5.2 tlha) in the
experiment conducted at Los Banos in the dry season of 1983, in which two-thirds of
urea and urea-PPDA were administered at day 26 and the remaining third at day 50 after
transplanting. In the experiment, the plants took up more N from urea-PPDA than from
urea, but the difference was only significant at day 60 after transplanting; 15N recovered
in plant and soil at grain harvest was 77.7% in the urea-PPDA treatment and only 65.8%
in the urea treatment. At Munoz, 15N recovery (in plant and soil) was 73.6% (urea
treatment) and 83.4% (urea-PPDA) at day 14 after fertilization and similar for both
treatments at grain harvest. As mentioned above, the grain yield was also similar.
Rao and Ghai (l986b) described a pot experiment aimed at studying the effect of
PPDA on rice grown on an alkali soil (sandy loam, pH 9.0). Soil samples (20 kg/pot), to
which 20 ppm P (as single superphosphate) and 12.5 ppm ZnS04 had been added, were
saturated with water, following which urea with or without PPDA was surface-mixed
and the pot watered to maintain 3.5 cm of standing water. Then four 33-day-old rice
seedlings were transplanted into each pot.
Urea-N (at a total rate of 150 ppm in all treatments) was applied by three modes:
M-1: one-half of N at transplanting and one-fourth each at weeks 3 and 6 after
transplanting;
M-2: one-third ofN each at transplanting and weeks 3 and 6 after transplanting;
M-3: one-fourth ofN each at transplanting and weeks 3,6, and 9 after transplanting.
PPDA was used at a rate of 5% relative to weight of urea.
At weeks 6 and 9 after transplanting and at maturity, the plant dry weight, N uptake
and N uptake efficiency were recorded.
At weeks 6 and 9, in urea-only treatments, dry matter, N uptake, and N uptake
efficiency in the three modes of application had the following order: M-2> M-1 > M-3.
At maturity, the three modes of application presented the orders: M-2 ~ M-3 > M-1
(grain yield); M-2 = M-1 ~ M-3 (straw yield); M-3 > M-2 ~ M-l (N in grain); M-l ~ M-
3 ~ M-2 (N in straw); M-3 > M-2 ~ M-l (N uptake efficiency). Thus, taking into
account the grain yield and N uptake efficiency at maturity, M-3 (Le., split application
284
of urea in four equal amounts) proved to be the most efficient mode of application and
M-1 the least efficient one.
At week 6, PPDA increased plant dry matter, N uptake and N uptake efficiency in
each mode of application. At week 9 and at maturity, the effect of PPDA remained
positive in M-3 and M-1, but became negative, depressive, though insignificantly, in M-
2. Under the influence ofPPDA, in the most efficient M-3, the grain yield and N uptake
efficiency at maturity increased by 14.3 and 17.0%, respectively. Therefore, a saving of
fertilizer can be achieved by applying urea and PPDA as in M-3.
In the field experiments of Buresh (1987) (see page 126), the rice grain yields were
not significantly different depending on the nature offertilizers (urea, urea-PPDA, urea
phosphate, urea-urea phosphate) applied at 30 kg Nlha rate and the methods of their
application (into soil or floodwater).
But in two other rice experiments on a clay soil at Pila, Buresh et al. (1988b,c)
found that PPDA, used at a rate of 2% relative to weight of urea, was efficient in
increasing N content in grains. One of the experiments was conducted in the 1985 dry
season (see page 126). Urea rates were 30, 60, and 120 kg N/ha. Two-thirds of the urea
was applied at day 18 after transplanting of seedlings and one-third at days 5-10 after
panicle initiation. In the other experiment, conducted in the 1986 dry season, urea rates
were 40, 80, and 120 kg Nlha, one half of which was administered at day 16 after
transplanting and the other half at days 5-10 after panicle initiation. In both
experiments, urea and urea-PPDA were broadcast into 5-cm standing floodwater.
Within the plots there were microplots (80 by 80 cm) installed. Each microplot was
surrounded by 30-cm deep border made of painted, galvanized metal that had been
pushed approximately 20 cm into the soil. Urea broadcast (with or without PPDA) into
the microplots was labeled with 5 atom% 15N. At rice maturity, grain yield was recorded
and total Nand 15N in grain, straw, and soil+root were determined. The unrecovered 15N
was assumed to represent total gaseous N loss, because runoff loss was prevented by the
metal border of microplots and leaching loss was negligible (as the fraction of added
15N recovered in the 15-30-cm soil layer was consistently less than 2%).
PPDA significantly reduced N losses from the higher urea-N rates but did not
eliminate them. Thus, in the first experiment, the N losses from the 30, 60, and 120 kg
urea-Nlha rates were reduced from 5 to 3 kg Nlha, from 14 to 10 kg Nlha and from 34
to 16 kg Nlha, respectively. In the second experiment, N losses from 40,80, and 120 kg
urea-Nlha in the absence and presence ofPPDA were 5 and 6, 27 and 16, and 44 and 30
kg Nlha, respectively.
Reduction of N losses was not associated by a significant increase in grain yield.
Thus, mean yields for the three urea-N rates were 6.0 tlha (urea-only treatments) and 6.2
tlha (urea-PPDA treatments) in the first experiment; the corresponding values in the
second experiment were 6.0 and 6.3 tlha, respectively. But significant increases
occurred in the N content of grains. For example, in the first experiment at the 120 kg
urea-Nlha rate, the grains contained 33% (urea treatment) and 44% (urea-PPDA
treatment) of the applied N. In the second experiment, the corresponding values were 23
and 32%, respectively.
Elimination of gaseous N loss could increased grain yield by a maximum of 6 and
8% in the first and second experiment, respectively. These percentages corresponded to
0.4 and 0.5 tlha increases in grain yield, respectively.
285
But in other rice field experiments, in which unamended and PPDA-amended urea
were broadcast to floodwater, amendment of urea with PPDA did not increase either
yields or grain uptake of N compared with unamended urea (Snitwongse et aI., 1988;
Raju et al. , 1989; Satrusajang et al., 1991).

Compounds
In most studies, the effect of PTA and TPTA compounds on rice plants was compared
The experiments performed by Buresh et af. (1988a), concerning comparison of the
inhibitory effectiveness of nBTPT A and PPDA on urea hydrolysis, ammonium
accumulation and pNH3 in the rice field at Pila and Munoz, are dealt with on page 211.
Comparison of the effiCiency of these inhibitors on grain yield and N content of rice is
described in another paper (Buresh et af.. 1988b).
Mean value of grain yields recorded following application of urea at different rates
was 5.5 t/ha at Pila and 6.2 tlha at Munoz. Under the influence of nBTPTA, the grain
yield increased insignificantly at Pila (5.6 t/ha) and significantly at Munoz (6.8 t/ha).
PPDA also increased insignificantly the grain yield at Pila (5.8 t/ha), but significantly at
Munoz (6.7 t/ha).
Total grain N content per hectare increased significantly only in the urea + nBTPTA
treatment at Munoz, whereas percent grain N content slightly increased under the
influence of both inhibitors at Pila and remained unaffected at Munoz.
In the same rice field, the effect of nBTPTA on grain yield and N content did not
significantly differ from that of PPDA. Thus, at Munoz agronomic efficiency for 50 kg
urea-N/ha was 41 kg grainlkg urea-N in urea-only treatment and 49 and 50 kg grain/kg
urea-N in the urea-PPDA and urea-nBTPTA treatments, respectively.
Cai et al. (1989) conducted a similar experiment on a grey clay soil in New South
Wales. The test plant was flooded rice. The experimental plots, covered by -1 O-cm deep
floodwater, were treated with solutions containing unlabeled or labeled (with 15N at 2
atom% excess) urea (50 kg N/ha) with or without nBTPTA or PPDA at a rate of 1%
relative to weight of urea. The inhibitors were added to the floodwater immediately
before urea application. At maturity, total N and 15N contents in soil and plants were
determined and the grain yield evaluated. The inhibitors increased, though
insignificantly, recovery of 15N in soil and plants. Application of nBTPTA did not lead
to increase of grain yield which was 3.37 t/ha in both urea and urea-nBTPTA
treatments. Under the influence ofPPDA, the grain yield increased to 3.60 t/ha, but this
increase was also insignificant.
In the greenhouse experiment performed by Byrnes et al. (1989b) and referred to on
page 248, nBTPTA and PPDA significantly increased recovery of 15N from urea at both
42 and 119 days after transplanting (DAT) of rice plants. Thus, at 119 DAT, i.e;. at
maturity, the 15N recovered from grain + straw + (soil + roots) was only 50.1 % of the
added 15N in the treatment with urea alone (49.9% was lost in gaseous form), and 90.4
and 72.2% in the urea-nBTPTA and urea-PPDA treatments, respectively. In other
words, N losses were reduced from about 50% to about 10% by nBTPTA and to about
28% byPPDA.
The effect of Cu-chelate on 15N recovery was insignificant when added to nBTPTA
(total 15N recovery: 92.2%) but significant when added to PPDA (total 15N recovery:
286
79.1 %). Ah(S04h insignificantly reduced and Cu-chelate + AIz(S04h insignificantly

enhanced the effect of PPDA, the total 15N recoveries being 69.2 and 76.1%,
respectively.
The effect of different treatments on 15N content in grains was similar to their effect
on total 15N recovery.
Grain yield was significantly higher in four treatments [urea+nBTPTA; urea
+nBTPTA+Cu-chelate; urea+PPDA +Cu-chelate; urea+PPDA +Cu-chelate +AIz(S04hJ
than in the treatment with urea alone. But the yield increases recorded in these four
treatments (about 40, 36, 36, and 32%, respectively) were not significantly different.
PPDA alone and with AIz(S04h did not significantly affect grain yield. Straw yields
were not significantly different in the urea-only treatment and the other treatments.
Water percolation of soil, in comparison with no percolation, had no significant
effect on either grain or straw yields.
In the field experiment of Phongpan and Byrnes (1990) (see page 148), the effects of
urea-N rate and nBTPTA on N uptake and yield of rice plants were also studied. The
floodwaters of plots, into which 0, 25, 50, and 75 kg urea-N/ha with or without 0.536 kg
nBTPTAlha was broadcast at day 10 after transplanting (DAT) of rice seedlings, were,
at day 5 before panicle initiation stage, again fertilized with half rates of the first urea
application. Thus, the total urea applications were 0,37.5,75, and 112.5 kg Nlha.
N content in plants was determined at 47 DAT (approximately 7 days before panicle
initiation stage) and in straw and grain of the mature, harvested plants. N uptake at 47
DA T and in straw at maturity was greater from plots receiving urea than from the
unfertilized, control plots, but nBTPTA did not significantly increase N uptake at any
rate of urea application. N contents and yields of grain in the control, urea- and
urea+nBTPTA-treated plots were not significantly different, apparently because of the
high availability of N in the soil.
In another field experiment of Phongpan and Byrnes (1993) (see page 149), similar
results were obtained. Thus, nBTPTA broadcast with urea into the floodwater of rice
plants did not result in increased N uptake and grain yield at harvest.
As described in a short report by Norman et al. (1991), 15N-Iabeled urea was applied
preflood on dry and saturated soil, with or without nBTPTA, at 0, 5 or 10 days prior to
establishment of the permanent flood or added into the floodwater the day of flooding.
Use of nBTPTA significantly decreased ammonia volatilization losses and increased
fertilizer N uptake by the rice plants only when the urea was applied on saturated soil 5
days prior to the flood. But in no situation did the use of nBTPTA significantly increase
grain yield.
For evaluation of the effect of nBTPTA on rice plants, Bollich (1991) conducted a
field experiment on a silt loam soil in Louisiana in 1989-1990. Urea was applied to
plants at rates of 67 and 134 kg Nlha at 3-daY intervals beginning 9 days preflood and
ending 6 days postflood. Maturity and plant height were only slightly influenced by
nBTPTA while grain yields were unaffected.
But in the rice field experiments of the Chinese investigators Chen and Lu (1997),
grain yields were higher in the urea + nBTPTA and urea + PPDA treatments, especially
at high N level, than in the treatment with urea alone. Based on 15N tracing in the urea,
it was found that nBTPTA and PPDA enhanced the N uptake by plants. Hydroquinone
was less effective than were nBTPTA and PPDA.
287

In the rice field experiments conducted by Raju et al. (1989) during the winter seasons
of 1985 and 1986 on a clay loam soil at the Agricultural Research Station, Maruteru,
Andhra Pradesh, India, neem cake-coated urea (containing 33.7% N) was compared
with prilled urea. Rates of N fertilizer addition were 0, 37.5, 75.0, 112.5, and 150.0
kglha. The average grain yields (t/ha) were not significantly different in the prilled urea
treatment (4.364) and the neem cake-coated urea treatment (4.3 70).
But in other experiments, neem cake and extracts manifested yield-increasing
effects.
John et al. (1989) applied prilled urea uncoated or coated with neem cake (0.2 g
powdered neem cake per g urea) at a rate of 58 kg N/ha for fertilization of lowland rice
fields on silty clay soils at the International Rice Research Institute, Los Banos,
Philippines. The experiments were carried out in 1986 and 1987. The results showed
that coating urea with neem cake had no effect on loss of urea-N in either years;
however, it significantly increased grain yield (0.4 t/ha) and total plant N (II kglha) in
1987 but not in 1986.
Alcoholic extract of neem kernel, as a urease and nitrification inhibitor, was used in
the pot experiment of Reddy and Chhonkar (l990b) (see page 280). It was added at a
rate of 10% of applied urea (100 mg urea-Nlkg soil) and resulted in the following grain
yield increases in the different treatments: 21 % (no flooding, no FYM addition), 22%
(no flooding, FYM addition), 43% (flooding, no FYM addition), and 72% (flooding,
FYM addition). Thus, the neem extract was more effective in flooded than nonflooded
soil planted to rice.
Comparing the effects of urea coated with neem extract and uncoated urea on rice
plants, Vyas et al. (1991) found that the plants took up 86.1 % of the applied coated
urea-N and 34.2-57.1 % from the uncoated urea-No
De et al. (1992) found in 2-year experiments (1989 and 1990) on rice fields that
three splits of 70 kg Nlha (30 kg as basal application + 20 kg applied at tillering + 20 kg
applied at panicle initiation), as neem extract-coated urea, saved more than 30 kg Nlha
compared to prilled urea (50 kg Nlha as basal application + 25 kg applied at tillering +
25 kg applied at panicle initiation) and gave higher grain yields (t/ha) which, in the two
years of experiments, were the fallowing: 3.370 (1989) and 3.347 (1990) in the neem
extract-coated urea treatment, and 3.170 (1989) and 3.160 (1990) in the treatment with
prilled urea.
7.3. 7. Effect qfCombined Urease. Nitrification and/or Algal Inhibitors

Rice was the test plant in a pot experiment conducted by Amberger and Gutser (1984).
Samples (9 kg/pot) of a sandy loam soil (pH 6.5) were amended with urea alone and
with urea + thiourea (TU) or DCD or with urea + TU + DCD. Total N addition was 1.8
g/pot. TU-N represented 3 or 6%, and DCD-N was equal to 5 or 10% of the total N. The
urea and inhibitors were applied either 3 weeks before flooding of the soil (system A) or
after flooding to the growing plants (system B).
Dry matter yield and N uptake by plants were higher in system B than in system A.
In system A, TU and DCD at their higher rate caused a significant increase in yield and
N uptake. In system B, only TU at 3% of total N was efficient, but TU at 6% was
phytotoxic. In both systems, combinations of TU and DCD did not show better results.
288
In the rice field experiment carried out in Fuzhu, China, prilled urea was applied
alone (control) and with a urease inhibitor (nBTPTA) or with a nitrification inhibitor (2-
ethynylpyridine, 2EP) or with both inhibitors. The following grain yields (tlha) were
obtained: 4.74 (control), 5.30 (nBTPTA), 5.15 (2EP), and 5.45 (nBTPTA + 2EP). The
grain N contents (%) showed the same order: 0.947 (control), 0.979 (nBTPTA), 0.968
(2EP), and 1.012 (nBTPTA + 2EP). All increases were significant (p=0.05). The best
results were obtained by combined use of the two inhibitors (Freney et aI., 1989).
In the experiments of Khanif and Husin (1992), urea + hydroquinone (HQ), as
compared to urea alone, brought about only an insignificant increase in rice yield, but
when HQ was used in combination with DCD, the plant uptake ofN from urea fertilizer
significantly increased.
In the rice field experiments conducted by Chaiwanakupt et al. (1996) and
Phongpan et al. (1995, 1997) in Thailand (see page 182), the grain yields increased in
parallel with the reduction of volatile ammonia losses. In the experiment carried out
during the 1991 wet season, the highest yield was recorded in the treatment with
algicide + mixed urease inhibitors (4.66 tlha), which is significantly higher than that
obtained in the control treated with algicide + urea (4.00 tlha). nBTPTA in single
application without algicide increased the yield (4.22 tlha) in comparison with that of
control (urea only) (3.57 tlha), but when nBTPTA was applied with algicide, it reduced
the yield from 4.00 to 3.87 tlha.
In the experiment, which was carried out during the 1992 dry season and in which
algicide was applied to all treatments, the following grain yields (tlha) were obtained:
3.6 (control), 3.7 (nBTPTA), 4.0 (PPDA), and 4.1 (nBTPTA+PPDA). In other words,
the nBTPTA +PPDA combination was most efficient in increasing the grain yield.
In the field experiment described by Freney et al. (1995) and Phongpan et al. (1997)
(see page 249), the rice grain yield of3.14 tlha in the urea-only treatment was increased
to 3.17-3.81 tlha in the treatments with inhibitors. The increase was not significant
when urease inhibitor was not used and significant (P<0.05) when the urease inhibitors
cyclohexyl-PTA and N-(n-butyl)-PTA with or without the nitrification inhibitor
phenylacetylene were applied together with the algicides copper sulfate and terbutryn.
At the same time, addition of CHPTA and nBPTA in combination with phenylacetylene
and algicides resulted in a 2- or 3-fold increase of applied urea-N in the grain.
7.4. EFFECT OF UREASE INHIBITORS ON BARLEY (Hordeum vulgare)
7.4.1. Effect a/Heavy Metal Compounds

One of the examples described in the patent of Lewis and Slater (1979a,b) (see page 9)
refers to a germination test with barley seeds. The product tested consisted of ferric
nitrate complexed with 6 moles of urea plus 3 moles of free urea and was applied at a
rate equivalent to 100 kg Nlha. It was placed together with seeds at a depth of I cm in a
moist silty loam soil (PH 8.1). For comparison, urea was applied alone at the same N
rate. After 8 weeks, it was found that germination in the case of the tested product was
significantly higher (249% higher) than when urea alone was used.
7.4.2. F;Uect o/Organic Mercury Compounds

See Table 61.
289
7.4.3. Effect of Urea Derivatives

In the pot experiments of Kolyada (1970, 1973) (see page 49), thiourea at the rate of 0.1
g Nlkg soil was toxic to the barley plants, especially at the beginning of the growing
season. The grain yield was highest in the treatment with 0.1 parts ofthiourea-N + 0.9
parts ofN~NOrN + PK; it exceeded by 18% the yield recorded in the NH4 NO r N +
PK treatment (the rate of N, P, and K addition was always the same: 0.1 glkg soil).
Malhi and Nyborg (1984) conducted field experiments on the influence of thiourea
on the reducing over-winter losses from autumn-applied urea on Canadian soils (10
sites) cropped to spring barley.
On 6.8 m x 1.8 m plots, urea prills and urea pelleted with thiourea (2:1 urea to
thiourea by weight), both prills and pellets at a rate of 56 kg Nlha, were applied in
October 1973 or 1974. In the mixed treatment, the fertilizers (U-mix and U+T-mix)
were incorporated into -10 cm of soil, while in banded treatments the fertilizers (U-
band and U+T-band) were placed 5 cm deep and 23 cm apart. In the spring, separate
plots were fertilized with U-mix and U-band. All plots received P, K, and S fertilizers I
week before sowing of barley in May. Prior to sowing, the soil (0-90-cm layer) was
sampled for determination ofNH4 +-N and N0 3--N. At harvest time (late August or early
September), grain yield and N uptake by grain were determined.
In the experiment started in October, percent recovery of the added fertilizer N as
ammonium-N + nitrate-N, grain yield (tlha) and percent uptake of the added N had the
following average values in the four treatments at the 10 sites:
62,2.24, and 31 (U-mix); 68, 2.42, and 37 (U-band); 75,2.61, and 44 (U+T-mix);
and 82, 2.91, and 57 (U+T-band).
Thus, the U+T-band treatment was most effective. But a little better results were
obtained when U-mix and U-band were applied in the spring: the grain yield was 3.17
and 3.26 tlha, and the percent grain uptake ofN was 63 and 67, respectively.
In two other field experiments, Malhi and Nyborg (1988) compared thiourea with
nitrapyrin and ATe (4-amino-l,2,4-triazole hydrochloride), and found that urea +
inhibitors applied in bands in autumn increased grain yield (tlha) of spring-sown barley
by 0.72 (thiourea), 0,60 (nitrapyrin), and 0.52 (ATC) over the banded urea applied
without inhibitors in autumn. Thus, thiourea was more effective than the other two
inhibitors. But also in these experiments, the yield" were higher when urea without
inhibitors was mixed into the soil in the spring.
7.4.4. Effect ofPolyhydriC Phenols and Quinones

See Table 61.
7.4.5. Effixt of Ph osphorodiamides

The results obtained in evaluation of the efficiency of urea+PPDA in 43 winter cereal
experiments carried out in Germany, in the 1978-1980 period, on large surfaces (each of
5 ha) were described by Kampfe et al. (1983). The test plants were barley and rye in 37
and 63% of the experiments, respectively. With the exception of some data referring to
mean grain yields, the barley and rye were considered together as winter cereals.
Conditioned urea (i.e .. urea priUs coated with a mineral oil-bitumen mixture) was
used. Rates of PPDA were 0.5 and 1% relative to urea-No Surfaces not fertilized and
surfaces fertilized with lime ammonium nitrate served for comparison. The fertilizers
290
were applied twice in each year. Finally, the grain yield and total N contents in green
matter and grain were assessed.
The mean values of grain yields obtained in the whole 1978-1980 period, in the N
fertilizer treatments, and expressed as percentage of those registered in the lime
ammonium nitrate treatments (100%) were the following in the barley and rye
experiments: 99 and 95% (urea), 104 and 100% (urea + 0.5% PPDA), and 103 and
106% (urea + 1% PPDA ), respectively. These data show that urea and urea + 0.5%
PPDA were more efficient, whereas urea+ 1% PPDA was less efficient, in the barley
than in the rye experiments.
These experiments also proved that, when fertilizer application is followed by dry
weather, the grain yield is significantly lower in urea treatment than with lime
ammonium nitrate, and PPDA improves the fertilizing efficiency of urea.
It is evident from Table 63 that the effect of urea on grain yield is greatly influenced
by soil texture. In sandy soils, urea was less efficient, whereas in loamy soils its
TABLE 63. Effect of nitrogen fertilizers on grain yield of winter cereals in experiments on large surfaces,
as influenced by soil texture (mean values for the 1978-1980 period)"
Sands Slightly loamed Sandy loams and
sands and loamy loams
Fertilizer sands
(13 experiments) (20 experiments) (10 experiments)
dtlha % dtlha % dtlha %
Unfertilized control 20.2 61 26.8 68 33.5 75
Lime ammonium nitrate 33.3 100 39.1 100 44.7 100
Conditioned urea 31.0 93 38.1 97 44.7 100
Conditioned urea + 0.5% PPDA 33.2 100 40.5 103 45.3 101
Conditioned urea + 1% PPDA 35.1 105 40.5 103 47.6 106
LSD 10% 2.8 8 2.7 7 3.4 8
LSD 20% 2.2 7 2.1 5 2.6 6
a From Kiimpfe et al. (1983).
efficiency was equal to that of lime ammonium nitrate. Under the action of PPDA,
especially of its 1% amount, urea became a fertilizer as efficient or even more efficient
as lime ammonium nitrate in all soils.
The higher efficiency of urea in the barley experiments than in those with rye is
explained by a greater number of sandy soils used in the rye experiments and not by a
differentiated urea action determined by the species of cereals, because the sandy soils
lost more urea-N as volatile ammonia than did the other soils.
The effects exerted by weather conditions and soil texture on total N contents in
green matter and grain were similar to those exerted by them on grain yield.
In conclusion, the experiments carried out on large surfaces confirmed the results
obtained in microplots and plots in respect of the fertilizing efficiency of urea-
PPDA.
The experiment in which Bremner and Krogmeier (1989) found that PPDA added to
soil samples completely eliminated the adverse effect of urea on germination of seeds of
four plant species, including barley, is dealt with on page 276. See also Table 61.
291
In the pot experiments described by Winiarski (1990), PPDA added at a rate of 1%

to urea fertilizer led up to a 26% increase in dry matter yield of barley. The plant uptake
ofN also increased in the urea-PPDA treatment.
Kucharski et af. (1990) and Kucharski (1994) described a 3-year field experiment on
22_m2 plots installed on a brown soil of loamy sand texture (pH 5.9-6.3). The test plant
was spring barley. The plots were fertilized yearly with 26 kg P/ha as triple
superphosphate, 83 kg Klha as KCI and with urea or urea + 1% PPDA (relative to urea
weight) at rates of 0, 40, 80, and 120 kg N/ha. Grain and straw yield as well as uptake
and utilization of N by the barley plants were not significantly affected by PPDA at any
rate ofurea+PPDA.

Compounds
Barley was one of the test plants on which Bremner and Krogmeier (1988) studied the
effect of 10 inhibitors, including four PTAs and one TPTA (see page 275). See also
Table 61.
The effects of nBTPTA on winter barley and winter wheat were studied by
Goodroad and Wilson (1989) under similar field conditions, and the results obtained
were also similar (see page 277).
Grant and Bailey (1997, 1999) conducted field experiments on a clay loam and a
fine sandy loam soil at two locations near Brandon, Manitoba, over 3 years (1994-1996)
to evaluate the effect of seed-placed urea (0,20,40, 60, 80, and 100 kg N/ha), with and
without addition of nBTPTA, on stand density, growth, and yield of barley.
Seedling damage, as indicated by reduction in stand density, occurred on both soils
at rates of seed-placed urea as low as 40 kg N/ha. Addition of nBTPTA to urea
increased stand density at N levels where damage occurred with the urea applied
without nBTPTA. Grain yields were increased by nBTPTA on both soils. The average
yields (t/ha) at the 0, 20, 40,60,80, and 100 kg N/ha rates of urea with no nBTPTA and
with nBTPTA were the following: 2.517 and 2.517, 2.691 and 2.699,2.555 and 2.905,
2.991 and 3.232, 2.652 and 3.401, and 2.856 and 3.172, respectively, on the clay loam
soil, and 2.870 and 2.870,3.091 and 3.095, 3.165 and 3.462, 3.244 and 3.283,3.008 and
3.298, and 2.741 and 3.910, respectively, on the fine sandy loam soil.
The conclusion was drawn that use of nBTPTA appears promising as a method of
reducing the risk of seedling damage from seed-placed urea fertilizer, thus increasing
the rate ofurea-N that can be placed with the seed.
7.4.7. Effoct q(Cyclotriphosphazatriene (CTPAT) Derivatives

See Table 61.
7.4.8. F;Uect of Combined Urease and Nitrification Inhibitors

Muravin et af. (1983) compared the effect of nitrapyrin and ATC (4-amino-l,2,4-
triazole hydrochloride) used without or together with hydroquinone (HQ) on the uptake
of urea-N by barley plants cultivated in pots containing a limed, previously acid soil
(soddy-podzol, pH 4.9). Urea, labeled with 15 N, was used at a rate of 0.1 g/kg soil;
nitrapyrin and ATC were applied at a rate of 2% and HQ at 10% relative to urea-No The
following analyses were carried out: total N in soil and plants, mineral N (NH4 + + N0 3·)
292
in soil, isotopic composition ofN in soil and plants. Crop yields were also assessed, and
the N balance was calculated.
In the soil treated with urea alone, the plants took up 51 % of the applied urea-N,
19% remained in soil, and 30% was lost through volatilization. The corresponding
values recorded in the other treatments were the following: urea + nitrapyrin: 54, 21,
and 25%; urea + nitrapyrin + HQ: 57, 23, and 20%; urea + ATC: 53,23, and 24%; urea
+ ATC + HQ: 54, 24, and 22%, respectively. Thus, the most favorable effect on N
uptake by plants and reduction of volatile N losses was exerted by the mixture of
nitrapyrin and HQ. Grain yield was also highest in the urea + nitrapyrin + HQ treatment
(see also Muravin, 1989).
Pisareva and Muravin (1988) and Pisareva (1989) described pot experiments in
which the effect of HQ applied together with the nitrification inhibitor, 3-
methylpyrazole-l-carboxarnide (MPC) and with 15N-labeled urea (25 atom% excess)
(urea + MPC + HQ) on barley plants was compared with those of urea + MPC and urea
alone. Two soils were used: a soddy-podzolic soil (PH 5.5) and a calcareous chernozern
(pH 7.8). Both received PK as basal fertilizers. Rates of application per kg of soil were:
P2 0 S: 50 mg; K2 0: 75 mg; urea-N: 100 mg; MPC: 2 mg; HQ: 5 mg. The experiments
were initiated in 1985 and repeated in 1986. Grain yields on the podzolic soil in 1986
and 00 the chemozern in both 1985 and 1986 were highest in the urea + MPC + HQ
treatment (the highest grain yield on podzolic soil in 1985 was recorded in the urea +
MPC treatment). But N contents in grains were always lower in the urea + MPC + HQ
treatment than in those with urea + MPC and urea alone. HQ decreased plant uptake of
N from soil (podzol in 1985; chernozem in both years) or increased it (podzol in 1986).
7.5. EFFECT OF UREASE INHIBITORS ON OATS (Avena sativa)
7.5.1. Effict Q{Heavy Metal Compounds

In the pot experiments ofWyszkowska et al. (2001) (see page 19), K2 Cr2 0 7 reduced soil
urease activity even at its lowest rate (40 mg Cr/kg soil), but had at this rate no
significant effect on dry matter yield (-35 glpot). At the rate of 80 mg Cr/kg soil, the
yield significantly decreased in the soil not amended with straw and remained
unaffected in the straw-amended soil. At the highest rate (120 mg Cr/kg soil), the dry
matter yield was -3 glpot in the soil not amended with straw and -25 glpot in the straw-
amended soil. Thus, phytotoxicity of Cr was significantly reduced by the straw
amendment.
7.5.2. Effict ofInorganic Boron Compounds

Geissler et af. (1970), who studied ammonia volatilization from urea pellets prepared
from urea melt and inhibitors, without any hydrophobic material (see page 179), also
described a pot experiment using oats as test plants. The soil (a loamy sand, pH 6.4) was
fertilized, at a practical rate, with urea or with urea containing 2, 4 or 6% borax or with
ammonium nitrate, all being applied on the surface of moist soil, and after 3 weeks oats
were seeded and allowed to grow for 8 weeks. Thereafter, the plants were harvested and
their dry weight determined. It has been established that the yield was highest in the
NH4 N0 3 treatment and lowest in the urea treatment. When borax-containing urea pellets
were administered, the yield increased and, at 6% borax content, was nearly the same as
in the NH4N0 3 treatment.
293
7.5.3. Effect of Fluorides

root growth of oat plantlets was strongly reduced at rates 2: 0.5 g F/kg soil.
7. 5.4. ~ffect of Urea Derivatives

In Kolyada's (1970, 1973) pot experiments (see page 49), the oat plants behaved
towards thiourea like the barley plants (see page 289). The average grain yield of oats
was 21% higher in the treatment with thiourea-N + NH4 N0 3-N + PK than in the
NH4 N0 3 -N + PK treatment, the N, P, and K addition being in both treatments 100
mg/kg soil.
In a pot experiment described by Germann-Bauer (1987), mixtures of 5 kg of a loess
brown earth with 1 kg of quartz sand were amended with 1 g P20 S as dicalcium
phosphate, 1.2 g K20as potassium sulfate, and 1,200mg N as urea or 1,171 mg urea-N
+ 60 mg guanylthiourea (GTU) or 1,143 mg urea-N + 120 mg GTU. The mixtures were
moistened to 50% ofWHC and preincubated at 17-25°C. After 2 and 3 (or 5) weeks, the
mixtures were percolated with water in amounts equivalent to 30 or 80 mm of rainfall.
In the 6th week, oats were sown in each pot. During the growth period, the pots were
kept at ambient temperature (8-35°C).
The grain yields (g fresh weight/pot) were 41.5-46.8 (urea-only treatment), 56.4-
59.0 (urea + 60 mg GTU), and 58.7-60.2 (urea + 120 mg GTU). Due to GTU,
significant increases occurred also in the N uptake by plants.
7.5. 5. ~ffect of Phosphorodiamides

Held et al. (1978) described pot experiments carried out in the 1973-1977 period. Urea
rate was 0.5 g of N per pot containing 8 kg of soil basally fertilized with P, K, and Mg
and moistened to about 50% ofWHC. Urea prills and urea conditioned with mineral oil-
bitumen mixture were tested. Phenylphosphorodiamidate (PPDA) was applied usually
as coating on conditioned urea at a rate of 0.5 or 1% relative to urea-No The fertilizers
were surface-applied or mixed into the soil. Thereafter, the volatilized ammonia was
measured. At days 10, 15 or 20 after fertilizer application, the undecomposed fertilizers
were washed into the soil with 400 ml of distilled water. Following the last washing,
oats were sown in all the pots. At the end of growing season, the yields (grain and
straw) were recorded.
The results obtained in various years did not differ essentially from one another. For
example, in 1976 the surface-applied prilled and conditioned urea lost about 26% of
their N content as volatile NH 3 • But NH3 loss was only approximately 1% when they
were mixed into the soil. PPDA reduced the 26% volatile NH3 loss from the surface-
applied urea to less than 5% and, consequently, caused significant (10-19%) increases
in oat yields. The 1% rate was not always more efficient than the 0.5% rate.
Heber et al. (1979) conducted, in 1976 and 1977, pot experiments on two soils:
loamy sand and sand (6 kg/pot). Three N sources: urea, urea + 1% PPDA (on urea-N
basis), and ammonium nitrate were compared. The total amount of 0.8 g N/pot was
surface-applied either after emergence of oat seedlings or when the plants reached 25
cm height, or 0.4 + 0.4 g N/pot were administered at both stages. In a variant, urea
(without PPDA) was mixed into the soil before sowing of oats. At day 14 after their
294
surface application, the fertilizers were washed into the soil with 200 ml of water. Dry
matter and total N contents in grains and straw were determined.
The results showed that in both soils and both years the grain and straw yields as
well as the total N contents increased, under the influence of the applied fertilizers, in
the following order:
urea < urea+ I % PPDA < NH4 N0 3 •
Utilization of N from urea, urea+ I % PPDA, and NH4N0 3 had the mean values of
30, 70, and 90%, respectively. In the variant in which urea was mixed into the soil
before sowing of oats, crop yields and utilization of N corresponded, in most cases, to
the values registered in the urea+ I % PPDA treatments. The rate of 0.8 g N/pot applied
after emergence of oat seedlings was more efficient than the same amount applied at 25-
cm high plants, which was explained by higher temperatures in the period when the
plants reached this height. The same N rate in divided application (0.4 + 0.4 g N/pot)
did not lead to increased oat yield.
In 1977 a similar pot experiment was carried out by Matzel et al. (1979b). Two soils
(a sandy loam and a sand) were used and oats served as test plants. In a variant, 15N_
labeled urea (0.8 g N/pot) without PPDA was mixed into the soil (6 kg) before sowing.
In the other variants, urea was surface-applied and washed into the soil with 200 ml of
water after 14 days. We specify these variants: labeled urea (0.8 g N/pot) with or
without 1% PPDA was applied immediately after emergence of seedlings; the same rate
of labeled urea with or without 1% PPDA was applied at 25-cm high plants; unlabeled
urea (0.4 g N/pot) with or without 1% PPDA was administered at emergence of
seedlings, whereas the labeled urea (also 0.4 g N/pot) with or without 1% PPDA was
applied at 25-cm high plants.
Analyses of total N and 15N in plants indicated that in both soils the crop yield (grain
and straw) and plant uptake of N were highest in the variant in which urea without
PPDA had been mixed into the soil. In the other variants, PPDA increased both crop
yield and uptake of N from urea. Under the influence of PPDA, the plants took up, on
an average, II % more N from urea.
7.5.6. Effect of' Phosphoric Triamide (PTA) and Thiophosphoric Triamide (TPTA)
Compounds
Bremner and Krogmeier (1988) used oats among the test plants on which they studied
the effect of 10 inhibitors, including four PTAs and one TPTA (see page 275).
7.6. EFFECT OF UREASE INHIBITORS ON RYE (Secale cereale)
7. 6.1. ~[rect qfPhosphorodiamides

In field experiments carried out in the 1977-1979 period, Karnpfe et al. (l982a) studied
the effect of phenylphosphorodiamidate (PPDA) on winter rye grown on a sandy soil.
Four N fertilizers, namely conditioned urea (prilled urea coated with mineral oil-
bitumen mixture), conditioned urea + 0.5 or 1% PPDA, and lime ammonium nitrate,
were compared in respect of their effect on grain yield and total N content in green
matter of plants under conditions of natural weather and simulated drought.
The N fertilizers were applied twice, at rates of 50 and 40 kg fha, respectively. PK
fertilizers were also administered, at rates of 26 kg P and 120 kg Klha. Drought was
simulated by means of plastic tents (6 by 4.5 m) put up over the plots. The simulated
295
drought began after addition of the first or second N rate and lasted 14 days.
Unfertilized plots served as controls.
TABLE 64. Effect of nitrogen fertilizers on grain yield of winter rye (mean
values for the 1977-1979 period)"
Grain yield ofwinter rye
Fertilizer
dtlha %
Unfertilized control 20.8 53
Lime ammonium nitrate 39.0 100
Conditioned urea 36.8 94
Conditioned urea + 0.5% PPDA 38.6 99
Conditioned urea + 1% PPDA 38.7 99
LSD 5% 2.0 5
"Adapted from Kiimpfe et al. (1982a).
Table 64 shows that diminution of grain yield in urea treatment in comparison with
the lime ammonium nitrate treatment could be prevented by using urea+PPDA. There
was no significant difference between the effects of the 0.5 and 1% PPDA rates.
The 14-day simulated drought after fertilization led to a decrease in grain yield as
compared to that recorded under natural weather conditions. In the 1977-1979 period,
the yield decreased, under the influence of simulated drought, to 96% (in the lime
ammonium nitrate treatment), 89% (urea), 92% (urea + 0.5% PPDA), and 94% (urea
+1%PPDA).
Total N contents in green matter of plants were determined twice, namely at days 14
and 28 after application of each of the two N fertilizer rates. During the first 14 days,
PPDA retarded N uptake by the plants, but at day 28 after N application there were no
significant differences between the lime ammonium nitrate treatment and the urea + 0.5
and 1% PPDA treatments as concerns total N contents in green matter. In the 1977-1979
period, neither were these contents influenced by simulated drought.
The results of the 136 field experiments reviewed by Kiimpfe et at. (1982b) (see
pages 271-272) are valid not only for winter wheat but also for winter rye. The results
of the 43 experiments on large surfaces (Kiimpfe et at.. 1983) refer not only to winter
barley but also to winter rye (see page 289).
The experiment in which Bremner and Krogmeier (1989) found that PPDA added to
soil samples completely eliminated the adverse effect of urea on germination of seeds of
four plants, including rye, is dealt with on page 276.

Compounds
Rye was one of the test plants on which Bremner and Krogmeier (1988) studied the
effect of 10 inhibitors, including four PTAs and one TPTA (see page 275).
7.7. EFFECT OF UREASE INHIBITORS ON SORGHUM (Sorghum bicotor)
7.7.1. Effect of Inorganic Sulfur Compounds

Lamond et at. (1986, 1987, 1991a) conducted 3-year field experiments on two Kansas
soils used for continuous, no-till, grain sorghum production. The experiments were
296
carried out in the 1986-1988 period on a silt loam and in the 1987-1989 period on a silty
clay loam. Urea-ammoniumnitrate (UAN) solution was the N fertilizer at rates of 0, 56,
and 112 kg N/ha. Ammonium thiosulfate (ATS) was added to UAN at a rate of 10%
(volume/volume). Placement methods were: surface broadcast, dribble surface band on
51-cm centers, and knifed-injected 15-18 cm deep on 51-cm centers.
Grain yields and N concentrations of leaf tissue and grain were consistently and
significantly increased by N fertilization of both soils. Knifed placement of UAN was
superior to broadcast and dribble application. The results obtained with ATS added to
UAN were inconsistent. For example, in 1986 the broadcast UAN + ATS, as compared
to broadcast UAN, produced a 5% grain yield increase (at 56 kg N/ha) and an 11.7%
increase (at 112 kg N/ha); in 1989 ATS broadcast with UAN induced an 11% grain
yield decrease (at 56 kg N/ha) and a 3.5% increase (at 112 kg N/ha). On overall
average, the grain yield increase due to ATS was 4.5% on silt loam, and 1.2% on the
silty clay loam, both increases being unsignificant at p=0.05.
7.7.2. F;[fect ofOrganic Mercury Compounds

See Table 61.
7.7.3. F;[{ect ofPolyhydric Phenols and Quinones

See Table 61.
7. 7. 4. Effect ofPhosphorodiamides
A pot experiment in which the effect ofphenylphosphorodiarnidate (PPDA) on sorghum
was studied, is mentioned in a report presented by the International Fertilizer
Development Center (IFDC, 1981). 15N-Labeled urea was used in form of large
granules (supergranules of 0.25-1 g) point-placed at 10-cm depth in soil (clay) or as
prills incorporated into the top 2 cm of soil or as prills with or without 2% PPDA
applied on soil surface. The plants were watered by simulated rainfalls (245 or 465
mm). Depending on the fertilizer and water managements, the plants took up 15N in the
following proportions: 47.2% (from surface-applied urea), 52.5% (from surface-applied
urea-PPDA), 56.5% (from urea incorporated into the soil), and 63.5% (from
supergranules) under the conditions of 245-mm simulated rainfalls. The corresponding
values in the case of more abundant (465 mm) simulated rainfalls were: 52.3, 54.7,
59.4, and 70.6%, respectively. Thus, PPDA increased, to some extent, efficiency of the
surface-applied urea. The supergranules point-placed at 10-cm depth were more
efficient than was the surface-applied urea-PPDA, but an eventual enhancing of
efficiency by PPDA added to supergranules was not studied. The data cited also show
that incorporation of urea prills is more advantageous than their surface application, and
each form of urea was more efficient with more abundant simulated rainfalls.
These investigations, performed at IFDC Headquarters (Muscle Shoals, Alabama),
were described, with more details, but with similar conclusions, by Buresh et al. (1984).
See also Table 61.
7.7.5. F;[tec/ of PiJo!;phoric Triamide (PTA) and Thiophosphoric Triamide (!,PTA)

Compounds
Sorghum was one of the test plants on which Bremner and Krogmeier (1988) studied
the effect of 10 inhibitors, including four PTAs, nBTPTA. and PPDA, and on which
297
Krogmeier et al. (1989a) studied the relation between the urea-caused leaf tip necrosis
and the soil urease-inhibiting effect of nBTPTA and PPDA (see page 276). See also
Table 61.
In the field experiments conducted by Goodroad and Wilson (1989) in Georgia, the
effects of urea and urea-nBTPTA on the grain yields of sorghum were not apparently
different, because drought conditions were most likely to reduce yields.
Lamond et at. (1993, 1994, 1998) evaluated the effect of nBTPTA on no-till and
conventional-till continuous grain sorghum in Kansas. The N sources [urea, urea +
nBTPTA (Agrotain), and ammonium nitrate (AN)] were surface-broadcast and were
incorporated in the conventional tillage system.
All N sources performed similarly in the conventional tillage, but AN and
urea+nBTPTA often outperformed urea in no-till. For example, urea, urea+nBTPTA,
and AN, applied at rates of 0, 56, 112, and 168 kg N/ha on the no-till sorghum fields at
Belleville, produced the following grain yields (bu/acre): 76 (no N added), 102, 100,
and 107 (at 56 kg N/ha), 116, 118, and 129 (at 112 kg NIha), and 127, 129, and 128 (at
168 kg N/ha), respectively, in 1993, and 37 (no N added), 85, 88, and 90 (at 56 kg
N/ha), 107, 116, and 108 (at 112 kg N/ha), and 112, 118, and 118 (at 168 kg N/ha),
respectively, in 1994. Thus, the results indicated that the use of nBTPTA can improve
efficiency of urea when surface-broadcast in no-till production system.
7. 7. 6. Effect of Cyclotriphosphazatriene (CTPA T) Derivatives

See Table 61.
7.8. EFFECT OF UREASE INHIBITORS ON GRASSES
Many grass species were studied. We nominalize them in alphabetical order:

bermudagrass (Cynodon dactylon) , bluegrass (Poa pratensis), bromegrass (Bromus
inermis), caucasian bluestem (Botriochloa caucasica), Italian ryegrass (Lotium
mult(florum), lemongrass (Cymbopogon flexuosus), lovegrass (Eragrostis curvula),
orchardgrass (Dactylis glomerata), perennial ryegrass (Lotium perenne), Saint
Augustine grass (Stenotaphrum secundatum), tall fescue (Festuca arundinacea).
The varieties and hybrids will be nominalized in the sections dealing with the effect
of urease inhibitors on them.
7.8.1. E.tfect ofAlkali Metal and Alkaline Earth Metal Salts

In the field experiments of Rappaport and Axley (1984), the plots (3 by 15 m) were
installed on a sandy loam soil in Maryland. The test plant was hybrid sorghum-sudan
grass (Sorghum sudanese). The solution of urea (8.4 g N/m2) and that of urea + KCl
(urea:KCl=I:1 by weight) were applied 3 days after germination of grass seeds or KCl
was applied 10 days after addition of urea. The hay yield increase from 574 to 660 and
662 glm2 , due to addition of KCl simultaneously with urea or 10 days later,
respectively, was significant (p=0.05).
In the field experiment performed by Rodgers et at. (1984) (see page 27), the slight
reduction of ammonia loss from eaClz-amended urea priUs applied on a perennial
ryegrass ley led to increases in N uptake by plants and in yields.
298
Fenn (1985) conducted experiments in 1979 and 1980 on bermudagrass growing on

a calcareous fine-textured sandy soil along golf course fairways. Urea and urea-calcium
nitrate (four parts urea-N + one part calcium nitrate-N) were compared. Rate of N
addition was 70 kglha in 1979 and 70 + 100 kglha in 1980. Color intensity and verdure
density (top growth below the 2.5 cm mowing height) were greater in the treatment with
urea-calcium nitrate than in the treatment with urea alone at both application rates.
The effect of calcium chloride on bermudagrass uptake of urea-N and yield was
studied by Sloan and Anderson (1987, 1998). Field and greenhouse experiments were
carried out on two Texas soils (a fine sandy loam and a clay). In the urea+CaCh
applied, the CaIN equivalent weight ratio was 0.25.
In the field experiments, urea and urea+CaCl 2 were surface-applied at a rate of 100
kg Nlha on the bermudagrass plots and allowed to remain on the surface until rainfall
moved them into the soil. The results of more than 31 trials throughout the growing
season showed that urea+CaCIz performed no better than the urea alone.
In the greenhouse experiments, 15N-labeled urea and urea+CaCh at a rate of 200 kg
Nlha were surface-applied to bermudagrass sod cores from the two soils. The sod cores
were subjected to either low or high rainfall regimes beginning 7 days after fertilizer
application. After 1 month, bermudagrass was harvested and analyzed for total Nand
15N contents. Calcium chloride significantly increased bermudagrass N use efficiency of
surface-applied urea by 33 to 47% on the sandy loam soil but had little effect on the
clay soil. At the same time, the bermudagrass dry matter yields were not significantly
different between the urea+CaCb and urea treatments, and tl1is finding was valid for
both soils and both low and high rainfall regimes.
7.8.2. E.tfect ofInorganic Boron Compounds

Sor et al. (1968, 1971) and Esso Company (1969) studied the efficiency of
commercially prepared urea pellets with borax and octadecylamine (ODA) under field
conditions by using tall fescue as test plant. A loamy sand soil from New Jersey was
used; its initial pH of about 6.4 was adjusted to 7.0 with CaCO) and MgC0 3 • Urea was
applied at a rate of 112 kg Nlha. The following green and dry matter yields (g/230 dm2 )
were recorded in the different treatments: 261 and 104 (no urea control); 579 and 140
(urea); 686 and 135 (urea + 4% borax); 983 and 220 (urea + 4% borax + 1% ODA),
respectively. It is clear that the urea pellets with 4% borax and 1% ODA were most
efficient in increasing the grass yield. It should be added that borax and ODA, applied
without urea, did not have any significant effect on the grass yield.
Efficiency of fertilization with urea-4.1 % borax-l % ODA was also studied by Sor et
al. (1971) in field experiments carried out under a variety of conditions (climate, soils,
plants), in New Jersey, California, Arizona, and Florida as well as in France, by using
bluegrass, tall fescue, bermuda grass, Saint Augustine grass as test plants. Under the
influence of the urea-borax-ODA, the crop yields increased by 1-66% in comparison
with those registered in the treatment with urea alone.
7.8.3. Etfect o/Inorganic Su((ur Compounds

Results of the laboratory investigations of Gascho (1986), according to which both
ammonium thiosulfate (ATS) and KCl or CaCh reduced volatile an1monia losses from
urea-ammonium nitrate (UAN) applied on samples of a loamy sand (PH 6.8), incubated
at 30 D C for 7 days (see page 38), were partially confirmed and, at the same time,
299
partially not confirmed through the field experiments conducted by Gascho and Burton
(1987), in Georgia, on a loamy sand (pH 4.7) covered with an established sod of Tifton
44 hybrid bermudagrass. Eleven days before starting the experiments, the soil was
limed with 3.36 tlha of broadcast agricultural-grade dolomite, which resulted in the
increase of pH to 6.2 in the 0-5-cm soil layer but only to 5.0 in the 5-10-cm layer.
Three experiments were carried out over two years. N was applied at a rate of 336
kglha/year in the first and third experiment and 168 kglhalyear in the second one. In
each experiment, VAN served as N source. In the second experiment, urea alone and
NH 4 NO} alone were also used. ATS was added to VAN at a rate of 5%
(volume/volume). Besides the treatments with VAN and VAN + ATS, there were other
treatments, namely VAN + ATS + KCI, VAN + ATS + KCl + ammonium
polyphosphate (APP), and UAN + ATS + KCI + APP + CaCb. Solutions were applied
as a dribble on a spacing of 15 cm.
Four parameters were estimated: dry matter yield, percent N content and N removed
in dry matter, and relative N efficiency, i.e.. (N removed in dry matterlN removed in dry
matter when VAN was applied alone) x 100.
The estimations showed that in experiments 1 and 2 ATS added to VAN had a
favorable, statistically significant or insignificant effect on these paran1eters. In all
experiments, KCl, APP, and CaCl 2 added to VAN with or without ATS as well as ATS
added to UAN in experiment 3 did not influence the parameters or did diminish them,
but the diminutions were, in general, insignificant. Dry matter yield was significantly
higher with NH 4 N0 3 than with urea, but in the VAN treatment the dry matter yield did
not significantly differ from yields obtained with NH4 N0 3 alone and urea alone. At the
same time, in respect of the other parameters, differences between the VAN, NH4 N0 3 ,
and urea treatments were not significant. The authors consider that these results are
related to the reduced NH3 volatilization which was due, probably, to soil pH remaining
acid even after dolomite broadcast.
In a short report, Lamond and Bonczkowski (1989) summarized the results of 2-year
field experiments (1987 and 1988) initiated to evaluate the effect of VAN (at rates of 0,
67, and 134 kg Nlha) applied with or without ATS on bromegrass. Addition of ATS to
VAN produced inconsistent results but in some cases increased the yields.
In the field and greenhouse experiments conducted by Sloan and Anderson (1987,
1998) and referred to in Section 7.8.1, ATS added to urea at a rate of 10%
(weight/weight) was also tested. In the field experiments, urea+ATS performed no
better than urea alone. In the greenhouse experiments, in which two soils were studied
under a high and a low rainfall regime, ATS did not affect bermudagrass N use
efficiency for either soil or rainfall regime. On both soils, bermudagrass dry matter
yields in the urea+ATS and urea treatments were not significantly different under low
rainfall regime, but under the high rainfall regime ATS significantly decreased the
yields.
7. 8.4. Ef.fi!Cl of Urea Deriva lives

Amberger and Gutser (1984) carried out a pot experiment with ryegrass. Samples (15
kg/pot) of a loamy sand soil (PH 6.2) were amended with 3 g of N in form of urea, urea-
N 95'Yo + thiourea-N 5% or urea-N 90% + thiourea-N 10%. N uptake by the plants was
highest when urea was amended with 10% thiourea-No
300
7. R. 5. Effect ofDithiocarbamates
Hyson (1963), who patented dithiocarbamates as inhibitors of soil urease activity (see
page 52), pointed out that the dithiocarbamate-amended urea fertilizer was applied by
broadcasting to areas of bermudagrass at a rate of 28-560 kglha with outstanding
beneficial results.
7.8.6. Effect ofXanthates

Ashworth et al. (1979) studied the effect of potassium ethyl xanthate (KEtX) on
germination and growth of ryegrass in vegetation pots. Germination was delayed by
KEtX, and it retarded the growth of plants for 3 weeks, but after 7 weeks mean dry
matter yields per pot were not significantly affected by KEtx.
7. R. 7. Effect ofBramo-nitro Compound5

In pot experiments carried out by Norden et af. (1985, 1986), samples of a loamy sand,
limed to pH 5.6 and basically fertilized with nitromagnesia and Thomas potash, were
additionally fertilized with urea (0, 0.5, 0.75, and 1.0 g N/pot as top dressing) with or
without a urease inhibitor (at a rate of 2% by weight ofurea-N) and sown with Italian
ryegrass. The inhibitor tested was a bromo-nitro compound (BNC), namely 2-bromo-2-
nitropropyl-N-methylcarbamate. N-Methyl-maleimide (N-methyl-MI) was the reference
inhibitor. The plants were analyzed after five cuttings in the period of June 23 -
November 27.
The analyses showed that the amount of N taken up by the plants like their dry
matter and raw protein contents in the experimental variants increased in the order: no
urea < urea < urea+N-methyl-MI < urea+BNC (at each urea rate).
7.8.8. F/Ject ofPolyhydric Phenols and Quinones

Tomlinson (1970) performed pot and field experiments. In pots filled with several soils
from England, Italian ryegrass was sown. Urea was applied twice, each application
being equivalent to 224 kg of Nlha. The inhibitors tested were hydroquinone (HQ) and
2,5-dimethyl-p-benzoquinone used at a rate of 1% relative to urea-No Ammonium
nitrate served for comparison. The results (Table 65) indicate that the inhibitors usually
increased the growth response of ryegrass; however, they did not increase the efficiency
of urea to that of ammonium nitrate (AN).
TABLE 65. Effe~t of hydroquinone (HQ) and 2,5-dimethyl·p-benzoquinone (DBQ) on growth of

Italian ryegrass on several soils in pot experiments"
Dry matter of plants (g/po! and %)
Treatment
Soil A Soil B Soil C Soil D SoilE
Urea alone 8.65( 100) 6.24(100) 4.58(100) 9.53(100) 10.61(100)
Urea + 1% HQ 9.13( 106) 6.93(111) 4.95(108) 9.63(101) 11.32(107)
Urea + 1% DBQ 8.89( 103) 7.53(120) 4.15( 90) 10.38(109) 11.43(108)
NH4 NO, 13.23(153) 11.53(184) 8.81(192)
"From Tomlinson (1970).
In the field experiments carried out in England, the same fertilizers and inhibitors
were applied on grasslands as in the pot experiments. Rate of fertilizer application was
301
50 or 100 kg N/ha, and that of inhibitors was 1% relative to urea-N. The inhibitors
increased the yields, but efficiency of urea + inhibitor seldom attained the efficiency of
AN. Similar results were obtained in field experiments performed in South Africa where
the fertilizers and inhibitors were applied to lovegrass plots.
Calancea et al. (1977) carried out pot and field experiments for studying the effect
of hydroquinone (HQ) and p-benzoquinone (BQ) on Italian ryegrass sown on a
chemozemic soil. The pots contained 1 kg of soil to which 1 kg of sand was mixed.
Each of the plots had a 30-m2 area.
In the pot experiments, in which urea-N rates were 200,600, and 1,800 mg/pot at a
constant HQ rate (30 mg/pot), ryegrass yield (dry matter of herbage), total N uptake by
plants, and coefficient of N utilization from urea tended to increase with increasing
urea-N rate. When urea-N rate was constant (200 mg/pot) and HQ rates were different
(60,90, and 120 mg/pot), the highest coefficient ofN utilization was recorded at the 90
mg/pot HQ rate, although yield and total N uptake showed a trend to increase with HQ
rate. These effects of HQ were very marked at the first harvest (cut) and less evident at
the second and third cuts.
Under field conditions, HQ was more efficient when urea was applied as a single
dressing (300 kg N/ha) than when it was administered as two or three divided dressings
(2 x 150 kg N or 3 x 75 kg N/ha). Thus, the coefficient of N utilization from urea
increased, due to HQ, by 18.4, 12.0, and 7.0%, respectively.
Of six treatments (100, 200, and 400 kg urea-N/ha + 0 or 1 kg BQ/ha), the best
results concerning coefficient of N utilization from urea were obtained with 200 kg
urea-N/ha + 1 kg BQ/ha.
In other pot experiments on the same chernozemic soil, Calancea et at. (1982) found
that HQ enhanced the uptake of N by ryegrass plants from both urea and glycoluril
(acetyleneurea; a slow-release N fertilizer), and this effect ofHQ was more marked with
glycoluril than with urea.
In the pot experiments described by Hera et al.( 1980), samples of a leached
chernozem and a brown forest soil were used. The soil (1 kg) and sand (0.5 kg)
mixtures were treated with 150 mg of urea, labeled with 15N (5.225 atom% excess), and
with 3 mg of HQ, then sown with ryegrass. Untreated mixtures and mixtures treated
with urea alone were the controls. After 6 weeks of ryegrass growth, it was established
that on the chernozem the yields increased by 57% in the urea + HQ treatment and by
31 % in the urea treatment as compared to the untreated control. The corresponding
values registered on the forest soil were 51 and 25%, respectively. Analysis of 15N in
plants indicated that uptake of urea-N was higher by 19% (chernozem) or by 23%
(forest soil) when urea was applied together with HQ than when urea was used alone.
All increases were statistically very significant.
A field experiment was also conducted on 30-m2 plots on the leached chernozem.
Urea was applied at rates of75, 150, and 300 kg N/ha with or without 2% HQ (on urea
weight basis). Under the influence of HQ, the ryegrass yield and N uptake from urea
increased by 30-37 and 15-20%, respectively.
The effect ofHQ on N uptake from urea and soil by ryegrass plants was also studied
in a pseudogleic podzolic soil (Calancea and Kiss, 1984). The pots contained mixtures
from 5 kg of soil and 1 kg of sand. Urea, 15N-Iabeled (5.225 atom% excess), was added
to the pots at a rate of 200 mg of N with or without 30, 60, 90, and 120 mg of HQ/pot.
These amounts are equivalent to 100 kg of urea-N/ha and to 15, 30, 45, and 60 kg of
302
HQ/ha. The soil-sand mixtures were sown with ryegrass, then moistened and kept at 65-
70% of WHC in greenhouse for 90 days, during which the herbage was harvested (cut)
three times, then analyzed for total Nand I'N contents.
One can see from Figure 78 that the ratio of N uptake from soil to N uptake from
urea was lower in the HQ-treated soil than in the untreated control during the whole
experimental period. The ratio increased linearly with the age of plants, and there was a
s,o
E
g JD
i
~ 2Sl
E
g
.l!
i 1.0
z
AtJe Ilf plantsldoysl
Figure 78. Elleet of hydroquinonc (IIQ) on the ratio of N uptake Irom soil to N uptake from urea by
ryegrass plants.
a - Control (no HQ), b- 30 mg HQ/pot. e - (i0 mg HQ/pot. d - 90 mg HQ/pot. e- 120 mg HQ/pot.
IFrom Calancea and Kiss (1984)./
parallelism between the increase of this ratio and the increase in the rate of HQ. The
ratio was higher than I, i.e., the plants took up more N from soil than from urea, except
in the lO-day-old plants growing in pots treated with 30 and 60 mg of HQ. These young
plants took up more N from urea than from soil, especially at 30 mg of HQ/pot. In
addition, of all treatments, that with 30 mg of HQ resulted in the lowest ratio between N
uptake from soil and N uptake from urea. Thus, the optimum rate for stimulation of N
uptake from urea was 30 mg of HQ/pot, i.e., 15 kg ofHQ/ha.
Gorelik et al. (1983) studied, in the 1979-1981 period, the effeet of hydroquinone
(HQ) on crop yield and coefficient of utilization ofurea-N in vegetation pots, eaeh filled
with 2.5 kg of soil. Details on the experiments are given below:
Soils Plants Rates

Loamy-sandy soddy-podzol Orchardgrass 0.75 or 1.5 g of urea-N/pot; 0, I or 5% HQ
and heavy loam (sierozem) relative to urea weight (in 1979)
Sandy soddy-podzol Orchardgrass 0.38 or 0.75 g of urea-N/pot; 0, I or 5% HQ

relative to urea weight (in 1980)
Sandy soddy-podzol Perennial ryegrass 0.38 or 0.75 g of urca-N/pot; O. I. 2 or 5% HQ

relative to urea weight (in 1981)
303
Rates of the urea and HQ, divided into 5 equal parts, were applied at intervals of
about 2 weeks on the soil surface immediately after each harvest (cut). Dry weight and
total N content of plants were determined.
The results showed that in the loamy-sandy podzol fertilized with urea at the higher
rate (1.5 g N/pot) and in sierozem, at both urea rates, HQ did not significantly affect the
efficiency of urea. When the loamy-sandy podzol was fertilized with urea at the lower
rate (0.75 g N/pot), the yields increased significantly under the influence of both HQ
rates. whereas the coefficient of utilization of urea-N by plants increased from 41 % (no
HQ) to 49% (1 % HQ). and 52% (5% HQ). In the sandy podzol under orchardgrass. HQ
increased significantly the yield only when urea was applied at the lower rate (0.38 g
N/pot) and HQ at the higher rate (5%); coefficient of utilization ofurea-N (35% in the
no HQ treatment) increased. under the influence of 1 and 5% HQ. to 40 and 48%,
respectively. In the sandy podzol under ryegrass. HQ was efficient in yield-increasing at
both urea rates; the coefficient of utilization of urea-N, in presence of 0, 1, 2, and 5%
HQ, had the following values: 40, 40. 45, and 48%, respectively, when urea rate was
0.38 g N/pot, and 36,41,44, and 49%, respectively. when urea rate was 0.75 g N/pot. It
appears evident from these results that HQ acted more efficiently in a light-textured
than in a heavier-textured soil.
Rodgers et al. (1984) studied the effect of HQ on the efficiency of urea in
experiments on ryegrass leys at Rotharnsted. HQ, added at a rate of 5 kglha to the
annual rate of 375 kg of urea-Niha applied as a single dressing in the 1981-1983 period,
increased the crop yield and N uptake by plants in 1981 and 1983, but did not affect
them in 1982. When HQ was added at a rate of 5 kglha to 375 kg of urea-Niha applied
annually in three equally divided dressings in 1982 and 1983, its effect on crop yield
and N uptake was depressive in 1982 and stimulating in 1983. HQ did not affect the
K/(Ca + Mg) ratio in the ryegrass plants.
7.8.9. Effect ofPh osphorodiamides

The effect of phenylphosphorodiamidate (PPDA) on a ryegrass variety (Latium
multif!orum var. westerwoldicum) was studied by Heber et aT. (1979). Pot experiments
were carried out in 1976 and 1977. Two soils, a loamy sand and a sand (6 kg/pot). were
used. Three N sources: urea, urea+l% PPDA (on urea-N basis). and ammonium nitrate.
were compared. A total amount of 2.4 g N/pot divided into 3 equal parts was surface-
applied immediately after emergence of seedlings and a few days after the first and
second harvest (cut). Dry matter and total N contents in plants were determined after
each of the three cuts.
The results showed that in both soils and both years the hay yields and total N
contents increased. under the influence of the applied fertilizers, in the following order:
urea < urea+ 1% < NH4N0 3.
Values of the hay yield and total N content, recorded in the NH4N0 3 treatment, were
reached and even exceeded. to some extent, only in the case of hay from the first cut in
the urea+ 1% PPDA treatment. Utilization of N from urea, urea+ 1% PPDA, and
~N03 ranged between 45 and 50.60 and 61, and 71 and 78%, respectively.
Kampfe et aT. (1982a) studied the effect of PPDA on ryegrass in field experiments
carried out on a loamy soil under conditions of natural weather and simulated drought in
the 1977-1979 period.
304
The N fertilized used were: conditioned urea (prilled urea coated with mineral oil-
bitumen mixture), conditioned urea + 0.5 and 1% PPDA (on urea-N basis), and lime
ammonium nitrate. They were applied twice, at rates of 100 and 60 kg Nlha,
respectively. TIle plots were also fertilized with P and K (35 kg P and 100 kg K/ha). The
control plots received no fertilizers.
Drought was simulated by means of plastic tents (6 by 4.5 m) put up over the plots.
The simulated drought began after addition of both N rates and lasted 14 days. Dry
matter content of ryegrass hay and total N content in green matter of plants were
determined.
The data in Table 66 show that diminution of hay yield in urea treatment in
comparison with the lime an1monium nitrate treatment could be partially or completely
prevented by using urea+PPDA. The effects of the 0.5 and 1% PPDA rates were not
significantly different.
TABLE 66. Effect of nitrogen fertilizers on dry matter content of rye grass hay (mean values for
the 1977-1979 period)"
Dry matter content of ryegrass hay from cuts I + 2
Fertilizer
dvha 0/0
Unfertilized control 57.4 62
Lime ammonium nitrate 92.3 100
Co nditi on ed urea 87.6 95
Conditioned urea + 0.5% PPDA 93.7 102
Conditioned urea + 1% PPDA 90.7 98
LSD 5% 6.5 7
"Adapted from Karnpfeetal. (1982a).
The 14-day simulated drought after fertilization led to no significant changes in the
hay yields from cuts 1+2 as compared to that recorded under natural weather conditions.
Total N contents in green matter of plants were determined twice, namely at days 14
and 21 after application of each of the two N fertilizer rates. During the fust 14 days,
PPDA retarded N uptake by the plants. But at day 21 after N application, there were no
significant differences between the lime ammonium nitrate treatment and the urea + 0.5
and 1% PPDA treatments as concerns total N contents in green matter of plants. In the
1977-1979 period, neither were these contents influenced by simulated drought.
The experiments of Gorelik et at. (1983) for studying the effect of hydroquinone
(HQ) on orchardgrass and perennial ryegrass (see page 3(2) also comprised treatments
with PPDA used at a rate of 1% relative to weight of urea. Thus, in a heavy-textured
sierozem, the effect of urea-PPDA on crop yield (dry matter of aboveground parts of
orchardgrass) and on plant utilization ofurea-N did not differ from that of urea alone. In
the other two soils studied (loamy sandy podzol and sandy podzol), PPDA significantly
increased yields of orchardgrass and ryegrass, and the coefficient of plant utilization of
urea-N showed, under the influence ofPPDA, 4-13% increases. In comparison with HQ
also used at 1% rate, PPDA was more efficient.
Linke et at. (1983) synthesized the results of 69 field experiments on natural
grasslands in the 1974-1979 period. The same fertilizers were used, the same aspects
were studied, and the same methods were applied as in the cereal experiments (Karnpfe
et 01., 1982b) (see pages 271-272) and, essentially, similar results were obtained. The N
305
fertilizers were administered, for the first emergence of plants, at a rate of 60 kg N/ha
(in 1974 and 1975) or 100 kg N/ha (in the 1976-1979 period), and for the second
emergence at a rate of 60 kg N/ha (in each year). At the beginning of growing season in
each year, 30 kg P/ha and 100 kg Klha were also administered. The plots, from which
plants were collected for determination of hay yield (dry matter) and total N content,
were of 15 m2 •
Figure 79 presents the results concerning the influence of the amount of rainfall
during the first 5 days after fertilization on the hay yield. In the drought period, the hay
yield was much lower in the urea treatment than in that with lime ammonium nitrate.
PPDA improved, to some extent, the efficiency of urea. Between some limits of the
rainfall amounts, all urea fertilizers were more efficient than lime ammonium nitrate. In
comparison with the results obtained in the cereal experiments (see Figure 77), the 0.5%
PPDA rate had a weaker effect, as the yield recorded in the urea+0.5% PPDA treatment
was nearly the same as in the urea-only treatment. PPDA increased evidently the
/-- ......,
2.5
/ '
I "
I \.
I ,
/ \.
I .~ ,
(ij'
S
10
/ .~\
~ / ~.
j 0.0 ----r-
E :::.:---",.."
~
0 -1.0
'- a
o
-2.0
i i i
o 5 10 15 20 25 30
Rainfalls (mm)
Figure 79. Influence of rainfalls on the difference between hay yield (dry matter) recorded in the
treatment with lime ammonium nitrate and hay yields obtained in the treatments with urea with or
without 0.5 or 1% PPDA addition.
a - Conditioned urea. b - Conditioned urea + 0.5% PPDA. c - Conditioned urea + 1% PPDA.
/From Linke et al. (1983)./
efficiency of urea when its rate was I %. However, even the effect of this PPDA amount
disappeared at rainfalls ~ 27 IllIll. whereas in the cereal experiments the effect persisted
at rainfalls of about 33 mm.
The yield-increasing effect of 1% PPDA addition to urea fertilizer applied in these
field experiments on natural grasslands was also pointed out by Linke and Kampfe
(1987) in their communication presented at the International Fair Symposium held in
Leipzig.
In the field experiment carried out on a grass ley in 1984 and 1985 (Rodgers et aI.,
1987; see page 125), the dry grass yields from three cuts (t/ha) in the urea and
urea+PPDA treatments were 7.96 and 8.97, respectively (single application of 375 kg
306
urea-Nlha), and 8.50 and 8.58, respectively (divided applications of urea: 3 x 125 kg
Nlha), in 1984. In 1985 the corresponding values were 9.11 and 11.36, and 12.46 and
12.32, respectively. In both years, the yield-increasing effect of PPDA was great only in
the single application of urea. Similar results were obtained concerning N uptake by
plants.
Joo and Christians (1986) conducted experiments in 1984 at the Iowa State
University Turfgrass Research area north of Ames. The treatments of plots (2.32 m 2)
installed on blueglass turf on a fine loamy soil (PH 7.5) included urea solution (at 0,49,
and 98 kg Nlha) amended or not amended with 1 and 2% PPDA and 25% Mg2+ as
MgCb.6H20 (all percentages mean weights relative to weight ofurea-N). The fertilizer
solutions were surface-applied monthly in June through September. Clipping yields
were determined weekly for 5 weeks after each treatment.
The average fresh weight of clippings over a 5-week period in plots treated with 2%
PPDA increased 28% at the lower N rate in August, 31 % at the lower N rate and 20% at
the higher N rate in September. On overall average, 2% PPDA significantly (p<O.OI)
increased the clipping yields, whereas the effect of 1% PPDA was not significant
(p>0.05). The MgCl 2 treatment showed a 13 to 25% increase of the fresh clipping yield
at the lower N rate in August and September but a 20% decrease at the higher N rate in
August.
PPDA reduced foliar bum significantly at the lower N rate in September, but the
decreasing effect was not consistent in each month. In the heat of July, the MgCb
treatment resulted in greater foliar bum at the higher N rate, but it reduced bum at both
N rates under the relatively wet conditions and cool temperatures of September.
7.8.10. EfJect of Phosphoric Triamide (PTA) and Thiophosphoric Triamide (TPTA)

Compounds
In some studies, the effect of PTA and TPTA compounds on grasses was compared with
that of other inhibitors.
In the field experiment conducted by Joo et al. (1987) and briefly described on page
146, bluegrass turf was fertilized monthly, in the period of June-September 1986, with
liquid urea (49 kg Nlha) with or without addition of 0.5, 1 or 2% N-(n-
butyl)thiophosphoric triamide (nBTPTA) (on urea-N basis). Clipping weights were
collected weekly for 4 weeks after each fertilizer treatment at a 5-cm mowing height on
1.5-m2 areas.
The results presented in Figure 80 show that clipping yield varied with month of
application, but it was always significantly higher in plots treated with urea + nBTPTA
than in those treated with urea only. At the same time, no significant differences were
recorded between effects of the three nBTPTA rates.
Joo et at. (1991a) summarized the results of investigations carried out in 1985 and
1986. Numerical data on clipping yields are presented for the following experimental
conditions: liquid urea was applied on the bluegrass turf at a rate of 49 kg Nlha; the
amendments and their rates relative to urea-N (weight/weight) were: PPDA 1, 2, and
3%; nBTPTA (applied only in 1986) 0.5, 1, and 2%; ammonium thiosulfate (ATS), K+
(as KCl), and Mg2+ (as MgCb) 5, 15, and 25%. Only urea was added to the control. On
average, the following clipping yields (g of fresh weight/1.5 m 2) were registered: 66
(control), 63 (PPDA), 65.7 (ATS), 68 (Kj, and 71.3 (Mg2+) in 1985, and 70 (control),
307
_ eam..1 c::z:a nBTPTA 0.5%

c::J nBTPTA 1% IB28BnBTPTA 2%
'.0<
Figure 80. Effect of nBTPTA on clipping yield of bluegrass in plots fertilized with liquid urea. /From
Joo et al. (1987),/
79 (PPDA), 80.7 (nBTPTA), 75.7 (ATS), 73 (KJ, and 72.3 (MiJ in 1986. In 1985 the
differences among amendments were small due to a very dry period in early summer
that restricted growth. In 1986 nBTPTA, PPDA, and ATS significantly increased the
yield, but ATS was less efficient than nBTPTA and PPDA. K+ and Mi+ had no
significant effect on the yield.
The investigations on the effect of urease inhibitors on the bluegrass turf were
continued (Joo et al., 1991 b, 1992, 1993).
Joo et al. (1991b, 1993) described the investigations carried out in 1987. The 2.32-
m 2 plots were treated with liquid urea (49 kg N/ha) amended with nBTPTA at rates of 0
(urea alone), 0.25, and 0.5% of the weight of urea-No Urea labeled with 5% 1~ was
applied at the center of each plot to an area measuring 0.2 m 2 • The rest of the plot
received the same rate of urea-N minus the 15N label. Clippings were collected weekly
at a 5-cm mowing height for 5 weeks; then shoots, thatch, and soil (0-7.5- and 7.5-
15-cm layers) were sampled and analyzed for total N and 15N.
During the 5-week experimental period, the N recovered in the five clippings
represented 7.5% of the added urea-N in the urea-only treatment and 7.9 and 8.1% in
the treatments with urea+0.25% nBTPTA and urea+0.5% nBTPTA, respectively. Total
N recovery (i.e.. recovery in clippings, shoots, thatch, and soil) was 28.8% (urea), 45%
(urea+0.25% nBTPTA), and 36.1% (urea+0.5% nBTPTA). Thus, N recovery in
clippings was highest with 0.5% nBTPTA and total N recovery with 0.25% nBTPTA.
The field experiments carried out in 1988 and 1989 were described by Joo et al.
(1992). Granular (and not liquid) urea was applied to plots (1.92 by 1.92 m) at rates of
0,49, and 73.5 kg N/ha (in 1988) and at rates of 0,24.5, and 49 kg N/ha (in 1989). In
these years, the averaged dried clipping weights of bluegrass were not significantly
different in the urea-only and urea+nBTPTA treatments, irrespective of the rates of urea
and nBTPTA. These findings are in contrast to the results of laboratory experiments of
Joo et al. (1992), as in these experiments nBTPTA significantly decreased loss of urea-
N through ammonia volatilization (see page 210). It was assumed that lack of yield
response to nBTPTA was due to other loss mechanisms and immobilization of urea-N
in the soil-bluegrass plant system.
308
The orchardgrass pasture experiments conducted by Bundy and Oberle (1988) are
referred to on page 201. In these experiments, urea (67.2 kg N/ha) with or without
urease inhibitors (2%) was surface-applied at Lancaster in 1983 and/or 1984, and,
besides ammonia volatilization, the orchardgrass dry matter yield and N uptake were
also assessed. In 1983 the significant reductions in NH3 losses due to addition of PPDA,
diethyl-PDA, and trichloroethyl-PDA to urea prills resulted in significantly higher dry
matter yields. However, only the urea-PPDA treatment showed significantly higher N
uptake than urea. In 1984 nBTPTA, which was more effective than PPDA in reducing
NH3 losses from urea, increased significantly (at p=0.1 level, but not at p=O.05 level)
the yield and N uptake. The effect of PPDA on yield and N uptake was insignificant
even at the p=O.llevel.
For comparing the effects of nBTPTA, PPDA, and hydroquinone (HQ) on perennial
ryegrass, Li et af. (1993) carried out a greenhouse experiment. Air-dried soil samples (1
kg) were placed in plastic pots (with a height of 12 cm and a diameter of 15 cm). The
soil moisture content was brought up to 20% on weight basis and kept constant by a
spray of water each day. Ryegrass seeds (200) were broadcast evenly in each pot and
covered with a thin layer of soil. Two days after germination, 100 mg urea-N labeled
with 15N, amended or not amended with urease inhibitors in solution was added to each
pot (rate of inhibitors is not mentioned in the paper). At days 15 and 30 after
fertilization, the grass shoots were harvested and their dry weight, total N and 15N
contents determined. The roots and soil were also analyzed for total N and 15N after
both harvests.
The results indicated that the dry matter weights of grass shoots were not
significantly different in the urea-only and urea+nBTPTA or urea+PPDA treatments at
any harvest. In contrast, the yields obtained at the first harvest were insignificantly
higher and those registered at the second harvest were significantly lower in the
urea+HQ than in the urea-only treatment. During the first 15 days of growth, the plant
uptake of N from urea was not significantly different among the treatments, but during
the second 15-day period of growth the urea+nBTPTA treatment, in comparison with
the other treatments, induced a significant (10%) increase in plant uptake ofurea-N.
In the field experiment of Watson et at. (1990a,b) (see page 150), the ryegrass plants
were harvested approximately 7 weeks after surface application of urea, urea+0.5%
nBTPTA, and calcium ammonium nitrate (CAN) at a rate of 100 kg N/ha. Herbage dry
matter yield of ryegrass (t/ha) was 0.644 (in unfertilized plots), 2.991 (in urea plots),
3.255 (in urea+0.5% nBTPTA plots), and 3.278 (in CAN plots). It is evident that the
yield performance ofurea+0.5% nBTPTA was comparable to that of CAN.
Watson et af. (1994a) studied the effect of different levels of nBTPTA in urea
granules on the herbage dry matter yield of ryegrass. A field experiment was carried out
in parallel with that for studying ammonia volatilization (see page 150). But the plots
were a little smaller (1.5 by 1.5 m), and unfertilized and CAN-fertilized plots were also
used. The ryegrass plants were harvested after 6 weeks of regrowth in plots installed for
each of the time periods I to 5. The dry matter yield (t/ha) in the differently treated plots
had the following average values: 2.02 (unfertilized), 4.57 (urea), 4.75-4.98
(urea+nBTPTA), and 4.82 (CAN). In the urea+nBTPTA-treated plots, the highest
yields: 4.98 and 4.87 t/ha were recorded at the 0.05 and 0.01% nBTPTA levels,
respectively, and the lowest ones: 4.75 and 4.77 tlha at the 0.1 and 0.5% nBTPTA
309
levels, respectively. In other words, the lower levels of nBTPTA affected more
positively the dry matter yield than did its higher levels.
As the growing season progressed, the yield decreased from 7.47 tlha (in time period
1) to 1.93 tlha (in time period 5).
In another experiment carried out in parallel with that mentioned above, 15N-labeled
urea granules (at approximately 18 atom% excess 15N) with or without nBTPTA (at five
levels) were applied to microplots (50 by 50 cm). Total N and 1~ in soil and plant were
determined and the 15N recovery was calculated.
nBTPTA increased inSignificantly the 15N recovery in soil at all depths down to 15
cm, but caused a significant increase in percent N derived from urea in the shoot and the
shoot recovery of 15N. Total 15N recovery in the soil-plant system was increased by up
to 17% by incorporating nBTPTA into urea granules. All these increasing effects of
nBTPTA were similar at its five levels.
In these experiments, the contents of macronutrients (P,K, Na, Ca, Mg, S) in dry
matter were not significantly different between the N treatments, and there were no
visual signs of phytotoxicity (leaf tip scorch) on any treatment.
A 3-year study was initiated by Joost (1995) in 1992 to compare the effectiveness of
ammonium nitrate, urea, and urea+O.l4% nBTPTA applied to caucasian bluestem in
Missouri. Rates offertilizers were 0, 78, 156, and 234 kg NlhaJyear, as either a single or
in two equal split applications. Forage dry matter yield and total N uptake of plants
were significantly lower in response to urea than either NH4N0 3 or urea+nBTPTA.
Applying half of the total N in the spring and half after the first harvest resulted in
significantly higher July dry matter yield and crude protein content in 1992 and 1993,
but regrowth was insufficient to produce a July harvest in 1994 due to drought
conditions.
Lamond et al. (1996) conducted field experiments on established bromegrass.
Nitrogen fertilizers (ammonium nitrate, urea, urea+nBTPTA, urea-ammonium nitrate
solution) were applied at rates of 0, 50, and 100 kg Nlha in mid to late March.
Ammonium nitrate and urea+nBTPTA consistently produced higher yield and often
higher forage protein levels than urea or urea-ammonium nitrate.
Watson and Miller (1996) studied the short -term plant physiological effects not only
of nBTPTA-amended urea but also of nBTPTA used without urea. The test plant was
perennial ryegrass. Four experiments were carried out.
In Experiment 1, field-moist samples (380 g at 21 % H2 0, weight/weight) of a clay
loam soil (pH 5.7), collected from under a permanent grassland sward at Baillies Mills,
County Down, Northern Ireland, were placed in pots (10 cm diameter, 9 cm depth). A
solution containing 20 mg N (as NH4 N0 3 ), 10.3 mg P (as KH 2 P0 4 ), and 39.3 mg K (13
mg K as KH 2 P0 4 and 26.3 mg K as K2 S04 ) was added to the soil in each pot; then four
I-week-old ryegrass seedlings were planted into the soil. The soil was maintained at
80% field capacity with distilled water. The plants grew under natural lighting
conditions in a greenhouse. After 6 weeks of growth, the soil was again fertilized as
before the planting. After 4 weeks, only 20 mg of NH4 N0 3 was added, and after a
further 4-week period, 10.3 mg P, 39.2 mg K, and 15N-labeled urea granules (the 15N
enrichment being 18 atom% excess 15N) at a rate of 78.5 mg N/pot (equivalent to 100
kg Nlha) were uniformly applied to the soil surface. The urea granules contained no or
0.01,0.1 or 0.5% nBTPTA (weight/weight). At intervals of 0.75, 1.75,4,7, and 10 days
after urea fertilization, the content of each pot was separated into three fractions: shoot,
310
root, and soil. The analyses to which the three factions were submitted and some of the
results obtained are briefly summarized below.
Urease activity in shoot significantly (p<0.05) decreased in the urea treatment over
the 10-day period. In the urea+nBTPTA treatments, shoot urease activity was strongly
reduced at day 0.75 compared with the urea treatment. Thus, during 0.75 days in the
urea+0.5% nBTPTA treatment, shoot urease activity was reduced by 90.2% compared
with 0.15% in the urea treatment. The time taken for shoot urease activity in the
urea+0.01, 0.1, and 0.5% nBTPTA treatments to return to that in the urea treatment was
4,7, and 10 days, respectively. Urease activity, which was higher in root than in shoot,
was not affected by nBTPTA.
15N recovery of urea in the shoot at days 7 and 10 was significantly lower (P<O.Ol)
than that ofurea+nBTPTA. 15N recovery in root was similar to that in shoot. At day 10
total 15N recovery (shoot + root + soil) of urea was significantly lower (P<0.05) than
that ofurea+nBTPTA.
After l.75 days, the residual urea-N in soil was 1.3 and 62% of the N applied in the
urea and urea+0.5% nBTPTA treatments, respectively. However, the effect of nBTPTA
was short-lived. Thus, after 4 days, less than 1% ofN applied was as urea with 0.01 and
0.1 % nBTPTA and only 6.8% with 0.5% nBTPTA. Urea and mineral N levels in the
shoot and root were very low or negligible during the whole 10-day period.
There was no significant difference in soluble protein and water-soluble
carbohydrate contents in shoot and root between urea and urea+nBTPTA. However,
nBTPT A produced significant changes in composition of free amino acids in shoot and
root, suggesting a difference in metabolism.
Experiment 2 was a field experiment which ran concurrently with Experiment 1.
The site was an established perennial ryegrass-white clover sward on a clay loam soil
(pH 6.3) at the Agricultural Research Institute, Hillsborough. The plots (1.5 by 1 m)
received 100 kg N/ha as unlabeled urea, urea+O.1 or 0.5% nBTPTA or calcium
ammonium nitrate as well as P and K fertilizers. The plots were harvested at intervals of
2, 4, 8, 14, and 30 days after N fertilization. The same analyses were made with
generally similar results as in Experiment 1.
However, in the field experiment, contrary to the pot experiment, substantial levels
of urea were measured in the shoot at days 2 and 4 following application of urea+O.l
and 0.5% nBTPTA.
In Experiment 3, nBTPTA was applied alone in a pot experiment similar to
Experiment 1. After 4-week growth of ryegrass seedlings, the soil in pot was treated
with 40 rn1 water or 40 ml aqueous solution containing 0.055, 0.55 or 2.77 !1g
nBTPTA/g oven-dry soil. Urease activity was determined in the three fractions (shoot,
root, and soil) at days 0.75,1.75,4,7, and 10 after nBTPTA application.
Urease activity in each fraction was significantly reduced by all concentrations of
nBTPTA compared to control (no nBTPTA) at day 0.75 and appeared recovered to that
of control by day 10.
In Experiment 4, the effect of urea and urea+nBTPTA on leaf tip necrosis was
studied in a pot experiment similar to Experiment 1. After 4-week growth of ryegrass
seedlings, the soil in pots was N-fertilized at rates of 0,23.9,47.8, and 95.6 mg N/pot as
urea amended with nBTPTA at 0.1 or 0.5% (weight/weight) applied in 15 rn1 solution.
Leaf tip necrosis (scorch) on living leaves was measured 15 days after fertilizer
application.
311
Leaf tip necrosis increased with increasing urea application rate and nBTPTA
concentration. Thus, increasing the urea-N applied from 23.9 to 95.6 mg N/pot
(equivalent to an increase from 50 to 200 kg Nlha), in the absence of nBTPTA, resulted
in a significant increase in the average tip damage per leaf from 0.92 to 3.04 mm.
Increasing the concentration of nBTPTA to 0.5% led to markedly increased leaf tip
necrosis, particularly at the high urea-N application rate. New developing leaves
showed no visual sign of tip necrosis.
The effect of nBTPTA-amended urea to cause some leaf tip necrosis, like its effect
to inhibit shoot urease activity, was transient and short-lived. Therefore, the conclusion
was drawn that the benefit of nBTPTA in reducing ammonia volatilization from urea
and, thus, in increasing dry matter production of ryegrass would appear to far outweigh
any of the observed negative, short-term effects.
In the 3-year field experiment conducted by Watson et af. (1998) (see page 152),
approximately 6 weeks after each fertilizer application, the central part of each plot was
harvested, and the herbage dry matter yield and N uptake by perennial ryegrass were
determined.
The results showed that repeated application of nBTPTA on the same soil exerted no
adverse effect on grass production.
7.8.11. Effect ofLignin

Garcia et af. (1997, 1998) coated urea granules with Kraft pine lignin and a resin
mixture. Granules with three coatings (lignin, resins, and linseed oil) were also
prepared. For testing their efficiency, a greenhouse experiment was carried out. Samples
of a sandy loam soil (PH 7.8) from the province of Toledo (Spain) were used. Two-kg
soil samples in pots were fertilized with uncoated or coated urea (300 mg N/pot),
superphosphate (60 mg P/pot), and K2 S04 (40 mg Klpot), moistened and sown with 4 g
of perennial ryegrass seeds. The plants were harvested (cut) three times at 25-day
intervals, and their dry matter weights and total N contents were determined.
The cumulative dry matter yields were higher with coated than with uncoated urea.
The urea coated with lignin+resins+linseed oil performed better than the urea coated
without linseed oil. Determination of total N contents showed that plant uptake of N
was highest from urea granules coated with lignin, resins, and linseed oil.
7.8.12. EfJectofPlant Materials

Rajeswara Rao and Chand (1996) conducted a field experiment during 199111992 at
Hyderabad on a light-textured red sandy loam soil (PH 7.8) to evaluate the effect of
neem cake-coated urea (NCCU) and neem oil-coated urea (NOCU) in comparison with
prilled urea (PU) on the yield and N use efficiency of lemongrass (Cymbopogon
flexuosus var. flexuosus) , an important medicinal and aromatic plant. The main
components of its essential oil are citral and geraniol. The three N fertilizers were
applied at rates of 0, 60, and 120 kg Nlhalyear.
Slips of lemongrass were planted at a spacing of 60 cm x 30 cm in June 1991. The
basal fertilizers were single superphosphate (17 kg Plha) and KCI (33 kg K/ha). The N
fertilizers, in three equal splits with one split dose for each harvest, were placed 5 cm
below the soil surface in holes. The plants were harvested in winter (December 1991),
summer (April 1992), and rainy (August 1992) seasons. Biomass yields and essential oil
yields were determined.
312
Total yields of the three harvests in the PU, NCCU, and NOCU treatments with the
60 and 120 kg Nlha/year had the following average values for biomass yields (tlha):
44.9 (PU), 58.4 (NCCU), and 58.6 (NOCU), and for essential oil yields (llha): 305.3
(PU), 413.1 (NCCU), and 407.3 (NOCU). Both biomass and essential oil yields and N
use efficiency of lemongrass were significantly (p<0.05) higher with coated ureas than
with uncoated urea, while the difference between the effects of the two coated ureas was
not significant. The contents of citral and geraniol in essential oil was not influenced by
rate and nature of N fertilizers.
7.8.13. Effect of Combined Urease Inhibitors

Calancea (1979) treated samples of a chemozemic soil with 0 or 0.5 g of urea + 0, 5, 15
or 45 mg of hydroquinone {HQ)/pot. Ryegrass yield, taken as 100% in the untreated
control, increased significantly (to 175.97%) in the treatment with urea alone. The
yields, relative to the treatment with urea alone (100%), increased in the urea + HQ
treatments. The increase was insignificant at 5 mg of HQ/pot (100.11 %) and very
significant at 15 and 45 mg of HQ/pot (112.38 and 114.48%, respectively). It was also
found that the yields decreased, but not significantly, when the soil in pots received 0.5
g of urea + 5 mg of HQ from an old preparation or 0.5 g of urea + 5 mg of thiourea or
0.5 g of urea + 5 mg of thiourea + 2.5 mg of triazine. The decrease became very
significant when the soil was treated with 0.5 g of urea + 5 mg of HQ + 5 mg of
thiourea + 5 mg of diethyldithiocarbamate. Another observation was that the inhibitors
caused only minor changes in the amino acid composition of total proteins in ryegrass.
7.8.14. Effect of Combined Urease and Nitrification Inhibitors

Calancea et al. (1977) studied the effect of HQ, p-benzoquinone (BQ) and two
nitrification inhibitors (nitrapyrin and sulfathiazole) on ryegrass plants growing in pots,
each containing 1 kg of chemozemic soil mixed with I kg of sand.
TABLE 67. Effect of bydroquinone and nitrapyrin on ryegrass yield and coefficient of N utilization
from urea"
Average Total N Coefficient ofN Difference
Treatment" herbage yield uptake utilization from urea
~~~t2 ~m~~t) ~%2 ~%2
No (control) 5.64 98.7
Urea 9.03 157.4 37.2
Hydroquinone (HQ) 6.56 114.2
Urea + HQ 9.76 180.3 50.7 +13.5
Urea + nitrapyrin (NP) 9.77 184.9 45.9 + 8.7
Urea + HQ+NP 10.05 189.0 51.4 +14.2
"From Calancea el ai. (1977); see also Hera and Eliade (1981).
b Rates of additions per pot were: urea 100 mg N; HQ 2 mg; nitrapyrin 2 mg.
Table 67 shows that HQ and nitrapyrin, applied separately at a rate of 2% relative to

urea-N, increased ryegrass yield (dry matter of herbage) to a similar extent, but HQ was
more efficient than nitrapyrin in increasing the coefficient of N utilization from urea (by
13.5 and 8.7%, respectively). The highest yield and the highest (14.2%) increase in
313
coefficient of N utilization from urea were registered when HQ and nitrapyrin were
applied together.
For studying the effect of BQ, the soil in pots was treated with 1 g of urea + or
0.01,0.1 or 1% BQ (relative to urea-N) or 0.1% BQ + 0.1% sulfathiazole. The urea +
°
0.01 % BQ treatment led to increased yield, N uptake by plants and coefficient of N
utilization from urea, but BQ at higher rates and, especially BQ + sulfathiazole, proved
to be phytotoxic.
The urea+DCD+ATS fertilizer patented by Sutton et al. (1991) (see page 244) was
applied on turf at rates of 40,80, 120,and 160 kg ofurea-Nlha. For comparison, urea (at
the same four rates) as well as ammonium nitrate, commercial fertilizers Sulfur Kote
and Osmocote (at 80 kg Nlha) were used. Color ratings were estimated for 78 days after
fertilizer application, and it was found that the patented fertilizer provided a greener turf
over a longer period of time than did the other fertilizers.
7.9. EFFECT OF UREASE INHIBITORS ON LEGUMINOUS PLANTS
The legume species studied include alfalfa (Medicago sativa), broadbean (Vicia [aba),
common bean (Phaseolus vulgaris), pea (Pisum sativum), red clover (Trifolium
pratense), soybean (Glycine max), and yellow lupine (Lupinus luteus).
7.9.1. E.fJect of Heavy Metal Compounds

In Kucharski and Niklewska's (1992) pot experiment, in which samples ofa brown soil
were treated with 0, 10, 100, and 1,000 ppm of Zn (as sulfate), soil urease activity was
inhibited only by the 1,000 ppm Zn rate (see page 16), but the dry mass of overground
parts and roots of broadbeans and N uptake by plants were reduced by the 100 and
1,000 ppm Zn rates, whereas dry mass of root nodules was reduced by all Zn rates.
According to Kucharski (1997), Zn, Pb, and Cd inhibited soil urease activity and
reduced the yield of yellow lupine (see Table 4).
7.9.2. Effect ofFluorides

inhibited in soil samples treated with NaF at rates ofO.1-3 g F/kg soil (see page 34), but
root growth of pea plantlets was strongly reduced at rates ~ 0.5 g F/kg soil.
7.9.3. Effect o[ Organic Mercury Compounds

For studying the effect of phenylmercuric acetate (PMA) on the germination of alfalfa
seeds, May and Douglas (1975, 1978) used the same two techniques also applied for
germination of wheat seeds (see page 264). According to the first technique, 100 alfalfa
seeds were placed on moistened paper towelling in Petri dishes and treated with 0,
0.003,0.015 or 0.03 g of powdered PMA/dish. The germination took place at 20D C and
lasted 8 days. The results showed that PMA, at each concentration, significantly
inhibited the germination. In the second technique, germination of 100 alfalfa seeds
took place in samples of two Australian soils (a grey clay and a solonized brown soil).
The samples received PMA at rates of 0, 10, 50, and 100 ppm relative to soil weight.
Germination was carried out under the same conditions as in the first technique. It was
found in both soils that germination was not inhibited by any PMA concentration.
314
The effect of PMA on germination of alfalfa seeds was also studied by Bremner and
Krogmeier (1990) (see page 277 and Table 61).
7.9.4. Effect of Thiuram Disu/jides

In the pot experiments of Borchaninova and Filippova (1978) (see page 56), tetra-
methylthiuram disulfide exerted a negative effect on the growth of the aboveground
biomass of red clover plants and reduced the weight of their roots and number of root
nodules.
7.9.5. Effect of Hydroxamic Acids

The experiments performed by May and Douglas (1975, 1978) and referred to in
Section 7.9.3 were also carried out with benzohydroxamic acid (BHA). It was found
that germination of alfalfa seeds on paper towelling was completely hindered by the
higher concentrations of BHA, but germination in the soils was not inhibited by any
BHA concentration.
7.9.6. Effect of Polyhydric Phenols and Quinones

In experiments similar to those described in Section 7.9.3, May and Douglas (1975,
1978) evaluated the effect of catechol, hydroquinone, p-benzoquinone, 2,5-dimethyl-p-
benzoquinone, and o-naphthoquinone on germination of alfalfa seeds.
At rotes of 0.003 and 0.Q15 gldish, inhibition of germination was not always
significant, but at the rote of 0.03 gldish, all these compounds significantly inhibited the
germination. All rotes of2,5-dimethyl-p-benzoquinone were strongly inhibitory.
Germination in soil was influenced to a lesser extent. Thus, at 10 ppm none ofthese
compounds inhibited significantly the germination in any of the two soils studied.
Significant inhibitions in germination were caused by 2,5-dimethyl-p-benzoquinone at
100 ppm in both soils and by hydroquinone at 100 ppm in the grey clay soil.
It was demonstrated by Arziani et al. (1985) that the exogenous 14C-Iabeled
hydroquinone taken up by pea and common bean seedlings in axenic cultures was
inactivated (detoxified) in the plant tissues mainly by glycosylation and, to a lesser
extent, by conjugation with peptides.
Bremner and Krogmeier (1990) studied the effect of catechol and 2,5-dichloro-l,4-
benzoquinone on germination of alfalfa seeds (see page 277 and Table 61).
Xu et al. (1994) reported on field experiments carried out in 1988, 1989, and 1990
for studying the effect of hydroquinone (HQ) on the yield of soybean. The plots were
treated usually with 1% HQ relative to weight of urea-No In each year, the yield
increased due to HQ. For example, in 1990 the yields obtained (kglha) in the differently
treated experimental plots were the following: 3,829.5 (control: no urea and no HQ);
4,105.5 (urea only, 3.75 g N/mz); 4,177.5 (urea + 37.5 mg HQ/mz), and 4,695.0 (urea +
75 mg HQ/mz). In pot experiments, in which the HQ rote was I, 2 or 3% relative to
urea-N, HQ increased the accumulation of plant biomass and the Nz-fixing activity by
enhancing the nodulation, and led to higher N contents in both stem and seed of
soybean.
7.9.7. Effect ofPhosphorodiamides

Krogmeier et al. (1989b) proved that the leaf tip necrosis commonly observed after
foliar fertilization of soybean plants with urea a) is due to accumulation of toxic
315
amounts of urea rather than formation of toxic amounts of ammonia in leaves and b) is
accordingly increased rather than decreased by addition of a urease inhibitor such as
phenylphosphorodiamidate (PPDA) to the urea fertilizer applied. It was also proved by
Krogmeier et al. (199Ia,b) that the leaves of soybean plants grown on media deficient
in Ni (which is an essential component of the urease molecule) have a lower urease
activity and, consequently, these plants are more susceptible to leaf tip necrosis when
they are foliar-fertilized with urea. Their susceptibility would be even higher after foliar
application of urea with a urease inhibitor.
Nevertheless, the potential of urease inhibitors for inducing phytotoxicity should not
preclude their use as additives to urea fertilizers, because the ammonia produced
through hydrolysis of urea by soil urease is much more detrimental to plant growth than
is urea accumulation induced by them (Krogmeier et al., 1989a; see also page 276).
The effect ofphosphorodiamidic acid, PPDA, and 4-chloro-PPDA on germination of
alfalfa seeds was studied by Bremner and Krogmeier (1990) (see page 277 and Table
61).

Compounds
Alfalfa was one of the test plants on which Bremner and Krogmeier (1988) studied the
effect of 10 inhibitors, including four PTAs and one TPTA (see page 275), and on
which Bremner and Krogmeier (1990) studied the effect of 23 urease inhibitors,
including 10 PTAs and two TPTAs (see page 277 and Table 61).
7.9.9. Effect of Cyclotriphosphazatriene (CTPAT) Derivatives

Two CTPAT derivatives were studied by Bremner and Krogmeier (1990) for evaluation
of their effect on germination of alfalfa seeds (see page 277 and Table 61).
7.10. EFFECT OF UREASE INHIBITORS ON OTHER PLANTS
Many plants were studied: cotton (Gossypium hirsutum), cucumber (Cucumis sativus),
geranium (Pelargonium graveolens), Japanese mint (Mentha arvensis), lettuce (Lactuca
sativa), oilseed-rape (Brassica napus), onion (Allium cepa), potato (Solanum
tuberosum), pumpkin (Cucurbita pepo), radish (Raphanus sativus), sugarbeet (Beta
vulgaris cv. altissima), sugarcane (Saccharum ojJicinarum), sweet potato (Ipomoea
batatas), tobacco (Nicotiana tabacum).
7.10.1. Effect ofHeavy Metal Compounds

In the sugarbeet field experiment of Bayrakli and Gezgin (1996), urea at a rate of 200
kg N/ha was applied alone or with pyrite at I and 2 tlha rates (see Section 4.9.l.2).
Refined sugar yield (t/ha) was 7.61 (urea alone), 8.63 (urea + pyrite at I tlha), and 8.l4
(urea + pyrite at 2 tlha). However, the yield-increasing effect of pyrite was not
significant at p=0.05.

Application of urea (200 kg N/ha) alone or with 540 and 1,080 kg KCl/ha on sugarbeet
fields (Bayrakli and Gezgin, 1996; see Section 4.9.1.2) led to insignificant changes (at
316
p=0.05) in refined sugar yields (t/ha), namely to changes from 7.61 (urea alone) to 8.14
(urea + KCl at the lower rate) and to 7.38 (urea + KCl at the higher rate).
Hallmark et al. (1996, 1998) carried out investigations to determine whether the
product N-HIB Ca (composed of 12% Ca as CaClz, 1.5% Mg as MgClz, and a urease
inhibitor) can increase N use efficiency and sugarcane yields. The results showed that
adding 45 kg Calha as liquid N-HIB Ca to 135 kg N/ha of liquid urea increased sugar
yields by 2.95 tlha across four years (1991-1994). This compares with a 2.25 tlha sugar
response from increasing N from 135 to 200 kg N/ha across four years where calcium
was not added.
7. J O. 3. Effect of Fluorides
root growth of cucumber and pumpkin plantlets was strongly reduced at rates ~ 1.5 g
F/kg soil.
7. J 0.4. Effect of Urea Derivatives

In the pot experiments described by Kolyada (1970) (see page 49), using 10 mg
thiourea-N + 90 mg NI-4NOr N instead of 100 mg NH4N0 3-N led to a 17% increase in
the onion yield.
Thiourea, which in the pot experiments described by Kolyada (1973) inhibited soil
urease activity when applied at the rate of 0.1 g thiourea-N/kg soil (see page 49), was
toxic to the plants, especially at the beginning of the growing season. The yield of
radish was highest in the treatment with 10 mg thiourea-N + 90 mg NI-4N0 3-N + 100
mg P/kg soil + 100 mg K/kg soil; it exceeded by 17% the yield registered in the
treatment with NH4NOr N + PK (in which the rate of N, P, and K addition was
identical: 100 mg/kg soil).
7. J0.5. Effect ofDithiocarbamates

It is specified in the description of Hyson's (1963) patent (see page 52) that, when the
dithiocarbamate-amended urea fertilizer was applied by broadcasting to areas of tobacco
cropland at a rate of 28-560 kg/ha, outstanding beneficial results were obtained.
7. J o. 6. Effect of Heterocyclic Sulfur Compounds

In the field experiment ofTu et al. (1995), treatment of soil with dazomet, followed by
fertilization and planting of tobacco seedlings, led to a decrease, then to an increase, of
urease activity during the growth period (see page 72) and to a significant increase of
tobacco yield. The yield increase was attributed mainly to the nematicide effect of
dazomet.
7. J0.7. Effect of Polyhydric Phenols and Quinones

Rodgers and Pruden (1984) and Rodgers et at. (1986) studied the effect of
hydroquinone (HQ) on the efficiency of urea under field conditions, the test plant being
oil-seed rape. Some details concerning these experiments, performed at Rothamsted, are
presented on page 94.
In these experiments, carried out in 1983-1984 and 1984-1985, HQ enhanced the
efficiency of urea: significantly increased the rape seed, oil, and protein yields in all
317
experimental plots. For example, in the 1984-1985 experiment, in plots fertilized with
50 kg ofurea-N + 0 or 3 kg ofHQlha applied to seedbed and with 75 + 75 kg ofurea-N
+ 0 or 5 + 5 kg of HQlha given as spring dressings, the following yield data were
recorded: seed yield (tlha at 90% dry matter): 2.40 (urea alone) and 2.72 (urea + HQ);
oil yield (tlha): 0.83 and 1.01; protein yield (tlha): 0.45 and 0.52, respectively.
Arziani et al. (1985) proved that the axenic cultures of pumpkin seedlings take up
the exogenous 14C_HQ and inactivate it by glycosylation and conjugation with peptides,
glycosylation being the principal inactivating reaction.
Sweet potato fields were amended by Xue et at. (1991) with 10 kg of urea + 5 kg of
KCl + 0, 50, 100 or 150 g of HQ or quinhydrone (QH) per mu (1/15 ha). The yield
increases were 17.6, 25.3, and 31.3%, and 17.3,31.2, and 40.2% in the treatments with
the three increasing rates ofHQ and QH, respectively.
7.10.8. Effect ofPhosphorodiamides

Kampfe et al. (1981) studied the effect of three N fertilizers: urea, urea +
phenylphosphorodiamidate (PPDA) (1 % on urea-N basis), and lime ammonium nitrate
as well as that of the herbicide betanil 70 on the emergence of sugarbeet glomerules,
under field conditions, on I_m2 microplots installed on a slightly loamed sand soil (in
1976-1978) and on a black loess soil (in 1976 and 1977). The fertilizers, at rates of 0,
150, and 220 kg Nlha, were applied at days 6, 10, and 15 before sowing of 50
glomerules per m 2 • Immediately after sowing, betanil was administered at a rate of 7-10
kg/ha. Microplots not fertilized and/or not treated with herbicide served for comparison.
The seedlings were counted at the stage of 2-4 leaves.
On the sandy soil, application of 220 kg Nlha led to a significant diminution (:S22%
on average) of the count of emerged seedlings. The toxic effect of urea was stronger
than that of the lime ammonium nitrate; PPDA did not attenuate the toxic effect.
Moreover, in 1976, urea and urea+ 1% PPDA, even at the rate of 150 kg Nlha,
manifested a toxic effect when they had been applied at day 10 or 15 before sowing.
Betanil administered alone also reduced the number of emerged seedlings and, when
applied together with fertilizers, strengthened their toxic effect. For example, in the
treatments with urea or urea+ 1% PPDA + betanil, 30-40% of the glomerules did not
emerge.
The fertilizers were also toxic on the black soil. Betanil alone had no effect on the
emergence of glomerules and enhanced less evidently the toxic effect of fertilizers,
which was attributed to the high adsorptive capacity of this soils.
In another experiment, on the day of fertilization with urea and urea+ 1% PPDA (at
rates of 0, 150, and 220 kg Nlha), the herbicide betanal (6 kg/ha) was applied on 1O_m2
plots, and immediately after sowing betanil (7-10 kg/ha) was administered. There were
also plots not treated with betanal. It was established that betanal did not bring about
any significant change in the number of emerged seedlings in the two soils studied. In
the treatments with urea+PPDA + betanil or urea+PPDA + betanil + betanal, as
compared to the urea + betanil or urea + betanil + betanal treatments, the counts of
emerged seedlings were higher, but the differences between treatments were not
significant.
Yeomans and Cerrato (1993) mentioned in a short report on their growth chamber
experiments that the oilseed rape plants treated with urea and urea+PPDA had lower
yields than those treated with nitrate and nitrate+PPDA.
318
7.J O. 9. Effect of Phosphoric Triamide (PTA) and Thiophosphoric Triamide (TPTA)

Compounds
In the sugarbeet field experiment of Bayrakli and Gezgin (1996), N-(n-
butyl)thiophosphoric triamide (nBTPTA) at the rate of 0.5% was most efficient in
reducing the volatile ammonia losses from plots treated with urea (200 kg N/ha) (see
Section 4.9.1.2). nBTPTA at this rate significantly increased (p<0.05) the refined sugar
yield (t/ha) from 5.61 to 8.87, whereas the yield-increasing effect of 0.25% nBTPTA
from 5.61 to 8.65 was not significant.
7.10.10. Effect ojHumic Substances

Pirogovskaya et al. (1993) conducted in 1989 and 1990 experiments in potato fields on
soddy-podzolic clayey and on soddy-podzolic loamy and sandy soils in several regions
in Belorussia. Total area of fields was 140 ha. Urea coated or mixed with humic
substances: hydro humate and oxyhumate extracted from peats and urea alone were
applied at rates of 90-120 kg N/ha together with P and K fertilizers before seeding
potatoes.
Urea amended with hydrohumate and oxyhumate produced higher yields than urea
alone. On overall average, the yield increase with urea-hydro humate was 8.5% in the
clayey soils and 13.3% in loamy and sandy soils. The corresponding yield increases
with urea-oxyhumate were 4.9 and 6.8%, respectively. The yield increases were
accompanied by increased crude protein and decreased nitrate contents in the tubers. All
these effects of humates were more marked in light- than in heavy-textured soils.

Parashar et al. (1980) compared the efficiency of five N sources: urea (U), neem cake
mixed with urea (NCMU), neem cake-coated urea (NCCU), sulfur-coated urea (SCU),
and nitrapyrin-amended urea (NU) for sugarcane. Field experiments were carried out in
the 197611977 and 197711978 crop seasons. In both years, the N rates were 75 and 150
kg N/ha. The weight ratio of name cake/urea was 20/100.
The refined canesugar yields were always higher at 150 than at 75 kg N/ha rate. The
yields presented the orders: NCMU ~ NCCU ~ NU > SCU > U in 197611977, and
NCMU> NU > NCCU ~ SCU ~ U in 197711978. Thus, the performance of NCMU
should be considered better than that of the more costly SCU or NU.
Urea amended with neem cake or neem oil also performed better than urea alone in
increasing the yield of Japanese mint (Ram et ai., 1988) and geranium (Rajeswara Rao
et ai., 1990).
7.10.12. Effect of Combined Urease Inhibitors

According to Ziyamukhamedov et ai. (1986), fertilization of cotton soils with urea, to
which copper sulfate + potassium ethyl xanthate had been added, led to 10-15%
increases in the coefficient of plant uptake of fertilizer N and to 7-17% increases in
cotton yields as compared with the results registered in the control (urea without
inhibitors).
7.10.13. Effect of Combined Urease and Nitrification Inhibitors

In the greenhouse experiment of Montemurro et al. (1998) (see page 246), fresh weight
oflettuce plants was not significantly affected by nBTPTA, DCD, and nBTPTA+DCD,
319
but the nitrate content in plants (mglg fresh weight), which was 2.182 in the urea-only
treatment, was reduced insignificantly to 2.143 by nBTPTA and significantly to 1.650
and 1.736 by DCD and nBTPTA+DCD, respectively.
321
Chapter 8. Effect of Urease Inhibitors on Other Enzyme Activities, Microbial

Counts and Biomass as well as on Respiration and Other Microbial
Processes in Soils
A primordial requirement with which the inhibitors of soil urease actlVlty should
comply is the specificity: they should inhibit urease activity effectively without
exhibiting negative side effects on the soil life; they should not affect the other
enzymatic activities, the microorganisms and microbial processes playing an important
role in the biological cycles of elements and in soil fertility.
8.1. EFFECT OF UREASE INHIBITORS ON OTHER ENZYME ACTIVITIES IN

SOILS
8.1.1. F;[fect ofHeavy Metal Compounds

In studies on the effects of pesticides on biochemical and microbiological properties of
Canadian soils, Tu (1981a,b, 1982, 1990, I 992a,b ) used HgCh as a reference compound
and determined, besides urease activity (see page 10), other enzyme activities as well.
Rate of HgClz addition was 70 or 80 flglg soil. and incubation was carried out at 28°C.
Tu (198Ia) found that HgClz significantly decreased (p=0.05) dehydrogenase and
phosphatase activities in a clay loam soil (PH 7.2). In an organic soil (PH 7.2), HgClz
caused a significant and an insignificant decrease in dehydrogenase activity after 7 and
14 days of incubation, respectively, whereas phosphatase activity was significantly
reduced (Tu, 198Ib). In another organic soil (PH 6.7-6.8) studied by Tu (1982, 1990),
HgCIz addition led to a significant decrease and to no significant changes in
dehydrogenase activity after 7- and 14-day incubation, respectively, but phosphatase
activity was not affected significantly. Invertase and amylase activities had significantly
increased values in the HgClrtreated samples after 2-day incubation, but no significant
differences were found between these activities and those measured in the control
samples after 3 days of incubation.
In a sandy loam (PH 7.6), Tu (1990, I 992a,b) registered significantly increased
dehydrogenase activity due to HgCl 2 after each incubation time (2, 7, and 14 days),
whereas the effect of HgClz on phosphatase activity was insignificantly inhibitory.
Invertase and amylase activities were significantly increased and not affected
significantly by HgClz after 2 and 3 days of incubation, respectively.
Skujins et at. (1986) found that CrCl J and CuCh were stronger inhibitors of
nitrogenase than of urease activity in samples of a forest soil (see page 14). Thus, er 3+
at 200 flglg soil rate inactivated the nitrogenase system, but Cu2+ at its lowest rate (50
flglg soil) exhibited a stimulatory effect on this enzyme system.
In the pot experiment of Kandeler et al. (1990) (see page IS), the heavy metal salts
inhibited not only urease activity but also dehydrogenase. ~-glucosidase, cellulase,
protease, nitrate reductase, alkaline phosphatase, phospholipase, and arylsulfatase
activities, in both soils studied (a sandy loam and a clay loam). Xylanase activity was
inhibited in the sandy loam but not in the clay loam. Dehydrogenase, alkaline
phosphatase, and arylsulfatase activities were most sensitive to the heavy metal salts.
In the experiment of Benedetti et al. (1990), Cr203 added to samples of an Italian
soil (at a rate of 100 mg Cr/kg soil) caused decreases not only in the urease activity (see
322
page 15) but also in phosphatase and casein-hydrolyzing and Na-benzoyl-L-

argininamide (BAA)-hydrolyzing protease activities.
Chernykh (1991), whose pot experiments are briefly described on page 15,
determined the effect of Cd, Pb, and Zn on urease and other, three enzyme activities and
drew the conclusion that sensitivity of the activities to these heavy metals when each
was applied alone presented the order: urease> invertase » catalase> phosphatase. A
partly other order was established when these heavy metals were applied in
combination: urease » catalase> invertase> phosphatase. Thus, urease activity was
always most sensitive and phosphatase activity least sensitive to Cd, Pb, and Zn.
Soil urease activity in the pot experiment of Kucharski and Niklewska (1992) was
inhibited only by the 1,000 ppm Zn rate (see page 16), but dehydrogenase and acid and
alkaline phosphatase activities were inhibited also by the 100 and 10 ppm Zn rates, and
the extent of inhibition was proportionate to the Zn rate.
Speir et at. (1995) studied the effect of Cr(VI) on soil urease activity (see page 16)
and also on phosphatase and arylsulfatase activities and established for these three
activities the following order of decreasing sensitivity to Cr(VI): arylsulfatase >
phosphatase> urease.
Soil urease activity, as studied by Hemida et al. (1997) (see page 17), was more
sensitive than nitrate reductase activity and much more sensitive than amidase activity
to Cu 2+ and Zn2+ in both clay and sandy soils studied.
In the pot experiments referred to by Kucharski (1997), Zn inhibited soil urease
activity and stimulated dehydrogenase activity, whereas Pb and Cd inhibited both
activities (see page 17 and Table 4).
Wyszkowska et al. (2001) found that in their pot experiments the effect of Cr on soil
dehydrogenase and acid and alkaline phosphatase activities was similar to the effect
exerted by Cr on soil urease activity (see page 19).
In the experiment of Moreno et al. (2001) (see page 19), dehydrogenase activity was
also determined. In both soils treated with CdS04, the ED50 (the Cd concentration
inhibiting dehydrogenase activity by 50%) was much higher after 3 hours of incubation
than after 28 days. This means that the initial sensitivity of dehydrogenase activity to Cd
decreased during the incubation.

Yarovenko et al. (1982) found that MgClz, NaCI, and MgClz+NaCI increased urease
activity in samples of a chemozern containing or not containing residues of vetch-oats
(see page 22), whereas the effect of these salts on other soil enzyme activities was
highly varied. Thus, in soil samples without crop residues, MgCl2 did not affect
invertase and neutral phosphatase activities, stimulated acid and alkaline phosphatase,
nitrate and nitrite reductase activities, and inhibited protease and hydroxylamine
reductase activities. NaCl stimulated neutral and alkaline phosphatase activities and
inhibited the other activities. MgCi 2+NaCI did not affect acid phosphatase activity,
stimulated neutral and alkaline phosphatase activities, and inhibited the other activities.
The inhibitory effect was attenuated or even replaced by stimulatory effect in soil
samples containing crop residues.
Garcia and Hernandez (1996) treated samples of a calcareous soil with 0.1 to 1.3 M
solutions ofNaCI and Na2S04 (see page 26), and found that dehydrogenase and catalase
activities were decreased by the 0.1 M NaCI and Na2S04 solutions and increased by the
323
more concentrated salt solutions. The increasing effect of Na2S04 was more marked
than that of NaCl. At the same time, both salts inhibited ~-glucosidase, BAA-
hydrolyzing protease, and phosphatase activities. NaCI was more inhibitory than
Na2S04. The degree of inhibition increased with increasing salt concentrations. The
inhibition caused by NaCl reached 40% (~-glucosidase), 56% (BAA-hydrolyzing
protease), and 75% (phosphatase).
8.1. 3. Effect ofFluorides

In the pot experiments of Abliwva and Tomina (1997) (see page 34), NaF, applied at
rates of 50 and 500 mg F/kg soil, increased dehydrogenase activity, did not affect
catalase activity, and decreased invertase activity.
8.1.4. Effect of Organic Mercury Compounds

Reddy and Chhonkar (1990b) carried out a pot experiment in which 3-kg samples of a
sandy loam soil (pH 7.6) were amended with phenylmercuric acetate (PMA) at a rate of
2 mgikg soil, planted to rice and flooded or not flooded. No PMA was added to the
control. During the growth period, nitrate reductase activity was measured periodically
in soil, and it was found that PMA decreased the activity throughout the growing
period. No activity was detectable in flooding water of either control or PMA-treated
variant. The inhibitory effect of PMA on soil nitrate reductase activity was confirmed in
a laboratory experiment in which Reddy and Chhonkar (1990c) treated 5-g soil samples
with a solution containing 0, 1 or 2 mg PMAlml, and, after 24-hour preincubation at
28°C, a nitrate solution was added to give final concentrations ranging from 100 to
1,000 !!g N0 3 --N Ig soil.
8.1. 5. E;Uect qf Urea Derivatives

In Kolyada's (1970, 1973) pot experiments, thiourea added to samples of a soddy-
podzolic soil at a rate of 0.1 g thiourea-N/kg soil inhibited urease activity (see page 49)
and also the catalase and invertase activities. Inhibition of each activity was strong at
day 5 after thiourea application but weakened during the growing season.
8.1. 6. Effect ofDithiocarbamates

The effect of manganese ethylene-l ,2-bisdithiocarbamate (maneb) on the urease activity
in Canadian soils was studied by Tu (1980, 1981 a,b) (see page 56). The effect of rnaneb
on other soil enzyme activities was also studied. Maneb was applied at rates of 0, 5, and
10 !!g!g soil.
In a clay loam soil (PH 7.2), maneb at both rates significantly increased (p=0.05)
dehydrogenase activity and insignificantly decreased phosphatase activity (Tu, 1981a).
In an organic soil (PH 7.2), maneb at the lower rate significantly increased
dehydrogenase activity after 7 and 14 days of incubation, but at the higher maneb rate
dehydrogenase activity was insignificantly decreased after 7 days and significantly
increased after 14 days of incubation. Phosphatase activity was significantly decreased
by both maneb rates (Tu, 1981 b).
In another organic soil (pH 6.7), invertase activity was not affected significantly by
the lower maneb rate after 1 and 2 days of incubation, but at the higher maneb rate
invertase activity significantly increased after 1 day and insignificantly decreased after 2
324
days of incubation. Amylase activity determined after I and 3 days of incubation was
not significantly affected by either 5 or 10 Ilg maneb/g soil (Tu, 1982).
8.1. 7. Effect q(Thiuram Disu!fides

Studies of Tu (1981a,b, 1990) on the effect of tetramethylthiuram disulfide (thiram) on
urease activity in Canadian soils (see page 56) were also accompanied by determination
of other soil enzyme activities. Rates of thiram addition were: 0, 5, and 10 Ilg/g soil.
In a clay loam soil (PH 7.2), dehydrogenase and phosphatase activities were
significantly increased by the lower thiram rate and insignificantly by the higher rate
(Tu, 1981a).
In an organic soil (PH 7.2), the two thiram rates had no significant effect on
dehydrogenase activity but significantly decreased phosphatase activity (Tu, 198Ib).
In another organic soil (PH 6.7), invertase and amylase activities were not
significantly affected by any thiram rate (Tu, 1982).
In a sandy loam soil (pH 7.6), thiram, applied exceptionally only at the rate of 10
Ilglg soil, significantly decreased dehydrogenase activity after both 7 and 14 days of
incubation but had no significant effect on phosphatase activity. Invertase activity was
significantly decreased, whereas amylase activity was not affected by thiram after either
2 or 3 days of incubation (Tu, 1990).
8.1.8. Ef]ect q(Heterocyclic Sulfur Compounds

Tu et at. (1995) studied the effect of 3,5-dimethyltetrahydro-l,3 ,5-thiadiazine-2-thione
(dazomet) on urease activity (see page 72) and also on dehydrogenase activity in a
Canadian loamy sand soil, under field conditions. Dazomet (0 and 56 kglha) was
applied on May 10, 1978. Dehydrogenase activity, determined in soil samples taken on
May 23 and June 27, was not significantly different in dazomet-treated and untreated
soil in the May samples, but it was significantly higher in the dazomet-treated than
untreated soil in the June samples.
8.1. 9. E.ffect q(Monohydric Phenols

Shen et al. (1997) determined dehydrogenase activity in untreated and nitrophenol-
treated samples of a Chinese soil. Depending on the rate of nitrophenol addition (ppm
on soil weight basis), the following relative activity values were registered: 100% (0
ppm), 208.3% (50 ppm), 100% (100 ppm), 66.7% (150 ppm), and 33.3% (200 ppm). In
other words, nitrophenol at the lowest rate stimulated and at the two highest rates
inhibited soil dehydrogenase activity.
8.1.10. Ej]ec! q(Polyhydric Phenols and Quinones

May and Douglas (1979) studied the effect of catechol, p-benzoquinone, and 2,5-
dimethyl-p-benzoquinone on several enzymatic activities in three Australian soils. Each
compound was used at a rate of 50 Ilglg soil. The data in Table 68 show that the three
compounds studied did really manifest a strong inhibitory effect on urease activity in
each soil, but they also inhibited, though to a lesser extent, the other activities, except in
the case of invertase which was not affected by any of the compounds in any of the
soils. Degree of inhibition varied depending on the nature of enzyme and soil type. On
325
TABLE 68. Effect of catechol (CT), p-benzoquinone (BQ), and 2,5-dimethyl-p-benzoquinone

(DBQ) on activity of soil enzymes"
Inhibition (%)
CT BQ DBQ
Enzyme
Soils·
A B C A B C A B C
Urease 96 96 93 93 96 92 96 94 92
Dehydrogenase 20 22 34 54 60 59 85 90 88
Invertase 0 () () () 0 () () () ()
Amylase 8 13 6 6 9 17 25 41 13
Protease 18 24 8 12 14 8 17 7 23
Phosphatase 4 () () 0 4 0 0 4 0
Arylsulfatase 7 5 23 7 2 7 4 8 0
"From May and Douglas (1979).
bA~Loam B~Silt loam C~Clay.
average, the enzymatic activities were inhibited in the following order:

urease » dehydrogenase> protease;::: amylase> arylsulfatase > phosphatase
(invertase, as mentioned above, was not inhibited).
2,4-Di-t-butylphenol and 24 quinones and polyhydric phenols, which were tested at
a rate of 50 ppm (on soil basis) (Mishra and Flaig, 1979) or at rates of 10 and 20 ppm
(Mishra et af.. 1980) as inhibitors of urease activity in two German soils (brown earth
and black earth) (see pages 79 and 91), were also studied for evaluating their effect on
soil actual dehydrogenase activity. Dehydrogenase activity, like urease activity, was
determined after 0, 7, 14, and 28 days of incubation of soil samples (200 g) with 5 rnl of
a solution containing the test compound (at rates of 50 and 10 or 20 ppm, respectively),
5 rnl of water or urea solution (100 ppm urea) and water up to 60% ofWHC.
It was established that all compounds that inhibited urease activity also inhibited
dehydrogenase activity. Thus, 4,6-di-t-butylcatechol was the strongest inhibitor of both
activities. Degree of inhibition increased with increasing inhibitor rate. Inhibition of
dehydrogenase activity was more marked in the black earth than in the brown earth
(urease activity was more strongly inhibited in the brown earth than in the black earth).
In general, inhibition of dehydrogenase activity decreased with time. 2,4-Di-t-
butylphenol, I-chloroanthraquinone, and 2,5-di-t-butylhydroquinone, which enhanced
urease activity, also enhanced dehydrogenase activity. 4,6-Di-t-butylresorcinol and 4,6-
di-f-butylpyrogallol, which did not inhibit urease activity, manifested an inhibitory
effect on dehydrogenase activity.
It was found in studies on Canadian soils that p-benzoquinone (BQ) used at a rate of
50 J.lg/g soil significantly inhibited dehydrogenase activity in samples of an organic soil
(pH 7.2) incubated for 7 days, but the activity recovered after 14 days of incubation.
Phosphatase activity was also inhibited as evidenced by a 2-hour test (Tu, 1981b). At
the same rate, BQ had no significant effect on invertase and amylase activities in
samples of an organic soil (PH 6.7) (Tu, 1982) and of a sandy loam soil (PH 7.3 7) (Tu,
1988). Incubation lasted 1 and 2 days (invertase) or 1 and 3 days (amylase).
Working with samples of two soils (leached chemozem and alluvial soil), Radulescu
ef al. (1984) studied the effect of hydroquinone (HQ) on several enzyme activities and
respiration of soil. HQ was used in amounts of 0, 1, 2, 4, and 10% relative to weight of
urea, corresponding to 0, 0.012, 0.024,0.048, and 0.120 mg HQ/g soil.
326
From the results presented in Table 69, one can see that urease activity decreased
greatly in the presence of HQ. In the leached chernozem, there was a parallelism
between the activity decrease and the rate of HQ. In the alluvial soil, the strongest
inhibition was produced, surprisingly, not by the highest HQ rate (0.120 mglg soil) but
by the 0.048 mg rate.
Actual dehydrogenase activity showed a trend to decrease with increasing HQ rate
in both soils. The HQ rates of 0.012-0.048 mglg soil enhanced potential dehydrogenase
activity in the leached chemozem but had an opposite effect in the alluvial soil. At the
same time, the 0.120 mg HQ rate markedly diminished potential dehydrogenase activity
of both soils. HQ did not bring about any changes in catalase activity of the two soils
studied. Invertase activity measured in the presence of buffer solution remained
unchanged in the HQ-treated samples of the leached chernozern but decreased, to some
extent, in similarly treated samples of the alluvial soil. When invertase activity was
measured without buffer, the activity values were a little lower in the HQ-treated
samples of chernozern than in the untreated sample, but the activity was practically
identical in all samples of the alluvial soil. Phosphatase activity in the chemozem was
not significantly influenced by any of the HQ amounts applied. In the alluvial soil, the
0.012-0.048 mg HQ rates did not affect phosphatase activity, while the highest rate
slightly increased it.
The conclusion can be drawn that, besides the inhibitory effect on urease activity,
HQ also manifested inhibitory or stimulatory effects on other enzymatic activities,
depending on its rate, soil type, and experimental conditions. However, all these side
effects of HQ were less marked than its strong inhibitory effect on soil urease activity.
Reddy and Chhonkar (1990a,b) studied the effect of three dihydric phenols and five
quinones on the nitrate reductase activity in a sandy clay loam (PH 7.6). Reddy and
Chhonkar (1990a) conducted a pot experiment in which hydroquinone, added to 3-kg
soil samples at a rate of 2 mglkg soil, decreased nitrate reductase activity. Reddy and
Chhonkar (1990b) determined the inhibition constant of the eight compounds, each
having been applied at rates of 1 and 2 !1g1g soil. The constants ranged from 5.3 to
12.7 x 10.9 M and their values (x 109 M) presented the following descending order:
catechol (12.7), l,4-benzoquinone (9.7), hydroquinone (9.6), l,4-naphthoquinone (9.5),
4-methy1catechol (8.9), 2-methyl-l,4-naphthoquinone (7.2), 2,6-dibromoquinone-4-
chloroimide (5.4), and 2,6-dichloroquinone-4-chloroimide (5.3).
Zhao and Zhou (1991) and Zhao et al. (1991) performed a long-term laboratory
experiment to study the effect of HQ applied with urea on several enzyme activities in a
meadow brown soil (silty loam, pH 6.55). Soil samples (100 g) were submitted to the
following treatments: no urea and no HQ; urea alone (480.8 mg); urea + HQ (0.96, 2.0,
4.81 or 9.61 mg). The moisture content was adjusted to 40% of WHC. Incubation took
place at 30°C. After 3, 10,25,55, and 88 days of incubation, subsamples were taken to
run specific enzyme assays.
The results showed that HQ had no effect on invertase activity but temporarily
promoted or inhibited the activities of polyphenol oxidase, dehydrogenase, protease,
and phosphatase through its direct effect on these enzymes or indirectly through causing
a delay in urea hydrolysis and changes in soil microbial activity. With time, the
promoting and inhibitory effects gradually decreased with no significant difference at
the end of incubation.
TABLE 69. Effect of hydro quinone on enzymatic activities and respiration of soil"
En~matic activities" Res2iration'
Hydroquinone
Soil Deh:tdrogenase Invertase With added Without added
(J.lglg soil) Urease Catalase Phosphatase
Actual Potential With buffer Without buffer glucose glucose
0 2.30 0.30 0.40 42.25 1.10 0.74 2.05 80.90 16.97
0.012 0.75 0.17 0.66 43.97 1.13 0.65 2.22 90.71 21.48
Leached
0.024 0.71 0.27 0.44 43.97 1.09 0.66 2.09 87.79 17.77
chernozem 2.19
0.048 0.52 0.25 0.50 45.69 1.18 0.47 81.69 9.54
0.120 0.27 0.09 0.31 43.28 1.13 0.60 2.19 63.12 10.07
0 1.89 0.42 0.89 40.54 1.05 0.64 0.85 55.44 30.58
0.012 0.70 0.34 0.71 38.48 0.71 0.61 0.84 77.44 19.14
Alluvial soil 0.024 0.63 0.37 0.67 38.13 0.56 0.65 0.87 77.88 21.98
0.048 0.00 0.30 0.85 40.88 0.74 0.63 0.93 102.08 26.62
0.120 0.63 0.22 0.47 38.82 0.72 0.61 1.05 206.58 26.84
"From Radulescu et al. (1984).
bEnzymatic activities are expressed as follows: urease in rng of ammonia produced by 5 g of soil during a 24-hour incubation at 37°C; dehydrogenase activity
in rng oftriphenylforrnazan/3 g of soill24 hours at 37°C; catalase in rng ofH20 212.5 g of soil! 1 hour at 20°C; invertase in difference of optical rotation (ilu°)/lO
g ofsoil/24 hours at 37°C; phosphatase in rng ofphenoll2.5 g ofsoill2 hours at 37°C.
'Respiration is expressed as rng of CO,!50 g of soil!7 days at 20°C.
w
N
-...I
328
8.1.11. Effect ofPh osphorodiamides

In the wheat experiment carried out under field conditions by Kucharski (1992) (see
page 273), phenylphosphorodiamidate (PPDA) inhibited not only soil urease activity
but also the acid and alkaline phosphatase activities (in both single and divided
applications of urea+PPDA) and stimulated the dehydrogenase activity (in divided
application ofurea+PPDA).

Compounds
Banerjee et af. (1997, 1999) applied urea (50 kg N/ha) with or without N-(n-
butyl)thiophosphoric triamide (nBTPTA) (0.25% weight/weight) to barley plots (5 by
1.8 m) under conventional and zero tillage at two sites in Manitoba, on a fine sandy
loam (pH 7.3) and a clay loam soil (PH 7.6), respectively. The experiments were
initiated in 1994. Soil samples were collected during the autumn of 1994, spring and
summer 1995 and spring 1996, and submitted to physical, chemical, enzymological, and
microbiological analyses. Acid and alkaline phosphatase and arylsulfatase activities
were determined.
Based on the results obtained, the conclusion was drawn that urea with and without
nRTPTA and tillage system had no significant impact on the three enzyme activities
determined.
8.1.13. Effect o/Humic Substances

In the experiments of Ablizova and Tomina (1997), ammonium humate (AH) applied at
rates of 0.3 and 0.6 tlha reduced urease activity in the dark chestnut soil studied (see
page 167). It was also found that AH markedly increased dehydrogenase and catalase
activities at its both rates, whereas invertase activity was slightly increased at the lower
AH rate and not affected at the higher AH rate.
8.1.14. Ef]ec! o/Lignosu((onates

Builov et af. (1979) treated 2_m 2 microplots installed on a solonetz soil with ammonium
lignosulfonate (ALS) in amounts of 0, 1.3, and 1.7% (relative to weight of the O-lO-cm
soil layer). Similar plots installed on a compact meadow soil were treated with ALS in
amounts of 0, OJ, and 1.0% (relative to weight of the 0-20-cm soil layer). Catalase,
polyphenol oxidase, invertase, and phosphatase activities were determined 3 months, 1
and 3 years after application of ALS to the solonetz soil. The compact meadow soil was
analyzed for the four enzyme activities 3 months and I year after ALS application.
In the solonetz soil, each activity was higher in the ALS-treated than in the control
(untreated) plots at each sampling time, and the increase was more marked at the 1.7
than at the 0.3% ALS addition, except polyphenol oxidase activity which was higher at
the lower than at the higher ALS addition. The maximum value of each activity was
recorded 1 year after ALS application.
In the compact meadow soil, the 0.3% ALS addition was more stimulatory on
polyphenol oxidase, invertase, and phosphatase activities than was the 1% addition, and
the reverse was true for the catalase activity.
329
8.1.15. E.tfect ofPlant Materials

In a pot experiment of Reddy and Chhonkar (1990b), urea alone (100 mg N/kg soil) and
urea-amended with alcoholic extract of neem kernel (14% relative to weight of urea)
were added to 3-kg samples of a sandy clay loam (PH 7.6). Farmyard manure (5%
relative to soil weight) was also added to some samples. All samples were then
incubated under flooded or non-flooded conditions.
The neem extract always decreased nitrate reductase activity in soil, and its
decreasing effect exceeded those of phenylmercuric acetate and hydroquinone, applied
at a rate of 2 mg/kg soil.
* *
*
Hickisch (1981) described the effect of 15 urease inhibitors (without giving their
names) on urease and dehydrogenase activities in a sandy soil. The fresh soil samples
(250 g in a 5-cm layer) were treated with standard (practical) dose of inhibitors, then
incubated at 25°C for 3 days. Untreated samples served for comparison. Incubation was
followed by determination of the enzyme activities. As expected, each of the 15
compounds inhibited urease activity; degree of inhibition varied between 23 and 80%
(average: 50.6%). Dehydrogenase activity was inhibited by 7 urease inhibitors (degree
of inhibition: 1-17%; average: 6.1%), stimulated by other 7 urease inhibitors (degree of
stimulation: 2-11 %; average: 5.1 %), and unaffected by one urease inhibitor.
8.2. EFFECT OF UREASE INHIBITORS ON MICROBIAL COUNTS AND

BIOMASS AS WELL AS ON RESPIRATION AND OTHER MICROBIAL
PROCESSES IN SOILS
8.2.1. Effect on Microbial Counts and Biomass
8.2.1.1. E.tfect o/Heavy Metal Compounds

In the pot experiment of Kandeler et al. (1990) (see page 15), the heavy metal salts
reduced the total microbial count in a sandy loam but did not affect it in the other soil
studied (a clay loam).
Tu (l992b) treated samples ofa sandy loam with 80 ~g HgCb/g soil. No HgCb was
added to the control. Urease activity determined after 7 days of incubation at 28°C and
number of bacteria determined after 7 and 21 days of incubation were not significantly
different (p=0.05) in the HgCh-treated and untreated soil samples. Only the number of
fungi was reduced after 7 days, but after 21 days the same number was registered in the
treated and untreated soil samples.
In the pot experiment of Kucharski and Niklewska (1992), in which the soil samples
were treated with 0, 10, 100, and 1,000 ppm Zn (see page 16), the counts of
heterotrophic bacteria, actinomycetes, Azotobacter cells, arnmonifying, proteolytic, and
cellulolytic microorganisms and number of fungi varied irregularly, showing
unchanged, decreased or increased values, depending on the Zn rate and time of
sampling (2 weeks after beginning of the experiment or after harvesting the test plant,
broadbean).
330
Kozdr6j (1995) determined urease activity in 200-g samples of a forest soil treated
with I or 2 mg Cu or Cd (as sulfates) (see page 17) and also the numbers of bacteria and
fungi tolerant to 0.1 or I mg of Cu or Cdil nutrient medium. Urease activity did not
correlate with the number of tolerant bacteria, but it correlated significantly and
positively with the number of fungi tolerant to 0.1 mg Cd when the soil sample was
treated with 1 mg Cd. The correlation remained significant but became negative when
the soil sample received 2 mg Cd.
In the complex study of the effects of Cr(VI) on soil biological properties, Speir et
af. (1995) determined urease, phosphatase, and arylsulfatase activities (see pages 16 and
322) and other parameters, including microbial biomass C, and it was found that
sensitivity of biomass C to Cr(VI) was similar to that arylsulfatase activity and higher
than those of phosphatase and urease activities.
Hemida et af. (1997) amended samples of a clay soil and a sandy soil with 0, 200 or
2,000 Ilg Cu or ZnJg soil. After L 4, and 12 weeks of incubation, the soils were
analyzed enzymologically (see pages 17 and 322) and also microbiologically.
The results obtained after 12 weeks will be summarized below. The number of
glucophilic fungi was not affected significantly (p=0.05) by Cu and Zn in the clay soil,
but it was increased in the sandy soil amended with 2,000 Ilg ZnJg soil. The number of
thermophilic and thermotolerant fungi was not affected by Cu and Zn in either clay or
sandy soil. The number of cellulose-decomposing fungi was reduced by both rates of
Cu and Zn in the clay soil and not affected in the sandy soil. The number of bacteria did
not suffer significant changes due to Cu and Zn in any soil. The number of
actinomycetes did not change significantly in the clay soil, but it was increased in the
sandy soil at both 200 and 2,000 Ilg ZnJg soil rates.
Potassium dichromate applied at rates of 80 and 120 mg Cr/kg soil, in the pot
experiments of Wyszkowska et al. (2001) (see page 19), adversely affected growth of
Azotobacter sp. and actinomycetes but stimulated the proliferation of oligotrophic,
copiotrophic, anl!llonifying, and Nrfixing bacteria.
8.2.1.2. Effect o/Fluorides

In the pot experiments of Ablizova and Tomina (1997) (see page 34), the effect ofNaF
on the soil microbial populations was not marked. For example, counts of the
anl!llonifying bacteria in the untreated soil and NaF -treated soil at rates of 10 and 50 mg
F/kg soil were 0.5.106 , 0.3.10 6 , and 0.2.106/g soil, respectively.
8.2. 1.3. F:ffect of Urea Derivatives

In Kolyada's (1970, 1973) pot experiments, thiourea applied at the rate of 0.1 g
thiourea-N/kg soil decreased urease activity (see page 49) and count of nitrifying
bacteria, but led to increases in the counts of ammonifying and butyric acid bacteria,
fungi, and cellulolytic microorganisms.
8.2.1.4. Effect q(Dithiocarbamates

Tu (1980, 1981a) studied the effect of manganese ethylene-1,2-bisdithiocarbamate
(maneb) on soil urease activity (see page 56) and also on counts of bacteria and fungi.
Tu (1980) treated samples of a sandy loam soil (pH 7.6) witl1 0, 100, and 200 ~lg
maneb/g soil. Number of N 2-fixing bacteria was determined after 1, 2, and 4 weeks of
incubation, and it was found at each incubation time that maneb significantly decreased
331
(p=0.05) the number of these bacteria at both rates. Number of fungi, determined after 1
and 2 weeks of incubation was significantly decreased by both rates of maneb for 1
week, but no significant effect was recorded after 2 weeks.
Lower maneb rates (5 and 10 Ilg/g soil) were applied by Tu (l981a) to samples of a
clay loam soil (pH 7.2). The incubation lasted 2 or 7 days. After 2 days, decreased
number of bacteria was found at the higher maneb rate and decreased number of fungi
at both maneb rates. After 7 days, no significant effect of maneb was recorded. The
number of non-symbiotic Nrfixing bacteria was not significantly affected by maneb
(except in a single case: the number of these bacteria was higher at the lower maneb rate
after 7 days of incubation).
8.2.1.5. Effect ofThiuram Disu!fides

The effect oftetramethylthiuram disulfide (thiram) on urease activity in a clay loam soil
(pH 7.2) was studied by Tu (1981a) (see page 58). The effect of thiram on counts of
bacteria and fungi was also studied under condition identical to those under which the
effect of maneb was studied (see the preceding section). Thiram at both 5 and 10 Ilg/g
soil rates increased the number of bacteria and fungi after 2-day incubation and the
number of non-symbiotic Nz-fixing bacteria after 7 -day incubation.
8.2.1.6. Effect oJHeterocyclic Sulfur Compounds

The field experiment conducted by Tu et al. (1995) for evaluating the effect of 3,5-
dimethyltetrahydro-l,3,5-thiadiazine-2-thione (dazomet) applied at a rate of 56 kglha on
soil urease activity is referred to on page 72. The effect of dazomet on number of
bacteria and fungi was also studied. Dazomet significantly (p<0.05) increased the
number of bacteria and did not affect the number of fungi.
8.2.1. 7. Effect of Monohydric Phenols

Shen et al. (1997) found that the numbers of ammonifying, nitrifying, and denitrifying
bacteria were reduced by addition of nitrophenol to samples of a Chinese soil. There
was a parallelism between degree of reduction and rate of nitrophenol (100, 200, and
300 ppm on soil weight basis). Sensitivity of bacteria to nitrophenol presented the order:
nitrifiers » denitrifiers > arnmonifiers. The effect of nitrophenol (m soil enzyme
activities was also studied (see pages 82 and 324).
8.2.1.8. E.Uect ofPho;.,phorodiamides

Held et at. (1978) mentioned, without presenting concrete data, that, according to soil
biological investigations, phenylphosphorodiamidate (PPDA) did not influence
microbial population and spectrum (counts, generic and specific composition of soil
microorganisms), and drew the conclusion that PPDA did not exert any effect on
microbial urease synthesis in soil.
Mai and Fiedler (1986), who studied the effectiveness of PPDA for inhibiting urea
hydrolysis in the soil of a -90-year-old spruce forest (see page 114), also dealt with the
effect of PPDA on soil microorganisms. In the urea + PPDA treatment, in comparison
with the urea-only treatment, a significant reduction occurred in the total number of
bacteria and actinomycetes as well as in the count of ureolytic bacteria. This effect of
PPDA lasted, generally, 2-3 weeks. PPDA did not bring about any changes in total
number and species composition of fungi and in count of nitrifying bacteria. It should
332
be mentioned that comparison of the plots that received urea alone with the untreated
control plots revealed that urea addition led to a significant increase in counts of soil
microorganisms from all groups studied.
In the wheat experiment carried out under field conditions by Kucharski (1992) (see
page 273), PPDA had no visible effect on total number of heterotrophic soil bacteria,
number of Azotobacter cells, actinomycetes, proteolytic, and cellulolytic micro-
organisms, but slightly stimulated the proliferation of fungi.
In the laboratory experiments of Kucharski (1994) (see page 129), the results
obtained concerning the effect of PPDA on the munber of heterotrophic soil bacteria,
actinomycetes, proteolytic and ammonifying microorganisms, and fungi indicated no
regularity: PPDA had no effect or exerted an inhibitory or stimulatory effect depending
on duration (2 up to 25 days) and/or temperature (l0, 20 or 30°C) at which the urea-
and urea+PPDA-treated samples were incubated.
8.2.1.9. F;[fect Qf PhosphoriC Triamide (PTA) and Thiophosphoric Triamide (TPTA)

Compounds
Ambroz et aZ. (1970) amended 400-g samples of a rendzina soil (PH 7) with phosphoryl
triarnide (PTA) at a rate of 0.04 g PTA-N and with 1 g of glucose. Diammonium
phosphate instead of PTA served for comparison. All samples were moistened and
incubated at 28°C for 14 days, during which the total numbers of bacteria and fungi
were determined at 2-3 day intervals. The results showed that PTA stimulated the
growth of both bacteria and fungi. In another experiment, the soil samples were
amended with PTA at rates of 0.02 and 0.04% PTA-N relative to weight of soil and
incubated at 28°C. It was found that PTA did not inhibit the growth of Azotobacter
chroococcum.
The effect of N-(n-butyl)thiophosphoric triamide (nBTPTA) on the soil microbial
biomass C and N was studied by Banerjee et aZ. (1997, 1999). Microplots under
conventional or zero tillage were fertilized with urea (50 kg Nlha) with and without
nBTPTA (0.25% relative to weight of urea). The soil was sampled for analyses 4 times
during the 1994-1996 period (see Section 8.1.12). The results showed that urea with and
without nBTPTA application and tillage system had no significant effect on microbial
biomass C and N. Nevertheless, biomass C and N had positive significant correlations
with soil moisture, organic C, and extractable C contents.
8.2.1.10. Effect QfCyc!otriphosphazatriene (CTPAT) Derivatives

In the experiments of Ambroz et al. (1970) briefly described in the preceding section,
hexaamino-CTP AT was also studied as was PTA. The results were also similar.
8.2.1.11. Effect ofHumic Substances

In the pot experiments of Abliwva and Tomina (1997) (see page 167), sodium humate
(SH) rather reduced the microbial counts at its rates of 0.5 and 1 tlha and increased them
at its rate of 1.5 tlha. For example, numbers of ammonifying bacteria (x J(t /g soil) were
0.50 (untreated soil), 0.32 (0.5 t SHlha), 0.10 (J t SHlha), and 0.80 (1.5 t SH/ha).
8.2.1.12. F;[fect QfLignosu(fonafes

The effect of ammonium lignosulfonate (ALS) applied at rates of 0, 1.3, and 1.7 tlha to
a solonetz soil and at rates of 0,0.3, and 1% to a compact meadow soil on soil enzyme
333
activities was studied by Builov et al. (1979) (see Section 8.1.3). Numbers of
heterotrophic bacteria were determined I, 2, and 3 years after ALS application to the
solonetz soil and 1 year after ALS application to the compact meadow soil. The
numbers (x 106/g soil) in the solonetz soil, untreated and treated with 1.3 and 1.7%
ALS, were the following: 18.2,31.8, and 54.3 (after 1 year), 6.8, 70, and 64 (after 2
years), and 126, 157, and 148 (after 3 years), respectively. The numbers recorded 1 year
after ALS application to the compact meadow soil were: 57.6 (untreated), 77.7 (treated
with 0.3% ALS), and 57.7 (treated with 1% ALS). Thus, in the solonetz soil the 1.7%
ALS rate was more efficient than the 1.3% rate 1 year after its application, but the
reverse was true 2 and 3 years after ALS application.
* *
*
Muller and Hickisch (1979) studied three urease inhibitors, marked by A, B, and C
(as they were not nominalized). Two experiments were carried out, both with a black
earth from Germany.
In the first experiment, fresh soil samples (50 g) were treated only with an inhibitor
at a rate of 1% relative to weight of urea-N if urea had been applied at a rate of 500 kg
N/ha. The next steps were incubation at 25°C for 7 days and, then, determination of the
counts of all bacteria, proteolytic, cellulolytic, and nitrifying bacteria as well as total
fungal mycelial mass and mycelial mass of penicillia, mucoraceae, and fusaria.
The microbiological analyses showed that inhibitor A significantly decreased the
count of proteolytic bacteria and the mycelial mass of mucoraceae and insignificantly
the count and mass of the other microorganisms, except mycelial mass of penicillia
which increased and count ofnitrifiers which remained unchanged. Under the influence
of inhibitor B, there was an insignificant decrease in the counts of bacteria and a
significant one in mycelial mass of all fungi. Inhibitor C did not bring about any
significant changes in bacterial counts and fungal biomass.
The second experiment was carried out in pots, each containing 5 kg of soil.
Inhibitors A and C were applied with or without urea four times, namely before
incubation and after 4, 8, and 12 weeks of incubation at 18°C. Each of the four
applications consisted of 0 or 200 kg urea-N/ha + I % inhibitor relative to N. During
incubation, total count of bacteria was determined periodically (in total, 20 times).
Inhibitor A had, also in this experiment, a decreasing effect on total bacterial count.
This effect became more and more pronounced during the incubation but it was
attenuated when urea was also applied. Inhibitor C led to an increase in total bacterial
count, and the increase was enhanced by urea application. In conclusion, inhibitor A
does not and inhibitor C does correspond for use in practice as a soil urease inhibitor.
Hickisch (1981) studied the effect of 15 urease inhibitors on counts of bacteria and
fungi in a sandy soil, under experimental conditions identical to those mentioned on
page 329. The results showed that growth of bacteria was inhibited by 10 urease
inhibitors (5-70% inhibitions; average: 24.8%) and stimulated by 5 urease inhibitors (2-
33% stimulations; average: l3.8%). Nine urease inhibitors inhibited and six stimulated
the growth of fungi; the degree of inhibition was 2-82% (average: 12.0%) and the
stimulation ranged from 4 to 36% (average: 10.0%). It was emphasized that under field
conditions even a 60% inhibition of microbial growth in the inhibitor-affected soil
334
layers disappeared in 4 weeks due to recolonization by microorganisms from the

unaffected deeper soil layers or from the neighboring fields.
8.2.2. Effect on Respiration
8.2.2.1. F;fJect ojHeavy Metal Compoundv

Kandeler et al. (1990), whose pot experiment is referred to on page 15, found that the
heavy metal salts inhibited both basal and substrate-induced respirations in both
Austrian soils studied (a sandy loam, pH 7.52 and a clay loam, pH 7.32). The inhibiting
effect of the heavy metal salts was stronger on substrate-induced respiration than on
basal respiration.
In Tu's (1992b) experiments, HgCh added to samples ofa Canadian sandy loam soil
(pH 7.6) at a rate of 80 l1g/g soil significantly increased (p<0.05) the respiration during
3 days of incubation at 30°e.
Studying the effect of Cr(VI) on soil biological parameters, Speir et af. (1995) (see
page 16) found that sensitivity of respiration to CreVI) was less than that of urease,
phosphatase or arylsulfatase activities.
8.2.2.2. F;/Ject qfAlkali Metal Salts

In the pot experiment of Garcia and Hernandez (1996) (see page 26), the inhibitory
effect of NaCl on soil respiration was stronger than that of Na2S04. Thus, respiration
was inhibited as much as 57% by a 1.3 M solution of NaC!.
8.2.2.3. F;/Ject q{1norganic Sulfur Compounds

Addition of ammonium thiosulfate to a flooded rice soil led, during the experimental
period (20 days), to 10-70% increases in soil respiration (Lim and Seo, 1994).
8.2.2.4. F;fJect ofDithiocarbamates

Tu (1980) added manganese ethylene-1,2-bisdithiocarbamate (maneb) to samples of a
sandy loam soil (pH 7.6) at rates of 0, 100, and 200 l1g/g soil. During the incubation (3
days at 28°C), the respiration increased about 7 and 9 times in samples treated with 100
and 200 Ilg maneb/g soil, respectively.
8.2.2.5. F;[fect qfPolyhydric Phenols and Quinones

Muller and Forster (1980) mentioned, without presenting experimental data, that
hydroquinone (HQ) drastically reduced soil respiration due to its microbicidal effect.
Riidulescu et af. (1984), determining respiration of soil samples treated with 0.012-
0.120 mg HQ/g soil (see Table 69), found that in the leached chernozem with added
glucose, respiration was enhanced by HQ except in its highest amount which had an
apposite effect. In the alluvial soil with added glucose, each HQ rate increased
respiration which was most intense at the highest HQ rate. In the chernozern without
added glucose, only the lowest HQ amount enhanced the respiration, whereas the 0.048
and 0.120 mg/g soil rates decreased it. Respiration of the alluvial soil without added
glucose was a little lower in the HQ-treated samples than in the untreated one.
By using 8-g samples of a Canadian sandy loam (PH 7.37) untreated or treated with
50 and 100 Ilg of p-benzoquinone (BQ)/g soil, Tu (1988) measured O2 consumption by
the samples at 30°C for 67 hours. The results indicated that respiration was significantly
335
higher in both BQ-treated samples (103 and 126 J1l O2 consumedlg soil, respectively)
than in the untreated control (75 III O2 consumed/g soil). In other words, BQ stimulated
the soil respiration.
8.2.2.6. F/Jixt of Phosphoric Triamide (PTA) and Thiophosphoric Triamide (TPTA)

Compounds
For studying the effect of phosphoryl triarnide (PTA) on respiration of two rendzina
soils (pH 7), Ambroz et al. (1970) used glucose and cellulose as respiratory substrates.
Soil samples (60 g each) were amended with 0.3 g glucose and PTA at a rate of
0.04% PTA-N relative to soil weight. No PTA was added to the control samples. All
samples wcre moistened and incubated at 28°C for 90 hours, during which the O2
consumption was measured. It was found that PTA markedly increased the respiration.
Ten-g soil samples were amended with 0.1 g cellulose powder and 0 or 0.04% PTA-
N, then moistened and incubated at 28°C for 18 days. Measurement of O2 consumption
indicated that the effect of PTA on the respiration on cellulose substrate was very weak.
In the experiments of Clay et al. (1990) (see page 244), the great reduction of the
volatilization of ammonia from urea amended with nBTPTA [N-(n-butyl)thio-
phosphoric triarnide] was associated with reduction of soil respiration.
8.2.2.7. E.tfect QrCyc!otriphosphazatriene (CTPAT) Derivatives

The experiments of Ambroz et al. (1970) (see the preceding section) were also carried
out with hexaarnino-CTPAT. The effect of hexaarnino-CTPAT on glucose-induced
respiration was weaker and on the cellulose-induced respiration stronger than that of
PTA.
8.2.2.8. Effect Qr Lignosu{{onates

Meier et al. (1993) found that ammonium lignosulfonate inhibited soil urease activity
and enhanced the respiration (see page 168).
8.2.3. Effect on Cellulose Decomposition
8.2.3.1. E.ffect QfAlkali Metal and Alkaline Earth Metal Salts

Yarovenko et al. (1982), who studied the effect of neutral salts on urease activity in
samples of a chernozern (see page 22), also dealt with the effect of these salts on
cellulose decomposition. The results obtained after 7 months of incubation showed that
MgCl 2 and MgClz+NaCl increased and NaCl decreased the degree of cellulose
decomposition in samples not amended with CTOP residues, whereas cellulose
decomposition in samples amended with crop residues was enhanced in the following
order: MgCh > MgC!2+NaCI > NaC!.
8.2.3.2. E.fJect of Ph osphorodiamides

According to the data obtained by Mai and Fiedler (1986), under the conditions of an
experiment in a -90-year-old spruce forest (see page 121), decomposition degree of
cellulose in soil in 9 months was 36% in unfertilized (control) plots, 54% urea-fertilized
plots, and 48% in plots treated with urea + 1% PPDA (phenylphosphorodiamidate).
336
8.2.4. Effect on Methane Emission and Oxidation
8.2.4.1. Effect of Combined Urease and Nitrification Inhibitors

Zhou et al. (1999) carried out pot experiments in which a flooded (anaerobic) rice soil
was fertilized with urea or with urea amended with hydroquinone (HQ), dicyandiamide
(DCD) or with HQ + DCD. Emission of methane (CH4) was measured during the
growth period of rice plants.
Both HQ and DCD reduced the C~ emission, and the highest (50%) reduction was
obtained in the HQ+DCD treatment.
8.2.4.2. Effect ofPhosphoroamides

It was mentioned in a short report by Bronson and Mosier (1992) that N-(n-
butyl)thiophosphoric triamide (nBTPTA) at a rate of 25 flg/g soil strongly inhibited
oxidation of methane in both soils studied. The inhibition was 83% in a fine sandy loam
and 60% in a sandy clay loam. The investigations were described in details by Bronson
and Mosier (1994). Besides nBTPTA, phenylphosphorodiamidate (PPDA) and the
oxygen analog of nBTPTA (nBPTA) were also studied. nBTPTA and PPDA were
applied at rates of 5 and 25 Ilg/g soil in both soils, but nBPTA was added only to the
fine sandy loam at the rate of25 flg/g soil.
The effect of these urease inhibitors on nitrification is dealt with on page 241. Their
effect of CH4 oxidation was studied under identical conditions. Percent inhibition of
C~ oxidation registered after 6 days of incubation at 28°C at 5 and 25 flg/g soil
inhibitor rates were 25 and 83 (nBTPTA), 28 and 64 (PPDA) in the fine sandy loam,
and 20 and 60 (nBTPTA), I and 28 (PPDA) in the sandy clay loam. nBPTA brought
about a 48% inhibition of C~ oxidation. It should be mentioned, however, that the
nitrification inhibitors acetylene and nitrapyrin were more effective inhibitors of CH4
oxidation than were the three urease inhibitors studied.
8.2.5. Effect on Nitrogen Mineralization
8.2.5.1. Effect ofHeavy Metal Compounds

In the pot experiment of Kandeler et al. (1990), the Zn, Cu, Ni, Cd, and V salts inhibited
soil urease activity (see page 15) and reduced by about 50% N mineralization in both
Austrian soil studied. The heavy metals also inhibited deamination of arginine. The
inhibition was -50% in a sandy loam (pH 7.52) and -20% in a clay loam (pH 7.32).
8.2.5.2. Effect ofphospho roam ides

Bremner et al. (l986b) used alanine as an organic N and tested nine phosphoroamides,
namely two phosphorodiamidates [phenylphosphorodiamidate (PPDA) and trichloro-
ethyl-PDA], six phosphorotriamides [N,N-dimethyl-, N,N-diethyl-, N-cyclohexyl-, N-
phenyl-, N-benzyl-N-methylphosphoric triamides and 4-fluoro-N-(diaminophosphinyl)
benzamide] and one thiophosphorotriamide [N-(n-butyl)thiophosphoric triamide
(nBTPT A)], with three Iowa soils (two clay loams and a sandy clay loam). Air-dried
soil samples (lOg) were treated with I ml of water or alanine solution containing 2 mg
Nand 3 ml of water or aqueous solution containing 0.1 or 0.5 mg of test compound.
After aerobic incubation (at 30°C for 21 days), the soil samples were analyzed for
N~-N and (N03- + NOn-N. The sum of the amounts of these three N forms was the
337
same in each soil treated with alanine with or without addition of phosphoroamides.
This means that the nine phosphoroamides tested did not affect mineralization of
alanine in any of the three soils studied (see also Bremner, 1986).
Banerjee et ai. (1997, 1999) conducted field experiments to evaluate the effects of
conventional and zero tillage, urea, and urea + nBTPTA [N-(n-butyl)thiophosphoric
triamide] fertilization on soil properties (see Section 8.1.12), including potential N
mineralization (No). The conclusion was drawn that No was not significantly affected by
either urea with or without nBTPTA or tillage system.
8.2. 6. Effect on Nitrification

See Subchapter 5.2.
8.2.7. Effect on Denitrification

First, the investigations in which several types of urease inhibitors were tested will be
dealt with.
Yeomans and Bremner (1986) evaluated the effect of 14 urease inhibitors (among
which there were six phosphoroamides) on denitrification of N0 3- in samples of a clay
loam soil from Iowa. The reaction mixtures were prepared from air-dried or field-moist
soil containing 30 g of oven-dry material + 5 m1 of water with or without 0.03, 0.3 or
1.5 mg of test compound + 5 ml of a solution containing KN0 3 (3 mg N) with or
without mannitol (4.5 mg C) and additional water to bring the total volume of water to
15 ml. The air from the flasks containing the reaction mixtures was evacuated and
replaced with helium for creating anaerobic conditions. After incubation (1-8 days at
30°C), the atmosphere in each flask was sampled for determination of the gaseous
products of denitrification (N20, NO, and N 2), and the soil was analyzed for N02- and
N0 3-.1t should be noted that only trace amounts of NO were detected.
None of the urease inhibitors tested had a significant effect on denitrification of
nitrate by soil microorganisms when applied at the rate of 1 or 10 J,lglg soil. The results
obtained with air-dried soil samples, which were treated with 50 J,lg of urease inhibitor/g
soil and incubated for 8 days, are presented in Table 70. One can see from this table that
2,5-dimethyl-BQ and 2,6-dimethyl-BQ strongly inhibited denitrification. The inhibitory
effect of PPTA was weaker. At the same time, eight compounds (PCMB, HQ, BQ,
2,6-dichloro-BQ, PTA, DAPBA, 4-F-DAPBA, and DAPBAA) enhanced denitrification
of nitrate and three compounds (CT, 2,5-dichloro-BQ, and PPDA) did not affect it (see
also Bremner, 1986).
When the urease inhibitors were applied at the rate of 50 J,lg/g soil to air-dried soil
amended with mannitol to promote denitrification of nitrate by microorganisms, the
strong inhibitory effect of 2,5-dimethyl-BQ and 2,6-dimethyl-BQ on denitrification
appeared again. Under these conditions, PCMB and CT also retarded denitrification,
whereas HQ, BQ, 2,5-dichloro-BQ, and 2,6-dichloro-BQ inhibited nitrite reduction and
decreased production of N 20. The six phosphoroamides did not significantly affect
denitrification.
Repetition of these experiments with field-moist soil samples, treated with 50 J,lg
urease inhibitor/g soil, gave results partially different from those obtained with air-dried
samples. Thus, the strongest inhibitors of denitrification were 2,5-dichloro-BQ and 2,6-
dichloro-BQ, whereas 2,5-dimethyl-BQ and 2,6-dimethyl-BQ together with five other
338
TABLE 70. Effect of various urease inhibitors on denitrification of nitrate in a cla~ loam soil"
N03·-Nlost N I2roduced !Jlg/g soiQ
COIIllound
(/tg!1l soil) NOi-N Nz()'N Nz-N (NOz·+NzO+Nz!;N
None 109 0 34 74 108
Na p-chloromercuribenzoate (PCMB) 151 85 8 58 151
Catechol (CT) 106 0 34 71 105
Hydroquinone (HQ) 122 3 62 59 124
2,5-Dimethyl-BQ 82 22 2 59 83
2,6-Dimethyl-BQ 85 31 3 52 86
2,5-Dichloro-BQ 106 0 35 72 107
2,6-Dichloro-BQ 115 0 42 75 117
Phenylphosphorodiamidate (PPDA) 107 0 15 93 108
Phosphoryl triamide (PTA) 126 0 26 100 126
N-Phenylphosphoric triamide (PPTA) 97 0 32 66 98
N-(Diaminophosphinyl)benzamide
(DAPBA) 128 0 9 120 129
4-Fluoro-N-(diaminophosphinyl)
benzamide (4-F-DAPBA) 121 0 21 102 123
N-(Diaminophosphinyl)
benzeneacetarnide (DAPBAA) 126 0 0 125 125
"From Yeomans and Bremner (1986), by courtesy of Marcel Dekker, Inc.
urease inhibitors (pCMB, HQ, PPDA, DAPBA, and DAPBAA) enhanced

denitrification. The other compounds did not affect denitrification.
In samples also treated with mannitol, 2,5-dimethyl-BQ and 2,6-dimethyl-BQ were
again the strongest inhibitors of denitrification; CT, 2,5-dicWoro-BQ, 2,6-dichloro-BQ,
PPDA, PPTA, and 4-F-DAPBA had a retarding effect on denitrification. An enhancing
effect was registered with PCMB, HQ, and BQ. Denitrification was not significantly
affected by PTA, DAPBA, and DAPBAA.
Based on these results, one can conclude that the effect of urease inhibitors on
denitrification is largely influenced by the initial moisture content of soil (air-dried or
field-moist) as well as by the presence or absence of an organic compound (mannitol)
promoting denitrification of nitrate by microorganisms. For example, depending on
these conditions, PPDA applied at the rate of 50 Ilg/g soil retarded or enhanced or did
not affect denitrification.
Bremner et al. (1986b) performed similar experiments by using the same three soils
and the same nine phosphoroarnides also used for evaluating their effect on
mineralization of alanine (see Section 8.2.5.2). The methods applied for studying their
effect on denitrification are identical to those described by Yeomans and Bremner
(1986), but only air-dried soil samples were used and there were no treatments with 1
Ilg of phosphoroamide/g soil and with mannitol. It was found that the nine
phosphoroamides studied at rates of 10 and 50 Ilg/g soil had no significant effect on
denitrification of nitrate, except PPTA and 4-F-DAPBA which slightly enhanced
denitrification when applied at the rate of 50 Ilg/g soil (see also Bremner, 1986).
The effect of nBTPTA, PPDA, and HQ on denitrification of N0 3- in a Belgian loam
soil was studied, in several laboratory experiments, by Wang et al. (1991c). The first
experiment was conducted without adding organic matter to the soil samples. Ten rn1 of
KN0 3 solution containing 2 mg ofN and 10 rn1 of water or aqueous solution containing
40 Ilg of test compound were added to 1O-g air-dried soil samples. The 1:2 soi1:water
339
ratio simulated flooded soil conditions. The bottles containing the mixtures were closed
and incubated at 25°C up to 20 days, during which time N03' and N02- contents in
mixtures were periodically determined. Only a small decrease of N03- content was
noted in both presence and absence of urease inhibitors. To show whether the low
denitrification was related to lack of carbon source, 0.03 mg of glucose was added to
each mixture after 20 days of incubation. Addition of glucose resulted in a rapid
decrease ofN03- content, irrespective of the presence of urease inhibitors, which clearly
shows that the original soil did not contain enough readily available carbon for
denitrification and that the urease inhibitors tested did not influence denitrification.
In the second experiment, 1% (on soil weight basis) of ground barley straw was
added to the soil samples. In continuation, the procedure was the same as in the first
experiment. In these mixtures, the loss of N03--N was rapid and, after 2 days of
incubation, reduction of N0 2- and N03- was almost complete in both absence and
presence of urease inhibitors. However, during the first 24 hours, HQ retarded
denitrification by about 20%.
In the third experiment (see also Zhou et al.. 1992), only the HQ was studied. Its
rates were 0, 40, 200, and 400 flgllO g soil. No or 1% barley straw was added to the
soil. Denitrification of N03- was followed by determination of N20. None of the HQ
rates affected emission ofN20 in the no-straw treatments. Thus, in 3 days about 5% of
the added N0 3--N was lost as N 20 from both untreated and HQ-treated soil samples.
When soil + 1% straw mixtures were used, HQ reduced, during the first day of
incubation, the emission ofN20 from 74.75% (no HQ) to 62.50% (40 flg HQ/lO g soil),
52.25% (200 flg HQ), and 36.75% (400 flg HQ). However, this inhibiting effect of HQ
was short-lived, as after 3 days of incubation the N20 loss was always about 98%.
The last experiment differed from the third by using less straw (0.1 %) and assessing
denitrification through analysis ofN03- and N02-. After 5 days of incubation, about half
of the added nitrate was lost in the treatments with 0, 40, and 200 flg HQ/IO g soil, but
only about 20% when the rate ofHQ was 400 flgllO g soil.

Speir et al. (1995) studied the effect of Cr(VI) on soil urease activity (see page 16) and
on other soil biochemical and microbiological properties, including denitrification. The
results indicated that denitrification was more sensitive to Cr(VI) than were
arylsulfatase, phosphatase, and urease activities, microbial biomass and respiration.
8.2.7.2. Effect of Organic Mercury Compounds

The effect of Na p-chloromercuribenzoate (PCMB) on denitrification was studied by
Yeomans and Bremner (1986) (see page 337 and Table 70).
8.2.7.3. Effect ofPolyhydric Phenols and Quinones

Two dihydric phenols (catechol and hydroquinone) and five quinones were studied by
Yeomans and Bremner (1986) (see page 337 and Table 70). Hydroquinone was studied
also by Wang et al. (l991c) (see page 338).
8.2.7.4. Effect of Phosphoroamides

Studying the effect of phenylphosphorodiamidate (PPDA) on the fertilizing efficiency
of urea in a flooded rice field on a clay soil, Simpson et al. (1985) concluded that the
340
major effect of PPDA was not so much to prevent ammonia volatilization but to reduce
the losses caused by denitrification to N2 of the nitrates derived from urea-N (see page
119).
Six phosphoroamides were tested by Yeomans and Bremner (1986) (see page 337
and Table 70), nine by Bremner et al. (1986b) (see page 337), and two by Wang et al.
(1991c) (see page 338).
8.2.7.5. Effict of Combined Urease and Nitrification Inhibitors

In the pot experiments of Zhou et at. (1999), emission of N20 from a flooded rice soil
was very markedly decreased, when the fertilizer urea applied was amended with both
hydroquinone (HQ) and dicyandiamide (DC D), as the N2 0 emission was only one third
of the N20 emission measured in the urea-only treatment. HQ alone and DCD alone
were less effective than the combined HQ and DCD.
8.2.8. Effect on Ndixation

The non-symbiotic Nz fixation was studied.
8.2.8.1. F;Uect ofHeavy Metal Compounds

In studies on the soil biological effects of pesticides, Tu (1990, 1992a,b) used HgCh as
a reference compound and determined its effect not mly on enzyme activities (see
pages 10 and 321), but also on Nz fixation capacity. The method of acetylene reduction
to ethylene (HC=CH ---> HzC=CH z) was applied in soil samples incubated for 2 and 7
days after HgCl z addition.
In samples of a sandy loam soil (PH 7.6) and an organic soil (PH 6.8), treated with
10 f.1g HgCh/g soil, no significant decrease (p=0.05) occurred in the N2 fixation capacity
during 2 and 7 days of incubation (Tu, 1990). When HgCh was added to samples of the
sandy loam soil at a rate of 80 f.1g1g soil, a significant decrease appeared in the N z
fixation after 2 days of incubation and an insignificant increase after 7 days (Tu,
1992a,b).
8.2.8.2. Effect o/Fluorides

In pot experiments, Ablizova and Tomina (1997) studied the effect ofNaF at rates of 0,
10, and 50 mg Flkg soil on urease activity (see page 34) and also on N z fixation capacity
of a chestnut soil. Nz fixation was determined in samples taken in spring and summer.
Increased N2 fixation was recorded in both spring and summer samples that had been
treated with NaF. In the spring samples, the increasing effects of the two NaF rates m
N2 fixation were not significantly different, whereas in the summer samples the
increasing effect of the lower NaF rate was much stronger than that of the higher NaF
rate.
8.2.8.3. F;/Ject ojDithiocarbamates

Tu (1981a) studied the effect of maneb (manganese ethylene-l,2-bisdithiocarbamate)
applied at rates of 0, 5, and 10 f.1g1g soil on urease activity (see page 58) and also on N2
fixation capacity of a clay loam soil (PH 7.2). N2 fixation was measured after 2 and 7
days of incubation. Maneb at 5 f.1g1g soil rate had no significant effect (p=0.05) on N z
fixation after either 2 or 7 days, but at the 10 f.1g/g soil rate maueb had a significant and
341
an insignificant decreasing effect on N2 fixation after 2- and 7-day incubation,

respectively.
8.2.8.4. E;fJect of Thiuram Disulfides

Tu (1981a) studied the effect of thiram (tetramethylthiuram disulfide) on N2 fixation
under conditions identical to those under which he studied the effect of maneb (see the
preceding section). At both 5 and 10 Ilg thiramlg soil rates, significantly and
insignificantly decreased N2 fixation was recorded after 2- and 7-day incubation,
respectively.
8.2.8.5. Effect ofHumic Substances

Ablizova and Tomina (1997) studied the effect of sodium humate (SH) applied at rates
of 0, 0.5, 1, and 1.5 t/ha on soil urease activity (see page 167) and also on the N2
fixation capacity, which was measured in spring and summer samples. Increased N2
fixation capacity was recorded in the spring samples at each rate of SH. In the summer
samples N2 fixation in the SH treatments presented the order: control (no SH added) >
SH 1.5 tlha > SH 1 t/ha > SH 0.5 t/ha.
8.2.8. 6. Effect ofLignosulfonates

Builov et al. (1979) amended 2_m2 microplots installed on a solonetz soil with
ammonium lignosulfonate (ALS) at rates of 0, 1.3, and 1.7% relative to weight of the
O-lO-cm soil layer. The N2 fixation was determined 2 years after ALS application. The
following order ofN2 fixation was recorded: control (no ALS added) < ALS 1.7% «
ALS 1.3%.
8.2.9. Effect on SulfUr Oxidation

Tu (1992b) used HgClz as a reference compound in his pesticide studies. The effect of
HgClz on the capacity of soil to oxidize elemental sulfur was also evaluated. Samples of
a sandy loam soil (PH 7.6) were treated with 80 Ilg HgC12/g soil and incubated for 2 and
4 weeks. It was found after both incubation times that HgCl2 exhibited no significant
changes (p=0.05) in the capacity of soil to oxidize elemental S.
8.2.10. Effect on Adenosine Triphosphate (ATP) Content

The effect of HgCh on ATP content was determined in several studies on the side
effects of pesticides on soil life as in these studies, HgCl2 served as a reference
compound (Tu, 1982, 1990, 1992a,b). Canadian soil samples treated with HgClz and
incubated were analyzed for ATP after 1 and 2 days of incubation.
Addition of HgCl 2 to samples of an organic soil (pH 6.7) at rates of 70 and 150 Ilg/g
soil led to significantly decreased (p=O.05) ATP contents after both 1 and 2 days of
incubation (Tu, 1982). In another organic soil (PH 6.8), HgCh at a rate of 10 Ilg/g soil
caused significant decreases in the ATP content after both incubation times. Addition of
HgCh to samples of a sandy loam soil (PH 7.6) resulted in a significant decrease and an
insignificant increase in the ATP content after 1- and 2-day incubation, respectively
342
(Tu, 1990). However, HgCh added to samples of the sandy loam soil (PH 7.6) at a
higher rate (80 Ilg/g soil) significantly decreased the ATP content after both incubation
times (Tu, 1992a,b).
The heavy metal salts tested by Kandeler et al. (1990) (see page 15) decreased the
ATP content in both Austrian soils studied. The decrease was -60% in a sandy loam
and -30% in a clay loam.
Besides urease and dehydrogenase activities, the ATP content was also determined
in CdS04 -treated samples of two Italian soils (a sandy loam and a sand) studied by
Morano et at. (2001) (see pages 19 and 322). Sensitivity of the microbial ATP
production to Cd in the sandy loam was lower after 7 days than after 3 hours or 28 days
of incubation, whereas this sensitivity in the sand decreased continuously during the
incubation.
8.2.10.2. Effect of Dithiocarbamates

Maneb applied at a rate of 5 Ilg/g soil to samples of an organic soil (PH 6.7) caused a
significant decrease and an insignificant increase in the ATP content after 1 and 2 days
of incubation. respectively. When the maneb rate was 10 Ilg/g soil, the decrease in ATP
content became significant after both incubation times (Tu, 1982).
8.2.10.3. Effoct of Thiuram Disulfides

Thiram was added to samples of an organic soil (pH 6.7) at rates of 5 and 10 Ilg/g soil.
ATP content was determined after 1 and 2 days of incubation. Thiram at the lower rate
led to a significant and an insignificant increase in the ATP content after 1- and 2-day
incubation, respectively. At the higher rate, thiram addition resulted in significant
decreases in ATP content after both incubation times (Tu, 1982). In another organic soil
(pH 6.8), thiram at the rate of 10 Ilg/g soil insignificantly decreased the ATP content
after both 1 and 2 days of incubation, whereas in a sandy loam soil (PH 7.6) the effect
of thiram at a rate of 10 Ilg/g soil on the ATP content was significantly and
insignificantly decreasing after 1- and 2-day incubation, respectively (Tu, 1990).
343
Chapter 9. Use of Urease Inhibitors in the Analysis of Urea and/or Ammonium

from Urea-treated Soils
The urease, present and active in soil, may cause errors in analysis of urea and/or
ammonium, because, acting on urea, the urease splits it and, thus, decreases the real
amount of urea and, by producing NH/, increases the real amount of NH/.
Consequently, it is necessary to inhibit urease, to stop the reaction catalyzed by urease
in soil samples to be analyzed. Only these conditions assure obtaining accurate data on
the amount of urea and/or NH4 + in soil samples submitted to analyses. In other words,
inhibition of urease activity constitutes a step in these analyses.
Table 71 specifies the inhibitors and methods of their application and refers to the
investigators who elaborated the methods of application of urease inhibitors for soil
analyses.
The inhibitor preferred by majority of the investigators for analysis of urea and/or
NH4+ in urea-treated soil is phenylmercuric acetate (5 llg per ml of 2 M KCI solution).
TABLE 71. Compounds used as inhibitors of urease activity in soil samples analyzed for determination of
their content in urea and/or NH/
Compound Method of application Reference
2 3
60 mg ofCuS04 is added to a suspension of Yolk (1966)
60 g of soil in 250 ml of solution made
alkaline with MgO.
0.01 N HCl The soil in an amount equivalent to 10 g of Paulson and Kurtz (1969a)
oven-dry matter is extracted with 100 ml of
IN KCl: 0.01 N HCl solution.
0.1 NHCl It serves as an extractant and inhibitor of Yolk (1970)
HgCh The reaction mixture, prepared from 10 g of Tanabe and Ishizawa (1969)
air-dried soil, 2-4 ml of toluene, 60 ml of
0.2 M phosphate buffer (pH 6.7), and 10 ml
of 10% urea solution, and incubated, is
treated with 10 ml of 1% HgCIz solution
and brought to 200 ml by adding a solution
containing 20 g of KCI.
HgCh The aqueous suspension containing 25 g of Gould et al. (1973)
soil is treated with 0.2 g of CaCh, 0.2 g of
decolorizing caIbon, and 5 ml of 1% HgCh
solution, then enough distilled water is
added to dilute to a total water volume of
100 ml.
To 10 g of soil in 20 ml of aqueous Uoyd and Sheaffe (1973)
suspension is added a HgCh solution with a
final concentration of 0.1 %; KCI is used as
an extractant at a rate of 0.8 glkg soil.
40 ml of2.5 M KCI solution containing 100 Tabatabai and Bremner (1972) (see
Ilg of Ag;,SOJml is added to 5 g of soil in also Tabatabai,1982, 1994)
10 ml of aqueous suspension.
344
TABLE 7L -continued-
2 3
The reaction mixture, prepared from I g of Bums et al. (1978),
air-dried soil, I ml of 0.2% NaN 3 solution, Lethbridge et al. (1980)
2 ml of 0.5 M Tris-maleate buffer (pH 7.0),
and I ml of 6 M urea solution, and
incubated, is treated with 0.5 ml of 10 mM
Ag2SO. solution.
The reaction mixture, prepared from 5 cm.! Schinner and Pfitscher (1978)
of air-dried soil, 0.7 ml of toluene, 9 ml of
0.05 M Tris buffer (pH 9.0), I ml of 0.2 M
urea solution in Tris buffer, and incubated,
is treated with about 35 ml of 2.5 M KCI
solution containing 0.01% AgN03 and
brought with distilled water to 50 ml.
Phenylmercuric 100 ml of2 M KCI solution containing 5 I1g Douglas and Bremner (1970) (see
acetate (PMA) of PMA/ml is added to 109 of soil. also Bremner, 1982;
Tabatabai, 1982, 1994)
20 g of soil is extracted with 2 M Na2S0. Singh et al. (1984)
solution containing 5 j1g of PM AIm I.
For determination of urea and NH.+ in Fillery et al. (1984)
floodwater from rice fields, 18 ml of
floodwater is treated with KCI to clarify the
solution and with PMA to inhibit urease
activity (final concentration: 2 M KCI and 5
j1g PMAlml).
For conservation of urea and NH4 + in Simpson et al. (1985)
floodwater from rice fields, I ml of 1%
H.lP04 and I ml of 1.5 mM PMA solution
are added to 8 ml of floodwater.
The mixture containing 5 g of soil and 2.5 Liao and Raines (1985)
ml of aqueous phase is extracted with 22.5
ml of I N K2S04 solution containing 10 I1g
of PM AIm I.
p-Hydroxymercuri- To 0.2 g of soil is added 0.1 ml of 19 mM Shih and Souza (1978)
benzoate (PHMB) PHMB solution in 25 mM phosphate buffer
Thiourea (TU) (pH 8.5) or 0.1 ml of 2 M TV solution in 50
mM phosphate buffer (pH 7.0).
Phenylmercuric The reaction mixture, prepared from 25 g of Perez Mateos and Gonzalez
borate (PMB) soil, 45 ml of 50 mM Tris buffer (pH 9.0) Carcedo (1988)
and 5 ml of 1-20 mM urea solution, and
incubated, is treated with 175 ml of 2.5 M
KCI solution containing 200 111 ofPMB/1.
Dimethyl-p-benzo- For measuring pH in suspensions of soil- Rachhpal-Singh and Nye (I 984b)
quinone (DBQ)O 0.0 I M CaCh solution 1:5, DBQ is added to
the CaCh solution at a rate of I 0 11g/1.
Hydroquinone (HQ) To I g of soil in 5 ml of aqueous suspension Lichko and Kiselev (1985, 1986)
is added 2.0-2.5 ml of I mM HQ solution.
Phenylphosphoro- I N KCI solution containing 10 I1g of Savant etal. (l987a,b, 1988b)
diamidate (PPDA) PPDAlml is used as an extractant and
urease inhibitor.
The soil sample (100-120 g) is extracted Medina and Sullivan (1986, 1987)
with 250 ml of 2 M KCI solution containing
5 j1g of PPDAlml.
The soil is extracted with 2 N KCI solution Christianson et al. (1990)
containing 0,01 g of PPDA/I. The
soil:solution ratio is 1:3.
'Position of the two methyl groups is not specified by Rachhpal and Nye (1984b).
345
TABLE 71. --cantinueti-

2 3
Phenylphosphoro- The reaction mixture; prepared from 5 g of McCarty et al. (1992)
diamidate (PPDA) soil, 0.2 ml of toluene, and 5 ml of water
containing 2 rng ofurea-N and incubated, is
treated with 45 ml of2 M KCl containing 1
rngPPDAlI.
For conservation of urea in soil extracts Hendrickson and Douglass (1993)
obtained with deionized water (10 ml H20/2
g soil), 0.1 ml of a PPDA solution (0.02 g
PPDAlI) is added to 5 ml soil extract.
347
Chapter 10. Urease Inhibitors Used with Another Purpose than Inhibition of Soil
Urease Activity
10.1. UREASE INHIBITORS AS FERTILIZERS
10.1.1. Inorganic Boron Compounds

In the fertilizer composition patented by Besekau et al. (1974), the boric acid added to
urea serves not only as a urease inhibitor but also as a micronutrient (see page 32).
10.1.2. Inorganic Sulfur Compounds

Ammonium thiosulfate (ATS) is a urease and nitrification inhibitor and an NS fertilizer
(12-0-0-26). Thus, Hensen et al. (1984) found that ATS improved winter wheat
production when topdressed and concluded that ATS is an acceptable S fertilizer source
for maize and winter wheat. Kissel (1984) obtained good results when ATS was banded
below the winter wheat seeds. According to Swan et al. (1986), ATS is an acceptable S
source for rapeseed. The conditions under which ATS improves the winter wheat yields
were studied by Mahler and Lutcher (1989) (see page 263). In a 3-year field experiment
conducted by Graziano and Parente (1996) (see page 253), plots fertilized with ATS
(22.8 kg ATS-Slha) gave 0.5 and 1.2 t maize grain yieldlha greater than plots fertilized
with equal rate of S from single superphosphate in the last 2 years of the experiment.
10.1.3. Hexamethylenetetramine
As shown on page 47, hexamethylenetetramine (HMTA) was patented as an inhibitor of
soil urease activity by Neumann and Richter (1976). In winter wheat field experiments
conducted by Verstraeten and Livens (1975, 1977), HMTA was found to be a slow-
release N fertilizer. HMTA displayed a fairly high N efficiency and reduced the
incidence of mildew disease (Erysiphe graminis). HMTA is gradually mineralized in
soils under both aerobic conditions (Verstraeten, 1977) and water-logged conditions
(Taslim and Verstraeten, 1977). Mineralization of HMTA added to samples of seven
Belgian soils in amounts of 100 to 200 ppm N and incubated under aerobic conditions at
10 or 30°C for 4 weeks ranged between 60 and 90% compared with urea mineralization.
Under water-logged conditions, a clay loam soil was studied. In one of the experiments,
rate ofHMTA or urea addition was 100 ppm N, the incubation took place at 30°C and
lasted 45 days. Release of ammonium from HMTA was highest at day 15 and
represented -50% of the added HMTA-N, whereas -90% of the added urea-N was
hydrolyzed during 3 days of incubation. In other words, HMTA is mineralizable also
under anaerobic conditions, but this process is slower than hydrolysis of urea.
In a 3-year (1991-1993) field experiment conducted by Borovskii and Yanishevskii
(1994), HMTA, applied as a nitrogen fertilizer for vegetable crops on an alluvial
meadow soil and a soddy-podzolic soil in the Moscow region, produced high crop
yields. Moreover, the crops produced with HMTA were of an excellent quality as their
nitrate content was smaller than that of crops obtained with urea or ammonium nitrate.
The three fertilizers were administered at the same yearly rate: 140 kg Nlha in 1991 and
120 kg Nlha in 1992 and 1993.
10.1.4. Urea Derivatives

Data will be presented on thiourea and biuret (urease and nitrification inhibitors).
348
Based on the results of pot experiments, Kolyada (1970, 1973) drew the conclusion
that thiourea was an efficient N fertilizer when applied 8-16 days before planting the
soil to barley, oats or radish. During this period, thiourea lost phytotoxicity due to its
decomposition.
In a laboratory experiment, Sahrawat (1981) proved that biuret was mineralized
under both water-logged and aerobic conditions. In soil samples amended with 100 mg
biuretlkg soil and incubated at 30°C for 5 weeks, the biuret-N was mineralized to
NH4 +-N in a proportion of 48.3% under water-logged conditions and to N0 3·-N in a
proportion of 18.3% under aerobic conditions. Thus, biuret may be considered a slow-
release N fertilizer.
10.1. 5. Phosphoroamides
Phosphoryl triamide [P(O)(NH 2)3; PTA] and some other phosphoroamides were
patented as NP fertilizers by Pellegrini (1965, 1968), Wanek et al. (1966), Fiedler et al.
(1974b) and studied by other investigators as well.
Pellegrini (1965) conducted a field experiment to evaluate the efficiency of PTA as
a NP fertilizer. The test plant was winter wheat. The plots were installed on a silt loam
soil (PH 8.4). Urea and Ca(H 2P04)2 served for comparison. Rates of additions were
191.4 kg P20s/ha and 102 kg N/ha. All fertilizers were surface-applied in mid-February
at the end of the emergence of young plants. The grain yields in the different treatments
presented the order: Ca(H2P04h < urea+ Ca(H 2P04)2 ::::: urea « PTA.
Based on the patent of Wanek et at. (1966), Ambroz et al. (1970) compared the
nitrification of PTA with that of (NH4)2HP04. These compounds as sole N sources were
added to a liquid nutrient medium at rates of 0.01, 0.02, and 0.04% N. The medium (20
ml) was inoculated with 0.5 g soil (rendzina, pH 7) and incubated at 28°C for 14 days,
during which the N03·-N content was determined at 3-day intervals. The results showed
that the two compounds were nitrified to the same extent.
Pellegrini (1968) commented on his own patent (Pellegrini, 1965) by pointing out
two disadvantageous properties of PTA: very different mobility in different soils and
weak stability on storage when exposed to air. In his new patent six
alkylphosphoroamides, namely two phosphoromonoamidate (PA) and four phosphoro-
diamidate (PDA) compounds are nominalized as NP fertilizers: dimethyl-PA, methyl-
ethyl-PA, methyl-PDA, ethyl-PDA, n-butyl-PDA, and 2-ethyl-n-hexyl-PDA. All these
compounds labeled with 32p showed a better mobility in the nine soils studied as
compared with the mobility of Ca(H 2P0 4 )2 or with that of PTA. In a pot experiment,
they were tested as P sources for common beans at a rate of 50 kg P20 5/ha, and it was
found that they performed better than Ca(H2P0 4 h.
Wakefield et al. (1971) carried out greenhouse experiments. In one of the
experiments, thiophosphoryl triamide [P(S)(NH 2)3; TPTA] was tested as a source of N
and P for two successive crops of maize on a silt loam limed to pH 6.4. The uptake ofN
and P from TPTA was the same as that from ammonium nitrate and superphosphate.
The yield of dry forage was less from TPTA, however, especially at the high application
rates because of a slight toxicity to the first crop. This toxicity soon disappeared, and no
abnormalities were observed in the second crop. It was found in another experiment that
three derivatives of TPTA, sodium diamidothiophosphate [NaOP(S)(NH 2)2], diam-
monium monoamidothiophosphate [(~OhP(S)NH2]' and diammonium thiophosphate
[(~O)2P(S)OH], were also effective Nand P sources, but condensation products of
349
the pyrolysis of TPT A were less effective and initially toxic. These investigations were
also referred to by Sheridan (1970).
Fiedler et al. (l974a) conducted pot experiments to evaluate ethylphosphoro-
diamidate (EPDA) - C2HsOP(O)(NH2)2; phenylphosphorodiamidate (PPDA) -:-
C6HsOP(O)(NH2h; diethylphosphoroamidate (DEPA) - (C2HsO)2P(O)NH2; and
dimethylthiophosphoroamidate (DMTP A) - (CH30)2P(S)NH2 as NP fertilizers and
triethylphosphate - (C 2HsO)}P(O) as a P fertilizer. Diammonium phosphate (DAP) -
(NH4)2HP04 was used as a reference NP fertilizer. The test plants were Italian rye grass,
oats, and mustard. Three soils were used: a neutral loess loam, and acid and a neutral
heath sand. The pots contained 3 kg loam + 4 kg quartz sand, 3.5 kg sand + 4 kg quartz
sand, and 3 kg sand + 3 kg quartz sand, respectively. Rates of P and N additions/pot
were: 120 mg P + 1,200 mg N (P:N=I:lO) or 240 mg P + 1,200 mg N (N:P=I:5). To
achieve these ratios, urea or ammonium nitrate was also added with the test compounds.
Each pot also received K (0.66 g), Mg (0.1 g), and micronutrients (Mo, Cu, Zn, B, and
Mo). The crop yields and P and N contents in plants were determined, and the
phytotoxicity of the compounds tested was also evaluated.
The results led to the following conclusions: a) EPDA exhibited the best fertilizer
effect; b) with increasing degree of esterification, replacement of ethyl group by the
phenyl group and of the PO group by the PS group, the compounds became less
effective in supplying the plants with physiologically active P and their phytotoxic
effect increased; c) if homogeneously distributed in acid to neutral, non-calcareous soil,
the effect of EPDA was comparable with that of DAP, whereas in calcareous soils
EPDA performed better than DAP due to its better mobility.
Fiedler et al. (1974b) patented several alkylphosphoric triamides (alkyl-PTAs) as
NP fertilizers and tested them in pot experiments.
In one of the experiments, the test plant was Italian ryegrass grown on three soils (a
loess loam, pH 7.0, a sand, pH 5.5, and another sand, pH 7.0). The compounds tested
were: dimethyl-PTA - (CH3hNP(O)(NH2h; diethyl-PTA - (C 2HshNP(O)(NH2)2; and
bisdimethyl-PTA - [(GI3hN)zP(O)NH2' Diammonium phosphate (DAP) - (NH4hHP04
was the reference NP fertilizer. All fertilizers were surface-applied. The crop yields
obtained with these three alkyl-PT As were comparable with that produced with DAP on
the loam and sand (pH 7.0). In the other sand (pH 5.5), the alkyl-PTAs performed a
little better than DAP. Imidodiphosphoric tetraamide - [(NH2)2POhNH was also tested
with Italian rye grass on the loam and sand (pH 5.5). This compound increased the yield
on the loam and decreased it on the sand in comparison with the yield registered with
DAP.
In another experiment, dimethyl-PTA and bisdimethyl-PT A were tested with oats
grown on a loess loam (pH 6.3). The grain yields had the following relative values:
100% (DAP), 151% (dimethy-PTA), and 140.5% (bisdimethyl-PTA). The P content of
grains was also increased by both PTA compounds.
Some alkyl-PTAs, e.g., bisdiethyl-PTA - [(CzHS)2NhP(O)NH2 and hexamethyl-PTA
- [(Oi,hNhP(O) were much less effective than DAP and very toxic and, therefore,
excluded as NP fertilizers. Thus, the effectiveness of alkyl-PTA compounds as NP
fertilizers and their toxicity are determined by the number and kind of their alkyl
groups.
The studies on the fertilizer effect of PTA compounds on oats, briefly described in
the patent of Fiedler et al. (1974b), were described in details by Fiedler et al. (1975).
350
Additionally, phosphoryl triamide (PTA) and triurea phosphate - (lhNCONH)3P(O)

were also tested, and mustard was also used as a second crop. The experimental
conditions were the same as in the experiments of Fiedler et al. (1974a) (see above in
this section).
In comparison with DAP, the readily hydrolyzable compounds, such as PTA and
dimethyl-PTA, performed better in neutral and slightly acid soils, respectively, whereas
the less hydrolyzable compounds, such as bisdimethyl-PTA and diethyl-PTA were more
effective in acid soils. Phytotoxicity of bisdiethyl-PTA and hexamethyl-PTA was
reiterated. Triurea phosphate was also less effective than DAP.
In comparison with oats, the mustard plants were less able to respond by increased
yields to any of the PTA compounds tested.
Calancea et al. (1990) and Calancea and Chiriac (1996) compared the effects of urea
and PTA on Italian ryegrass in pot experiments. Each pot contained 2 kg of soil
(alluvial podzolized clay, pH 5.8) mixed with 1 kg of sand. 15N-Labeled urea and PTA
(at 4.885 and 5.000 excess% of 15N, respectively) were surface-applied at rates of 0, 50
up to 500 mg N/pot (equivalent to 75 up to 750 kg Nlha). In the urea treatment, P
fertilizer was also applied in form of Ca(H2 P04 )z in a single dose (100 mg P/pot). Soil
humidity was maintained at 65-70% of WHC.
During the growth period, no phytotoxic effect was observed in either urea or PTA
treatment. Dry matter yields increased proportionately with rates of urea and PTA; the
yield-increasing effect of PTA was similar to that of urea. Total N uptake by plants and
N uptake (and N loss) from the soil reserve were higher and N uptake from fertilizer
was lower in the PTA than in the urea treatment. However, at the highest fertilizer rate
(500 mg N/pot), the coefficient ofN utilization by plants was higher in the PTA than in
the urea treatment.
10.1.6. Cyclophosphazene Compounds

Hexaaminocyclotriphosphazatriene (HA-CTPAT), also called phosphonitrilic hexamide
- [NP(NH 2 )Zh, was patented as an NP fertilizer by Wanek et al. (1966) and Illarionov et
al. (1968). Pechkovskii et al. (1983) patented a technology for cogranulating urea and
HA-CTPAT or octaaminocyclotetraphosphazatetraene - [NP(NH2)z]4. The product
obtained contained 0.3 -2.0% HA-CTPAT and was more stable than the urea granules.
Ondracek et al. (1970a,b) found, in comparative studies, that performance of HA-
CTPAT was similar to that of other NP fertilizers. Ambroz et al. (1970) proved in
microbiological experiments that HA-CTPAT was nitrifiable to the same extent as
diammonium phosphate. Wakefield et al. (1971) also evaluated HA-CTPAT and the
octaamino derivative as NP fertilizers.
Davidescu and Chirca (1977) presented a new complex NPK fertilizer containing
potassium salts of cyclophosphazanic acids - nK[HNP(O)O]n, with n=3 or 4. The new
fertilizer was tested in a pot experiment. A mixture of 4 kg chernozemic soil and 4 kg of
sand was added to each pot. There were four variants: I. control (no fertilizer added); II.
ammonium nitrate (AN) + superphosphate (SP); III. AN + SP + phosphazanic fertilizer;
and IV. AN + phosphazanic fertilizer. The test plants were soybean and common bean.
Their growth period was 4 May-l October.
The best result (a 10% increase in the yield of beans) was obtained in variant III, i.e.,
when the phosphazanic fertilizer was applied together with ammonium nitrate and
superphosphate. The phosphazanic fertilizer did not bring about any changes in the
351
chemical composition of beans, with the exception of a slight decrease in the lipid
content.
At the Tennessee Valley Authority, Muscle Shoals, Alabama, HA-CTPAT was
studied initially, more precisely before discovering its capacity to inhibit soil urease
activity (Medina and Sullivan, 1986, 1987) as a potential NP fertilizer (Anonymous,
1985b).
In a pot experiment, Calancea et al. (1986) used HA-CTPAT labeled at the level of
NH2 groups or at the cycle, P3N3C5NH2)6 or P3 15 N3(NH2)6. Maize was sown in a weakly
podzolized soil (1,4, and 7 kg soil/pot). Rates of HA-CTPAT addition were 50, 100,
200, 300, and 400 mg N/pot. Beginning with the 12th day of growth up to the 69th day,
the plants were systematically sampled for determination of their dry weight and total N
and 15N contents.
It was found that in the soil treated with HA-CTPAT the plants grew slowly and
took up less 1~ during the first 45 days, but in the 46-69-day period they grew
vigorously and took up more 15N.
In the next growing season, the experiment was repeated by submitting the same soil
samples to the same treatments as in the first experiment. The maize plants in this
second culture were examined like those of the first culture.
It became evident that plant growth and uptake of N were more marked in the
second culture than in the first and significantly correlated with the rate ofHA-CTPAT.
Of the 15N taken up, 70-80% represented the NH2-N and 20-30% the cyclic-No But the
plants always took up more N from the soil reserve than from HA-CTPAT. One can
deduce from these findings that HA-CTPAT is decomposed slowly in the soil; at the
beginning, the compound is deaminated which is followed by decomposition of the
cycle. Nitrogen (and phosphorus) are released in plant-available forms.
This conclusion was confirmed in a newer pot experiment using the two 15N-labeled
forms of HA-CTPAT (Calancea and Chiriac, 1993). Total N content in the maize plants
(100%) originated from the soil reserve and HA-CTPAT in proportions of
67.50±10.94% and 32.50±10.94%, respectively. Of the total N, N from 1~2 and
cyclic- 15N represented 20.58±7.56% and 11.92±3.87%, respectively.
Italian ryegrass was the test plant in another pot experiment (Calancea et al., 1990).
The effect ofHA-CTPAT on dry matter yield was compared with that of urea. Rates of
additions/pot containing 2 kg of alluvial podzolized clay, pH 5.8 mixed with 1 kg of
sand were 0,50, 100,200,300,400, and 500 mg N. In the urea treatment, Ca(H2P04)2
(100 mg P/pot) was also added.
Dry matter yields increased with increasing rate of both fertilizers, but the increase
was higher with urea than with HA-CTPAT at rates of 50-400 mg N/pot, and the same
increase was recorded with urea and HA-CTPAT at the highest N rate. When HA-
CTP AT was applied at lower rates, the plants took up N mostly from the soil reserve but
at its higher rates the plant uptake of N from this fertilizer was more pronounced than
from urea.
10.1. 7. Plant Materials

Ketkar and Ketkar (1995) consider neem seed crush and deoiled cake as both
nitrification (and urease) inhibitors and as organic fertilizers.
352
10.2. UREASE INHIBITORS USED FOR CONTROLLING AMMONIA (AND

ODOR) EMISSION FROM LNESTOCK WASTES
The observations of Berg (1977) that sarsaponin reduced volatilization of ammonia

from poultry manure are mentioned in Section 2.31.3.2.
Strumpf et al. (l981a,b) patented ro-(naphthoxy)alkanohydroxamic acids and
dihydroxamic acids as inhibitors of soil urease activity (see pages 64 and 65) and also
for controlling ammonia (and odor) emission from livestock wastes. However, no
example is given in the patent descriptions on the urease-inhibiting capacity of these
compounds when added to livestock wastes.
Sallade and Sims (1992) used sodium thiosulfate (STS) for inhibiting mineralization
of N and nitrification in poultry manure. A laboratory incubation experiment was
carried out.
Two poultry manure (PM) samples were used: PM-l and PM-2 had pH 8.7 and 7.4,
and a total N content of 5.5 and 4.6%, respectively. PM was added to a soil-sand
mixture at a rate of 2.7 Wkg soil-sand, and the rate of STS addition was 64 mg S/kg
soil-sand. The controls received no PM and/or no STS. The mixtures were moistened to
field capacity and incubated at 25°C for 12 weeks. During incubation, the mixtures were
analyzed periodically for NH/-N, N03~, and N02~' The analytical data served for
calculation of the percent inhibition of net mineralization of N and nitrification.
Net mineralization of N from PM was reduced by the use of STS, particularly from
PM-1 where the percentage of N mineralization decreased from 42% after 4 weeks of
incubation to 3% after 12 weeks. Inhibition of nitrification by STS averaged in manures
89% after 4 weeks and only 20% at week 12.
In another experiment, Sallade and Sims (1994) used ammonium thiosulfate (ATS).
Soil columns (75 cm long and inner diameter 19.6 cm) were packed with a reconstituted
profile of a loamy sand (PH 6.5 in the 0-30-cm, 4.3 in the 30-60-cm, and 4.0 in the 60-
90-cm layers) and amended with PM (PH 8.7; total N content 5.49%) or urea-
ammonium nitrate (UAN). PM was incorporated into the top layer of columns at the
time of packing at a rate of 3 Wkg soil. ATS provided 14 mg Nand 32 mg S/kg soil.
The UAN columns received per kg soil 10 mg N as AN and 100 mg N as DAN, while
the UAN+ATS columns were treated with 10 mg N as AN + 86 mg N as DAN + 14 mg
N + 32 mg S as ATS. Maize was grown in the columns to create a dynamic soil/plant
system. Three leaching (simulated rain) events were used during the experimental
period: 28 and 49 days after planting and the day after the plants were harvested (10
weeks). The column leachates and soil were analyzed for NH.t+-N and N0 3-N, and the
total N content in plants was also determined.
The analytical data indicated that the use of ATS with PM and DAN did not result in
significantly decreased N leaching or increased N uptake by plants. These results are in
contrast with those obtained by Sallade and Sims (1992) in the laboratory incubation
experiment in which PM and STS were tested (see above in this subchapter).
Varel (1996) obtained unsatisfactory results with acetohydroxarnic acid (AHA),
because for stopping of urea hydrolysis in cattle waste slurries gram quantities of AHAIl
slurry were required; thus, use of AHA was not practical. In contrast, Varel (1997)
registered good results with phenylphosphorodiamidate (PPDA) and cyclohexyl-
phosphoric triamide (CHPTA).
353
One-l slurries of cattle and swine wastes (1: 1 g:g feces to urine) were treated with
PPDA or CHPTA and incubated at ambient temperature (22-25°C). No inhibitor was
added to the controls. During incubation, all slurries were analyzed periodically for urea
and ammonia. Different amounts of inhibitors were applied per 1 slurry: 10 mg PPDA,
10 and 40 mg CHPTA, and 10 mg CHPTA weekly - incubation time: 28 days (both
cattle and swine wastes); 10,40, and 100 mg PPDA applied weekly for 7 weeks -
incubation time: 70 days (cattle waste) and 84 days (swine waste).
With cattle waste (3.3 g ureall) and swine waste (4.8 g urea/l), both inhibitors at 10
mg/l slurry prevented hydrolysis of urea for 4-11 days, and then a gradual hydrolysis
occurred until complete at day 28. Hydrolysis of urea in untreated cattle and swine
wastes (controls) was complete within 1 day. Addition of inhibitors once per week was
the most effective method of preventing urea hydrolysis. Weekly addition of 10,40 or
100 mg PPDAIl cattle waste slurry (5.6 g urea/I) prevented 38, 48, and 70% of the urea
from being hydrolyzed after 28 days, respectively. With swine waste slurry (2.5 g
ureall), these PPDA concentrations prevented 72, 92, and 92% of the urea from being
hydrolyzed after 28 days, respectively. As additions of PPDA were stopped after 7
weeks, all urea at the three PPDA concentrations was hydrolyzed after 70 and 84 days
for the cattle and swine wastes, respectively.
The conclusion was drawn that use of the inhibitors makes possible a significant
control of ammonia emission from livestock wastes and an increase in fertilizer value of
wastes by improving the N to P ratio for plant growth.
Sotomura et al. (2000) patented a deodorant composition comprising extracts from
plants (especially pine or green tea), a urease inhibitor (e.g., thiourea, boric acid,
p-benzoquinone or tannic acid), a lipase, and a lower alcohol such as methanol, ethanol
or propanol. The composition is durable and effective for removing odorous gases
(especially, ammonia, trimethylamine, butyric acid or propionic acid) from pet manure,
etc.
10.3. UREASE INHIBITORS USED FOR PREVENTING TOXICITY OF UREA-

AMENDED FODDERS
Baintner (1964) and Tang! and Baintner (1969) used acetohydroxamic acid (AHA) to
reduce susceptibility of goats to ammonia poisoning induced by the feeding of large
quantities of urea. Larger amounts of urea were tolerated by the animals when urea was
fed with AHA than when it was fed alone. Thus, the goats to which the lethal dose of
urea (150 g) was added with AHA did not die. It was also found that in the presence of
AHA only a small proportion of urea-N was absorbed as ammonia; this suggested that
AHA inhibited the activity of rumen urease. The inhibiting effect of AHA on rumen
urease was also directly demonstrated in in vitro experiments. In other experiments,
sheep with fistulated rumen were used. When urea in a dose of 30 g without AHA was
infused into the rumen, the animals showed symptoms of severe poisoning or died. No
symptoms of poisoning were observed when 30 g of urea was introduced with 13 g of
AHA. In this case, hydrolysis of urea was retarded from 4 hours 15 minutes to 13 hours
15 minutes. Finally, it was emphasized that only pure AHA should be used because
hydroxylamine present as an impurity has poisoning and even lethal effect as it causes
methaemoglobinemia. The pure AHA is a costly product. Therefore, the perspective for
using AHA as an additive to fodder of ruminants is questionable on economic grounds.
354
Jones (1968) performed in vitro experiments. Washed suspensions of bovine rumen

microorganisms were used. When these suspensions were incubated with AHA, the
urease activity was depressed. It was also found that AHA was slowly degraded by the
rumen microbiota. Another finding was that AHA inhibited production of volatile fatty
acids by the rumen microorganisms cultured on cellulose-containing nutrient media.
In the experiments of Jones and Milligan (1975) and Whitelaw et at. (1991), AHA
was not used successfully to block urease activity in sheep rumen, because the
microbiota adapted to it. Absorption of AHA by the rumen was also demonstrated.
Hartbrich et al. (1976a) and Chemische und Pharmazeutische Fabriken (Magdeburg,
Germany) (1977) patented 27 phosphoroamides (3 phosphoroamidates, 18 phosphoro-
diamidates, and 6 thiophosphorodiamidates) as additives to solid and fluid urea-
containing fodders and fodder mixtures for ruminants. By inhibition of urease of rumen
bacteria, the too rapid hydrolysis of urea is prevented. Thus, the possibility of poisoning
of animals due to ammonia released in great amounts from urea is eliminated, and, at
the same time, the utilization of urea-N becomes more efficient. These additives are
recommended for practice at the preferred rates of 0.05-1 % relative to urea-No
The examples given in the patent descriptions refer to an artificial rumen system
using coarsely filtered natural rumen fluid taken from fistulated cows. Testings of the
capacity to inhibit rumen urease activity are described for 15 inhibitors patented. In
most cases, phenylphosphorodiamidate (PPDA) was tested.
Voigt et al. (1980a,b) used lactating dairy cows with fistulated rumen and
duodenum. Urea solution with and without PPDA was infused into the rumen. lsN_
Labeled urea was used. Besides determination of urea and ammonia contents and pH in
the rumen fluid, the contents of trichl oroacetate (TCA)-soluble and TCA-precipitable N
compounds, as well as total N contents in urine and feces and protein-N content in milk,
were also determined.
Three treatments were applied: I. urea alone (180 glday); II. urea + PPDA (1 % of
urea-N) added to animals not adapted to PPDA; and III. urea + PPDA added to animals
after their 30-day adaptation to PPDA.
The urea content in the rumen fluid 2 and 6 hours after addition of urea and
urea+PPDAhad the following values (mgllOO ml fluid) in the three treatments: 6.0 and
1.5 (I), 57.9 and 17.2 (II), and 20.9 and 0.9 (III), respectively. The corresponding values
for the ammonia content (mgllOO rnl fluid) were: 61.5 and 3.5 (I), 5.3 and 2.6 (II), and
23.9 and 1.3 (III), respectively. The pH values measured 2 hours after urea and
urea+PPDA additions were: 6.8, 6.2, and 6.3, respectively. These data prove that the
effect of PPDA to inhibit rumen urease activity was stronger in animals not adapted to
PPDA than in those adapted to PPDA during 30 days.
It resulted from the other determinations that utilization of urea-N was most
effective in the animals adapted to PPDA, i.e., when the rumen urease was not strongly
inhibited by PPDA. The conclusion was drawn that in adapted animals efficiency of
urea can be improved by PPDA.
Whitelaw et al. (1991) found that PPDA used at concentrations inhibiting urease
activity in sheep rumen was not toxic to animals, had no antibacterial effect, and did not
present any environmental problems.
Selivanov (1986) used cyanuric acid (H2N-CO-NH-CO-N=C=O; CUA) for
inhibition of urease activity in sheep rumen. Rumen fluid was collected from animals
adapted and non-adapted to CUA. Their fodder had been amended with urea or with
355
urea + CVA. Reaction mixtures, prepared from rumen fluid and urea (53 mg%) or urea
+ CUA (4 mg%), were incubated for 6 hours and analyzed for urea and ammonia at 1-
hour intervals. In the absence of CVA, urea was completely hydrolyzed in 4 and 5 hours
in rumen fluid of the non-adapted and adapted animals, respectively. In reaction
mixtures with urea+CVA, urea at concentrations of 8.6 and 3.4 mg% remained
unhydrolyzed in the rumen fluid of the non-adapted and adapted animals, respectively,
after 4 hours of incubation. In other words, CUA was more inhibitory in the rumen of
non-adapted than adapted animals.
There are reports showing that Yucca schidigera extracts, containing sarsaponin and
sarsaponin fractions, improved performance and health in ruminants by inclusion of
these extracts in fodders at rates of 100-250 glt fodder, and the effect was attributed to
inhibition of urease activity (e.g., Ellenberger et al., 1984). Killeen et al. (1994) studied
the effect of a commercial Yucca schidigera extract on urease of a bacterium (Bacillus
pasteurii) and on the ~-galactosidase of the fungus Aspergillus oryzae. It was found that
the effect of the extract on urease was not specific as ~-galactosidase was also inhibited.
Another finding was that the urease-inhibiting effect of the extract was much too low to
account for the in vivo effects of the Y. schidigera extracts at fodder inclusion levels as
little as 100 giL
Ludden ef al. (2000a,b) conducted in vitro and in vivo experiments using N-(n-
butyl)thiophosphoric triamide (nBTPTA) as a urease inhibitor.
The in vitro experiments were carried out with steer rumen fluid. The reaction
mixtures, prepared in test tubes, contained rumen fluid, ground fescue hay or ground
fescue hay and ground maize in 1:1 mixture, urea with and without nBTPTA and were
incubated at 39°C for 6 or 48 hours. During and after incubation, they were analyzed for
urea, ammonia, and volatile fatty acid (VFA) contents and fiber digestibility (FD).
nBTPTA decreased rate of urea hydrolysis and, consequently, formation of ammonia.
Total VFA concentration was not affected, but acetate/propionate ratio and FD were
decreased by nBTPTA. The conclusion drawn from the in vitro experiments was that
nBTPTA can be used to decrease the rate of ammonia release from dietary urea and
offers a way to improve urea-N utilization in ruminants.
In the in vivo experiments, ruminally cannulated lamb wethers were used to
investigate the chronic effect of nBTPTA on ruminal N metabolism and N balance. In
one of the experiments, the animals were given into the rumen 0 or 0.125 up to 4 g
nBTPTA daily and fed a cracked maizelcotton seed hull diet containing 2% urea twice
daily at 2.5% of initial body weight for 15 days. nBTPTA inhibited ruminal urease
activity and thus decreased the rate of ammonia formation, but this effect of nBTPTA
diminished as the experiment progressed. On day 15, no differences were found in the
VFA concentration and in FD between the treatments with and without nBTPTA.
However, nBTPTA increased urinary N excretion and thus decreased N retention. In
contrast to the results obtained in the short-term in vitro experiments, the results of the
in vivo experiments indicated that the nunen microbiota was able to adapt to chronic
nBTPTA administration, thereby limiting its practical use in improving the utilization of
dietary urea.
356
10.4. UREASE INHIBITORS USED FOR TREATMENT AND PREVENTION OF

SOME HUMAN DISEASES
The bacteria producing urease implicated in the genesis of some human diseases infect
the urinary tract (containing substantial amounts of urea) or the gastrointestinal tract
(containing limited amounts of urea - its concentration in normal stomach is of about 3
rnM).
The urease of urophathogenic bacteria is directly associated with the formation of
infection stones [urinary and renal stones (calculi); urolithiasis and nephrolithiasis] and
contributes to the pathogenesis of acute pyelonephritis and urinary catheter encrustation.
Urease of the bacteria infecting the gastrointestinal tract generates gastritis and
peptic ulcer and contributes to the pathogenesis of hyperammonemia, hepatic
encephalopathy, and hepatic coma.
Some of these diseases also affect animals.
The infection stones are a mixture of struvite (MgNH4P04.6H20) and carbonate
apatite [Ca\O(P04)6.C03]. The ammonia released by bacterial urease-catalyzed
hydrolysis of urea increases the pH from 6.5 to 9.0, at which the normally soluble
bivalent cations become supersaturated and crystallize forming the stones. In humans,
two urinary tract bacteria, Proteus mirabilis (P. mirabilis) and Ureaplasma urealyticum
(U urealyticum), are the most common bacteria implicated in stone formation. Other
Proteus species (e.g., P. morganii) and bacteria belonging to other genera such as
Klebsiella. Morganella, Providencia are also implicated in stone formation.
In dogs, struvite stone formation is associated with Staphylococcus aureus. Bovine
pyelonephritis is caused by Corynebacterium renale that possesses a potent urease.
In humans, the bacterium implicated in development of gastritis and peptic
ulceration is Helicobacter pylori (H pylori) (formerly Campylobacter pylori and C.
pyloridis). It is an acid-sensitive bacterium which multiplies in a pH range of 6.9 to 8.0.
H pylori is uniqely adapted for surviving in the highly acidic environment of human
TABLE 72. COJ1llounds patented as inhibitors of bacterial urease implicated in the genesis of some human
diseases
Urease inhibited/ Disease
COIl1lound(s) Reference
treated and prevented
[[(4-Aminophenyl)sulfonyl)aroino)phenyl P. mirabilis urease! Alaimo et al. (1980)
phosphorodiamidates Urinary tract infections
8-[(4-Aminophenyl)sulfonyl)amino-2- P. mirabilis urease! Alaimo and Millner
naphthalenyl phosphorodiamidate Urinary stones (1980)
N-(Diaminophosphinyl)arylcatboxarnides P. morgan;; urease! Bayless and Millner
Urinary stones (1980a)
Phosphorotriamides P. mirabilis urease! Bayless and Millner
Urinary stones (1980b)
Hydroxamic acid glycoside derivatives Fecal urease! Ito et al. (1994)
Hyperamrnonemia
2-Benzamido-3-carbostyrylpropanoic acids and H. pylori urease! Yamazaki and
their salts (rebamipide) Gastric IlUICOSal disoniers Oosaka (1995)
2-[4-(3-Methoxypropoxy)-3 -methylpyridin-2-yl)- H. pylori urease! Tsuchiya et al.
methylsulfinyl-IH-benzimidazole Hyperamrnonemia, (l995a)
hepatic encephalopathy
Proanthocyanidins (e.g.• from pine bark or grape H. pylori ureaselUlcer (rat Salo (2000)
seed) + catechin and epicatechin model)
357
stomach. It produces a potent urease. Due to a very high affmity for urea, the H pylori
urease is able to hydrolyze the limited amounts of urea in stomach and to produce a
"cloud" of ammonia that protects the bacterium from stomach acid and enables it to
colonize and damage the gastric mucosa. Being adjacent to mucosa, H pylori becomes
able to scavange urea from the blood. Thus, urease in an important virulence factor for
Hpylori.
As the bacterial urease is implicated in the genesis of the diseases mentioned, for
their treatment and prevention inhibitors of urease activity were used (Rosenstein and
Hamilton-Miller, 1984; Mobley and Hausinger, 1989; Park et al., 1996).
First, the compounds patented as inhibitors of bacterial urease implicated in the
genesis of some human diseases are presented (Table 72). Then examples are cited from
TABLE 73. Studies on the inhibition of bacterial urease and therapeutic and prophylactic effect of urease
inhibitors in some human diseases
Urease inhibited/Disease treated
Inhibitor(s) Reference
and prevented
Acetohydroxamic acid (AHA) Hyperammonemia Fishbein et al. (1965)
AHA Mucosal and fecal urease Aoyagi and Summerskill
(1966)
AHA Hyperammonemia Summerskill et al. (1967)
AHA Renal stones Griffith et al. (1979)
AHA Renal stones Martelli et af. (1981)
AHA Renal stones Williams et al. (1984)
AHA Urinary catheter encrustation Bums and Gauthier (1984)
AHA Urinary stones Griffith et al. (1988)
AHA H. pylori urease Goldie el al. (1991)
Hydroxamic acid derivatives (HADs) Urease of T -strain mycoplasmas Ford (1973)
(u. urealyticum)
HADs Urinary stones Kobashi et al. (1980)
HADs Urinary stones Munakata et al. (1980)
HADs H. pylori urease/Gastritis, Odake el al. (1994)
peptic ulcer
HADs Urinary stones Abou-Sier et al. (1995)
Hydroxyurea (HU) Urinary stones Carmignoni et al. (1980)
HU Renal stones Martelli et af. (1981)
N-(Diaminophosphinyl)-4-tluoro- Urinary stones Millner et al. (1982)
benzamide (tlurofamide)
Flurofamide H. pylori urease Kohler et al. (1995)
N-Acyl phosphoric triamides Urinary stones Takabe et al. (1984)
4-Substituted H. pylori urease Faruci el al. (1995)
phenylphosphorodiamidates
Proton pump inhibitors (omeprazole, H. pylori urease, P. mirabilis Tsuchiya et al. (l995b)
lansoprazole. rabeprazole) urease/Gastritis, peptic ulcer
Omeprazole H. pylori urease Bugnilo et af. (1993)
Lansoprazole, omeprazole H. pylori urease Nagata et al. (1993)
Omeprazole, lansoprazole H. pylori urease KOhler et al. (1995)
Rabeprazole H. pylori urease Park et al. (1996)
Ebrotidine (antiulcer drug) H. pylori urease Pietrowski et al. (1995)
Ecobet (antiulcer drug) H. pylori urease Ito et al. (1998)
Ethylalcoholic herb extracts H. pylori urease Imamura et al. (1995)
Peptides (a 24-mer and a 6-mer H. pylori urease Houimel et al. (1999)
peptide)
358
a great number of experimental and clinical studies using different compounds for
inhibition of bacterial urease and thus for treatment and prevention of urease-induced
human diseases (Table 73).
10.5. UREASE INHIBITORS USED FOR PREVENTING ACID MINE DRAINAGE
Of the 1,3,4-thiadiazoline-2-thiones proved to be inhibitors of soil urease activity (see

Section 2.l4.2), 2.5 -dimercapto-l,3 ,4-thiadiazole and 5-amino-I,3 ,4-thiadiazole-2-thiol
(both being relatively non-toxic) were used by Stichbury et al. (1995) to prevent acid
mine drainage (AMD). AMD is an aqueous pollutant formed as a result of chemical and
biological oxidation of sulfide minerals. It is produced in tailing ponds and in waste
rock dumps and is characterized by a low pH and high concentrations of sulfate and
dissolved metals. Controlling AMD is important because of its detrimental effects on
the biota of receiving waters.
In one of the experiments, fresh, unoxidized and oxidized tailings were sampled at a
zinc and lead mine (Bathurst, New Brunswick). The fresh tailings had the following
chemical composition by %: S 33.5; Fe 31.5; cot 4.45; Zn 1.65; Pb 1.40; Mg 0.59; Na
0.15; and Ni < 0.06. The main minerals were pyrite, quartz, chlorite, calcite, and
dolomite. A mineral nutrient medium was prepared, to which an inhibitor or the
equimolar mixture of the two inhibitors was added in concentrations of 0, 100, 200 or
500 mgll. pH of the medium was adjusted to 7.0. An 85-ml volume of the medium and
5 rnl of a Thiobacillus thioparus (Th. thioparus) culture were added to 250-rnl
Erlenmeyer flasks containing 20-g aliquots of fresh tailings and 0.5 g of oxidized
tailings. The flasks were incubated at 28°C for 42 days. During incubation, pH and
sulfate content were measured periodically. The inhibitors were considered successful if
there was no decrease in pH and no sulfate generation occurred.
2,5-Dimercapto-l,3,4-thiadiazole and the inhibitor mixture were more effective than
5-amino-l,3,4-thiadiazole-2-thiol in inhibiting sulfide oxidation. The effect increased
with inhibitor concentration.
Th. thioparus is a neutrophilic bacterium that plays a role in the initial oxidation of
metal sulfides. By the acidity produced, it creates acidic conditions necessary for
growth of sulfur- and iron-oxidizing acidophiles such as Th. ferrooxidans which
produce more acidity solubilizing the metal sulfides and causing further pollution of
AMD. Thus, by inhibition of Th. thioparus, growth of Th. ferrooxidans is also
prevented.
The conclusion arrived at was that the most likely treatment would be to add the
inhibitor to the fresh tailings as a slurry.
359
CONCLUSIONS
The investigation performed with the aim to increase the efficiency of urea fertilizers
led to identification of many chemical compounds able to inhibit urease activity in soils.
A great part of these compounds were tested not only in laboratory and vegetation
pots but also under field conditions, on experimental plots. A small number of
compounds were also tested on large agricultural areas.
The investigations were multidisciplinary. Taking part in the investigations were the
chemists who synthesized these compounds, the microbiologists and biochemists who
tested the compounds in laboratory, the agronomists who conducted the experiments in
greenhouse and experimental fields.
N-(n-Butyl)thiophosphoric triamide, phenylphosphorodiamidate, and hydroquinone
can be considered as the most thoroughly studied soil urease inhibitors.
At present, only two urease inhibitors have gained commercial importance.
Hydroquinone is being used on large agricultural areas in China since the 1990s. The
other urease inhibitor, N-(n-butyl)thiophosphoric triamide, was introduced under the
registered trade product name of Agrotain to the United States agricultural market by
IMC-Agrico Company (Bannockbum, Illinois) in spring 1996.
For the future, it is necessary to continue and even to intensify the concerted efforts
of investigators because a) other soil urease inhibitors; b) dual (both urease and
nitrification) inhibitors; c) urease inhibitors in combined use with nitrification
inhibitors; and d) urease inhibitors in natural products also present perspectives to have
commercial importance and to exhibit more advantageous properties than the inhibitors
tested in the past. Research of these compounds and their metabolites as well as
development of large scale production processes deserve further attention as the
increased use of urease (and nitrification) inhibitors in world agriculture will lead to
increased world food production and, even more significantly, to more efficient
environmental protection, both effects being of major importance for the future of
mankind.
361
REFERENCES
Abdel Hadi, A.H., Hamissa, M.R., and Khadr, M.S. (1980) Effect of biuret on the transtormation of urea in two
Egyptian soils, Z. Pf/anzenem. Bodenk. 143,257-261.
Abdel Magid, H.M. and EI Mahi, Y.E. (1986) In vitro effects of urease inhibitors on rate of urea hydrolysis in soil,
Fer!. Res. 8,203-212.
Abdel Magid, H.M., EI Mahi, Y.E., and Gammar, Y. (1983) Potential of urease inhibitors in reducing ammonia
losses in some tropical soils, Trop. Agric. (Trinidad) 60, 301-302.
Abdullatif, EA. and Stroehlein, J.L. (1990) Evaluation of Dwell as a nitrification inhibitor and its interaction with
nitrogen source and soil properties, Commun. Soil Sci. Plant Anal. 21,2153-2161.
Ablizova, Z.Ya. and Tomina, T.K. (1997) Changes in biological activity of a dark-chestnut soil under conditions of
fluoride contamination, Izv. Minist. Nauki-Akad. Nauk Resp. Kilzakhslana, Ser. Biol. Med. (4), 13-19 (in
Russian).
Abou-Sier, A.H., Essawi, M.Y.H., Khalifa, M., Ramadan, M.A., and Megahed, S. (1995) Synthesis and bacterial
urease inhibition activity of3,4,5-trihydroxy/methoxybenzohydroxamic acid and related derivatives, Bull. Fac.
Pharm. Cairo UnN. 33,19-23.
Abramyan, S.A. (1982) Effect of sodium salts on enzymatic activity of soil, Bioi. Zh. Annenii 35, 187-192 (in
Russian).
Abramyan, S.A. and Galstyan, A.Sh. (1981) Effect of exchangeable cations on enzymatic activity of soils, Bioi. Zh.
Armenii 34,142-148 (in Russian).
Agrawal, M.P., Shukla, A., and Singh, M. (1985) Nitrification inhibition of added nitrogenous fertilisers by
potassium chloride in soil, Plant Soil 86, 135-139.
Agrawal, M.P., Singh, R.D., Jafri, S.M.H., and Rajpai, P.K. (1988) Effect of placement of urea supergranule and
nitrogenous inhibitors with urea on mineral nitrogen pattern in soil, J. Indian Soc. Soil Sci. 36, 804- 808.
Alaimo, RJ. and Millner, O.E.jr. (1980) 8-[(4-Aminophenyl)sulfonyIJamino-2-naphthalenyl phosphorodiamidate,
us. Patent No. 4,225,526.
Alaimo, RJ., Sheffer, 1.B., and Millner, O.E.jr. (1980) [[(4-Aminophenyl)sulfonyIJaminoJphenyl phosphorodi-
amidates, U.S. Patent No. 4,222,948.
A1biach, R., Canet, R., Pomeres, E, and Ingelmo, E (2000) Microbial biomass content and enzymatic activities
after the application of organic amendments to a horticultnral soil, Bioresource Technol. 75, 43-48.
Aliev, S.A. (1988) Nitrogen Fixation and Physiological Activity of Organic Matter of Soils. Izv. Nauka,
Novosibirsk, pp. 53-56 (in Russian).
Alkanani, T. and MacKenzie, A.F. (1996) Banding urea and lignosulfonate in corn (Zea mays L.) production and
15N recovery, Can. J. Soil Sci. 76,365-371.
Al-Kanani. T., MacKenzie, A.F., and Blenkhorn, H. (1990a) The influence of fonnula modifications and additives
on ammonia losses from surface-applied urea-ammonium nitrate solutions, Fer!. Res. 22,49-59.
Al-Kanani, T., MacKenzie, A.F., and Blenkhom, H. (1990b) Volatilization of ammonia from urea-ammonium
nitrate solutions as influenced by organic and inorganic additives, Fer!. Res. 23, 113-119.
Al-Kanani, T., MacKenzie, A.E, Fyles, 1.W., Ghazala, S., and O'Halloran, LP. (1994) Ammonia volatilization from
urea amended with lignosulfonate and phosphoroamide, Soil Sci. Soc. Am. J. 58, 244-248.
AI-Rashidi, R.K. and AI-Jabri, M.M. (1990) Nitrification and urea hydrolysis in amended calcareous saline soils,
Arid Soil Res. Rehabil. 4,243-252.
Amberger, A. (1989) Research on dicyandiamide as a nitrification inhibitor and future outlook, Commun. Soil Sci.
Plant Anal. 20, 1933-1955.
Amberger, A. and Gutser. R. (1978) Transformation and effect of urea-dicyandiamide and ammonium sulfate-
dicyandiamide products with ryegrass and rice, Z. Pj/anzenem. Bodenk. 141,553-566 (in German).
Amberger, A. and Gutser, R. (1984) Effect of urea in combination with thiourea or dicyandiamide in different
cropping systems of rice, VDLUFA Schriftenreihe (11),200-211 (in German).
Amberger, A. and Vilsmeier, K. (1979) Researches on the effect of cyanamide, dicyandiamide, guanylurea,
guanidine and nitrite on the urease activity, Landw. Forsch. 32,409-415 (in German).
Arnbroz, Z., Vagner, M., Sroubkova. L., Ondracek, L., and Wanek, W. (1970) On the influence of certain nitrogen-
phosphorus compounds with a direct P-N bond on the soil microflora, Rostl. Vyroba 16, 609-614 (in Czech).
Anderson, 1.R. (1962) Urease activity, ammonia volatilisation and related microbiological aspects in some South
African soils, Proc. 36th Congr. S. Aft Sugar Technol. Assoc., pp. 97-105.
Anderson, 1.R. (1969) Fertilising process and composition, Blitish Patent No. 1,142,245.
Anderson, 1.R. (1970) Fertiliser compositions, US. Patent No. 3,515,532.
362
Andersson, P. and Bengtsson, M. (1975) Effects of copper and zinc on urease and phosphatase activities in spruce
needle mor, Medd. Avd. Ekol. Bot. Lunds Univ. 3 (6),1-18 (in Swedish).
Anello, L.G., Van Der Puy, M., Hendrickson, L.L., Rogic, M.M., Swerdloff, M.D., and Kole, J.E (1985)
Phosphorodiamide urease inhibitors and urease inhibited urea based fertilizer compositions, US. Patent No.
4,517,002.
Anonymous (1973) Inhibition of urease activity in soils, Soil-Fel1ilizer-Plant Research 1969-1972, Tennessee
Valley Authority, Muscle Shoals, Alabama, p. 20.
Anonymous (1981) Phenylphosphorodiamidate (1942-1981), Tennessee Valley Authority, Muscle Shoals,
Alabama.
Anonymous (1983) Use of nitrification and urease inhibitors to improve efficiency of nitrogen fertilizers,
Tennessee Valley Authority Bull. (Y-181), 38-42.
Anonymous (l985a) New urease inhibitors, Tennessee Valley Authority Bull. (Y-191), 37-43.
Anonymous (1985b) New urease inhibitors, Farm Chemicals 148 (11),51.
Anonymous (1986) Chemically modified urea, Annual Repol1 1985, International Fertilizer Development Center,
Muscle Shoals, Alabama, p. 3-4.
Anonymous (1987) Continued development of the urease inhibitor thiophosphOlyl triamide, Tennessee Wtlley
Authority Bull. (Y-198),63-66.
Anonymous (1998) Improving N fertilizer efficiency, Annual Report 1997-1998, South African Sugar Association
Experiment Station, Mount Edgecombe, p. 18.
Anonymous (2000) Urea fertilisers - new soil test values, Annual Repol1 1999-2000, South African Sugar
Association Experiment Station, Mount Edgecombe, p. 26.
Aoyagi, T. and Sumrnerskill, W.HJ. (1966) Inhibition by acetohydroxamic acid of human mucosal and mecal
urease-specific activity, Lancet i, 296-297.
Arziani, B.A., Ugrekhelidze, D.Sh., and Mitaishvili, T.!. (1985) Conjugation of exogenous hydroquinone with
peptides in plants, Soobshch. Akad. Nauk Gruz. SSR 118,609-611 (in Russian).
Ashworth, J., Briggs, G.G., Evans, A.A., and Matula, J. (1977) Inhibition of nitrification by nitrapyrin, carbon
disulphide and trithiocarixmate, J. Sci. Food Agric. 28,673-683.
Ashworth, J., Rodgers, G.A., and Briggs, G.G. (1979) Xanthates as inhibitors offertiliser nitrogen transfonnation in
soil, Chem. Ind. (London), (3),90-92.
Ashworth, J., Akerboom H.M., and Cn'lpin, J.M. (1980) Inhibition by xanthates of nitrification and urea hydrolysis
in soil, Soil Sci. Soc. Am. J. 44, 1247-1249.
Austin, E.R, Bradford, T.J., and Lupin, M.S. (1984) High-perfoIIlllUlce liquid chromatographic determination and
hydrolysis studies of phenyl phosphorodiamidate, a urease inhibitor, J. Agric. Food Chem. 32, 1090-1095.
Avramchuk, P.P. (2000) Method for fertilization of soil, Russian Patent No. 2,143,795.
Badr EI-Din, S.M.S., Moawad, H., Mahmoud, S.A.Z., Garnal R.E, and Enany, M.H. (1985) Urease inhibition by
metals and its relation to nitrogen transfonnations, Z. Pjlanzenern. Bodenk. 148,551-558.
Badura, L., Eckert, L., and Galimska-Stypa, R. (1986) Influence of zinc and cadmium on the bacteria active in
transfonnations of nitrogen in soil, Acta Bioi. Siles. 3, 9-19 (in Polish).
Baintner, K. (1964) Prevention of urea poisoning with acetohydroxamic acid, Allattenyesztes 13, 373-378 (in
Hungarian).
Baird, J.V. and Ngueguim, M. (1991) The effect of urease inhibitor, NBPT, on efficiency of nitrogen uptake by
maize, Agron. Abstr., p. 358.
Banerjee, M.R., Burton. D.L., and Grant, CA. (1997) Influence of soil type, urea fertilization and urease inhibitor
on soil biological quality under conventional and zero tillage, Agron. Abstr., p. 201.
Banerjee, M.R., Burton, D.L., and Grant, C.A. (1999) Influence of urea fertilization and urease inhibitor on the size
and activity of the soil microbial biomass under conventional and zero tillage at two sites, Can. J. Soil Sci. 79,
255-263.
Barth, A., Rollka, w., Michel, H.-J., and Franke, R. (1978) Researches on the potential effectiveness of
diamidophosphoric acid aryl esters as urease inhibitors, Spez. Agrochem. Forsch. Praxis (CUnnersdorf) 8, 15-
19 (in Gennan).
Barth, A., Rollka, W., and Michel, H.-J. (1980) Phosphoric acid ester diamides as urease inhibitors. A contribution
to the research of active compounds for development of tight-binding inhibitors of selected enzymes, Wiss.
Beitr. Mal1in-Luther-Univ. Halle-Wittenberg (2),5-10 (in German).
Baumgartner, M. and Ottow, J.c.G. (1985) Influence ofa fluorine compound (NaF) on the nitrification in soil, Mitt.
Deutsch. Bodenk. Ges. 43, 519-524 (in German).
Bayan, M.R. and Eivazi, E (1999) Selected enzyme activities as affected by free iron oxides and clay particle size,
lommun. Soil Sci. Plant Anal. 30,1561-1571.
Bayless, A.V. and Millner, O.Ejr. (1980a) N-[Diaminophosphinyl]ary\carboxamides, Us. Patent No. 4,182,881.
Bayless, A.V. and Millner, O.Ejr. (1980b) Phosphorotriamides as urease inhibitors, us. Patent No. 4,242,325.
363
Bayrakli, F. (1990) Anunonia volatilization losses from different fertilizers and effect of several urease inhibitors,
Cach and phosphogypsum on losses from urea, Fen. Res. 23, 147-150.
Bayrakli, F. and Gezgin, S. (1996) Controlling ammonia volatilization from urea surface applied to sugar beet on a
calcareous soil, Commun. Soil Sci. Plant Anal. 27, 2443-2451.
Benedetti, A., Ceccanti, B., Cafcinai, M., and Tarsitano, R. (1990) Decomposition of chromium-containing leather
residues in a sandy soil, Suelo Planta I, 15-24.
Berg, R.w. (1977) Now, ammonia can be controlled, Turkey World (July), p. 1.
Besekau, E., Harzfeld, G., Kiimrnel, R., Malzel, w., Miihlenbruch, D., and Thomas, G. (1974) Nitrogen fertilizer,
(East) German Patent No. 108,730.
Beyrouty, CA., Nelson, D.W., and Sommers, L.E. (l988a) Effectiveness of phosphoroamides in retarding
hydrolysis of urea surface-applied to soils with various pH and residue cover, Soil Sci. 145,345-352.
Beyrouty, CA., Sommers, L.E., and Nelson, D.W. (1988b) Anunonia volatilization from surface-applied urea as
affected by several phosphoroamide compounds, Soil Sci. Soc. Am. J. 52. 1173-1178.
Bhavanandan, v.P. and Fernando, V. (1970) Studies on the use of urea as a fertilizer for tea in Ceylon. 2. Urease
activity in tea soils, Tea Quan. 41, 94-106.
Bhupinderpaf-Singh, Bijay-Singh, and Yadvinder-Singh (1992) Urease activity and kinetics ofurea hydrolysis in
some soils from semiarid regions of northwestern India, Arid Soil Res. Rehabil. 6, 21-32.
Biau, E.E., Zaiko, A.S., and Muravin, E.A. (2000) Influence of systematic application of nitrification inhibitors on
the bafance ofJabeled nitrogen of fertilizers in pot experiments, Agrokhimiya (3), 30-40 (in Russian).
Blaise, D. and Prasad, R. (1995) Effect of blending urea with pyrite or coating urea with polymer on ammonia
volatilization loss from surface-applied prilled urea, Bioi. Fert. Soils 20,83-85.
Blaise, D. and Prasad, R. (1997) Dynamics of N-release from differently coated urea under flooded soil
conditions, Proc. Indian Natl. Sci. Acad.. Part B 63, 73-78.
Blaise, D., Amberger, A., and von Tucher, S. (1996) Ammonia volatilization and denitrification losses from
nitrogen fertilized flooded soil as affected by the addition of iron pyrites, Curro Sci. 71, 142-144.
Blaise, D., Amberger, A., and von Tucher, S. (1997) Influence of iron pyrites and dicyandiamide on nitrification
and ammonia volatilization from urea applied to loess brown earths (luvisols), BioI. Fen. Soils 24,179-182.
Bock, B.R. and Williams, H.M. (1984) Effects of thiourea on transformation and ammonia volatilization from
surface-applied urea, Agron. Abstr., p. 268.
Bollich, P.K. (1991) Influence ofa urease inhibitor and timing of pre flood nitrogen on rice,Agron. Abstr., p. 283.
Bopaiah, M.G. and Biddappa, CC (1987) Studies on the nitrogen release pattem by ureaform and coated fertilizers
in acid soils, J. Plantation Crops 15,42-46.
Borchaninova, M.L and Filippova, KF. (1972) Effect of pesticides on accumulation of nitrogen in plants and
rhizosphere of red clover. in V. Tohver and J. Simisker (eds.), Ecological and PhySiological-biochemical Bases
ofthe Microbiological Transformation ofNitrogen, State Univ., Tartu, pp. 266-270 (in Russian).
Borchaninova, M.L and Filippova, KF. (1978) Effect of seed disinfection with TMTD on the activity of certain
enzymes in the rhizosphere of red clover, in A.A. Boiko (ed.), Effict of Environmental Physicochemical
Factors on the Plants, State Univ., Perm, pp. 56-65 (in Russian).
Borovskii, E.V. and Yanishevskii, F.V. (1994) Effectiveness of complex polymeric fertilizers, urotropin and 1-
carbamoyl-3(5)-methylpyrazole applied to vegetable crops on afluviaf meadow and soddy-podzolic soils of the
Moscow region, Agrokhimiya (9), 43-64 (in Russian).
Bouldin, D.R., Hongprayoon, C, Lindau, Cw., and Patrick, W.H.jr. (1991) Urea transformations in flooded soil
columns. II. Derivation of model and implications to ammonia volatilization, Soil Sci. Soc. Am. J. 55, 1135-
1142.
Bremner, J.M. (1982) Nitrogen-urea, inA.L. Page, R.H. Miller, and D.R. Keeney (eds.), Methods of Soil Analysis.
2. Chemical and Microbiological Propenies (2nd Edition), Am Soc. Agron.-Soil Sci. Soc. Am, Madison, pp.
699-709.
Bremner. J.M. (1986) Inhibition of nitrogen transformations in soil, in F. Megusar and M. Gantar (eds.),
Perspectives in Microbial Ecology, Slovene Soc. Microbioi., Ljubljana, pp. 337-342.
Bremner, J.M. and Bundy. L.G. (1974) Inhibition of nitrification in soils by volatile sulfur compounds, Soil Bioi.
Biochem.6,161-164.
Bremner, J.M. and Bundy, L.G. (1976) Effects of potassium azide on transformations of urea nitrogen in soils, Soil
BioI. Biochem. 8, 131-133.
Bremner, J.M. and Chai, H.S. (1986) Evaluation of N-butyl phosphorothioic triamide for retardation of urea
hydrolysis in soil, Commun. Soil Sci. Plant Anal. 17,337-351.
Bremner, lM. and Chai, H.S. (1989) Effect of phosphoroamides on ammonia volatilization and nitrite
accumulation in soils treated with urea, Bioi. Fen. Soils 8, 227-230.
Bremner, J .M. and Douglas, L.A. (1971) Inhibition of urease activity in soils, Soil Bioi. Biochem. 3, 297-307.
364
Bremner. J.M. and Douglas. L.A. (1973) Effects of some urease inhibitors on urea hydrolysis in soils. Soil Sci. Soc.
Am. Proc. 37.225-226.
Bremner. J.M. and Krogmeier. M.J. (1988) Elimination of the adverse effects of urea fertilizer on seed gennination.
seedling growth. and early plant growth in soil. Proc. Natl. Acad. Sci. USA 85. 4601-4604.
Bremner, J.M. and Krogmeier, M.J. (1989) Evidence that the adverse effect of urea fertilizer on seed gennination in
soil is due to ammonia fonned through hydrolysis of urea by soil urease, Proc. Natl. Acad. Sci. USA 86, 8185-
8188.
Bremner, J.M. and Krogmeier, MJ. (1990) Effects of urease inhibitors on gennination of seeds in soil. Commun.
Soil Sci. Plant Anal. 21, 311-321.
Bremner, J.M. and Martens, D.A. (1987) Decomposition of phenylphosphorodiamidate in soil, Agron. Abstr., p.
179.
Bremner, J.M. and McCarty, G.W. (1989) Formation of phosphoryl triarnide by decomposition of
thiophosphoryl triamide in soil, Agron. Abstr.. p. 211.
Bremner, J.M., McCarty, G.W., and Chai, H.S. (1986a) Evaluation of ammonium thiosulfate as a soil nitrification
and urease inhibitor, Agron. Abstr.. p. 175.
Bremner, J.M., McCarty, G.W., Yeomans, J.C., and Chai, H.S. (I986b) Effects ofphosphoroamides on nitrification,
denitrification, and mineralization of organic nitrogen in soil, Commun. Soil Sci. Plant Anal. 17,369-384.
Bremner, J.M., McCarty, G.W., and Krogmeier, M.J. (1990) Assessment of the potential value of ammonium
thiosulfute as a soil urease inhibitor. Agron. Abstr., p. 244.
Bremner, J.M., McCarty, G.W., and Higuchi, T. (1991) Persistence of the inhibitory effects ofphosphoroamides on
urea hydrolysis in soils, Commun. Soil Sci. Plant Anal. 22, 1519-1526.
Briggs, M.H. and Segal. L. (1963) Preparation and properties ofa free soil enzyme, Life Sci. 2, 69-72.
Broadbent, F.E., Nakashima, T., and Chang, G,Y. (l985) Perfonnance of some urease inhibitors in field trials with
com, Soil Sci. Soc. Am. J. 49,348-351.
Bronson, K.F. and Mosier, A.R. (l992) Suppression of methane oxidation in soil by nitrogen fertilizers, nitrification
inhibitors, and urease inhibitors, Agron. Abstr., pp. 250-251.
Bronson, K.E and Mosier, A.R. (l994) Suppression of methane oxidation in aerobic soil by nitrogen fertilizers,
nitrification inhibitors, and urease inhibitors, Bioi. Ferl. Soils 17, 263-268.
Bronson, K.E. Touchton, J.T., Hiltbold, A.E., and Hendrickson, L.L. (1989) Control of ammonia volatilization with
N-(n-butyl) thiophosphoric triamide in loamy sands, Commun. Soil Sci. PlantAllal. 20,1439-1451.
Bronson, K.E, Touchton, J.T., Cummins, C,G., and Hendrickson, L.L. (1990) Use of the urease inhibitor N-(n-
butyl) thiophosphoric triamide in com production on a loamy sand,J. Ferl. Issu£s 7 (2), 31-34.
Brunner, 1. and Schinner, F. (1984) Influence of lead and cadmium on the microbial activity of a soil,
Bodenkultur 35, 1-12 (in German).
Bugnilo, M., Bayeli, P. E, Rappuoli, R., Pennatini, c., Figura, N., and Crabtree, J.E. (1993) Inhibition of
Helicobacter pylori urease by omeprazole, Eur. J. Gastroenterol. Hepatol. 5, 683-685.
Builov, v.v., Lichko, R.P., and Volokitin, M.P. (1979) Improvement of the properties of solonetz soils with
ammoniumlignosulfunate, Pochvovedenie (7),81-91 (in Russian).
Bundy, L.G. and Bremner, J.M. (l973a) Effects ofsubstituted p-benzoquinones on urease activity in soils, Soil Biol.
Biochem. 5, 847-853.
Bundy, L.G. and Bremner, J.M. (1973b) Inhibition of nitrification in soils, Soil Sci. Soc. Am. Proc. 37, 396-398.
Bundy, L.G. and Bremner, J.M. (1974a) Effects of urease inhibitors on nitrification in soils, Soil Bioi. Biochem. 6,
27-30.
Bundy, L.G. and Bremner, J.M. (l974b) Effects of nitrification inhibitors on transformations of urea nitrogen in
soils, Soil Bioi. Biochem. 6, 369-376.
Bundy, L.G. and Oberle, S.L. (l988) Evaluation of methods for control of ammonia volatilization from surface-
applied urea-containing fertilizers, J. Ferl. Issues 5 (I), 24-30.
Buresh, R.J. (1987) Relative susceptibility of conventional and experimental nitrogen sources to ammonia loss
from flooded rice fields, Ferl. Res. 13,139-153.
Buresh, R.J., Vlek, P.L.G., and Stumpe, J.M. (1984) Labeled nitrogen fertilizer research with urea in the semi-arid
tropics. 1. Greenhouse studies, Plant Soil 80,3-19.
Buresh, R.J., De Datta, S.K., Padilla, J.L., and Samson, M.I. (1988a) Effi:ct of two urease inhibitors on floodwater
ammOnia following urea application to lowland rice, Soil Sci. Soc. Am. J. 52, 856-861.
Buresh, R.J., De Datta, S.K., Padilla, l.L., and Samson, M.I. (1988b) Field evaluation of two urease inhibitors with
transplanted lowland rice, Agron. J. 80, 763-768.
Buresh, R.J., De Datta, S.K., Padilla, l.L., and Chua, T.T. (1988c) Potential of inhibitors for increasing response of
lowland rice to urea fertilization, Agron. J. 80,947-952.
Bums, J.R. and Gauthier, J.E (1984) Prevention of urinary catheter incrustations by hydroxamic acid, J. Urol. 132,
455-456.
365
Burns, RG., Gregory, L.J., Lethbridge, G., and Pettit, N.M. (1978) The effect of y-irradiation on soil enzyme
stability, Experientia 34,301-302.
Byrnes, B.H. and Amberger, A. (1989) Fate of broadcast urea in a flooded soil when treated with N-(n-butyl)
thiophosphoric triamide. a urease inhibitor, Fen. Res. 18.221-231.
Byrnes, 8.H. and Christianson, C8. (1988) Development of a urease inhibitor from N-(n-butyl) thiophosphoric
triamide. Agron. Abstr., p. 212.
Byrnes. B.H., Savant, N.K., and Craswell, E.T. (1983) Effect of a urease inhibitor phenyl phosphorodiamidate on
the efficiency of urea applied to rice, Soil Sci. Soc. Am. J. 47.270-274.
Byrnes, 8., Vilsmeier. K., Austin, E., and Amberger, A. (1989a) Degradation of the urease inhibitor phenyl
phosphorodiamidate in solutions and floodwaters, J. Agric. Food Chem. 37, 473-477.
Bymes, B.H., Gutser, R, and Amberger, A. (1989b) Greenhouse study on the effects of the urease inhibitors phenyl
phosphorodiamidate and N-(n-butyl) thiophosphoric triamide on the efficiency of urea applied to flooded rice,
Z. Pjlanzenern. Bodenk. 152,67-72.
Cabrera, M.L. and Kissel, D.E. (1984) Rate of urea hydrolysis in soil as affected by urea concentration,
Agron. Abstr., p. 268.
Cabrera, M.L.. Kissel, D.E., and Bock, B.R. (1991) Urea hydrolysis in soil: Effects of urea concentration and
soil pH, Soil BioI. Biochem. 23, 1121-1124.
Cai, G.x., Freney, J.R., Muirhead, W.A., Simpson, J.R, Chen, D.L., and Trevitt, A.C.F. (1989) The evaluation of
urease inhibitors to improve the efficiency of urea as a N-source for flooded rice, Soil Bioi. Biochem. 21, 137-
145.
Calancea, L. (1979) Effect of urease and nitrification inhibitors on the content of amino acids in Lolium
multijlorum, Bul. Inst. Agron. ClUj. Ser. Agric. 33, 33-38 (in Romanian).
C.alancea, L. and Chiriac, M. (1993) Utilization by maize ofphosphonitrilic hexamide labelled with 1'N at cyclic
and at amino positions, Isotopenpraxis Environ. Health Stud. 29,357-364.
Calancea, L. and Chiriaco M. (1996) Comparison of the efficiencies of urea nitrogen and phosphoryl triamide
nitrogen for Lolium multijlorum and their influence on the soil nitrogen reserve, Isotopes Environ. Health Stud.
32,79-86.
Calancea, L. and Kiss. S. (1984) Effect of hydroquinone on the uptake of nitrogen from soil reserve and urea by
Lolium multiflorum, Fifth Symposium on Soil Biology (Ia~i, 1981), Ed. Ceres, Bucharest. pp. 65-68.
Calancea, L., Bologa, M., and Firu. V. (1977) Use of 15N_urea in the study of the effect of soil urease inhibitors,
ESNA (European Society of Nuclear Methods in Agriculture) Newslell. (Uppsala) (15 Nov.). 63-70.
Calancea, L., Firu, V., and L.. Bologa, M. (1982) Effect of nitrification and urease inhibitors on the efficiency of
urea and glycoluril as fertilizers, Separation and Utilization of Isotopes. Inst. Isotopic Molec. Techno!., Cluj.
pp. 354-356 (in Romanian).
Calancea, L., Bologa, M., Chiriac, M, and Firu, V. (1986) Use of phosphonitrilic hexamide as a fertilizer, in C
Bodea (ed.), Progress in Biochemical Research, Romanian Acad. Sci .• Cluj Branch. pp. 127-137 (in
Romanian).
Calancea, L.. Bologa, M.,and Chiriac, M. (1990) Utilization of 15N to evaluate the availability of nitrogen from
different fertilizers for Lolium multijlorum, Stud. Univ. BabeJ-Bo/yQ/, BioI. 35 (1). 37-44.
Cao. CM., Zhou, L.K. et al. (1987) The technology of hydroquinone-containing fertilizer production, Chinese
Patent Publ. 3 (10). Cited by Zhao et al. (1992b).
Carmona, G., Christianson. C.B., and Bymes, B.H. (1988) Effect of urease inhibitor concentration in urea on N
losses via ammonia volatilization, Agron. Abstr.. pp.212-213.
Carmona, G., Christianson, CB., and Byrnes, B.H. (1990) Temperature and low concentration effilcts of the urease
inhibitor N-(n-butyl) thiophosphoric triamide (n-BTPT) on ammonia volatilization from urea, Soil BioI.
Biochem. 22,933-937.
Cannignani, G., Belgrano, E., Puppo, P., Cichero, A., and Giuliani. L. (1980) Hydroxyurea in the management of
chronic urea-splitting urinary infections, British J. Urol. 52, 316-320.
Carrier, D. and Bernier, B. (1976) Influence of the application of superphosphate and K2S04 .MgSO. with urea to a
jack pine forest humus. I. Ureolysis and ammonia volatilization. Can. J. Forest Res. 6, 287-292 (in French).
Ceccanti, 8., Nannipieri, P., Cervelli. S., and Sequi. P. (1978) Fractionation of humus-urease complexes, Soil BioI.
Biochem. 10, 39-45.
Chai, H.S. and Bremner, J.M. (1985) Evaluation of some phosphoroamides as soil urease inhibitors, Agron. Abstr.,
p.154.
Chai. H.S. and Bremner, J.M. (1986) Effects of phosphoroamides on ammonia volatilization and nitrite
accumulation in soils treated with urea, Agron. Abstr., p. 176.
Chai. H.S. and Bremner, J.M. (1987) Evaluation of some phosphoroamides as soil urease inhibitors, Bioi. Fer/.
Soils 3. 189-194.
366
Chai, H.S., Bremner, J.M., and McCarty, G.w. (1988) Effect of N-(n-butyl) thiophosphoric triamide on
hydrolysis of urea by plant, microbial, and soil urease, Agron. Abstr., p. 213.
Chaiwanakupt, P., Freney, J.R., Keerthisinghe, D.G., Phongan, S., and Blakeley, R.L. (1996) Use of urease,
algal inhibitors. and nitrification inhibitors to reduce nitrogen loss and increase the grain yield of flooded
rice (Orvza sativa L.), BioI. Fert. Soils 22, 89-95.
Chauhan, H.S. and Mishra, B. (1989) Ammonia volatilization from a flooded rice field fertilized with
amended urea materials, Fen. Res. 19,57-63.
Chemische und Pharmazeutische Fabriken (Magdeburg, Germany) (1977) Improvements in or relating to
animal fodder, British Patent No. 1.494,774.
Chen, W. and Lu, W. (1997) Effects of paddy urease inhibitors on fate of 15 N_urea, Henong Xuebao 11 (3),
151-156 (in Chinese). Chem Abstr., 1998, 128, 140185a.
Chen, L.J.. Shi, Y., Li, R.H., Hu. L.S., and Zhou, L.K. (1995) Synergistic effect of urease inhibitor and
nitrification inhibitor on urea-N transformation and N2 0 emission, Chinese J. Appl. Ecol. 6, 368-372 (in
Chinese).
Chen, L.. Boeckx, P., Zhou, L., Van Cleemput, 0., and Li, R. (1998) Effect of hydroquinone, dicyandiamide
and encapsulated calcium camide on urea N uptake by spring wheat, soil mineral N content and NzO
emission, Soil Use Manage. 14 (4),230-233.
Chemykh, N.A. (1991) Effect of heavy metals on enzymatic activity of soils, Khim. Sel'sk Khoz. (1),40-42
(in Russian).
Chien, M.M., Savant, N.K., and Engel, N.G.H. (1988) Evaluation of cyclohexyl phosphoric and thiophosphoric
triamides as sustained-action urease inhibitors in submerged soil, Agron. Abstr.. p.213.
Christianson, c.B. and Howard, R.G. (1994) Use of soil thin-layer chromatography to assess the mobility of the
phosphoric triamide urease inhibitors and urea in soil, Soil BioI. Biochem. 26, 1161-1164.
Christianson, C.B. and Vlek, P.L.G. (1991) Alleviating soil fertility constraints to food production in West Africa:
Efficiency of nitrogen fertilizers applied to food crops, Fert. Res. 29,21-33.
Christianson, C.B., Byrnes, B.H., and Carmona, G. (1990) A comparison of the sulfur and oxygen analogs of
phosphoric triamide urease inhibitors in reducing urea hydrolysis and ammonia volatilization, Fert. Res. 26,
21-27.
Christianson, C.B., Baethgen, W.E., Carmona, G., and Howard, R.G. (1993) Microsite reactions ofurea-nBTPT
fertilizer on the soil surface. Soil Bioi. Biochem. 25, 1107-1117.
Christianson, C.B., Carmona, G., Klein, M.D .. and Howard, R.G. (1995) Impact on ammonia volatilization losses
of mixing KCI of high pH with urea, Fen. Res. 40,89-92.
Clark, N.G., Croshaw, B., and Spooner, D.E (1967) New antibacterial and antifungal compositions, British Patent
No. 1,057,131.
Clark, N.G., Croshaw, B., and Spooner, D.E (l970) Control of micro-organisms, British Patent No. 1.193,954.
Clay, D.E., Malzer, G.L., and Anderson, J.L. (1990) Ammonia volatilization from urea as influenced by soil
tempemture, soil water content, and nitrification and hydrolysis inhibitors, Soil Sci. Soc. Am. J. 54, 263-266.
Conrad, J.P. (1940) The nature of the catalyst causing the hydrolysis of urea in soils, Soil Sci. 50, 119-134.
Cmswell, E.T. and Vlek, P.L.G. (1982) Nitrogen management for submerged rice soils, U;rtisols and Rice Soils in
Iivpics. 12th Int. Congr: Soil Sci. (New Delhi. 1982), Symp. Pap., pp. 158-181.
Creason. G.L., Schmitt, M.R .. Douglass, E.A., and Hendrickson, L.L. (1990) Urease inhibitory activity associated
with N-(n-butyl)thiophosphoric triamide is due to fonnation of its oxon analog, Soil Bioi. Biochem. 22, 209-
211.
Czapla, J. and Humiecki, W. (1998a) Effect of urea applied with ureolysis inhibitor on spring wheat yield, Acta
Acad. Agric. Tech. Ols.. Agric. (Olsztyn) (65),89-98 (in Polish).
Czapla, J. and Humiecki, W. (l998b) Yield of spring wheat subjected to foliar application of urea with addition of
ureolysis inhibitor,Acta Acad. Agric. Tech. Ols., Agric. (Olsztyn) (65), 99-107 (in Polish).
Daif, M.A. and van Beusichem, M.L. (1981) Effects of some tmce elements on urea hydrolysis in soil, Neth. J.
Agric. Sci. 29, 249-257.
Dalton. D.A., Evans, H_l., and Hanus, 1'.1. (1985) Stimulation by nickel of soil microbial urease activity and urease
and hydrogenase activities in soybeans grown in a low-nickel soil, Plant Soil 88,245-258.
Davidescu, V. and Chi rca, M. (1977) Researches concerning the effect of potassium salts of cyclophosphazanic
acids as a new type of fertilizers, Stud. Cere. Bioi.. Sel: Bioi. U;g. 29,173-178 (in Romanian).
De, G.c., Das. S., and Pal, D. (1992) Efficiency of neem-extract-coated urea on tmnsplanted summer rice (Oryza
sativa), IndianJ. Agron. 37,163-165.
De Boer, w., Duyts, H., and Laanbroek, H.J. (1989) Urea stimulated autotrophic nitrification in suspension of
fertilized, acid heath soiL Soil Biol. Biochem. 21. 349-354.
De Datta, S.K., Fillery, I.R.P., and CmswelL E.T. (1983) Results from recent studies on nitrogen fertilizer efficiency
in wetland rice, Outlook Agric. (Oxford) 12, 125-134.
367
Dick, W.A. and Tabatabai, M.A. (1978) Hydrolysis of organic and inorganic phosphorus compounds added to soils,
Geoderma,21,175-182.
Dobrotvors 'ka, K.M. (1956) Effect of brown coal on the activity of urease enzyme, Dop. Akad. Nauk Ukr. RSR (6),
591-593 (in Ukrainian).
Doelman, P. and Haanstta, L. (1986) Short- and 10ng-tenn effects of heavy metals on urease activity in soils, Bioi.
Fen. Soils 1,213-218.
Dornb6vari, J. and Kiss, A.S. (1988) The magnesium ion activates hydrolysis of urea in soil, HlI1Igagrochem. Kon/.
(Keszthely, 1988), pp. 276-279 (in Hungarian).
Douglas, LA. and Bremner, J.M. (1970) Extraction and colorimetric detennination of urea in soils, Soil Sci. Soc.
Am. Proc. 34, 859-862.
Douglas, L.A. and Bremner, J.M. (1971) A rapid method of evaluating different compounds as inhibitors of
urease activity in soils, Soil Bioi. Biochem. 3,309-315.
Douglass, E.A. and Hendrickson, L.L. (1989) Urease inhibition by N-(n-butyl)thiophosphoric triamide and its
oxon analog in diverse soils, Agron. Abstr., p. 214.
Douglass, E.A. and Hendrickson, L.L. (1991) HPLC method for the analysis of the urease inhibitor N-(n-
butyl)thiophosphoric triamide and its metabolites, J. Agric. Food Chem. 39, 2318-2321.
Eckert, D.J., Check, A.M., and Martin, V.L. (1990) Ammonium thiosulfate and ammonium sulfate effects on
com performance and inorganic soil nitrogen, J. Fert. Issues 7 (2), 24-30.
Edwards, A.P. (1982) The efficiency of urea as a direct and indirect source of nitrogen to wheat as influenced
by a urease inhibitor and sulfur coatings, Agron. Abstr., p. 187.
Elbe, W., Hackert, H .. Jasche, K., JOdicke, R., Oertel, M., and Thieme, H. (1979) Procedure for production of
urea with additives. (East) German Patent No. 138,935.
EIkholei, M.S., Wasif, S., and Hamdi, H. (1982) Comparative effects of varying levels of chlorides, sulfates
and carbonates of sodium, potassium, calcium and magnesium on hydrolysis and nitrification of urea in
soils, Egypt. J. Soil Sci. 11, 179-187.
Ellenberger, M.A., Rumpler. W.V .• Johnson, D.E., and Goodall, S.R. (1984) Evaluation of the extent of
ruminal urease inhibition by sarsaponin and sarsaponin fractions, J. Anim. Sci. 61 (Suppl.), 491-492.
EI-Sayed, MH., Allam, N.A., and Daif, M.A. (1976) Studies on urea hydrolysis. 1. Effect of biuret on urease
activity in vitro, Ann. Agric. Sci. (Egypt), 5, 269-280.
E1-Shahawy, R.M.and Mashhady, A.S. (1984) Nitrification ofammoniurn sulphate and urea fertilizers under saline
condition, Zbl. Mikrobiol. 139,343-347.
E1-Shinnawi, M.M. and EI-Shimi, S.A. (1981a) Enzymatic activities in salt-affected soils, Zbl. Bakteriol. 136, 106-
116.
EI-Shinnawi, M.M. and EI-Shimi, S.A. (1981b) Enzyme activities of soil in relation to moisture content and
salinity, Zbl. Bakteriol.. Abt. II 136,471-477.
Esso (Research and Engineering Company) (1966) Agricultural nutrient, British Patent No. 1,022,793.
Esso (Research and Engineering Company) (1969) Fertilizer composition, British Patent No. 1,151,813.
Fairlie, T.E. and Goos, R.J. (1986) Urea hydrolysis and anunonia volatilization characteristics of liquid fertilizer
mixtures. II. Studies under modified field conditions, J. Fert. Issues 3 (3), 86-90.
Fan, M.x. and MacKenzie, A.F. (1993a) Urea and phosphate interactions in fertilizer microsites: Ammonia
volatilization and pH changes, Soil Sci. Soc. Am. J. 57, 839-845.
Fan, M-X. and MacKenzie, A.F. (l993b) Interaction of urea with triple superphosphate in a simulated fertilizer
band, Fert. Res. 36,35-44.
Fan, Y.c. and Yeo H.Z. (1995) The efficiency of applying urease inhibitor to rice, Chinese J. Soil Sci. 16, 230-231
(in Chinese).
Faruci, W.S., Yang, B.v., O'Rourke, D., and Spencer, R.W. (1995) Inhibition of Helicobacter pylori urease by
phenyl phosphorodiamidates: Mechanism of action, Bioorg. Med. Chem. 3,605-610.
Fenn, L.B. (1982) New composition of matter and method of use for nitrogen fertilization, Eur. Patent Appl.. Publ.
No. 0 055 493 A 1.
Fenn, LB. (1985) Composition of matter and method of use for nitrogen fertilization, U.S. Patent No. 4,500,335.
Fenn, L.B. (1988) Effects of initial soil calcium content on ammonia losses from surface-applied urea and calcium-
urea, Fert. Res. 16.207-216.
Fenn, LB. and Hossner, LR. (1985) Ammonia volatilization from ammonium or ammonium-fonning nitrogen
fertilizers, Adv. Soil Sci. 1, 123-169.
Fenn, LB., Taylor, R.M., and Matocha, J.E. (1981a) Ammonia losses from surface-applied nitrogen fertilizer
as controlled by soluble calcium and magnesium: General theory, Soil Sci. Soc. Am. J. 45, 777-781.
Fenn, L.B., Matocha, J.E., and Wu, E. (1981b) A comparison of calcium CIIlbonate precipitation and pH depression
on calcium-reduced anunonia loss from surface-applied urea, Soil Sci. Soc. Am. J. 45, 1128-1131.
368
Fenn, L.B., Matocha, J.E., and Wu, E. (1982) Substitution of ammonium and potassium for added calcium in
reduction of anunonia loss from surface-applied urea, Soil Sci. Soc. Am. J. 46,771-776.
Fenn, L.B., Malstrom, H.L., and Wu, E. (1987) Anunonia losses from surface-applied mea as related to urea
application rates, plant residue and calcium chloride addition, Fen. &s. 12, 219-227.
Fenn, L.B., Tatum, G., and Horst, G. (1990) Ammonia losses from surface-placed mixtures of urea-calcium-
potassium salts in the presence of phosphorus, Fen. Res. Z1, 125-131.
Fernandes da Mata, MJ. and Drauschke, W. (1996) Researches on NHJ volatilization as influenced by loss-
reducing fertilization technologies in flooded rice soils, Tropenlandwin97, 177-184 (in Gennan).
Fernando, V. and Roberts, G.R. (1976) The partial inhibition of urease by natura1ly occurring polyphenols, Plant
Soil 44, 81-86.
Fiedler, H.J., Huth, J., Mai, H., Reissbrodt, R., and Richter, W. (1974a) On the fertilizer effect of phosphoric acid
alkyl ester amides and related compounds, Arch. Acker- Pjlanzenbau Bodenk. 18, 791-802 (in German).
Fiedler, H.I., Mai, H., Reissbrodt, R., Richter, W., and Riesel, L. (1974b) Procedure for fertilization of plants, (East)
German Patent No. 107,245.
Fiedler, HJ., Huth, J., Mai, H., and Reissbrodt, R. (1975) Effect of alky1ated derivatives of phosphorus oxide
triamide on yield and nutrient content of oats, Arch. Acker- Pjlanzenbau Bodenk. 19, 49-59 (in German).
FillelY, I.R.P. and De Datta, S.K. (1986) Ammonia volatilization from nitrogen sources applied to rice fields. I.
Methodology, ammonia fluxes, and nitrogen-IS loss, Soil Sci. Soc. Am. J. 50,80-86.
FillelY, I.R.P. and Vlek, P.L.G. (1986) Reappraisal of the significance of anunonia volatilization as an N loss
mechanism in flooded rice fields, Fen. Res. 9, 79-98.
Fillel}', I.R.P., Simpson, J.R., and De Datta, S.K. (1984) Influence of field environIllllllt and fertilizer management
on ammonia loss from flooded rice, Soil Sci. Soc. Am. J. 48,914-920.
Fillel}', I.R.P., Roger, P.A., and De Datta, S.K. (1986a) Anunonia volatilization from nitrogen SOUlCes applied to
rice fields. II. Floodwater properties and submerged photosynthetic biomass, Soil Sci. Soc. Am. J. 50, 86-91.
Fillel}', I.R.P., De Datta, S.K., and Cmswell, E.T. (l986b) Effect of phenyl phosphorodiamidate on the fate of urea
applied to wetland rice fields, Fen. Res. 9, 251-263.
Fishbein, W.N., Carbone, P.P., and Hochstein, H.D. (1965) Acetohydroxamate: Bacterial urease inhibitor with
therapeutic potential in hyperammonaemic states, Nature (London) 208, 46-48.
Flaig, W. (\976) The organic soil substance as a supplimental}' nitrogen SOUICe for nutrition of plants and some
models for technical realization, Landbalfforschung Vol/amrode 26, 117-121 (in German).
Fleisher, Z. and Hagin, 1. (1981) Lowering anunonia volatilization losses from mea applications by activation of
nitrification process, Fen. Res. 2,101-107.
Ford, O.K. (1973) T-stmin mycoplasmas and genital chlamydiae: Inhibition of T-stmin urease by hydroxamic
acids, J. Infect. Dis. 127, S82-S83.
Fox, R.H. and Piekielek, W.P. (1987) Comparison of surface application methods of nitrogen solution to no-
till corn (lea mays L.), J. Fen. Issues 4 (I), 7-12.
Fox, R.H. and Piekielek, W.P. (1993) Management and urease inhibitor effects on nitrogen use efficiency in
no-till corn, J. Production Agric. 6, 195-200.
Fox, R.H., Kern, J.M., and Piekielek, W.P. (1986) Nitrogen fertilizer SOUlCe, and method and time of
application effects on no-till com yields and nitrogen uptakes, Agron. J. 78, 741-746.
Frankenberger, W.T.jr. and Bingham, F.T. (1982) Influence of salinity on soil enzyme activities, Soil Sci. Soc.
Am.J. 46, 1173-1177.
Freney, J.R., Simpson, J.R., Zhu, Z.L., and Bidin, A. (1989) Gaseous nitrogen loss from urea fertilizers in
Asian cropping systems, ACIAR (Australian Centre for International Agricultural Research, Canberra)
Tech. ReponNo.9, 1-16.
Freney, J.R., Keerthisinghe, D.G., Chaiwanakupt, P., and Phongpan, S. (1993) Use of urease inhibitors to
reduce ammonia loss fullowing application of urea to flooded rice fields, Plant Soil 1551156, 371-373.
Freney, J.R., Keerthisinghe, D.G., Phongpan, S., Chaiwanakupt, P., and Harrington, KJ. (1995) Effect of
mease, nitrification and algal inhibitors on ammonia loss and grain yield of flooded rice in Thailand, Fen.
Res. 40, 225-233
Gameh, M.A., Angle, J.S., and Axley, J.H. (1990) Effects of urea-potassium chloride and nitrogen
transformations on ammonia volatilization from urea, Soil Sci. Soc. Am. J. 54, 1768-1772.
Gamzikova, 0.1. and Gamzikov, P.G. (1971) Effect of microelements on the enzymatic activity of soils, in
V.R. Filippov (ed.), Microelements in the Biosphere and Their Agricultural and Medicinal Use in Siberia
and in the Far East, Acad. Sci. USSR, Siberian Section, Buryat Branch, Ulan-Ude, pp. 303-306 (in
Russian).
Gaponyuk, E.I. and Kuznetsova, M.V. (1984) Effect of sodium fluoride on soil properties and development of
some agricultural plants, Gigiena Sanitariya (6),77-79 (in Russian).
369
Garcia, C. and Hernandez, T. (1996) Influence of salinity on the biological and biochemical activity of a
calciorthid soil, Plant Soil 178, 255-263.
Garcia, e., Vallejo, A., Diez, J.A., Garcia, L., and Cartagena, M.e. (1997) Nitrogen use efficiency with the
application of controlled release fertilizers coated with Kraft pine lignin, Soil Sci. Plant Nutr. 43, 443-
449.
Garcia, C., Garcia, L., Vallejo, A., Cartagena, M.e., and Diez, J.A. (1998) Forecasting by laboratory tests of
nitrogen leached and adsorbed in soil-plant system with urea-based controlled-release fertilizers coated
with lignin, Commun. Soil Sci. Plant Anal. 29, 2479-2491.
Garcia, C., Hernandez, T., Pascual, J.A., Moreno, lL., and Ros, M. (2000) Microbial activity in soils of SE
Spain exposed to degradation and desertification processes. Strategies for their rehabilitation, in C. Garcia
and T. Hernandez (eds.), Research and Perspectives of Soil EnzymoloKJi in Spain, Cons. Sup. Invest. Cie.,
Madrid, pp. 93-143.
Gascho, G.J. (1986) Improving the fertilizer efficiency of urea ammonium nitrate solutions by adding other
nutrients, J. Fert. Issues 3 (2), 62-65.
Gascho, GJ. and Burton, G.W. (1987) Nutrient additions to urea ammonium nitrate for improving N
management of Tifton 44 bermudagrass, J. Fert. Issues 4 (3), 86-90.
Gauthier, S.M., Ashtakala, S.S., and Lenoir, lA. (1976) Inhibition of soil urease activity and nematocidal action of
3-amino-l,2,4-triazole, HortScience 11, 481-482.
Gautney, J. (1987) Fluid fertilizers containing thiophosphoryl triamide, U.S. Patent No. 4,676,822.
Gautney, J., Kim, Y.K., and Gagen, P.M. (198.],) Feasibility of incorporating the nitrogen loss inhibitors
dicyandiamide, thiourea, phenyl phosphorodiamidate, and potassium ethyl xanthate into granular urea,
Tennessee Valley Authority Bull. (Y-182), 1-27.
Gautney, J., Kim, Y.K .. and Gagen, P.M. (1984) Feasibility of cogranulating the nitrogen loss inhibitors
dicyandiamide, thiourea, phenyl phosphorodiamidate, and potassium ethyl xanthate with urea, Ind. Eng. Chem.
Prod. Res. Del~ 23,483-489.
Gautney, J., Kim, Y.K., and Barnard, A.R. (1985) Solubilities and stabilities of the nitrogen loss inhibitors
dicyandiamide, thiourea, and phenyl phosphorodiamidate in fluid fertilizers, Ind. Eng. Chem. Prod. Res. Dell.
24,155-161.
Gautney, J., Barnard, A.R., Penney, D.B., and Kim, Y.K. (1985) Solid-state decomposition kinetics of phenyl
phosphorodiamidate, Soil Sci. Soc. Am. 1. 50, 792-797.
Gautney, J., Worley, S.D., and Ash, D.H. (1990) N,N'-Dihalo-2-imidazolidinones and N-halo-2-
oxazolidinones as urease and nitrification inhibitors, u.s. Patent No. 4,954,156.
Geissler, P.R., Sor, K., and Rosenblatt, T.M. (1970) Urease inhibitors, U.S. Patent No. 3,523,018.
Germann-Bauer, M. (1987) Effect and Decomposition of the Nitrification Inhibitor Guanylthiourea, Tech.
Univ. Munich, Wei hen stephan (in German).
Gezgin, S. and Bayrakli, F. (1995) Anunonia volatilization from ammonium sulfate, ammonium nitrate, and
urea surface applied to winter wheat on a calcareous soil, J. Plant Nutr. 18, 2483-2494.
Gill, J.S., Singh, B., Khind, e.S., and Singh, Y. (1999) Efficiency of N-(n-butyl)thiophosphoric triamide in
retarding hydrolysis of urea and ammonia volatilization losses in a flooded sandy loam soil amended with
organic material, Nutr. Cycling Agroecosyst. 53, 203-207.
Goldie, J., Veldhuyzen van Zanten, SJ.O., Jalai, S., Richardson, H., and Hunt, R.H. (1991) Inhibition of
urease activity but not growth of Helicobacter pylori by acetohydroxamic acid, J. Clin. Pathol. 44, 695-
697.
Gomab, A.H.M., Al Nahidh, S.I., and Amer, H.A. (1990) Amidase and urease activity in soil as affected by
sludge, salinity and wetting and drying cycles, Z. Pflanzen ern. Bodenk. 153,215-218.
Gonzalez Carcedo, S. and Perez Gonzalez, J.A. (1980) Influence of humic acids on the soil urease activity, Bull.
Liaison-Groupe Polyphenols (Narbonne) (9), 311-316.
Goodroad, L.L. and Wilson, D.O. (1989) Response of corn, grain sorghum, and winter small grains to NBPT-
treated urea, Agron. Abstr., p. 240.
Goos, R.J. (l985a) Identification ofanunonium thiosulfate as a nitrification and urease inhibitor, Soil Sci. Soc.
Am. J. 49,232-235.
Goos, RJ. (l985b) Urea hydrolysis and ammonia volatilization characteristics of fluid fertilizer mixtures. I.
Laboratory studies, J. Fert. Issues 2 (2), 38-41.
Goos, R.J. (1987) Ammonium thiosulfate as a urease inhibitor. A suggested mechanism, Proc. 17th North
Central Extension-Industry Soil Fertility Work.shop (St. Louis, Missouri, 1987), Potash and Phosphate
Inst., West Lafayette, Indiana, pp.l03-105.
Goos, RJ. and de Padua Cruz, A. (1999) Effect of ammonium sulfate pretreatment on ammonia volatilization
after urea fertilization, Commun. Soil Sci. Plant Anal. 30, 1325-1336.
370
Goos, RJ. and Fairlie, T.E. (1988) Effect of ammonium thiosulfate and liquid fertilizer droplet size on urea
hydrolysis, Soil Sci. Soc. Am. J. 52, 522-524.
Goos, R.J. and Johnson, B.E. (1992) Effect of ammonium thiosulfate and dicyandiamide on residual
ammonium in fertilizer bands, Commun. Soil Sci. Plant Anal. 23, 1105-1117.
Goos, RJ., Voss, R.P., and Fairlie, T.E. (1986a) Ammonium thiosulfate as a possible nitrification and urease
inhibitor, Sulphur Agric. 10,2-5.
Goos, R.I., Voss, R.P., Fairlie, T.E., and Johnson, B.E. (1986b) Containing N loss, Solutions 30 (1), 40-45.
Goos, R.I., Johnson, B.E., and Ahrens, W.H. (1990) Ammonium thiosulfate research 1988-1990, in J.L.
Havlin and J.S. Jacobsen (eds.), Proc. Great Plains Soil Fertility Con! (Denver, Colorado, 1990), Kansas
State Univ., Manhattan, pp. 63-67.
Gorelik, L.A., Gritsevich, Yu.G., and Paevskaya, N.!. (1983) Effect of urease inhibitors on hydrolysis and
effectiveness of surface-applied urea, Agrokhimiya (3), 9-15 (in Russian).
Goring, c.A.1. (1962) Control of nitrification by 2-chloro-6-(trichloromethyl) pyridine, Soil Sci. 93,211-218.
Gould, W.O. (1979) Inhibition of urease activity, Effictive Use of Nutrient Resources in Crop Production.
Proc. Alberta Soil Sci. Workshop, Alberta Agric., Edmonton, pp. 143-158.
Gould, W.O., Cook, F.D., and Webster,G.R. (1973) Factors affecting urea hydrolysis in several Alberta soils,
Plant Soil3S, 393-40 1.
Gould,WD., Cook, F.D., and Bulat, J.A. (1978) Inhibition of urease activity by heterocyclic sulfur
compounds, Soil Sci. Soc. Am. J. 42, 66-72.
Grant, CA. and Bailey, L.D. (1997) Effectiveness of urease inhibitors on the Canadian prairies, Proc. 40th
Annu. Manitoba Soc. Soil Sci. Meeting, pp. 191-200.
Grant, CA. and Bailey, L.D. (1999) Effect of seed-placed urea fertilizer and N-(n-butyl)thiophosphoric
triamide (NBPT) on emergence and grain yield ofbarley, Can. J. Plant Sci. 79,491-496.
Grant, CA., Jia, S., Brown, K.R., and Bailey, L.D. (1996a) Volatile losses of NH3 from surface-applied urea
and urea ammonium nitrate with and without the urease inhibitors NBPT or ammonium thiosulphate,
Can. J. Plant Sci. 76, 417-419.
Grant, CA., Ferguson, R., Lamond, R., Schlegel, A., and Thomas, W. (1996b) Use of urease inhibitors in the
Great Plains, Proc. Great Plains Soil Fertility Con! (Denver, Colorado, 1996), 14 pp.
Graziano, P.L. (1990) Improvement of nitrogen efficiency by addition of ammonium thiosulfate in the liquid
fertilization of maize, Feri. Res. 24, 111-114.
Graziano, P.L. and Parente, G. (1996) Response of irrigated maize to urea-ammonium nitrate and ammonium
thiosulphate solutions on a sulphur deficient soil, Nutr. Cycling Agroecosyst. 46, 91-95.
Grego, S., De Cezare, F., Marchi, G., and Badalucco, L. (1995a) NBPT inhibition of urea degradation in field
trials and its environmental implications, Riv. Agron. (Bologna) 29, 409-414 (in Italian).
Grego, S., Badalucco, L., De Cezare, F., Moscatelli, C, Gioacchini, P., and Vittori Antisari, L. (1995b)
Efficacy ofNBPT to slow down the degradation of urea in field conditions, Fresenius Environ. Bull. 4,
475-480.
Griffith, D.P., Gibson, J.R., Clinton, C.W., and Musher, D.M. (1979) Acctohydroxamic acid: Clinical studies
of a urease inhibitor in patients with staghom renal calculi, J. Urol. 119,9-15.
Griffith, D.P., Khonsari, F., Skumick, J.H., and James, K.E. (1988) A randomized trial of acetohydroxanic
acid for treatment and prevention of infection induced urinary stones in spinal cord injury patients, J.
Urol. 140, 318-324.
Grigoryan, K.V. (1982) Effect of the ions of heavy metals on activity of soil enzymes, Bioi. Zh. Armenii 35,
653-656.
Gupta, S.K. and Chaudhry, M.L. (1994) Influence of heavy metals and temperature on urea transformation in
a Typic Ustochrept, J. Indian Soc. Soil Sci. 42, 203-208.
Guthrie, T.F. and Bornke, A.A. (1981) Effects of low temperature and nitrification inhibitors on urea
hydrolysis, Can. J. Soil Sci. 61, 529-532.
Hallmark, WE., Brown, L.P., and Hawkins, G.L. (1996) Effect ofN-HIB calcium on nitrogen use efficiency,
Hallmark, WE., Brown, L.P., and Hawkins, G.L. (1998) Effect ofN-HIB calcium on nitrogen use efficiency,
Commun. Soil Sci. Plant Anal. 29. 1376.
Hamid, A. and Ahmad, M. (1987) Ammonia volatilization losses from nitrogen fertilizers in alluvial alkaline soils,
PakistanJ. Agric. Sci. 24,129-139.
Hartbrich, H.-J., von Lenge.xen, J., Klepel, M., Lang, S., Wetterau, H., Hoh, G., Held, P., and Michel, H.-J. (l976a)
Agent for improving utilization of urea in animal fodder, (East) German Patent No. 117,153.
Hartbrich, H.-J., Heinze, M., Jasche, K., Kiihne, V., Rothe, G., Thieme, H., Hiickert, H., Held, P., KJepel, M, Lang,
S., and Siegling, S. (1976b) Agent for reducing the loss of plant-available nitrogen in cultivated soils, (East)
371
Hartbrich, H.-J., Oertel, M, Held, P., Rothe, G., Thieme, H., Klepel, M., and Siegling, S. (1978) Application of
urease inhibitors - a possibility for reducing the ammonia nitrogen losses in urea fertilization. 1. Researches on
the urease-inhibiting potencies of chemical substances, Spez. Agrochem. Forsch. Praxis (Cunnersdorf) 8, 20-
24 (in German).
Heber, R, Miiller, S., Matzel, W., and Ansorge, H. (1979) Ammonia losses in urea fertilization. 4. Effect of the
urease inhibitor phosphoric acid phenyl ester diamide on the amount of NH3 losses and the fertilizing action of
urea in model and pot experiments, Arvh. Acker- Pjlanzenbau Bodenk. 23,231-240 (in German).
Heiseler. G., Huth, w., Jasche, K., Oertel, M., Opel, M., Paul, D., Rauchfu~, G., Thieme, H., and Voigt, U.
(1980) Procedure for production ofN-unsubstituted phosphoric acid ester amides, (East) German Patent
No. 142,714.
Held, P., Lang, S.. Klepel, M., Liebmann, R. Hartbrich, H.-J .• and Rothe, G. (1974) Agent for reducing the
loss of plant-available nitrogen in cultivated soils, (East) German Patent No. 107,664.
Held P., Lang, S., Tradler, E., Hoh, G., Klepel, M., Hartbrich, H.-J., and Rothe, G. (1976a) Agent for reducing
the loss of ammonia nitrogen in urea fertilization, (East) German Patent No. 121,457.
Held P., Lang, S., Tradler, E., Klepel. M., Drohne, D., Hartbrich, H.-J.,Rothe, G., Scheler, H., Grundmeier, S.,
and Trautmann, A. (l976b) Agent for reducing the loss of plant-available nitrogen in cultivated soils,
(East) German Patent No. 122,177.
Held, P., Lang, S., Oertel, M., Rothe, G .. Klepel, M, Thieme, H., and Hartbrich, H.-J. (1978) Application of urease
inhibitors - a possibility for reducing the ammonia nitrogen losses in urea fertilization. 2. Researches on
directed synthesis and biological control of phosphoric acid ester amides with urease-inhibiting effect, Spez.
Agrochem. Forsch. Praxis (Cunnersdorf) 8, 25-30 (in German).
Hemida, S.K., Omar, S.A.. and Abdel-Mallek, A.Y. (1997) Microbial populations and enzyme activity in soil
treated with heavy metals, Water. Air. Soil Pollut. 95, 13-22.
Hendrickson, L.L. (1992) Com yield response to the urease inhibitor NBPT: Five-year summary, J. Production
Agric. 5. 131-137.
Hendrickson, L.L. and Douglass, E.A. (1993) Metabolism of the urease inhibitor N-(n-butyl) thiophosphoric
triamide (NBPT) in soils, Soil Bioi. Biochem. 25,1613-1618.
Hendrickson, L.L. and O'Connor. M-J. (1987) Urease inhibition by decomposition products of
phenylphosphorodiamidate, Soil Bioi. Biochem. 19, 595-597.
Hendrickson, L.L., Omholt, T.E., and O'Connor, M-J. (1983) Role of urease inhibitors in increasing efficiency of
surface-applied urea, Agron. Abstr., p. 156.
Hendrickson, L.L., arnholt, T.E., and O'Connor, M-J. (1987) Effect of phenylphosphorodiamidate on
immobilization and ammonia volatilization, Soil Sci. Soc. Am. J. 51, 1067-1071.
Hensen. M.A., Westfall. D.G., and Ludwick, A.E. (1984) Sulfur nutrition of center-pivot irrigated com and dryland
winter wheat, J. Fert. Issues 1 (1),69-74.
Hera, C. and Eliade, G. (1981) Nitrogen losses through leaching of soils in Romania, Prob!. Agrofitoteh. Teor. Api.
(Fundulea) 3 (4), 309-324 (in Romanian).
Hera, C., Kiss, S., Calancea, L., Dragan-Bularda. M., Riidulescu, D., Bologa, M., Firu, v., Ghinea, L., Heidel, H.,
and Popescu. A. (1980) Inhibitor of urea hydrolysis in soil, Romanian Patent No. 68,084.
Hera, C .• Chiri\ii. v., and Ghinea, L. (1986) Optimization of nitrogen regime in soil by using enzyme inhibitors,
Prabl. Agrofitoteh. Teor. Api. (Fundulea) 8 (2),135-152 (in Romanian).
Hickisch, 8. (1981) Side effects of agrochemicals on soil microorganisms, Wiss. Z. Martin-Luther-Univ. Halle-
Wittenberg, Math. Naturwiss. Reihe 30 (3),127-132 (in German).
Hoeft, RG. (1986) Urease inhibitors - how well did they work?, in RG. Hoeft (ed.), Proc. Illinois Fert. and Chem.
Assoc. Con! (Urbana, 1986), pp. 1-15.
Hong, T. and Chen, D. (1997) Preparation of slow-release granular urea fertilizer, Chinese Patent No. 1,173,482.
Chern. Abstr., 2000, 132, P 78008x.
Hongprayoon, c., Lindau, C.W., Partick, W.H.jr.• Bouldin, D.R, and Reddy, K.R (1991) Urea transfonnations in
flooded soil columns. 1. Experimental results, Soil Sci. Soc. Am. J. 55, 1130-1134.
Honza, J. and MiruIl; J. (1990) Ammonia volatilization after application of nitrogen fertilizers and possibilities for
its reduction by addition of hexaaminocycIotriphosphazatriene (HA-CTPAT), Rostl. Vyroba 36, 791-796 (in
Czech).
Houimel, M, Mach, J.-P., Corthesy-Theulaz, I., Corthesy, 8., and Fisch, I. (1999) New inhibitors of Helicobacter
pylori urease holoenzyme selected from phage-displayed peptide libraries, Eur. J. Biochem. 262, 774-780.
Howard, D.D., Mullen, M.D., and Revelle, MA. (1992) Inhibition of urea hydrolysis by N-(n-butyl)
thiophosphoric triamide, Agron. Abstr., pp. 259-260.
Huang, Z., Wu, G., Li, W. et al. (1993) Method for producing long-acting urea fertilizer containing urease inhibitor,
Chinese Patent No. 1,073,933. Chern. Abst., 1994, 120, 7722s.
372
HuIagur, B.F. and Shinde, J.E. (1984) Effect of nonedible oil cakes and neem extract on urea hydrolysis and
ammonia volatilization loss in flooded soils, Bull. Indian Soc. Soil Sci. (New Delhi) (13),138-144.
Hyson, AM. (1963) Fertilizer composition comprising urea and dithiocarbamates, U.S. Patent No. 3,073,694.
IFDC (International Fertilizer Development Center, Muscle Shoals, Alabama) (1981) Nitrogen research, Annual
Report 1981, pp. 23-33.
Illarionov, V.V., Pogodilova, E.G., Lenskii, A.S., Sokolov, A.V., Koritskaya, T.S., and Azbikina, Yu.V. (1968)
Phosphorus-nitrogen fertilizer, USSR Patent No. 216,760.
Imamura, L., Tsuchiya, M., Inada, A, Nakanishi, T., and Kobashi, K. (1995) Inhibition of urease and growth of
Helicobacter pylori by hem extracts, Wakou Iyakugaku Zasshi 12, 129-\36. Chem Abstr., 1996, 124,
140618m
IMC (International Minerals and Chemical Inc.) (1995) Agrotain Urease Inhibitor Research Data. Corn
Research Data, pp. 1-14.
Ioppolo,A., D'Arrigo, C.M., and Radaelli, L. (1989) Influence of sodium chloride on urease activity,
Agrochimica 33, 61-68 (in Italian).
lto, H., Myazaki, T., Nakamura. T., and Kobashi, K. (1994) Preparation of hydroxamic acid derivatives of
hexoses as urease inhibitors, Japanese Patent No. 06,156,375. Chem Abstr., 1995,122, 133675h.
Ito, Y., Shibata, K., Hongo, A, and Kinoshita, M. (1998) Ecabet sodium. a locally acting antiulcer drug,
inhibits urease activity ofHelicobacter pylori, Eur. J Pharmacal. 345,193-198 .
.lanzen, H.H. and Bettany, J.R. (1986) Influence of thiosulfate on nitrification of ammonium in soil, Soil Sci.
Soc. Am. J. 50, 803-806.
Jasche, K., Kom R., Oertel, M., Rauchfu~, G., Stoye, H., Thieme, H., and Voigt, U. (1977) Procedure for
production of phosphoric acid phenyl ester diamide, (East) German Patent No. 128,315.
Jasche, K., Thieme, H., Oertel, M., and Haeckert, H. (1978) Use of urease inhibitors in urea fertilization, Proc.
Sci. and Tech. Conf on Mineral Fertilizers and Their Use in the "Agricultural Economy" (Varna, 1978),
W·276-282 .
.lohn, P.S., Buresh, RJ., Pandey, R.K., Prasad, R., and Chua, T.T. (1989) Nitrogen-IS balances for urea and
neem coated urea applied to lowland rice following two cowpea crowing systems, Plant Soil 120, 233-
241.
.lones, G.A. (1968) Influence of acetohydroxamic acid on some activities in vitro of the rumen microbiota,
Can. J Microbiol.14, 409-416 .
.lones, G.A. and Milligan, J.D. (1975) Influence on some rumen and blood parameters of feeding
acetohydroxamic acid in a urea-containing ration for lambs, Can. J Anim. Sci. 55, 39-47.
Joo, Y.K. and Christians, N.E. (1986) The response of Kentucky bluegrass turf to phenylphosphorodiamidate
(PPD) and magnesium (Mg++) applied in combination with urea, J, Fert. Issues 3(1),30-33.
Joo, Y.K., Christians, N.E., and Bremner, J.M. (1987) Effect ofN-(n-butyl) thiophosphoric triamide (NBPT)
on growth response and ammonia volatilization following fertilization of Kentucky bluegrass (Poa
pratensis L.) with urea, J. Fert. Issues 4 (3),98-102.
Joo, Y.K., Christians. N .E .. and Bremner. J.M. (1989) Effectiveness of urease inhibitors and cationic materials for
reduction of anunonia volatilization from turfgrasses treated with urea, Proc. 6th Int. TUfjgrass Res. Conf
(Tokyo, 1989), pp. 209-211.
Joo, Y.K., Christians, N.E., and Bremner, J.M. (1991a) Turf response to urease inhibitors and cationic materials
applied in combination with urea, HortScience 26, 537-539.
Joo, Y.K., Christians, N.E., and Blackmer, AM. (l991b) Kentucky bluegrass recovery of urea-derived nitrogen-15
amended with urease inhibitor, Soil Sci. Soc. Am. J. 55, 528-530.
Joo, Y.K., Christians, N.E., Spear, G.T., and Bremner, J.M. (1992) Evaluation of urease inhibitors as urea
amendments for use on Kentucky bluegrass turf, Crop Sci. 32. 1397-1401.
Joo, Y.K., Christians N.E., and Blackmer. AM. (1993) Kentucky bluegrass recovery of urea-derived nitrogen-15
amended with urease inhibitor, Agron. Abstr., p. 159.
Joost, R.E. (1995) Management of urea for efficient N fertilization of caucasian bluestem, Agron. Abstr., p. 252.
Joseph, P.A. and Prasad. R. (1995) Effect of dicyandiamide and neem cake blending ofprilled urea on growth
and yield of wheat, Proc. Indian Natl. Sci. Acad., Part B 61, 149-153.
Kamire, K.K. and Sonar. K.R. (1979) Mineralization of urea-N with neem (Azadirachta indica Juss.) and
karanj (Pongamia g/abra Vent.) cakes, J Maharashtra Agric. Univ. 4 (I). 10-13.
Kiimpfe, K., Jauert, R., and Lippert, J. (1981) Influence of the application of PPDA-urea and hemicides on the
emergence of sugaIbeet glomeruies, Arch. Acker- Pjlanzenbau Bodenk. 25, 783-787 (in German).
Kiimpfe, K., Linke, E., Herold, L.. Hartbrich, H.-.l .• and Tomer. D. (1982a) Results of the evaluation of PPDA-urea
under natural and simulated weather conditions in microplot experiments, Arch. Acker- Pjlanzenbau Bodenk.
26,483-490 (in German).
373
Kiirnpfe, K, Linke, E., Herold, L., and Tomer, D. (1982b) Results of the evaluation of PPDA-urea in field
experiments with winter cereals, Arch. Acker- Pjlanzenbau Bodenk. 26, 714-724 (in German).
Kiimpfe, K, Muller, S., Beer, K, and Oertel, M. (1982c) International trials on PPDA-urea, Mezhdunar.
Sel'skokhoz. Zh. (4),46-48 (in Russian); Results of the detennination of gaseous ammonia nitrogen losses and
of the estimation of the effect of PPDA-urea on yield, within an international project for fertilizer evaluation,
Int. Z. Landw. (4), 351-353 (in German).
Kiimpfe, K, Linke, E., Schnee, H., and Herold, L. (1983) Results of the evaluation of PPDA-urea in winter cereal
field experiments on lruge surfaces, Arch. Acker- Pjlanzenbau Bodenk. 27, 177-184 (in German).
Kandeler, E., Mentler, A., Pfeffer, M., and Horak, O. (1990) Soil biological estimation of the toxicity of heavy
metals in artificially contaminated soils, VDLUFA-Schrifienreihe 32,621-626 (in German).
Katyal, J.e., Singh, B., Vlek, P.L.G., and Buresh, RJ. (1987) Efficient nitrogen use as affected by urea application
and irrigation sequence, Soil Sci. Soc. Am. J. 51,366-370.
Keeney, D.R. and Sahrawat, KL. (1986) Nitrogen transformations in flooded rice soils, Fert. Res. 9, 15-38.
Keerthisinghe, D.G. and Freney, J.R. (1994) Inhibition of urease activity in flooded soils: Effect of
thiophosphoric triamides and phosphoric triamides, Soil Bioi. Biochem. 26, 1527-1533.
Ketkar, C.M. and Ketkar, M.S. (1995) Neem seed crush and deoiled cake as manure and as nitrification
inhibitors, in H. Schmutterer (ed.), Neem Tree Azadirachta indica A. Juss. and Other Meliaceous Plants,
VCH Verlagsges., Weinheim, pp. 531-539.
Khabirov, I.K, Khaziev, F.Kh., Prostyakova, Z.G., Latypov, Sh.A., and Kaspranskii, N.N. (1992) Regulation of the
nitrogen regime of soils by applying nitrification inhibitors, Pochvovedenie (8), 67-76 (in Russian).
Khadzhiev, T.Kh. (1985) Researches of the soil microbiology and biochemistry laboratory over the last decade
(Soil Sci. and Agrochem Ins!. Acad. Sci. Uzb. SSR), The Soils of Uzbekistan and Ways ofIncreasing Their
Fertility, Izd. Mekhnat, Tashkent, pp. 89-99 (in Russian).
Khanif, Y.M. and Husin, A. (1992) Improvement of fertilizer N recovery of rice by dicyandiarnide and
hydroquinone, Proc. Int. Symp. on Paddy Soils (Nanjing, 1992), Acad. Sin., Beijing, pp. 132-137.
Killeen, G.F., Buggie, KA., Hynes, MJ., Walsh, G.A., Power, R.F., and Headon, D.R. (1994) Influence of
Yucca schidigera preparations on the activity of urease from Bacillus pasteurii, J. Sci. Food Agric. 65,
433-440.
Kiss, S. and Pintea, H. (1987) Testing of the inhibitory capacity of some organic compounds on soil urease
activity, Publ. Rom. Natl. Soc. Soil Sci. (Bucharest) (23 B), 103-116 (in Romanian).
Kissel. D.E. (1984) Effects of P source and method ofN-P application on phosphorus availability to winter
wheat, J. Fert. bsues 1 (I), 125-129.
Knappe, S. and Carrion, N (1987) Use of PPDA urease inhibitor to improve the N effect of urea in tropical
soils, Abstr., Int. Fair Symp. "Agrochem" (Leipzig, 1987), pp. 26-27.
Knop, K (1982) Gaseous nitrogen losses from fertilizers in form of NH 3 , Rostl. Vyroba 28, 935-946 (in
Czech).
Kobashi, K, Munakata, K-I., Takabe, S., and Hase, J.-I. (1980) Therapy for urolithiasis by hydroxamic acids.
II. Urease inhibitory potency and urinary excretion rate of hippurohydroxamic acid derivatives, J.
Pharmacobiodyn. 3, 444-450.
Kole, J.F., Swerdloff, M.D., Rogic, M.M., Hendrickson, L.L., and Van Der Puy, M. (1984) Novel N-aliphatic and
N,N-aliphatic phosphoric triarnide urease inhibitors and urease inhibited urea based fertilizer compositions,
Eur. Patent Appl.. Publ. No. 0 119 487 A J.
Kole, J.F., Swerdloff, M.D., Rogic, MM., and Hendrickson, L.L. (1985a) N-Acyl phosphoric triarnide urease
inhibitors and urease inhibited urea based fertilizer compositions, Us. Patent No 4,517,003.
Kole, J.F., Swerdloff, M.D., Rogic, MM., and Hendrickson, L.L. (l985b) Arninophenol urease inhibitors and
urease inhibited urea based fertilizer compositions, US. Patent No. 4,517,005.
Kole, J.F., Swerdloff, M.D., Rogic, M.M., and Hendrickson, L.L. (1985c) Urease inhibited urea based fertilizer
compositions, Us. Patent No.4, 528,020.
Kole, J.F., Swerdloff, M.D., Rogic, M.M., Hendrickson, L.L., and Van Der Puy, M. (1985d) N-Aliphatic and
N,N-aliphatic phosphoric triamide urease inhibitors and urease inhibited urea based fertilizer
compositions, U.S. Patent No. 4,530,714.
Kolyada, T.I. (1970) Effect of thiourea on the biological activity of soil, in S.A. Samtsevich (ed.), Physiology
and Biochemistry ofMicroorganisms, Izd. Nauka i Tekhnika, Minsk, pp. 226-230 (in Russian).
Kolyada, T.I. (1973) Effect of thiourea, applied with mineral fertilizers, on soil biological activity and crop
yield, Soil Investigations and Application ofFertilizers, lzd. Uradzhai, Minsk, pp. 136-142 (in Russian).
Komai, Y., Yamada, K, and Yamaguchi, M. (1981) Effects of heavy metals on several microbial activities in
soil. I. Effects on carbon dioxide evolution, nitrogen immobilization, and urease activity, J. Sci. Soil
Manure 52, 305-310 (in Japanese).
374
Koren'kov, D., Gryzlov, Y., and Kaptsynel', Yu. (1980) Study of the effectiveness of new kinds and fonns of
mineral fertilizers, Mezhdunar. Sel'skokhoz. Zh. (4),25-28 (in Russian); Int. Z. Landw. (4), 337-341 (in
German).
Kozdr6j, J. (1995) The effect of cower and cadmium on urease activity and the development of tolerant
bacteria and fungi in an acid forest soil, Arch. Ochrorry Srodowiska (2),31-38.
Kozlovskaya, N.A, Nikitina, G.D., and Runkov, S,V. (1972) Rapid microdiffusion method for determination
of urease activity in peat bog soils, Agrokhimiya, (5),144-149 (in Russian).
Krasnova, N.M. (1983) Enzyme activity as a bioindicator of soil contamination with heavy metals, Dokl.
VASKhNlL (Moscow) (7), 41-43 (in Russian).
Krasnova, N.M. (1985) Effect of metals on soil biological activity in unsown plots and in root zone of barley
(field trial), Soil Chemistry. Microelements in Soils and Modern Methods for Their Study, V.V.
Dokuchaev Soil Sci. Inst., Moscow, W. 26-29 (in Russian).
Krishnapillai, S. (1979) Inhibition of nitrification by waste tea (tea fluff), Plant Soil 51,563-569.
Krogmeier, MJ., McCarty, G.W., and Bremner, J.M. (1989a) Potential phytotoxicity associated with the use
of soil urease inhibitors, Proc. Natl. Acad. Sci. USA 86,1110-1112.
Krogmeier, M.J., McCarty, G.W., and Bremner, J.M. (1989b) Phytotoxicity of foliar-applied urea, Proc. Natl.
Acad. Sci. USA 86,8189-8191.
Krogmeier, M.J., McCarty, G.W., Shogren, D.R., and Bremner, J.M. (I991a) Effect of nickel deficiency in
soybeans on the phytotoxicity of foliar-applied urea, Agron. Abstr., p. 270.
Krogmeier, M.J., McCarty, G.W., Shogren, D.R., and Bremner, J.M. (1991b) Effect of nickel deficiency in
soybeans on the phytotoxicity of foliar-applied urea, Plant Soil 135, 283-286.
Kubade, G.N. and Mohite, A.V. (1994) Transformation of urea and neem cake coated urea (NCU) in amended
saline sodic and normal soils, J. Maharashtra Agric. Univ. 19,332-335.
Kucharski, J. (1991) The effect of CMP, N-Serve, and ATC on the process of nitrification and on soil
microorganism, Polish J. Soil Sci. 24, 49-56.
Kucharski, J. (1992) The effect of urease inhibitor (PPDA) on winter wheat yield and on soil microorganism
activity, Polish J. Soil Sci. 25,171-176.
Kucharski, J. (1994) Efficiency of the phenyl ester of diamidophosphoric acid in controlling the rate of urea
hydrolysis in soil. Acta Acad. Agric. Tech. Olst.. Agric. (Olsztyn) (57), 83-90 (in Polish).
Kucharski, J. (1997) Relation between activity of enzymes and fertility of soil, Microorganisms in the
Environment - Occurrence, Activity and Role, Dept. Microbiol., Agric. Acad. Cracow, pp. 327-347 (in
Polish).
Kucharski, J. and Niklewska, T. (1992) The influence of zinc on the yields of broadbean and microbiological
activity of soil. Polish J. Soil Sci. 25,71-77.
Kucharski, 1, Czapla, 1, Los, Z., and Procyk, Z (1990) Effect ofthe temporary inhibition of urea hydrolysis
on the yield of spring barley, Fragm. Agron. (Pulawy) 9 (3), 31-39 (in Polish).
Kiihler, T.C., Fryklund, J., Bergman, N.-A, Weilitz, I., Lee, A, and Larson, H. (1995) Structure-activity
relationship of omeprazole and analogues as Helicobacter pylori urease inhibitors, J. Med. Chem. 38,
4906-4916.
Kumar, V. and Wagenet, R.J. (1984) Urease activity and kinetics of urea transformation in soils, Soil Sci. 137,
263-269.
Kumar, V. and Wagenet, R.J. (1985) Salt effects on urea hydrolysis and nitrification during leaching through
soil columns, Plant Soil 85, 219-227.
Kumar, V., Yadav, D.S., and Singh, M. (1988) Effects of urea rates, farmyard manure, CaC03, salinity and
alkalinity levels on urea hydrolysis and nitrification in soils, Australian J. Soil Res. 26,367-374.
Kuprevich, Y.F. (1951) Biological activity of soil and methods for its determination, Dokl. Akad. Nauk SSSR 79,
863-866 (in Russian).
Kurze, R. (1981) Agent for improving the fertilizing action of urea, (East) German Patent No. 146,173.
Kurze, R. and Richter, H. (1978) Procedure for production of phosphoric acid aryl ester amides, (East)
Kurze, R., Thieme, H., Voigt, U., lasche, K., Oertel, M., and Michel, H.-l. (1985) Procedure for production of
metal-phosphoric acid aryl esters with covalent P-N bonds, (East) German Patent No. 218,102.
Lamond, R.E. and Bonczkowski, L.c. (1989) Nitrogen fertilizer management for smooth bromegrass
production, Agron. Abstr., p. 317.
Lamond, R.E., Whitney, D.A., Hickman, 1.S., and Bonczkowski, L.c. (1986) Comparisons of nitrogen rates
and placement methods on no-till grain sorghum, Kansas Fertilizer Research 1986, Report of Progress
(S09), 148-149.
375
Lamond, R.E., Whitney, D.A., Hickman, J.S., and Bonczkowski, L.e. (1987) Comparisons of nitrogen rates
and placement methods on no-till grain sorghum, Kansas Fertilizer Research 1987, Report of Progress
(531),108-111.
Lamond, RE., Whitney, D.A., Hickman, J.S., and Bonczkowski, L.e. (1991a) Nitrogen rate and placement
for grain sorghum production in no-tillage systems, J Production Agric. 4, 531-535.
Lamond, R.E., Havlin, J.L., and Ojiem, J.O, (1991b) Nitrogen fertilizer management for no-till corn
production, Agron. A bstr. , p. 360.
Lamond, R,E" Whitney, D.A., Maddux, L.D., Gordon, 8., Key, D., and Koch, S. (1993) Nitrogen
management for no-till corn and grain sorghum production, Kansas Fertilizer Research 1993. Report of
Progress (697), 93-98.
Lamond, R.E., Whitney, D.A., Maddux, L.D., Gordon, W.B., Martin, V.L., Key, D., and Koch, S. (1994) Grain
sorghum, corn, and soybean fertilization studies, Kansas Fertilizer Research 1994, Report of Progress
(719), 73-80.
Lamond, RE., Thomas, W.L., Wesley, T.T., and David, M.A. (1996) Nitrogen management on bromegrass,
Lamond, R.E., Whitney, D.A., and Pierzynski, G.M. (1998) Nitrogen management and tillage effects on grain
sorghum production, Agron. Abstr., p. 313.
Lang, S., Held, P., Hartbrich, H.-I., Klepel, M., Rothe, G., Hackert, H., and Thieme, H. (1976) Agent for
reducing the ammonia nitrogen loss in urea fertilization, (East) German Patent No. 122,621.
Laura, R.D. and Parshad, M. (1991) Transfonnations of urea in sodic soils. 1. Chemical kinetics of urea
hydrolysis, Int. J Trop. Agric. 9,313-321.
Lebedeva, L.A" Lebedev, S.N., and Edemskaya, N.L. (1995) Effect of heavy metals and lime on urease
activity in soddy-podzolic soil, Vestn. Mosk. Gos. Univ., Ser. 17. Pochvovedenie (2), 68-71 (in Russian).
Lee, Y.G. and Radel, R.J. (1988) Degradation of urease inhibitors in soils, Tennessee Valley Authority Bull.
(Y-205), 1-18.
Leiros, M.C., Trasar-Cepeda, e., Garcia-Fernandez, F., and Gil-Sotres, F. (1999) Defining the validity of a
biochemical index of soil quality, Bioi. Fert. Soils 30, 140-146.
Leis, A.K., Varsa, E,e., and Klubek, B.P. (1989) Effect of anunonium thiosulfate and N-(n-butyl)
thiophosphoric triamide as urease inhibitors on no-till corn, Agron. Abstr., p. 317.
Lethbridge, G .. Bull, AT., and Burns, R.G. (1980) 1,3-~-Glucanase activity in soil and its response to soil
amendment, Rev. teol. BioI. Sol 17, 479-489.
Lewis, A. and Slater, RM. (1979a) Reducing NH,loss from urea, British Patent Appl. No.2 010 797 A.
Lewis, A. and Slater, RM. (l979b) New compositions and procedures for their production and application,
German Patent No. 28 55 036 A I.
Li, S.L. and Xue, Y.8. (1991) Study on the screening and use of urease inhibitors, Turang (Nanjing) 23,96-102 (in
Chinese). Chem Abstr., 1992,116, 105123j.
Li, L.T., Wang, Z.P.. Van Cleemput, 0., and Baelt, L. (1993) Urea N uptake efficiency of ryegrass (Lolium perenne
L.) in the presence of urease inhibitors, Bioi. Fert. Soils 15, 225-228.
Liao, C.F.-H. and Raines, S.G. (l982) Relationship between soil urease inhibition and chemical structure of
selected compounds, Agron. Abstr., p. 191.
Liao, C.F.-H. and Raines, S.G. (1985) Inhibition of soil urease activity by amido derivatives of phosphoric and
thiophosphoric acids, Plant Soil 85, 149-152.
Lichko, RP. and Kiselev, G.G. (1985) Method for detennination of urease activity in soils, USSR Patent No.
1,198,430.
Lichko, RP. and Kiselev, G.G. (1986) Rapid method for detennination of urease activity in soils, Pochvovedenie
(4),132-136 (in Russian),
Liesegang, A., Rothe, G., Hartbrich, H.-J., Lang, S., Siegiing, S., Hackert, H., Thieme, H., Kochmann, W., Pallas,
M., Hesse, 8., Miiller, S., Kramer, W, Weidner, K-F., and Thomas,P. (1976) Agent for reducing the loss of
plant-available nitrogen in cultivated soils, (East) German Patent No. 122,243.
Lightner, J.W., Mengel, D.B., and Rhykerd, e.L. (1990) Anunonia volatilization from nitrogen fertilizer
surface applied to orchardgrass sod, Soil Sci. Soc. Am. J. 54, 1478-1482.
Lim, S.-U. and Seo, Y.-H. (1994) Effect of ammonium thiosulfate on biological activity in a paddy soil,
Han 'guk T'oyang Piryo Hakhoechi 27,40-47 (in Korean). Chern. Abstr., 1995, 123, 54903y.
Lin, XJ., Chen, J,C., Zhen, S.L., Liu, ZZ., and Freney, J.R (1997a) Purification and kinetics of urease inhibitor
from Aspergil/us ochraceus,Acta Microbiol. Sinica 37, 276-280 (in Chinese).
Lin, XJ., Chen, J.e., Zhen, S.L., Liu, Z.Z" and Freney, J.R (1997b) Studies on characteristics, crystal structure and
effectiveness of urease inhibitor from Aspergillus ochraceus Wilh., Chinese J Struct. Chern. 16, 471-474 (in
Chinese).
376
Linke, E. and Kiimpfe, K. (1987) Results of testing PPDA urea in plot experiments on winter grain and grassland,
Abstr., Int. Fair Symp. "Agrochem" (Leipzig, 1987) p. 26.
Linke, E., K3.mpfe, K., Herold, L., and Hartbrich, H.-J. (1982) Results of the detennination of gaseous ammonia
nitrogen losses after PPDA-urea fertilization under conditions of field experiments, Arch. Acker- Pjlanzenbau
Bodenk. 26,477-482 (in German).
Linke, E., K3.mpfe, K., Herold, L., Tomer, D., and Hartbrich, H.-J. (1983) Results of the evaluation of PPDA-urea
in grassland experiments, Arch. Acker- Pjlanzenbau Bodenk. 27,143-149 (in German).
List'anska, J. (1982) Effect of nitrification and urease inhibitors on transformation of nitrogen in soil, Rostl Vyroba
28,577-585 (in Czech).
List'anska, J. (1984) Transformation of nitrogen in soil as affected by nitrification inhibitors, Agrochemia
(Bratislava) 24, 296-298 (in Czech).
List'anska, J. (1993) The transformation of nitrogen of liquid fertilizer N-Sol in soil in model laboratory
experiments, Rostl. Vyroba 39, 471-480 (in Czech).
Lloyd, A.B. and Sheaffe, MJ. (1973) Urease activity in soils, Plant Soil 39, 71-80.
Lu, w., Lindau, Cw., Pardue, J.H., Patrick, W.H.jr., Reddy, K.R., and Khind, C.S. (1989) Potential of
phenylphosphorodiamidate and N-(n-butyl) thiophosphoric triamide for inhibiting urea hydrolysis in simulated
oxidized and reduced soils, Commun. Soil Sci. Plant Anal. 20, 775-788.
Ludden, P.A, Harmon, D.L., Larson, B.T., and Axe, D.E. (2000a) Influence of the novel urease inhibitor N-(n-
butyl) thiophosphoric triamide on ruminant nitrogen metabolism. I. In vitro urea kinetics and substrate
digestion,J Anim. Sci. (Savoy, Illinois) 78,181-187.
Ludden, P.A, Harmon, D.L., Huntington, G.B., Larson, B.T., and Axe, D.E. (2000b) Influence of the novel urease
inhibitor N-(n-butyl) thiophosphoric triamide on ruminant nitrogen metabolism. II. Ruminal nitrogen
metabolism, diet digestibility, and nitrogen balance in lambs, J Anim. Sci. (Savoy, Illinois) 78, 188-198.
Luo, Q.-X., Freney, J.R., Keerthisinghe, D.G., and Peoples, M.B. (1994) Inhibition of urease activity in flooded
soils by phenylphosphorodiamidate and N-(n-butyl) thiophosphoric triamide, Soil BioI. Biochem. 26, 1059-
1065.
Ma, L., Hongprayoon, c., Lindau, Cw., and Selim, H.M. (1995) Transport of urea in flooded soil profiles as
affected by water percolation rates and NBPT, Soil Sci. t60, 101-110.
MacKenzie, AF., Phillip, L.E., and Kirby, P.C (1988) Effect of added urea and potassium chloride on yields of
com over four years and on soil potassium, Agron. J. 80, 773-777.
Maddux. L.D. and Barnes, P.L. (1991) Nitrogen placement on reduced tillage com,Agron. Abstr., p. 361.
Maddux, L.D., Kissel, D.E., Ball, J.D., and Raney, R.I. (1985) Nitrification inhibition by nitrapyrin and volatile
sulfur compounds, Soil Sci. Soc. Am. J. 49, 239-242.
Mago, G.S. and Totawat, K.L. (1989) Efficacy of coating materials on urea applied under saline water irrigation. I.
Mineralization ofurea-N, Ann. Arid Zone 28, 79-87.
Mahajan, KK and Tripathi, B.R. (1991) Leaching losses of nitrogen from different forms of urea in columns of
Alfisols, J Indian Soc. Soil Sci. 39,618-624.
Mahendrappa, M.K. and Odgen, E.D. (1973) Patterns of ammonia volatilization from a forest soil, Plant Soil 38,
257-265.
Mahler, R.L. and Lutcher, L.K. (1989) Urea-ammonium nitrate-ammonium thiosulfate applications for
dryland winter wheat, J. Fert. Issues 6 (3), 50-55.
Mai, H. and Fiedler, H.l. (1986) On the effect of the urease inhibitor PPDA on the soil microflora in a spruce
ecosy~tem, Arch. Acker- J>flanzenbau Bodenk. 30, 51-60 (in German).
Malhi, S.S. and Nyborg, M. (1979) Rate of hydrolysis of urea as influenced by thiourea and pellet size, Plant
Soil SI, 177-186.
Malhi, S.S. and Nyborg, M. (1982) An evaluation of carbon disulphide as a sulphur fertilizer and as a
nitrification inhibitor, Plant Soil 65, 203-218.
Malhi, S.S. and Nyborg, M. (1984) Inhibiting nitrification and increasing yield of barley by band placement of
thiourea with fall-applied urea, Plant Soil 77, 193-206.
Malhi, S.S. and Nyborg, M. (1988) Effect of ATC, N-Serve 24E and thiourea nitrification inhibitors on yield
and N uptake of barley fertilized with fall-applied N. Plant SoiltOS, 223-229.
Markert, S. (1974) On the possibility of regulating urea transformations in soil by some commercial
pesticides, Trans. 10th Int. Congr. Soil Sci. (Moscow, 1974) 9, 133-140 (in German).
Martelli, A, Bulli, P., and Cortecchia, V. (1981) Urease inhibitor therapy in infected renal stones, Eur. Urol. 7,
291-293.
Martens, D.A. and Bremner, I.M. (1982) Use of phosphoroami des for retardation of urea hydrolysis in soils,
Martens, D.A and Bremner, I.M. (1983a) Determination of phenylphosphorodiamidate in soil extracts and
urea fertilizer solutions, Commun. Soil Sci. Plant Anal. 14,915-923.
377
Martens, D.A. and Brenmer, J.M. (l983b) Fate of phenyl phosphorodi amidate added to soils, Agron. Abstr., p.
IS8.
Martens, D.A. and Bremner, J.M. (1984a) Urea hydrolysis in soils: Factors influencing the effectiveness of
phenylphosphorodiamidate as a retardant, Soil Bioi. Biochem. 16, SIS-SI9.
Martens, D.A. and Bremner, J.M. (l984b) Etlectiveness of phosphoroamides for retardation of urea hydrolysis in
soils, Soil Sci. Soc. Am. J. 48, 302-30S.
Marusina, N.M. (1981) Effect of heavy metals on enzymatic activity of soil and development of barley plants,
Byuli. Pochv. Inst. im. V V Dokuchaeva (Moscow) (27), 27-30 (in Russian).
Matzel, W. and Heber, R. (1979) Effect of urease inhibitors on NH3 losses following application of urea,
Tagungsber. Akad. Landwirlschafiswiss. DDR (162), 231-238 (in Russian).
Matzel, w., Heber, R., Ackennann, W., and Teske, W. (1978) Ammonia losses following urea fertilization. 3.
Ammonia volatilization as influenced by urease inhibitors, Arch. Acker- Pjlanzenbau Bodenk. 22, 18S-191 (in
Gennan).
Matzel, W., Lippold, H., and Heber, R. (1979a) Nitrogen uptake and balance of the fertilizers urea, urea with urease
inhibitor, and ammonium nitrate applied to spring wheat at stem elongation growth stage, Isotopes and
Radiation in Research on Soil-Plant Relationships, Int. Atomic Energy Agency, Vienna, pp. 67-82.
Matzel, w., Miiller, S., Lippold, H., and Heber, R. (1979b) Effect of the urease inhibitor diamidophosphoric
acid phenyl ester on the utilization of urea nitrogen in topdressing of spring wheat and oats, Arch. Acker-
Pjlanzenbau Bodenk. 23,469-477 (in German).
May, P.B. and Douglas, L.A. (197S) Gennination of wheat and alfalfa seeds as affected by some soil urease
inhibitors, Agron. J. 67, 718-720.
May, P.B. and Douglas, L.A. (1976) Assay for urease activity, Plant Soil 45, 301-30S.
May, P.B. and Douglas, L.A. (1978) Use of soil urease inhibitors to increase the efficiency of urea as a fertilizer,
Proc. 8th Int. Colloq. on Plant Analysis and Fertilizer Problems (Auckland, New Zealand, 1978), pp. 339-34S.
May, P.B. and Douglas, L.A. (1979) Effect of catechol, p-benzoquinone and 2,S-dimethyl-p-benzoquinone on soil
enzymes, Studies about Humus. Humus et Planta VII, Trans. Int. Symp. (Bmo, 1979), pp. 256-260.
McCarty, G.W. and Bremner, J.M. (1989a) Laboratory evaluation of dicyandiamide as a soil nitrification inhibitor,
Commun. Soil Sci. Plant Anal. 20, 2049·206S.
McCarty, G.W. and Bremner, 1.M. (1989b) Formation of phosphoryl triamide by decomposition of
thiophosphoryl triamide in soil, Bioi. Fert. Soils 8,290-292.
McCarty, G.W. and Bremner, 1.M. (1990a) Evaluation of2-ethynylpyridine as a soil nitrification inhibitor, Soil Sci.
Soc. Am. J 54, 1017-1021.
McCarty, G.w. and Bremner, 1.M. (l990b) Evaluation of 3-methylpyrazole-I-carboxamide as a soil nitrification
inhibitor, Boil. Fert. Soils 9,252-256.
McCarty, G.w., Bremner, 1.M., and Chai, H.S. (1989) Effect ofN-(n-butyl) thiophosphoric triamide on hydrolysis
of urea by plant, microbial, and soil urease, Bioi. Fert. Soils 8, 123-127.
McCarty, G.W., Bremner, 1.M., and Krogmeier, MJ. (1990) Evaluation of ammonium thiosulfate as a soil
urease inhibitor, Fert. Res. 24, 135-139.
McCarty, G.W., Brenmer, 1.M., and Krogmeier, MJ. (1991) Effects of thiosulfate and tetrathionate on urease
activity and seedling growth, Commun. Soil Sci. Plant Anal. 22, 7SS-768.
McCarty. G.W., Shogren, D.R.. and Bremner. J.M. (1992) Regulation of urease production in soil by
microbial assimilation of nitrogen, Bioi. Fert. Soils 12. 261-264.
McClung. G. and Frankenberger. W.T.jr. (1985) Soil nitrogen transtormations as affected by salinity, Soil Sci. 139.
40S-4I1.
McVay, K.A., Stecker, 1.A., Buchholz, D.D., and Odhiambo, G.D. (1991) Nitrogen source and placement in
no-till corn, Agron. Abstr., p. 361.
Medina, R. and Sullivan, 1.M. (1986) Cyclotriphosphazatriene-derivatives, u.s. Patent No. 4,618.691.
Medina. R. and Sullivan, 1.M. (1987) Cyclotriphosphazatriene-derivatives as soil urease activity inhibitors,
us. Patent No. 4.670,038.
Meier, 1.N .. Fyles, 1.W., MacKenzie, A.F., and O'Halloran, LP. (1993) Effects of lignosulfonate-fertilizer
applications on soil respiration and nitrogen dynamics, Can. J Soil Sci. 73. 233-242.
Mekhtiev, S.Ya., Marinescu, C.M., and Kravchuc. I.A. (1988) Effectiveness of the nitrification inhibitors.
Effictiveness of the Use of Ferlilizers in Agriculture of Moldavia, Izd. ~tiinta. Kishinev, pp. 149-160 (in
Russian).
MergeI', A.A., Semenov. V.M., and Sokolov, O.A. (1987) Effect of focally applied, concentrated nitrogen fertilizers
on the nitrogen regime and enzyme activity in grey forest soil, Pochvovedenie (12), S5-63 (in Russian).
Michaud, H" Rock. H.• and Seeholzer, 1. (1978) Procedure for production of odor-free prilled urea-dicyandiamide
fertilizers, GermanPatentNo.2714601 A I.
378
Millner, O.E.jr., Andersen, J.A., Appler, M.E., Benjamin, C.E., Edwards, J.G., Humphrey, D.T., and Shearer, E.M.
(1982) Flurofamide: A potent inhibitor of bacterial urease with potential clinical utility in the treatment of
infection induced urinary stones,J. Uro1.127, 346-350.
Miruii; J., Honza. 1.. and Zehmilek. J. (1990) Hexaaminocyciotriphosphazatriene (HA-CTPAn - an effective
inhibitor of soil urease, Pol'nohospodarstvo 36, 691-697 (in Czech).
Mishra, M.M. and Flaig, W. (1979) Inhibition of mineralization of urea nitrogen in soil. Plant Soil 51. 301-309.
Mishra, M.M., Flaig, W., and Soechtig, H. (1980) The effect of quinoid and phenolic compounds on urease and
dehydrogenase activity and nitrification in soil. Plant Soil 55, 25-33.
Mishra, B., Shanna. R.D., and Murthy, J.R. (1990) Effect of neem cake and shellac coating of urea and green
manure on ammonia volatilization and nitrogen use efficiencies for rice. J. Indian Soc. Soil Sci. 38. 224-228.
Mitsui, S., Namioka, H., and Mukai, N. (1960) Soil adsorption of urea. I. On the mechanism of adsorption of urea.
Soil Plant Food (Tokyo) 6,25-29.
Moawad, H .• Zohdy. LI., Garnal EI-Din. H., and Badr EI-Din, S.M.S. (1979a) Transformations of soil nitrogen
associated with urea fertilization combined with pesticide application, Egypt. J. Microbial. 14,21-28.
Moawad, H., Zohdy, LI.. Badr EI-Din. S.M.S., and Garnal EI-Din. H. (l979b) Changes in certain biological and
chemical properties of the urea-treated soil as affected by application of some pesticides, Egypt. J. Microbial.
14,29-36.
Moawad, H., Mahmoud, S.AZ .• Badr EI-Din, S.M.S., Garnal R-F., and Enany, M.H. (1984) Transformations and
effects of urea derivatives in soil, Z Pjlanzenem. Bodenk.147. 785-792.
Mobley, H.LT. and Hausinger. R-P. (1989) Microbial ureases: Significance. regulation, and molecular
characterization. Microbial. Rev. 53, 85-108.
Moe, P.G. (1967) Nitrogen losses from urea as affected by altering soil urease activity, Soil Sci. Soc. Am. Proc.31,
380-382.
Monreal, c., McGill, W.B., and Nyborg, M. (1986) Spatial heterogeneity of substrates: Effects on hydrolysis,
immobilization and nitrification ofurea-N, Can. J. Soil Sci. 66, 499-511.
Montemurro, F., Capotorti, G., Lancertosa, G., and Palazzo, D. (1998) Effects of urease and nitrification
inhibitors application on urea fate and nitrate accumulation in lettuce, J. Plant Nutr. 21, 245-252.
Moreno, J.L, Garcia, c., Landi, L.. Falchini. L. Pietramellara, G., and Nannipieri, P. (2001) The ecological
dose value (EDso) for assessing Cd toxicity on ATP content and dehydrogenase and urease activities of
soil, Soil Bioi. Biochem. 33, 483-489.
Mullen, M.D. and Revelle. M.A. (1992) Inhibition of urea hydrolysis and arnrnonia volatili7.ation by the
urease inhibitor N-(n-butyl) thiophosphoric triamide, Agron. Abstr., Appendix 1, p. 19.
Mullen, M.D .. Howard, D.O., and McClure. J.M. (1991) Effect ofN-(n-butyl)thiophosphoric triamide on urea
hydrolysis in high residue soil systems, Agron. Abstr., p. 362.
Muller, G. and Forster, I. (1980) Contributions to the problem of microbially induced urea transformation in
soil. II. Influence of nitrogen on the carriage and the effect of urease inhibitors. Zbl. Bakteriol., Abt. II
135. 5-11 (in German).
Muller, G. and Hickisch, B. (1979) Influence ofureolysis and nitrification inhibitors on soil microorganisms,
Arch. Acker- Pjlanzenbau Bodenk. 23, 539-546 (in German).
Mulvaney, R-L and Bremner, J.M. (1977) Evaluation of antimetabolites for retardation of urea hydrolysis in soils,
Soil Sci. Soc. Am. J. 41, 1024-1027.
Mulvaney, R.L and Bremner, J.M. (1978) Use of p-benzoquinone and hydroquinone for retardation of urea
hydrolysis in soils, Soil BioI. Biochem. 10, 297-302.
Munakata, K.-I., Kobashi, K., Takabe, S., and Hase, J.-1. (1980) Therapy for urolithiasis by hydroxamic acids. III.
Urease inhibitory potency and urinary excretion rate of N-acylglycinohydroxamic acids, J. Pharmacobiodyn.
3,451-456.
Muravin, E.A. (1989) Inhibitors ofNitrification, Agropromizdat, Moscow, pp. 77-79 (in Russian).
Muravin, E.A., Mochkova, T.V., Neigebaur. E.F., and Khrushkova, T.A. (1983) Comparative study of ATC and
nitrapyrin-based preparations as nitrification inhibitors, Izv. Timiryazev. Sel'skokhoz. Akad. (Moscow) (5). 172-
176 (in Russian).
Murphy, T.L. and Ferguson, R.B. (1992) The effect of a urease inhibitor on volatile ammonia loss and urea
hydrolysis on irrigated, ridge-till com, Agron. Abstr., p. 358.
Murphy, T.L. and Ferguson, R.B. (1997) Ridge-till com and urea hydrolysis response to NBPT, J. Production
Agric. 10,271-282.
Mutatkar, V.K. and Pritchett, W.L (1967) Effects of added aluminum on some soil microbial processes and on the
growth of oats (Avena sativa) in Arredondo fine sand, Soil Sci. 103,39-46.
Muthuswamy, P., Raju, G.S.N .. and Krishnamoorthy, K.K. (1975) Mineralization of urea coated with nitrification
inhibitors, J. Indian Soc. Soil Sci. 23, 332-335.
Nagata, K., Satoh, H., Iwahi. T., Shimoyarna, T.. and Tamura, T. (1993) Potent inhibition of the gastric pump
379
inhibitor lansoprazole against urease activity of Helicobacter pylori: Unique action selective for H pylori cells,
Antimicrob. Agents Chemother. 37, 769-774.
Nair, K.P.P. and Sharma, P.R (1979) Mineralization and field effectiveness of ordinat)' and coated urea, urea-
aldehyde condensation product and urea treated with nitrification inhibitor, J. Agric. Sci. 93,623-627.
Nelson, D.W., Beyrouty, C.A., and Schlegel, A.J. (1986) Effects of phosphoroamide urease inhibitors on nitrogen
transformations in soil and maize yields, Trans. 13th Congr. Int. Soc. Soil Sci. (Hamburg, 1986) 3, 880-881.
Neumann, R and Richter, H. (1976) Procedure for production of urea-containing compositions as fertilizers, (East)
Nornrnik, H. (1973) The effect of pellet size on the ammonia loss from urea applied to forest soil, Plant Soil 39,
309-318.
Nor, Y.M. (1982) Soil urease activity and kinetics, Soil Bioi. Biochem. 14,63-65.
Norden, J., Aigner, H.. Schindler. E, and Samblebe, R. (1985) Urea-containing fertilizers with urease inhibitors,
German Patent No. 33 22724 A 1.
Norden, J., Aigner, H., Schindler, E, and Samblebe, R. (1986) Fertilizers containing urea with urease inhibitors,
US. Patent No. 4,576,625.
Norman, RJ., Wolf, D.C., and Wells, RR (1991) Effect of application time, soil moisture, and NBPT on recovery
of 15N-labeled urea in rice, Agron. Abstr., p. 362.
Nosier, H.G., Bellinger, H., and Wessendorf, R (1971) Washing and cleaning products containing antimicrobial
agents, German Patent No.1 943 112.
O'Connor, M-J. and Hendrickson, L.L. (1987) Effect of phenylphosphorodiamidate on ammonia volatilization as
affected by soil temperature and rates and distribution of urea, Soil Sci. Soc. Am. J. 51, 1062-1066.
O'Connor, M-J., Falk, P.M., and Hendrickson, L.L. (1983) Effect of soil temperature and plant residue on ability of
phenylphosphorodiamidate (PPD) to reduce ammonia volatilization, Agron. Abstr., p. 159.
Odake, S., Morikawa, T., Tsuchiya, M., Imamura, L, and Kobashi, K. (1994) Inhibition of Helicobacter pylori
urease activity by hydroxamic acid derivatives, BioI. Pharm. Bull. 17, 1329-1332.
Oertel, M., Thieme, H., Jasche, K., and Rothe, G. (1978) Urease inhibitor usage in urea application, Chem.-Tech.
Umschau (Piesteritz) 10 (2), 47-55 (in German).
Ornholt, T.E. and Hendrickson, L.L (1983) Ability of phenylphosphorodiamidate (PPD) to reduce ammonia
volatilization and increase com yields in the field, Agron. Abstr., p. 215.
Omilinsky, B.A., Lindsay, A.D., Sutton, A.R, and Thornsberry, W.LJr. (1997) hnproved formulation for urease
inhibitor fertilizer additive concentrate, PCT Int. Appl. WO 97 22,568. Chern. Abstr., 1997, 127, 108531d.
Ondracek, L. Hampl, L, and Wanek, W. (1970a) Testing of the agrochemical effectiveness of trimeric phosphorus
nitride amide, Rostl. Vyroba 16,439-443 (in Czech).
Ondracek, L., Rezac, Z .. Mondry, E, Hampl, L, and Wanek, W. (1970b) The agrochernical effectiveness of
some phosphorus-nitrogen compounds with direct P-N bond, BioI. Plant. (Prague) 12, 159-166.
Osminkina, L.N. and Mirzaev, EM. (1979) Effect of copper on the hydrolysis of urea, Uzb. Khim. Zh. (2), 36-
38 (in Russian).
Ostromecka, M. (1968) The velocity of urea transformation under various soil conditions, Zesz. Probl.
Postepow Nauk Rain. (84), 391-398.
Otey, EH., Trimmell, D., Westhoff, RP., and Shasha, B.S. (1984) Starch matrix for controlled release of urea
fertilizer, J. Agric. Food Chem. 32, 1095-1098.
Ouyang, D.S., MacKenzie, A.F., and Fan, M.X. (1998) Nitrite accumulation and denitrification during urea
hydrolysis with added triple superphosphate, Commun. Soil Sci. Plant Anal. 29, 2333-2346.
Palazzo, D., Capotorti, G., Montemurro, E, Sunseri, E, and Vanadia, S. (1995) Urease inhibitor application:
Ammonia loss reduction and maize (Zea mays L.) yield enhancement, Riv. Agron. (Bologna) 29, 147-151.
Palazzo, D., Capotorti, G., Montemurro, E, and Sunseri, E (1996) Fertilization trials with urea amended with
urease and nitrification inhibitors, Inform. Agrar. (Verona) 52(30), 30-32 (in Italian).
Parashar, K.S., Prasad, R., Sharma, R.P., Sharma, S.N., and Singh, S. (1980) Efficiency of urea, nitrification
inhibitor-treated urea and slow release nitrogen fertilizers for sugarcane, Z. Pjlanzener. Bodenk. 143.262-
267.
Park, l-B.. Imamura, L., and Kobashi, K. (1996) Kinetic studies of Helicobacter pylori urease inhibition by a novel
proton pump inhibitor, rabeprazole.Biol. Pharm. Bull. 19, 182-187.
Patti, A.E, Verhey en, T.V., Douglas, L., and Wang, X. (1992) Nitrohurnic acids from Victorian brown coal,
Sci. Total Environ. 113,49-65.
Paulson, K.N. and Kurtz, L.T. (1969a) Locus of urease activity in soil, Soil Sci. Soc. Am. Proc. 33, 897-90l.
Paulson, K.N. and Kurtz, L.T. (1969b) Evaluation of urea-hydrocarbon complexes as "slow release" nitrogen
carriers, Soil Sci. Soc. Am. Proc. 33, 973.
Pechkovskii, V.V., Cherches, G.Kh., Plyshevskii, S.V., and Morozik, V.S. (1983) Procedure for obtaining
nitrogen fertilizer, USSR Patent No. 1,010,046.
380
Pedrazzini, F., Tarsitano, R., and Nannipieri, P. (1987) The effect of phenyl phosphorodiamidate on urease
activity and ammonia volatilization in flooded rice, Bioi. Fert. Soils 3, 183-188.
Pellegrini, G. (1965) New fertilizers containing phosphorus and nitrogen, French Patent No. 1,405,795.
Pellegrini, G. (1968) Fertilizers containing phosphorus and nitrogen, French Patent No. 1,551,475.
Pel'tser, A.S. (1972) Kinetics of 14C_urea decomposition in soil depending on soil moisture, addition of toxic
chemicals, liming and value of soil pH, Agrokhimiya (10),32-37 (in Russian).
Perez Mateos, M. and Gonzalez Carcedo, S. (1982) Compared action of phenylmercuric borate and acetate on
urease activity of soil, An. Edafol. Agrobiol. 41, 2245-2252 (in Spanish).
Perez Mateos M. and Gonzales Carcedo S. (1988) Assay of urease activity in soil columns, Soil Bioi.
Biochem. 20, 567-572.
Peterson, A.F. and Walter, C.R.jr. (1970) Regulation of urea hydrolysis in soil, U.S. Patent No. 3,547,614.
Phongpan, S. and Byrnes, B.H. (1990) The effect of the urease inhibitor N-(n-butyl) thiophosphoric triamide
on the efficiency ofurea application in a flooded rice field trial in Thailand, Fen. Res. 25, 145-151.
Phongpan, S. and Byrnes, B.H. (1993) Effect of methods of application on the efficiency of urea broadcast
onto lowland rice (Oryza sativa L.), Bioi. Fen. Soils 15,235-240.
Phongpan, S., Vacharotayan, S., and Kumazawa, K. (1988) Dynamics of ammonia volatilization losses in
wetland soil following the broadcasting application of urea amended with phenyl phosphorodiamidate,
Soil Sci. Plant Nutr. 34, 127-137.
Phongpan, S., Freney, lR., Keerthisinghe, D.G., and Chaiwanakupt, P. (1995) Use of
phenylphosphorodiamidate and N-(n-butyl)thiophosphoric triamide to reduce ammonia loss and increase
grain yield following application of urea to flooded rice, Fen. Res. 41, 59-66.
Phongpan, S., Freney, lR., Keerthisinghe, D.G., and Chaiwanakupt, P. (1997) Use of urease inhibitors to
reduce ammonia loss from broadcast urea and increase grain yield of flooded rice in Thailand, Soil Sci.
Plant NutI'. 43, 1057-1060.
Pietrowski, .T., Siomiany, P., and Siomiany, B.L. (1995) Inhibition of Helicobacter j7Ylori urease activity by
ebrotidine, Biochem. Malec. BioI. Int. 37, 247-253.
Pirogovskaya, G.V., Bogomaz, l.A., Shagieva, E.!., Kovalenok, M.F., Kuleshova, S.I., Naumova, G.v., and
Raitsina, G.I. (1993) Utilization of humic preparations in production of mineral fertilizers, in D.S. Orlov
(ed.), Humic Substances in the Biosphere, Izd. Nauka, Moscow, pp. 166-173 (in Russian).
Pisareva, E.S. (1989) Transformation and balance of urea nitrogen in soils following use of nitrification
inhibitor (MPC) and hydroquinone, in B.A. Yagodin (ed.), Ways for Increasing the Efficiency of
Fertilizers in the Nonchernozemic Zone, Timiryazev. Sel'skokhoz. Akad. (Moscow), pp. 91-101 (in
Russian).
Pisareva, E.S. and Muravin, E.A. (1988) Effect of the nitrification inhibitor MPC on transformation and
balance of urea nitrogen in soil, Abstr.. All-Unional Conj "Pedological. Agrochemical, and Ecological
Problems of the Formation of High(v Productive Agrocoenoses" (Pushchino, 1988), pp. 220-221 (in
Russian).
Pommer, E.-H. and Wessendorf. R. (1972) Fungicides for plant protection, German Patent No.2 054 887.
Popov, A.I., MergeI', A.A., Kurakov, L.v., Semenov, V.M., and Guzev, V.S. (1990) Microbiological state and
nitrogen regime ofa grey forest soil after application of nitrification inhibitors, Pochvovedenie (6),78-86
(in Rus sian).
Powlson, D.S. and Jenkinson, D.S. (1971) Inhibition of nitrification in soil by carbon disulphide from rubber
bungs, Soil Bioi. Biochem. 3, 267-269.
Prakasa Rao, E.V.S. and Puttanna, K. (1987) Nitrification and ammonia volatilization losses from urea and
dicyandiamide-treated urea in a sandy loam soil, Plant Soil 97, 201-206.
Prasad, R. and Power, J.F. (1995) Nitrification inhibitors for agriculture, health, and the environment, Adv.
Agron. 54, 233-281.
Praveen-Kumar and Aggarwal, R.K. (1988) Reduction of ammonia volatilization from urea by rapid
nitrification, Arid Soil Res. Rehabil. 2, 131-138.
Praveen-Kumar, Chhonkar, P,K., and Agnihotri, N.P. (1987) Mobility of urease inhibitors: Application of soil
thin-layer chromatography, Soil BioI. Biochem. 19, 687-688.
Praveen-Kumar, Chhonkar, P.K., and Burman,U. (1990) Effect of urease inhibitors on energy barriers of urease
activity in soils, J Indian Soc. Soil Sci. 38, 34-39.
Prusinkiewicz, Z. and Jozefkowicz- Kotlarz, J. (1982) Dynamics of ammonia volatilization from the urea applied in
fertilization of poor forest soils and the possibility of reducing the nitrogen losses by simultaneous application
of potassium chloride, Roczn. Glebozn, 33,19-35.
Pugh, K.B. and Waid, J.S. (1969a) The influence of hydroxarnates on ammonia loss from an acid loamy sand
treated with urea, Soil BioI. Biochem. 1, 195-206.
381
Pugh, K.B. and Waid, J.S. (1969b) The influence ofhydroxamates on ammonia loss from various soils treated with
urea, Soil Bioi. Biochem.I,207-217.
Puttanna, K., Nanje Gowda, N.M., and Prakasa Rao, E.y'S. (1999) Evaluation of nitrification inhibitors for use
under tropical conditions, Commun. Soil Sci. Plant Anal. 30,519-524.
Rachhpal-Singh and Nye P.H. (I 984a) The effect of soil pH and high urea concentrations on urease activity in soil,
J. Soil Sci. 35, 519-527.
Rachhpal-Singh and Nye P.H. (I 984b) Diffusion of urea, ammonium and soil alkalinity from surface applied urea,
J. Soil Sci. 35, 529-538.
Radel, RJ. (1990) Dual purpose urease and nitrification inhibitors, U.S. Patent No. 4,932,992.
Radel, RJ. and Crenshaw, M.D. (1990) Thiopyridine-N-oxides, thiopyridines, and thiopyrimidines as urease
inhibitors, U.s. Patent No. 4,932,991.
Radel, RJ., Gautney, 1., Randle, A.A., Cochran J.E., Miles, R.M., Williams, H.M., Bock, B.R., and Savant, N.K.
(1987) Evaluation ofthiophosphoryl triamide as a urease inhibitor, 194th Am. Chem. Soc. Natl. Meeting (New
Orleans, 1987), Paper No. 17; Abstr., p. 461.
Radel, RJ., Crenshaw, M.D., Bock, B.R., and Williams, H.M. (1989) Mercapto-pyridines and mercapto-
pyrimidines as urease inhibitors, Fert. Res. 20,123-127.
Radel, RJ., Randle, A.A., Gautney, J., Bock, B.R., and Williams, H.M. (1992) Thiophosphoryl triamide: A dual
purpose urease/nitrification inhibitor, Fert. Res. 31, 275-280.
Radel, RJ., Randle, A.A., and Kim, Y.K. (1994) Soil urease inhibition by thermal polymers of thiophosphoryl
triamide and phosphoryl triamide, Fert. Res. 39,153-160.
Rlidulescu, D., Pintea, H., Kolozsi, E., Cri~an R., Drilgan-Bularda, M., and Kiss, S. (1984) Effect of a urease
inhibitor on the enzymatic activities and respiration of soil, Fifth Sympo$jum on Soil Biology (I~i, 1981), Ed.
Ceres, Bucharest, pp. 69-73.
Raguotis, A.D. and Shleinys, R.1. (1986) Gaseous losses of nitrogen fertilizers in forest and possibilities for their
reduction, Pochvovedenie (9), 105-111 (in Russian).
Rajeswara Rao, B.R. and Shand, S. (1996) Relative efficiency ofprilled urea, neem cake-coated urea and neem oil-
coated urea as sources of nitrogen to lemongrass (Cymbopogon/lexuosus var./lexuosus), Indian J. Agric. Sci.
66,725-727.
Rajeswara Rao, B.R., Singh, K., Bhattacharya, A.K., and Naqvi, A.A. (1990) Effect ofprilled urea and modified
urea materials on yield and quality of geranium (Pelargonium graveolens L. Her.), Fert. Res. 23, 81-S5.
Raju, RA, Reddy, K.A., and Husain, M.M. (1989) Urease inhibitors and coated urea fertilizers for rice (Oryza
sativa) in deltaic alluvials, Indian J. Agric. Sci. 59,817-819.
Ram, M., Chatteljee, B.N., Yadav, R.L., and Singh, D.V. (1988) Minimizing volatilization and leaching losses of
nitrogen by different nitrogen carriers in Japanese mint (Mentha arvensis L.), J. Agric. Sci. 110,415-418.
Rao, D.L.N. and Ghai, S.K. (I 986a) Urease inhibitors: Effect on wheat growth in an alkali soil, Soil Bioi. Biochem.
18,255-258.
Rao, D.L.N. and Ghai, S.K. (I986b) Effect of phenylphosphorodiamidate on urea hydrolysis, ammonia
volatilization and rice growth in an alkali soil, Plant Soil 94, 313-320.
Rappaport, B.D. and Axley, J.H. (1984) Potassium chloride tor improVed urea fertilizer efficiency, Soil Sci. Soc.
Am. J. 48,399-401; Potash Rev. (Berne) (9), 1-5.
Reddy, M.S. and Chhonkar, P.K. (1990a) Effect of flooding, addition of organic matter and certain chemicals on
urea hydrolysis in soil and flood water, J. Res. APA U (Andhra Pradesh Agric. Univ.) 18, 81-83.
Reddy, M.S. and Chhonkar, P.K. (199Ob) Dissimilatory nitrate reductase in soil and flood water as influenced by
regulatory chemicals and oxygen stress, J. Indian Soc. Soil Sci. 38, 658-662.
Reddy, M.S. and Chhonkar, P.K. (l99Oc) Nature of inhibition of dissimilatory nitrate reductase activity in an
Inceptisol, J. Indian Soc. Soil Sci. 38, 742-744.
Reddy, M.S. and Chhonkar, P.K. (1991) Urease activity in .soil and flood water as influenced by regulatory
chemicals and oxygen stress, J. Indian Soc. Soil Sci. 39, 84-8S.
Reddy, Y.R.M. and Mishra, B. (1983) Effect of altered soil urease activity and temperature on ammonia
volatilization from surface applied urea,J.lndian Soc. Soil Sci. 31,143-145.
Reddy, R.N.S. and Prasad, R. (1975) Studies on the mineralization of urea, coated urea, and nitrification inhibitor
treated urea in soil, J. Soil Sci. 26, 304-312.
von Rheinbaben, W. (1987) Effect of magnesium sulphate addition to urea on nitrogen loss due to ammonia
volatilization, Fert. Res. II, 149-159.
Richards, I. (1995) Urea fertilizer containing urease-inhibiting metal nitrate, PCT Int. Appl. WO 95 IS,081. Chern.
Abstr., 1995, 123, 168759m.
Richter, H., Liedloff, B., Neumann, R., and Schiilke, U. (I97S) Procedure for improving the storage and/or
fertilizer properties of urea, (East) German Patent No. 129,896.
382
Rodgers, G.A. (1983) Effect ofdicyandiamide on ammonia volatilisation from urea in soil, Fer!. Res. 4, 361-
367.
Rodgers, G.A. (l984a) Action of nitrification inhibitors, in M. Woodbine (ed.), Antimicrobials and
Agriculture, Butterworths, London, pp. 33-43.
Rodgers, G.A. (l984b) Inhibition of soil urease activity by aminocresols, Plant Soil 79, 155-158.
Rodgers, G.A. and Pruden, G. (1984) Field estimation of ammonia volatilisation from lSN-labelled urea
fertiliser, J Sci. Food Agric. 35, 1290-1293.
Rodgers, G.A .. Widdowson, F.V., Penny, A., and Hewitt, M.Y. (1984) Comparison of the effects of aqueous
and of prilled urea, used alone or with urease or nitrification inhibitors, with those of 'Nitro-Chalk' on
ryegrass leys, J Agric. Sci. 103,671-685.
Rodgers, G.A., Wickramasinghe, K.N., and Jenkinson, O.S. (1985) Mineralization of dicyandiamide, labelled
with lSN, in acid soils, Soil Bioi. Biochem.17, 253-254.
Rodgers, G.A., Penny, A., and Hewitt, M.V. (1986) A comparison of the effects ofprilled urea, used alone or with a
nitrification or urease inhibitor, with those of 'Nitro-Chalk' on winter oil-seed rape, J Agric. Sci. 106, 515-526.
Rodgers, G.A., Penny, A., Widdowson, F.V., and Hewitt, M.V. (1987) Tests of nitrification and of urease
inhibitors, when applied with either solid or aqueous urea, on grass grown on a light sandy soil, J Agric.
Sci. 108, 109-117.
Rosenstein, I.J.M. and Hamilton-Miller, J.M.T. (1984) Inhibitors of urease as chemotherapeutic agents, Crit.
Rev. Microbiol. 11, 1-12.
Rotini, O.T. (1935) The enzymatic transformation of urea in soil, Ann. Lab. Ferment. "Lazzaro Spallanzani"
(Milano) 3,179-192 (in Italian).
Rozycki, M. and Bartha, R (1981) Problems associated with the use of azide as an inhibitor of microbial activity in
soil, Appl. Environ. Microbiol. 41, 833-836.
Runkov, S.V., Nikitina, G.O., and Kozlovskaya, N.A. (1974) Method for determination of urease activity of
peat soil, USSR Patent No. 445,749.
Sachdev, M.S., Oza, A. M., and Subbiah, B.Y. (1977) Possibilities of improving the efficiency of nitrogenous
fertilizers, Soil Organic Matter Studies, Int. Atomic Energy Agency, Vienna 1, 371-382.
Sahrawat, K.L. (1977) Effect of biuret content on transformation of urea nitrogen in soil, Soil Bioi. Biochem.
9,173-175.
Sahrawat, K.L. (1979) Evaluation of some chelating compounds for retardation of urea hydrolysis in soil,
Fert. Technol. 16, 244-245.
Sahmwat, K.L. (1981) Mineralization ofbiuret nitrogen in soil, Plant Soil 62, 469-471.
Sahrawat, K.L. (1989) Effects of nitrification inhibitors on nitrogen transfonnations, other than nitrification,
in soils, Adv. Agron. 42,279-309.
Sahmwat, K.L. and Mukerjee, S.K. (1977) Nitrification inhibitors. I. Studies with kamnjin, a furanolflavonoid
from karanja (Pongamia glabra) seeds, Plant Soil 47, 27-36.
Said. M.B. (1972) Adsorption of urea by some Sudan soils, Plant Soil 36, 239-242.
Sainz Rozas, H., Echeverria, H.E., Studdert, G.A., and Andmde, F.H. (1997) Ammonia volatilization from
urea in maize under direct drilling, Cie. Suelo 15, 12-16 (in Spanish).
Sallade, Y.E. and Sims, J.T. (1992) Evaluation of thiosulfate as a nitrification inhibitor for manures and
fertilizers, Plant Soil 147, 283-291.
Sallade, Y.E. and Sims, J.T. (1994) Nitrate leaching in an Atlantic coastal plain soil amended with poultry
manure or urea-ammonium nitrate: Influence of thiosulfate, Water. Air. Soil Pollut. 78, 307-316.
Sanikidze, G.S., Gamkrelidze, K.I., and Gogorikidze, N.I. (1987a) Effect of new forms of coated and inhibited
nitrogen fertilizers on count of microorganisms and activity of enzymes in red soils, Subtrop. Kul't. (2),
139-143 (in Russian),
Sanikidze, G.S., Gogorikidze, N.I., Gamkrelidze, K.I., and Zaalishvili, M,B. (1987b) Effect of micro elements
(B, Zn) on count of microorganisms and activity of enzymes in red soils, Subtrop. Kul't. (3),145-150.
Santra, G.H .. Oas, O.K., and MandaI, L.N. (1988) Loss of nitrogen through ammonia volatilization from
flooded rice fields, J Indian Soc. Soil Sci. 36, 652-659.
Sato, T. (2000) Urease inhibitors containing proanthocyanidins, their uses as drugs and fOod, and
determination of urease. Japanese Patent No. 159,669. Chern. Abstr., 2000, 133, P 42604v.
Satrusajang, A., Snitwongse, P., Buresh, RJ., and Friesen, O.K. (1991) Nitrogen-IS and sulfur-35 balances for
fertilizers applied to transplanted rainfed lowland rice, Fer!. Res. 28, 55-65.
Savant, N.K., James, A.F., and McClellan, G.H. (1987a) Effect of soil bulk density on hydrolysis of surface-
applied urea in unsaturated soils, Fer!. Res. 11, 221-229.
Savant, N.K., James, A.F., and McClellan, G.H. (1987b) Effect of amounts and sequence of additions of urea
and water on hydrolysis of surface-applied granular urea in unsaturated soils, Fer!. Res. 11, 231-243.
383
Savant, N.K., James, A.F., and McClellan, G.H. (1988a) Effect of soil bulk density on inhibition of hydrolysis
of surface-applied granular urea containing phenyl phosphorodiamidate in unsaturated soils, Commun.
Soil Sci. Plant Nutr. 19, 193-204.
Savant, N.K., James, A.F., Peters, G.E., and Medina, R. (l988b) Evaluation of
phenoxyaminocyclotriphosphazatrienes as sustained-action soil urease inhibitors, J. Agric. Food Chem.
36, 390-392.
Scharf. P.e. and Alley, M.M. (1987) Nitrogen loss inhibitors in intensively managed winter wheat, Agron.
Abstr., p. 270.
Schinner, F.A. and Pfitscher, A. (1978) Urease and catalase activities and CO2 evolution in different soils of
the upper subalpine zone, in A. Cernusca (ed.), Ecological Analyses of Alpine Meadow Areas in the
Gastein Valley, Univ.-Verlag Wagner, Innsbruck, pp. 259-273 (in German).
Schinner, F.A., Niederbacher, R., and Neuwinger, I. (1980) Influence of compound fertilizer and cupric sulfate
on soil enzymes and CO2-evolution, Plant Soil 57, 85-93.
Schlegel, AJ. (l991a) Reduction of phytotoxicity from urea fertilizer by the urease inhibitor NBPT, Agron.
Abstr., p. 362.
Schlegel, AJ. (1991b) Radical ammonia phytotoxicity from UAN solution by the urease inhibitor N-(n-butyl)
thiophosphoric triamide, J. Fert. Issues 8 (2), 40-44.
Schlegel, AJ., Nelson, D.W., and Sommers, L.E. (1986) Field evaluation of urease inhibitors for corn
production, Agron. J. 78, 1007-1012.
Schlegel, AJ., Nelson, D.W., and Sommers, L.E. (1987) Use of urease inhibitors and urea fertilizers on winter
wheat, Fert. Res.H, 97-111.
Schroth, w., Schmiedl, D., and Hildebrandt, A. (1974) Fertilizer compositions, (East) German Patent No.
110,031.
Selivanov, A.M. (1986) Effect of cyanuric acid on urease activity, Vestn. Sel'skokhoz. Nauki Kazakhstana (4),
62-64 (in Russian).
Sen, H.S. and Bandyopadhyay, B.K. (1986) Effect of placement, urease inhibitors and acidic amendments on
anunonia volatilization from wetland rice field fertilized with urea, J. Indian Soc. Soil Sci. 34, 514-519.
Sengik, E. and Kiehl, J.C. (l995a) Effect of organic residues and monocaIcium phosphate on ammonia
volatilization from urea-treated soil, Rev. Bras. Cie. Solo 19, 321-326 (in Portuguese).
Sengik, E. and Kiehl, J.e. (1995b) Controlling ammonia volatilization from soil treated with urea and peat by
application of inorganic salts, Rev. Bras. Cie. Solo 19, 455-461 (in Portuguese).
Sharma, B.D. and Gupta, I.e. (1989) Effect of rate and source of nitrogen and moisture content of soil on
anunonia volatilization from sandy soils, J. Indian Soc. Soil Sci. 37, 665-669.
Sharma, S.N. and Prasad, R. (1996) Use of nitrification inhibitors (neem and DCD) to increase N efficiency in
maize-wheat cropping system, Fert. Res. 44, 169-175.
Shcherbakov, A.P. and Stakhurlova, L.D. (1990) Nitrogen metabolism in soil following band application of
urea and MPC nitrification inhibitor, Dokl. VASKhNIL (Moscow) (12),25-28 (in Russian).
Shehata, S.M., EI-Shinnawi. M.M., and EI-Shimi, S.A. (1982) Activity of enzymes in organic carbon-
amended soils treated with varying levels of salts, Zbl. Mikrobiol. 137,76-85.
Shen, B., Li, S.P., Zhao, S.W., and Liu, JJ. (1997) Effect of chlorobenzene and nitrophenol on soil biological
activity, Acta Pedol. Sinica 34 (3), 309-314 (in Chinese).
Sheridan, R.e. (1970) Chemistry ofthiophosphoryl triamide, Sulphur Inst. J. 6, 11-13.
Sheudzhen, A.Kh., Aleshin, E.P., and Doseeva, O.A. (1991) Enzymatic activity in rice field soil as affected by
fertilization with rnicroelements, Dokl. VASKhNIL (Moscow) (8), 16-18 (in Russian).
Shih, K.L. and Souza, K.A. (1978) Degradation of biochemical activity in soil sterilized by dry heat and
gamma radiation, Origins Life 9,51-63.
Simihaian, M. (1998) OrganiC Inhibitors of Soil Urease Activity, Babe~-Bo1yai Univ., Cluj (in
Romanian).
Simihliian, M. and Silberg, LA. (1996) Effect of crysean and benzoyl crysean on soil urease activity, An. Univ.
Oradea, Bioi. 3,149-156 (in Romanian).
Simibaian, M. and Silberg, I.A. (1999) Determination of the inhibition constants of some soil urease
inhibitors, in R.P. Dick (ed.), Enzymes in the Environment: Activity. Ecology and Applications. Abstr., Int.
Con! (Granada, Spain, 1999), p. 24.
Simihiiian, M., Kiss, S., Pa§ca, D., and Suciu, C. (1992) Testing of some synthetic derivatives of L-glutamic
acid as inhibitors of soil urease activity, Stud. Univ. Babe$-Bolyai, BioI. 37 (2), 99-10 1.
Simihliian, M., Silberg, LA., and Kiss, S. (1999) Testing of some classes of compounds for inhibition of soil
urease activity, in R.P. Dick (ed.), Enzymes in the Environment: Activity, Ecology and Applications.
Abstr., Int. Can! (Granada, Spain, 1999), p. 25.
384
Simon, L. and Bergerova, E. (1986) Effect of N-Serve [2-chloro-6-(trichloromethyl) pyridine] on nitrogen

transformations and activity of some enzymes in the soil, Acta Fac. Rerum Nat. Univ. Comen., Microbiol.
(Bratislava) 14, 43-56.
Si~son, J.R., Freney, J.R., Muirhead, W.A., and Leuning, R. (1985) Effects of phenylphosphorodiamidate
and dicyandiamide on nitrogen loss from flooded rice, Soil Sci. Soc. Am. J. 49, 1426-1431.
Singh, B. and Bajwa, M.S. (1986) Studies on urea hydrolysis in salt-affected soils, Fen. Res. 8, 231-240.
Singh, B. and Bajwa, M.S. (1987) Ammonia volatilization from salt-affected soils fertilized with urea under
submerged conditions, J. Indian Soc. Soil Sci. 35, 358-364.
Singh, M. and Singh, T.A. (1986) Leaching losses of nitrogen from urea as affected by awlication of neem
cake, J. Indian Soc. Soil Sci. 34, 763-773.
Singh, M., Yadav, D.S., and Kumar, V. (1984) Leaching and transfurmation of urea in dry and wet soils as
affected by irrigation water, Plant Soil 81, 411-420.
Singh, M., Kundu, S., Tripathi, A.K., and Takkar, P.N. (1996) Influence of modified urea materials and
methods ofawlication on ammonia volatilization losses, J. Indian Soc. Soil Sci. 44, 512-514.
Siva, K.B., Aminuddin, H., Husni, M.H.A., and Manas, A.R. (1999) Ammonia volatilization from urea as
affected by tropical-based palm oil mill eflluent (POME) and peat. Commun. Soil Sci. Plant Anal. 30,
785-804.
Sivapalan, K., Fernando, v., and Thenabadu, M.W. (1983) Humified phenol-rich plant residues and soil urease
activity, Plant Soil 70, 143-146.
Sivapalan, K., Fernando, V., and Thenabadu, M.W. (1985) N-Mineralization in polyphenol-rich plant residues
and their effect on nitrification of applied ammonium sulfate, Soil Bioi. Biochem. 17, 547-551.
Skujins, J., Nohrstedt, H.-b., and Oden, S. (1986) Development of a sensitive biological method for the
determination of a low-level toxic contamination in soils. 1. Selection of nitrogenase activity, Swedish J.
Agric. Res. 16, 113-118.
Skvortsova, LN., Obukhovskaya, T.D., and Zaslavskaya, N.V. (1984) Microbiological testing of soil pollution
with mercury, Vestn. Mosk. Gos. Un;v, , Ser. 17, Pochvovedenie (2),32-35 (in Russian).
Sloan, J.J. and Anderson, W.B. (1987) Bermudagrass yield as affected by urea fertilizers amended with
ammonium thiosulfate and calcium chloride, Agron. Abstr., p. 270.
Sloan, JJ. and Anderson, W.B. (1995) Calcium chloride and ammonium thiosulfate as ammonia volatilization
inhibitors for urea fertilizers, Commun. Soil Sci. Plant Anal. 26, 2425-2447.
Sloan, J.1. and Anderson, W.B. (1998) Influence of calcium chloride and ammonium thiosulfate on
bermudagrass uptake ofurea nitrogen, Commun, Soil Sci. Plant Anal. 29, 435-446.
Snitwongse, P., Satrusajang, A., and Buresh, RJ. (1988) Fate of nitrogen fertilizer applied to lowland rice on a
Sulfic Tropaquept, Fen. Res. 16, 227-240.
Sommer. K. and Rossig, K. (1978) Effect of the kind of nitrification inhibition on the yield in different N
fertilizations and proposal for a classification, Landw, Forsch. 31, 291-299 (in Gel1Ilan).
Sor, K.M. (1968) Fertilizer composition consisting of urea, a urease inhibitor, and a hydrocatbon binder, U.S.
Patent No.3 ,888,989.
Sor, K.M. (1969) Urea-containing fertilisers, British Patent No.1, 157,400.
Sor, K., Stansbury, R.L., and De Ment, J.D. (1966) Agricultural nutrient containing urea, U,S. Patent No.
3,232,740.
Sor, K.M., Pelissier, J.A., and Latham, R.jr. (1968) Nitrogen fertilizer, French Patent No. 1,519,208.
Sor, K.M., Pelissier, J.A., and Latham, R.jr. (1971) Urease inhibited urea-containing co~ositions, U.S,
Patent No. 3,565,599.
Sotomura, M., Takahashi, Y., and Kotani, T. (2000) Deodorant composition for treating odorous gases,
Japanese Patent No. 3,076,840. Chern, Abstr., 2000, 133, P 167775h.
Speir, T.W.. Kettles, H.A., Parshotam, A., Searle, P.L., and Vlaar, L.N.C. (1995) A simple kinetic aWroach to
derive the ecological dose value, EDso, for the assessment of Cr(VI) toxicity to soil biological processes,
Soil Bioi, Biochem. 27, 801-8lO.
Stark, J.D. and Walter, J.E (1995) Persistence ofazadirachtin A and B in soil: Effects of temperature and
microbial activity, J. Environ, Sci. Health, Pan B 30,685-698.
Stichbury, M., Bechard. G., Lortie. L., and Gould, W.D. (1995) Use of inhibitors to prevent acid mine
drainage, in T.P. Hynes and M.C. Blanchette (eds.), Proc. Con! Mining and the Environment (Sudbury,
Canada, 1995) 2, 613-622.
Strumpf, T., Zanke, D., and Lyr, H. (1981a) Urease-inhibiting agent, (East) German Patent No. 149,504.
Strurnpf, T., Zanke, D., and Lyr, H. (1981b) Urease-inhibiting agent, (East) German Patent No. 149,505.
Subramoney, N. and Ramanathan, K.L. (1979) Slow-acting urea fertilizer, Indian Patent No. 146,213. Chern,
Abstr., 1980,92, 5562u.
385
Sullivan D.M. and Havlin, J.L. (l992a) Soil and environmental effects on urease inhibition by ammonium
thiosulfate, Soil Sci. Soc. Am. J. 56, 950-956.
Sullivan D.M. and Havlin, J.L. (1992b) Thiosulfate inhibition of urea hydrolysis in soils: Tetrathionate as a
urease inhibitor. Soil Sci. Soc. Am. J. 56, 957-960.
Sullivan D.M. and Havlin, J.L. (1992c) Field microplot and laboratory evaluation of urea hydrolysis inhibition
by ammonium thiosulfute, Soil Sci. 153,373-381.
Sulzer, G.M., Cheng. C.H., Klobucar, W.O., and Kolich, C.H. (1998) Preparation of N-
hydrocarbylthiophosphoric triamides, u.s. Patent No. 5,770,721.
Summerskill, W.H.J., Thorsell, F., Feinherg, J.H., and Adrete, J. (1967) Effect of urease inhibition in
hyperammonemia: Clinical and experimental studies with acetohydroxamic acid, Gastroenterology 54,
20-26.
Sundaram, KM.S. (1996) Azadirachtin biopesticide: A review of studies conducted on its analytical
chemistry, environmental behaviour and biological effects, J. Environ. Sci. Health. Part B 31,913-948.
Sutton, A.R., Weston, C.W., and Balser, R.L. (1991) Homogeneous granular nitrogen fertilizer, U.S. Patent
No. 4,994,100.
Swan, M., Soper, R.J., and Morden, O. (1986) The effect of elemental sulfur, gypsum and ammonium
thiosulfate as sulfur sources on yield of rapeseed, Commun. Soil Sci. Plant Anal. 17, 1383-1390.
Swerdloff, M.D., Van der Puy, M., Kolc, J.F., Rogic, M.M., Anello, L.G., and Hendrickson, L.L. (1984)
Urease inhibited urea based fertilizer compositions containing phosphorus compounds, Eur. Patent Appl.•
Publ. No. 0 119 494 A I.
Swerdloff, M.D., Kolc, J.F., Rogic, M.M., and Hendrickson, L.L. (1985a) Aryl phosphoric triamide and aryl
phosphorodiamidate urease and nitrification inhibitors and urease and nitrification inhibited urea and
reduced nitrogen based fertilizer compositions, U.S. Patent No. 4,517,004.
Swerdloff, M.D., Kolc, J.F., Rogic, M.M., and Hendrickson, L.L. (1985b) Phosphoroamide urease inhibitors
and urease inhibited urea based fertilizer compositions, u.s. Patent No. 4,517,007.
Swerdloff, M.D., Kole, J.F., Rogic, M.M., and Hendrickson, L.L. (1985c) Poly-phosphorodiamide urease
inhibitors and urease inhibited urea based fertilizer compositions, u.s. Patent No. 4,518,413.
Swerdloff, M.D., Van der Puy, M., Kole. J.F., Rogic. M.M.• and Hendrickson. L.L. (1985d) Urease inhibited
urea based fertilizer compositions containing N-aryl-N-aliphatic phosphorotriamide compounds, u.s.
Patent No. 4,539,037.
Swerdloff, M.D., Rogic, M.M., and Hendrickson, L.L. (1986a) O-Diaminophosphinyl derivatives of oximes
as urease inhibitors, U.S. Patent No. 4,624,695.
Swerdloff, M.D., Rogic, M.M., and Hendrickson, L.L. (1986b) Oxidized sulfur derivatives of
diaminophosphinyl compounds as urease inhibitors and urease inhibited urea based fertilizer
compositions, U.S. Patent No. 4,629,491.
Swerdloff, M.D.. Rogic, M.M., and Hendrickson, L.L. (1987) Oxidized sulfur derivatives of
diaminophosphinyl compounds as urease inhibitors and urease inhibited urea based fertilizer
compositions, U.s. Patent No. 4,696,693.
Tabatabai, M.A. (1977) Effects of trace elements on urease activity in soils, Soil Bioi. Biochem. 9. 9-13.
Tabatabai. M.A. (1982) Soil enzymes, in A.L. Page, R.H. Miller, and D.R. Keeney (eds.), Methods of Soil
Analysis. Part 2. Chemical and Microbiological Properties, (2nd Edition), Am. Soc. Agron.-Soil Sci. Soc.
Am., Madison, pp. 903-947.
Tabatabai, M.A. (1994) Soil enzymes, in R.W. Weaver, J.S. Angle, and P.S. Bottomley (eds.), Methods of Soil
Analysis. Part 2. Microbiological and Biochemical Properties, Soil Sci. Soc. Am., Madison, pp. 775-833.
Tabatabai, M.A. and Bremner, J.M. (1972) Assay of urease activity in soils, Soil Bioi. Biochem. 4, 479-487.
Takabe, S., Numata, A., and Kobashi, K. (1984) Stone formation by Ureaplasma urealyticum in human urine
and its prevention by urease inhibitors, J. CZin. Microbiol. 20, 869-873.
Tan, K.H. (1982) Studies on nitrogen in Malaysian soils. II. Urea hydrolysis and transformations, J. Rubber
Res. Inst. Malaysia 30,19-30.
Tanabe, l. and Ishizawa, S. (1969) Microbial activity of soil, Bull. Natl. Inst. Agric. Sci., Ser. B (21),115-253
(in Japanese).
Tangl, H. and Baintner, K. (1969) increasing the assimilation of urea by ruminant animals through application
of urease inhibitors. Chemical Substances in Protein Feeding ofRuminant Animals, Borovsk, pp. 184-187
(in Russian).
Taslim, H. and Verstraeten, L.MJ. (1977) Mineralization of slow-release N-compounds in water-logged
conditions, Z. Pjlanzenem. Bodenk.140, 183-191.
Thieme, H., Hackert, H., Tradler, E., Hartbrich, H.-J., Rothe,G., Klepel, M., and Held, P. (1976) Fertilizer
composition, (East) German Patent No. 122,178.
386
Thomas, J. and Prasad, R. (l983) Mineralization of urea, coated urea and nitrification inhibitor treated urea in
different rice growing soils, Z. Pjlanzenern. Bodenk. 146, 341-347.
Thormiihlen. EP.J. and du Preez, C.C. (1991) Inhibition of urease activity in certain soils from the central
irrigation areas in South Africa, S.-Afr. Tydskr. Plant Grond 8,212-214 (in Mrikaans).
Thormihlen, EP.J. and du Preez, c.c. (1992) Effect of urea concentmtions on the urease activity in soils of
the central irrigation areas in South Africa, S.-Aft: '(ydskr. Plant Grond 9,58-63 (in Afrikaans).
Todorov, Ts., Dimkov, R., Koteva, Zh., and Moncheva, P. (1987) Effect of lead contamination on the
biological properties of alluvial meadow soil, Pochvozn. Agrokhim. Rast. Zashchita (5), 33-40 (in
Bulgarian ).
Tomar, I.S. and MacKenzie, A.E (1984) Effect of catechol and p-benzoquinone on the hydrolysis of urea and
enelgy barriers of urease activity in soils, Can. J. Soil Sci. 64,51-60.
Tomar, I.S., Kirby, P.C., and MacKenzie, A.E (1985) Field evaluation of the effects of a urease inhibitor and
crop residues on urea hydrolysis, ammonia volatilization and yield of com, Can. J. Soil Sci. 65, 777-787.
Tomlinson, T.E. (1964) Some mctors which influence the formation of free ammonia from urea in soils,
Trans. 8th Int. Congr. Soil Sci. (Bucharest, 1964) 4,791-799.
Tomlinson, T.E. (1967) Method of controlling the hydrolysis of urea in soils and compositions therefor,
British Patent No. 1,094,802.
Tomlinson, T.E. (1970) Urea-agronomic implications, Proc. Fert. Soc. (113),1-76.
Tsuchiya, M., Imamum, L., and Kobashi, K. (1995a) Urease inhibitors, hyperammonemia inhibitors, and
hepatic encephalopathy inhibitors contalmng 2-[4-(3-methoxypropoxy)-3-methylpyridin-2-yl)
methylsulfinyl-lH-benzimidazole,Japanese Patent No. 07,118,153. Chern. Abstr., 1995,123, 74916h.
Tsuchiya, M., Imamura, L., Park, J.-B., and Kobashi, K. (1995b) Helicobacter pylori urease inhibition by
mbepmzole, a proton pul1tl inhibitor, Bioi. Pharm. Bull. 18, 1053-1056.
Tu, C.M. (1980) Influence of five pyrethroid insecticides on microbial populations and activities in soil,
Microb. Eco/. 5, 321-327.
Tu. C.M. (198Ia) Effects of pesticides on activities of enzymes and microolg3nisms in a clay soil, J. Environ.
Sci. Health. Part B 16, 179-191.
Tu, C.M. ( 1981 b) Effects of some pesticides on enzyme activities in an organic soil, Bull. Environ. Contam.
Toxicol. 27,109-114.
Tu. C.M. (1982) Influence of pesticides on activities ofinvertase, amylase and level of adenosine triphosphate
in organic soil, Chemosphere 11, 909-914.
Tu, C.M. (1988) Effect of selected pesticides on activities of invertase, amylase and microbial respimtion in
sandy soil, Chemosphere 17, 159-163.
Tu, C.M. (l990) Effect of four experimental insecticides on enzyme activities and levels of adenosine
triphosphate in mineral and organic soils, J. Environ. Sci. Health. Part B 25, 787-800.
Tu, C.M. (l992a) Effect of technical and formulated insecticides on activities of enzymes and level of
adenosine triphosphate in soil, Int. J. Environ. Health Res. 2, 76-83.
Tu, C.M. (l992b) Effect of three newer pesticides on microbial and enzymatic activities in soil, Bull. Environ.
Contam. Toxico/. 49, 120-128.
Tu, C.M. (1993a) Effect of fungicides, captafol and chlorothalonil, on microbial and enzymatic activities in
mineral soil, J. Environ. Sci. Health. Part B 28, 67-80.
Tu, C.M. (l993b) Influence of ten herbicides on activities of microorganisms and enzymes in soil, Bull.
Environ. Contam. Toxicol. 51, 30-39.
Tu, C.M., Marks, C.E, Johnson, P.W., and Elliot, J.M. (1995) Effects of nematicides on populations of soil
microflom and Pratylenchu.< penetrans, soil enzymes and flue-cured tobacco, J. Environ. Sci. Health.
Part B 30,95-111.
Tyalli, P.G. (1982) Ammonia volatilization following surface-application of urea under forest conditions,
Agrokhimiya (6), 23-27 (in Russian).
Ushikubo, A., Oyama, G., Ishimaru, K., and D'ltri, F.M. (1985) The effect of heavy metal concentration on
microbial activity in paddy soil, J. Agl'ic. Sci. (Tokyo) 30, 44-59.
Van Cleemput, O. and Wang, Z.P. (1991) Urea transformations and urease inhibitors, Trends Soil Sci. 1, 45-52.
Van Cleemput, 0., Wang, Z.P., Hassan Elnasikh, M., Bholah, M.A., and Takoudjou, C. (1991)
Transfurmations of 15N labelled urea in different types of soil, Stable Isotopes in Plant Nutrition. Soil
Fertility and Environmental Studies, Int. Atomic Enelgy Agency, Vienna, pp. 307-316.
Van der Puy, M., Kole, I.F., Anello, L.G., Rogic, M.M., and Swerdlolf, M.D. (1984a) S-Aryl and S-aliphatic
diamidophosphorothiolates as urease inhibitors and urease inhibited urea based fertilizer compositions,
Eur. Patent Appl.. Publ. No. 0 119495 A I.
Van der Puy, M., Gatrone, R.C., Robinson, M.A., and Hendrickson, L.L. (l994b) Urease inhibited urea based
fertilizer compositions containing 01gaR0 boron acid compounds, U.S. Patent No. 4,462,819.
387
Van der Puy, M., Kole, J.F., Anello, L.G., Hendrickson, L.L., Rogic, M.M., and Swerdloff, M.D. (1985a) S-
Aryl and S-aliphatic diamidophosphorothiolates as urease inhibitors and urease inhibited urea based
fertilizer compositions, U.S. Patent No. 4,537,614.
Van der Puy. M., Hendrickson, L.L., Kole, J.F., Rogic, M.M., and Swerdloff, M.D. (1985b)
Phosphorotriamidate urease inhibitors and urease inhibited urea based fertilizer compositions, US Patent
No. 4,540,428.
Varel, V,H. (1996) Effect of urease inhibitors on reducing ammonia emissions in cattle waste, Int. Con! on Air
Pollution from Agricultural Operations, Iowa State Univ., Ames, pp. 459-464.
Varel, Y.H (1997) Use of urease inhibitors to control nitrogen loss from livestock waste, Bioresource Technol.
62,11-17,
Varsa, E.C. (1991) The effect ofN fertilizers and N-(n-butyl) thiophosphoric triamide-amended N sources on
no-till corn ~ 1990 results, Annual Report. Plant and Soil Sci. Dept., Southern Dlinois Univ., Carbondale,
6pp,
Varsa, E.C. and Ebelhar, S.A. (1992) A comparison of Super Urea, Super N, and NBPT-treated N sources
with non-amended N fertilizers on no-till corn as influenced by placement and rates ~ 1991 results at
Belleville and Dixon Springs, Annual Report, Plant and Soil Sci. Dept.. Southern Illinois Univ. and Dixon
Springs Agric. Center, 11 pp.
Varsa, E.C. and Jan, N. (1997) Effect ofNBPT- and DCD-amended urea, alone and in combination on no-till
corn, Agron. Abstr., p. 230.
Varsa, E.C., Leis, A.K., and Jemison, J.M.jr. (1989) Response of no-till corn to urease inhibitors and
placement of N sources, Proc. 19th North Central Extension-Industry Soil Fertility Can! (St. Louis,
Missouri, 1989), pp. 131-138,
Varsa, E.C., Jemison, J.M., Osborn, M.W., Leis, A.K., Hnetkovsky S.W., and Jan, N. (1993) Effect ofNBPT-
amended urea and UAN on no-till corn in southern Illinois, Proc. 23rd North Central Extension-Industry
Soil Fertility Corif. (St. Louis, Missouri, 1993) 9, 72-80.
Verstraeten, L.M.J. (1977) Mineralization of hexamethylenetetramine in soils, Z. Pjlanzenem. Bodenk. 140,
175-181.
Verstraeten, L.MJ. and Livens. J. (1975) Effect of slow-release nitrogenous fertilizers on growth, yield and
quality of winter wheat, Z. Pjlanzenern. Bodenk. (4/5),435-446.
Verstraeten, L.M.J. and Livens, J. (1977) Effect ofa combined formula of slow-release N-fertilizers on winter
wheat, Proc. Int. Seminar on Soil Environment and Fertility Management in Intensive Agriculture (Tokyo,
1977), pp. 433-440.
Verstraeten, L.M.J., Seghers. I.. and Taslim, H. (1976) Effect of formaldehyde on N-mineralization in soils,
Agricultura (Leuven) 24,337-346.
Vilsmeier, K., Bomemisza, E., and Amberger, A. (1987) Urea, ammonium sulfate and dicyandiamide
transformations in Costa Rican soils, Fert. Res. 12,255-261.
Vittori Anti sari, L., Marzadori, C., Gioacchini, P., Ricci, S., and Gessa, C. (1996) Effects of the urease
inhibitor N -(n-butyl)thiophosphoric triamide in low concentrations on anunonia volatilization and
evolution of mineral nitrogen, Bioi. Fert. Soils 22, 196-201.
Vlek, P.L.G. and Craswell, E. T. (1981) Ammonia volatilization from flooded soils, Fert. Res. 2, 227-245.
Vlek, P.L.G., Stumpe, J.M., and Byrnes. B.H. (1980) Urease activity and inhibition in flooded soil systems,
Fert .Res. 1, 191-202.
Voigt, J., Piatkowski, B., and Bock, J. (1980a) Investigations on the effect of phosphoric acid phenyl ester
diamide as inhibitor of rumen urease of dairy cows. 1. Influence on urea hydrolysis, ammonia release and
fermentation in rumen, Arch. Tierem. 30,811-824 (in German).
Voigt, J., Krawielitzki, K., and Piatkowski, B. (1980b) Investigations on the effect of phosphoric acid phenyl
ester diamide as inhibitor of rumen urease of dairy cows. 2. Transformation of 15N_urea, Arch. Tierem. 30,
825-834 (in German).
Volk. G.M. (1961) Gaseous loss of ammonia from surface-applied nitrogenous fertilizers, J.Agric. Food
Chern. 9, 280-283.
Volk, G.M. (1966) Efficiency of fertilizer urea as affected by method of application, soil moisture, and lime,
Agron. J. 58, 249-252.
Volk, G.M. (1970) Gaseous loss of ammonia from prilled urea applied to slash pine, Soil Sci. Soc. Am. Proc.
34,513-516.
Volk, G.M. and Sweat, A.W. (1955) Mobility of urea nitrogen applied to Florida soils, Soil Sci. Soc. Florida
Proc. 15, 117-123.
Vostal. J., Knop, K.• and Vanek, V. (1975) Effect of salts on the transformation of urea nitrogen in soil. Sb.
JYs. Sk. Zemed. Praze, Fak. Agron., Rada A, pp. 87-102 (in Czech).
388
Vostal, J., Knop, K, and Vanek, V (1976) Effect of fertilizers on the transformation of urea nitrogen in soiL
Sb. Tj>s. Sk Zemed. Praze, Fak. Agron .. RadaA (1),121-137 (in Czech).
Vyas, B.N., Godrej, J.N.B., and Mistry, KB. (1991) Development and evaluation ofneem extract as a coating
for urea fertilizer, Fen. News 36 (2), 19-25.
Waid, J.S. (1975) Hydroxamic acids in soil systems, in E.A. Paul and A.D. McLaren (eds.), Soil Biochemistry,
Vol. 4, M. Dekker, New York, pp. 65-101.
Waid, J.S. and Pugh, KB. (1967) Acetohydroxamate inhibition of the urease activity of an acid soil, Chem.
Ind. (London) (2), 71-73.
Wakefield, Z.T., Allen, S.E., McCullough, J.F.. Sheridan, R.c., and Kohler, J.J. (1971) Evaluation of
phosphorus-nitrogen compounds as fertilizers, J. Agric. Food Chem. 19, 93-103.
Wanek, W, Ondracek, L., Jager, L., Jara, V, Rezac, Z., and Skiivanek, 1. (1966) New highly concentrated
fertilizers, Czechoslovak Patent No. 119,228.
Wang, X.J. and Douglas, L.A. (1996) Effect ofphosphoroamides on soil urease activity, and plant dry matter
production and urea-N uptake by wheat plants, Agrochimica 40, 209-215.
Wang, Z.P. and Van Cleemput, O. (1992) Effects of urease inhibitors on nitrogen transformations, Agron.
Abstr., p. 360.
Wang, Z.P., Van Cleemput, 0., and Baert, L. (l990) Effect of urease inhibitors on nitrification in soil, Soil Use
Manage. 6 (1), 41-43.
Wang, Z.P., Van Cleemput, 0., Demeyer, P., and Baert, L. (199Ia) Effect of urease inhibitors on urea
hydrolysis and ammonia volatilization, Bioi. Fen. Soils 11, 43-47.
Wang, Z.P., Van Cleemput, 0., Li. L.T., and Baert, L. (l991b) Effect of organic matter and urease inhibitors
on urea hydrolysis and immobilization of urea nitrogen in an alkaline soil, Bioi. Fen. Soils 11. 101-104.
Wang, Z.P., Li, L.T., Van Cleemput, 0., and Baert, L. (199Ic) Effect of urease inhibitors on denitrification in
soil, Soil Use Manage. 7 (4),230-233.
Wang, X.B., Xin, J.E, Grant, C.A., and Bailey, L.D. (1995) Effects of placement of urea with a urease
inhibitor on seedling emergence, N uptake and dry matter yield of wheat, Can. J. Plant Sci. 75, 449-452.
Wang, Z.P., Van Cleemput, 0., and Baert, L. (1996) Movement of urea and its hydrolysis products as
influenced by moisture content and urease inhibitors, Bioi. Fert. Soils 22, 10 1-108.
Watkins. S.H .. Strand, R.E, DeBell, D.S., and Esch, J.jr. (l972) Factors influencing ammonia losses from urea
applied to northwestern forest soils, Soil Sci. Soc. Am. Proc. 36, 354-357.
Watson C.P. (1990) The influence of soil properties on the effectiveness of phenylphosphorodiamidate (PPD)
in reducing ammonia volatilization from surface applied urea, Fert. Res. 24, 1-10.
Watson, CJ. and Miller, H. (1996) Short-term effects of urea amended with the urease inhibitor N-(n-butyl)
thiophosphoric triamide on perennial ryegrass. Plant Soil 184, 33-45.
Watson, c.J., Stevens, R.J., and Laughlin, R.J. (1990a) Use of the urease inhibitor NBPT [N-(n-butyl)
thiophosphoric triamide] to improve the efficiency of urea for grassland, in A. Scaife (ed.), Pro£'. First
Congr. Eur. Soc. Agron. (Paris, 1990), Session 3, Poster 55.
Watson, c.J., Stevens, RJ., and Laughlin, RJ. (l990b) Effect of the urease inhibitor NBPT [N-(n-butyl)
thiophosphoric triamide] for improving the efficiency of urea on ryegrass production, Fen. Res. 24, 11-
15.
Watson, CJ., Poland, P., Miller, H., Allen, M.B.D., Garrett, M.K, and Christianson, C.B. (1994a) Agronomic
assessment and 15N recovery of urea amended with the urease inhibitor nBTPT [N-(n-butyl)
thiophosphoric triamide] for temperate grassland, Plant Soil 161, 167-177.
Watson, c.J., Miller, H., Poland, P., Kilpatrick, DJ., Allen, M.B.D., Garrett, M.K, and Christianson, C.B.
(1994b) Soil properties and the ability of the urease inhibitor N-(n-butyl) thiophosphoric triamide
(nBTPT) to reduce ammonia volatilization from surface-applied urea, Soil Bioi. Biochem. 26, 1165-1171.
Watson, CJ., Poland, P., and Allen, M.B.D. (l998) The efficiency of repeated applications of the urease
inhibitor N-(n-butyl) thiophosphoric triamide for improving the efficiency of urea fertilizer utilization on
temperate grassland, Grass Forage Sci. 53, 137-145.
Wenzel, K-D., Dedek, W., and Thieme, H. (1981) Spectrophotometric determination of phosphoric acid
phenyl ester diamide in fertilizers in the presence of additives, Fresenius Z. Anal. Chem. 305, 287-288 (in
German).
Weston, C.W., Peacock, L.A., and Thornsberry, W.L.jr. (l994) Fluid urea-containing fertilizer, u.s Patent No.
5,364,438.
Whitelaw, EG., Milne, J.S., and Wright, D. (1991) Urease (EC 3.5.1.5) inhibition in the sheep rumen and its
effect on urea and nitrogen metabolism, British J. Nutr. 66, 209-225.
Wickremasinghe, K.N., Siva subramanian, S., and Nalliah, P. (l981) Urea hydrolysis in some tea soils, Plant
Soil 62, 473-477.
389
Williams, J.J., Rodman, J .S., and Peterson, C.M. (1984) A randomized double-blind study of acetohydroxamic
acid in struvite nephrolithiasis, N. Engl. J. Med. 311, 760-764.
Winiarski, A. (1990) Investigations on Reducing the Nitrogen Losses from Urea by Inhibitors of Ureolysis,
Jnst. Uprawy Nawozenia i Gleboznawstwa, Pulawy (in Polish).
Wyszkowska, J., Kucharski, J., Jastrzebska, E., and Hlasko, A. (2001) The biological properties of soil as
influenced by chromium contamination, Polish J. Environ. Stud. 10, 37-42.
Xie, R.J., Meier, J., Fyles, J.W., MacKenzie, A.F., O'Halloran, I.P., and Russell, E. (1993) Effect of calcium
lignosulphonates on urea hydrolysis and nitrification in soil, Soil Sci. 156,278-285.
Xie, R.J., MacKenzie, A.F., O'Halloran, J.P., and Fyles, J.W. (1994) Concurrent transformation of
lignosulfonate carbon and urea in clay soil, Soil Sci. Soc. Am. J. 58, 824-828.
Xu, F.H., Sui, W.S., Ying, R. Z., Wang, P., and Tang, S.D. (1994) Studies on protective nitrogen fertilization in
soybean. 1. Effect of hydroquinone allllication, Acta Pedol. Sinica 31 (2), 130-137 (in Chinese).
Xu, X.K., Zhou, L.K., Van Cleemput, 0., and Wang, ZJ. (2000) Fate ofurea- I5 N in a soil-wheat system as
influenced by urease inhibitor hydro quinone and nitrification inhibitor dicyandiamide, Plant Soil 220,
261-270.
Xue, Y.B. and Li, S.L. (1987) Effect of urease inhibitors on the rate of urea hydrolysis in soils, J. Fujian
Agric. Coli. 16,229-235 (in Chinese).
Xue, Y.B., Li, S.L., and Li, Q.L. (1991) Effect of urease inhibitors on the coefficient of urea utilization, J.
Fujian Agric. Coli. 20, 333-339 (in Chinese).
Yadav, D.S., Kumar, V., and Singh, M. (1986) Inhibition of soil urease activity and nitrification with some
metallic cations, Australian J. Soil Res. 24, 527-532.
Yamazaki, K. and Oosaka, T. (1995) Urease inhibitors containing 2-benzamido-3-carbostyrylpropanoic acids
for treatment of gastric mucosal disorders, Japanese Patent No. 07,101,862. Chern. Abstr., 1995, 123,
25680t.
Yarovenko, A.F., Vasil' eva, L.I., and Mussa, K.F. (1982) Effect of magnesium and sodium cations on the
humus and biological activity in a typical chemozem, Sb. Nauch. Tr. Khar'kov. Sel'skokhoz. Inst. 284, 39-
43 (in Russian).
Yeomans, J.e. and Bremner, J.M. (1986) Effects of urease inhibitors on denitrification in soil, Commun. Soil
Sci. Plant Anal. 17, 63-73.
Yeomans, J.e. and Cerrato, M.E. (1993) Growth and nitrogen nutrition of plants amended with chemical
inhibitors ofN transformations, Agron. Abstr., p. 264.
Youngdahl, LJ., Lupin, M.S., and Craswell, E.T. (1986) New developments in nitrogen fertilizers for rice,
Fert. Res. 9,149-160.
Zadak, C.G., Randall, G.W., and Malzer, G.L. (1987) N availability of surface-applied VAN to com as
affected by ammonium thiosulfate and simulated rainfall, Agron. Abstr., p. 271.
Zehmilek, 1.. Minai J., and Honza. J. (1990) Reduction of the volatilization losses of nitrogen from DAM 390
by addition ofhexaaminocyclotriphosphazatriene (HA-CTPAT), Rostl. V:yroba 36, 797-804 (in Czech).
Zhan. J., Lin, D.. and Guo, P. (1993) Manufacture of urea fertilizers containing boron, Boron Agric. 13 (2), 8.
Zhang, M., Nyborg, M., and Feng, Y. (1994) Release mechanism of coated urea in soil: Existence of thin
water film around urea granules, Agron. Abstr., p. 324.
Zhao. P. (1993) Long-acting composite fertilizers, Chinese Patent No. 1,077,445. Chern Abstr., 1994, 121,
8194d.
Zhao. X.Y. and Zhou, L.K. (1991) Effect of urease inhibitor, hydroquinone on soil enzyme activities, Soil
Bioi. Biochem. 23, 1089-1091.
Zhao, X.Y., Zhou, L.K., and Xie, C.G. (1988) Environmental impact assessment of hydroquinone used as a
urease inhibitor (I), Agroenviron. Prot. 7 (6), 18-20 (in Chinese).
Zhao, X.Y., Zhou, L.K.. and Wu. G.Y. (1989) Environmental impact assessment of hydroquinone used as a
urease inhibitor (2). Agroenviron. Prot. 8 (4),28-30 (in Chinese).
Zhao, X.Y., Zhou, L.K., Li, R.H., Xie, C.G., and Li, S.D. (1991) Evaluation of the environmental effect of the
urease inhibitor hydroquinone, Chinese J. Appl. Ecol. 2, 316-322 (in Chinese).
Zhao, X.Y., Zhou, L.K., and Wu, G.Y. (J992a) Vrea hydrolysis in a brown soil: Effect of hydroquinone, Soil
Bioi. Biochem. 24,165-170.
Zhao, X.Y., Xie, C.G.. Zheng, Q.Y., and Wu, Z.H. (1992b) Determination of hydroquinone and biuret in
hydroquinone-containing urea fertilizer, Fert. Res. 33, 93-96.
Zhao, X.Y., Li, S.D., Zhou, L.K., Meng, H.G., Li. R.H., and An, G.R. (1993a) Fate ofC 4 C]hydroquinone and
[ 15Nlurea in a soil-rice system: A pot trial, Soil Bioi. Biochem. 25, 143-146.
Zhao, X.Y., Zhou, L.K.• Li, R.H., An, G.R., and Zhang, B. (1993b) Effect of hydro quinone on maize yield and
urea efficiency. Soil BioI. Biochem. 25, 147-148.
390
Zhao, P., Wang, B.S., Han, Y.Q., and Gao, S.Q. (1993c) Effect of humic acids and urea on urease activity,
Chinese J Soil Sci. 24, 135-136 (in Chinese).
Zhou, L.K., Wu, G.Y., Zhang, Z.M., Cao, C.M., and Li, Y.H. (1988) Effect of urease inhibitor hydroquinone
on the efficiency of urea fertilizer, Acta Pedol. Sinica 25 (2), 191-198 (in Chinese).
Zhou, L.K., Zhao, X.Y., Li, R.H., Wu, G.Y., and Wang, Z.P. (1992) Effect of the urease inhibitor
hydroquinone on urea-N tmnsformation in soil, Chinese J Appl. Ecol. 3,36-41 (in Chinese).
Zhou, L.K., Xu, X.K., Chen, L.l., Li, R.H., and Van Cleemput, O. (1999) Effect of hydroquinone and
dicyandiamide on the emission ofN2 0 and CH. from a lowland rice soil. Chinese J Appl. Ecol. 10. 189-
192 (in Chinese).
Ziyamukhamedov, LA., Sultanov, S.A., Poberezhskaya, S.K., and Khudzhanazarova, D.A. (1986)
Effectiveness of sulfur-containing nitrification inhibitors for cotton, Abstr.. All-Unional Con!
"Perspectives of the Utilization of Nitrification Inhibitors for Increasing Efficiency of Nitrogenous
Fertilizers" (Moscow, 1986), pp. 102-103 (in Russian).
Zukowska-Wieszczek, D. (1979) Influence of lead addition on enzymatic activity and mineralization of
organic matter in soils of green areas in Warsaw, Czlowiek Srodowisko (3/4), 125-141 (in Polish).
Zyrin, N.G., Ras'kova, N.V:, and Platonov, G.V. (1980) Effect of heavy metals on enzymatic activity of soils,
Abstr.. All-Unional Con! "Melioration, Use and Protection of Soils of the Nonchernozemic Zone"
(Moscow, 1980), p. 186 (in Russian).
Appendix
The review papers listed below were also valuable sources of information for elaboration of the Introduction
and Chapters 1-9 of this book.
Beaton, J.D. (1978) Urea: Its popularity grows as a dry source of nitrogen, Crops Soils Mag., 30 (6), 11-14.
Bremner, lM. (1990) Problems in the use of urea as a nitrogen fertilizer, Soil Use Manage. 6 (2), 70-71.
Bremner, 1.M. (1995) Recent research on problems in the use of urea as a nitrogen fertilizer, Fert. Res. 42, 321-329.
Bremner, 1.M. and Hauck, R.D. (1974) Perspectives in soil and fertilizer nitrogen research, Trans. 10th Int. Congr.
Soil Sci. (Moscow, 1974) 9, 13-29.
Bremner, 1.M. and Mulvaney, R.L. (1978) Urease activity in soils, in R.G. Bums (ed.), Soil Enzymes, Acad.
Press, London, pp.149-196,
Buresh, R.J. and Baanante, CA. (1993) Potential economic benefits of modifications to urea that increase yield
through reduction in nitrogen losses, Agron. J 85,947-954.
Byrnes, B.H. and Freney, J.R. (1995) Recent developments on the use of urease inhibitors in the tropics, Fert. Res.
42,251-259.
Christianson, C.D. and Schultz, JJ. (1989) Potential for alternative nitrogen fertilizers in developing country
agriculture, Proc. AKro-Econ. Meeting Int. Fert. Ind. Assoc. (Budapest, 1989), pp. 24-72.
Faurie, G. and Bardin, R. (1979) Volatilization of ammonia. I. Influence of the nature of soil and nitrogen
compounds, Ann. Agron. 30, 363-385 (in French).
Fertiliser Association ofIndia (1977) Fertiliser urea- its efficient use, Fert. News 22 (9), 3-18.
Gasser, J.K.R. (1964) Urea as a fertilizer, Soils Fert. 27, 175-180.
Gorelik, L.A. and Gritsevich, Yu.G. (1984) Inhibitors of soil urease activity, Agrokhimtya (2), 103-126 (in Russian).
Gould, W.O., Hagedorn, C., and McCready, R.G.L. (1986) Urea transformations and fertilizer efficiency in soil,
Adv. Agron. 40, 209-238.
Hargrove, W.L. (1988) Soil, environmental, and management factors influencing ammonia volatilization under
field conditions, in B.R. Bock and D.E. Kissel (eds.), Ammonia Volatilization jrom Urea Fertilizer.s, Natl. Fert.
Dev. Center. Muscle Shoals, Alabama, pp. 17-36.
Harre, E.A. and Bridges, J.D. (1988) Importance of urea fertilizers, in B.R. Bock and D.E. Kissel (eds.), Ammonia
Volati/izationjrom Urea Fertilizer.s, Natl. Fert. Dev. Center, Muscle Shoals, Alabama, pp. 1-15.
Hauck, R.D. (1972) Synthetic slow-release fertilizers and fertilizer amendments, in CA.L Goring and 1.W.
Hamaker (eds.), Organic Chemicals in the Soil Environment, M. Dekker, New York, pp. 633-690.
Hauck, R.D. (1983) Agronomic and technological approaches to minimizing gaseous nitrogen losses from
croplands, in J.R. Freney and J.R. Simpson (eds.), Gaseous Loss of Nitrogen from Plant-Soil Systems, M.
Nijhofl7Dr. W. Junk, The Hague, pp. 285-312.
Hauck, R.D. (1984) Technological approaches to improving the efficiency of nitrogen fertilizer use by crop plants,
in R.D. Hauck (ed.), Nitrogen in Crop Production, Am Soc. Agron.-Crop Sci. Soc. Am-Soil Sci. Soc. Am,
Madison, pp. 551-560.
Hauck, R.D. (1985) Slow-release and bioinhibitor-amended nitrogen fertilizers, in O.P. Engelstad (ed.),
Fertilizer Technology and U~e (3rd Edition), Soil Sci. Soc. Am., Madison, W. 293-322.
391
Kiss, S., Driigan-Bularda, M., and Riidulescu, D. (1975) Biological significance of enzymes accumulated in
soil, Adv. Agron. 27, 25-87.
Kiss, S., Pllljca, D., Driigan-Bularda, M., Zborovschi, E., and Cri§llIl, R. (1991) Inhibition of soil urease
activity for increasing the efficiency of urea fertilizer, Environ. Enzymol. (Bucharest) 1, pp. 93-288 (in
Romanian).
KoelJiker, J.K. and Kissel, D.E. (1988) Chemical equilibria affecting ammonia volatilization, in B.R. Bock
and D.E. Kissel (eds.), Ammonia Volatilization from Urea Fertilizers, Nat!. Fert. Dev. Center, Muscle
Shoals, Alabama, pp. 37-52.
Ladd, J.N. and Jackson, R.B. (1982) Biochemistry of ammonification, in FJ. Stevenson (ed.), Nitrogen in
Agricultural Soils, Am. Soc. Agron.-Crop Sci. Soc. Am.-Soil Sci. Soc. Am., Madison, pp. 173-228.
Matzel, W., Ackermann, W., Brinschwitz, W., Buchler, D., Hannusch, L. Heymann, W., Kretschmar, M.,
Uppold, H., Runge, P., and Teske, W. (1974) Effective application of urea as a fertilizer in agriculture,
Fortschrittsber. Landw. Nahrungsguterw. (Berlin) 11. (7-8), 1-68 (in German).
Medina, R. and Radel, R.J. (1988) Mechanisms of urease inhibition, in B.R. Bock and D.E. Kissel (eds.),
Ammonia Volatilizationfrom Urea Fertilizers, Natl. Fer!. Dev. Center, Muscle Shoals, Alabama, pp. 137-
174.
Mulvaney, R.L. and Bremner, J.M. (1981) Control of urea transformations in soils, in E.A. Paul and J.N. Ladd
(eds.), Soil BiochemiStry, Vol. 5, M. Dekker, New York, pp. 153-196.
Nelson, D.W. (1982) Gaseous losses of nitrogen other than through denitrification, in F.J. Stevenson (ed.),
Nitrogen in Agricultural Soils, Am. Soc. Agron.-Crop Sci. Soc. Am.-Soil Sci. Soc. Am., Madison, pp.
327-363.
Radel, R.J., Gautney, J., and Peters, G.E. (1988) Urease inhibitor developments, in B.R. Bock and D.E. Kissel
(eds.), Ammonia Volatilization from Urea Fertilizers, Natl. Fert. Dev. Center, Muscle Shoals, Alabama,
pp.I11-136.
Sahrawat, K.L. (1979) Nitrogen losses in rice soils, Fert. News 24 (12), 38-48.
Sahrawat, K.L. (1980) Control of urea hydrolysis and nitrification in soil by chemicals - prospects and
problems, Plant SoilS7, 335-352.
Scharf; P.C. and Alley, M.M. (1988) Nitrogen loss pathways and nitrogen loss inhibitors: A review, J. Fert.
Issues S (4), 109-125.
Sullivan D.M. and Havlin, J.L. (1988) Agronomic use of ammonium thiosulfate to improve fertilizer
efficiency, J. Fert. Issues S (2), 37-44.
Terman, G.L. (1979) Volatilization losses of nitrogen as ammonia from surface-applied fertilizers, organic
amendments, and crop residues, Adv. Agron. 31, 189-223.
Tisdale, S.L., Nelson, W.L., and Beaton, J.D. (1985) Soil Fertility and Fertilizers (4th Edition), Macmillian,
New York, pp. 161-168 and 177-178.
Trenkel, M.E. (1997) Improving Fertilizer Use Efficiency. Controlled-Release and Stabilized Fertilizers in
Agriculture, In!. F ert. Ind. Assoc. (IFA), Paris.
Tucker. T.C. and Westerman, R.L. (1989) Gaseous losses of nitrogen from desert region soils, Arid Soil Res.
Rehabil. 3, 267-280.
Voss, R.D. (1984) Potential for use of urease inhibitors, in R.D. Hauck (ed.), Nitrogen in Crop Production,
Am. Soc. Agron.-Crop Sci. Soc. Am.-Soil Sci. Soc. Am., Madison, pp. 571-577.
Watson, C.J., Stevens, R.J., Garrett, M.K., and McMurray, C.H. (1990) Efficiency and future potential of urea
for tempemture gmssland, Fert. Res. 26,341-357.
Yeomans, J.e. (1991) Inhibition of nitrogen transfurmations in soils. Potentials and limitations for agriculture,
Trends Soil Sci. 1, 127-158.
393
SUBJECT INDEX
A CT Conventional till (Till/tillage,

Abbreviations conventional)
ADP Adenosine diphosphate CTPAT Cyclotriphosphazatriene
AH Ammonium humate CUA Cyanuric acid
AHA Acetohydroxamic acid CX Cellulose xanthate
ALS Ammonium lignosulfonate (DAP) (Diaminophosphinyl)
AM 2-Amino-4-cWoro-6-methyl- DAP Diammonium phosphate
pyrimidine DAPBA N-(Diaminophosphinyl)
AMD Acid mine drainage benzamide
AN Ammonium nitrate DAPBAA N-(Diaminophospbinyl)
ANH Ammoniated nitrohumate benzeneacetamide
APP Amidophosphoric acid mono- DAPT Diamidophosphorothiolate
phenyl ester DAT Days after transplanting
APP Ammonium polyphosphate DATPA Diammonium thiophos-
AS Ammonium sulfate phoroamidate
AT Aldrithiol DBC 2,6-Dibromoquinone-4-chloro-
ATC 4-Amino-l ,2,4-triazole hydro- imide
chloride DBQ Dimethyl-p-benzoquinone
ATP Adenosine triphosphate DCC 2,6-Dichloroquinone-4-chloro-
ATPDA Ammonium thiophosphoro- imide
diamidate DCD Dicyandiamide
ATS Ammonium thiosulfate DCDU Dicyandiamide-urea
AZ Azadirachtin DCMI Dichloromaleimide
BA Boric acid DEP A Diethylphosphoroamidate
BAA Na-Benzoyl-L-argininamide DMTP A Dimethylthiophosphoro-
BD (CaLS) Bulk-digested (CaLS) amidate
BHA Benzohydroxamic acid DNP 2,4-Dinitrophenol
y-BHC y-Benzene hexachloride DP 2,2' -Dipyridil
BNC Bromo-nitro compound DPPA Diphenylphosphoroamidate
nBPTA N-(n-Butyl)phosphoric DPTA Diethylphosphoric triamide
triamide DTB Desthiobiotin
BQ p-Benzoquinone DTNP 2,2' -Dithiobis-5-nitropyridine
nBTPT A N-(n-Butyl)thiophosphoric DTPNO 2,2' -Dithiobis-pyridine-N-
triamide oxide
CABA o-Chloro-p-aminobenzoic DTU Dicyandiamide-treated urea
acid EC Electrical conductivity
CaLS Calcium lignosulfonate ED50 Ecological dose - 50%
CAN Calcium ammonium nitrate EDTA Ethylenediaminetetraacetate
CHPTA Cyclohexylphosphoric 2EP 2-Ethynylpyridine
triamide EPDA Ethylphosphorodiamidate
CHTPT A Cyclohexylthiophosphoric E. U. Enzyme unit
triamide FD Fiber digestibility
CMI Chloromaleimide FYM Farmyard manure
CT Catechol GTU Guanylthiourea
394
HA humic acid PNO Pyridine-N-oxide

HA-CTPAT Hexaaminocyclotri- PP A Phenylphosphoroamidate
phosphazatriene PPDA Phenylphosphorodiamidate
HAD Hydroxamic acid derivative PPTA Phenylphosphoric triamide
HMTA Hexamethylenetetramine PRPyrite
HPLC High-performance liquid PSA Pyridine-3-sulfonic acid
chromatography PTA Phosphoric/phosphoryl triamide
HQ Hydroquinone PU Prilled urea
HS Humic substance QH Quinhydrone
HU Hydroxyurea SA Sulfanilamide
IPP Imidodiphosphoric acid diphenyl SCU Sulfur-coated urea
ester SF (CaLS) Sugar-free (CaLS)
KEtX Potassium ethyl xanthate SH Sodium humate
LCU Lac-coated urea SP Superphosphate
LIT Long time test ST Sulfathiazole
MCT 4-Methylcatechol STS Sodium thiosulfate
MH Maleic hydrazide SIT Short time test
MI Maleimide T A Titratable acidity
MNQ 2-Methyl-p-naphthoquinone TCA Trichloroacetate
MP 2-Mercaptopyridine TD Thiadiazole
MPC 3-Methylpyrazole-l-carbox- TLC Thin-layer chromatography
amide TPDA Thiophosphorodiamidate
MPM 2-Mercaptopyrimidine TPDA Trichloroethylphosphoro-
MPNO 2-Mercaptopyridine-N-oxide diamidate
MPOL 2-Mercapto-3-pyridinol TPTA Thiophosphoric/thiophospho-
NCNeemcake ryl triamide
NCCU Neem cake-coated urea TUThiourea
NCMU Neem cake mixed with urea UAN Urea-ammonium nitrate
NHA Nitrohumic acid UMM Urea-MgS04.1H20 mixture
NKE Neem kernel extract USG Urea supergranule
NOCU Neem oil-coated urea VFA Volatile fatty acid
NP Nitrapyrin WHC Water-holding capacity
NQ p-Naphthoquinone Acacia 170
NT No-till (Till/tillage, no-) Acacia decurrens 169
NTA Nitrilotriacetate Acetaldehyde 180
NU Nitrapyrin-amended urea Acetate 15,45. 134, 155,355
ODA Octadecylamine Acetic acid 32, 134
OTC Oxythiamine chloride Acetohydroxamic acid 60,61, 162,
P A Phenylacetylene 163, 180, 182, 191, 192, 195, 196,
P A Phosphoroamidate 220, 352-354, 357
PCMB p-Chloromercuribenzoate Acetone 92, 94, 110
PDA Phosphorodiamidate 3' -N-Acetylaminophenyl-p-benzo-
PG Phosphogypsum quinone 84
PHMB p-Hydroxymercuribenzoate Acetylene 182,244,249,336,340
PM Poultry manure Acetylenes, disubstituted 226
PMA Phenylmercuric acetate -, monosubstituted 226
PMB Phenylmercuric borate Acetyleneurea 30 I
395
Acidifying agents 247, 248 2-Amine-2-oxide-I,3 ,2-benzodioxa-

Acid mine drainage 358 phosphole 138
Actinomycetes 329, 330, 331, 332 -, phosphorodiamidic acid esters of
N-Acyl phosphoric triamides 240, 357 138
Adenosine diphosphate 209 2-Amine-2-thio-l ,3,2-benzodioxa-
Adenosine triphosphate 209, 341, 342 phosphole 138
Agrotain 154,260,297,359 -, phosphorodiamidic acid esters of
Alanine 336-338 138
Aldehydes 2, 179 Amino acids 310, 312
Aldrithiol 101 1-Aminoanthraquinone-2-sulfonate 91
Alfalfa 275,277,313-315 3-Aminobenzeneboronic acid hemi-
Algae 116, 120,209,247-249,282 sulfate 46
-, blue-green 247 4-Aminobenzeneboronic acid hydro-
Algal inhibitors 220, 243, 249, 287 chloride 46
Algicides 182, 183,247,248,250,288 1-Amino-2-chloroanthraquinone 91
Aliphatic amines 32 2-Amino-4-chloro-6-methylpyrimidine
- dihydroxamic acids 65, 66 222,237
Alkali metal salts 20-30, 72, 234, 251, 4-Amino-m-cresoI80
297,315,322,334,335 3-Amino-o-cresoI80
Alkaline earth metal salts 20-30, 188, 4-Amino-o-cresoI80, 195
234,251,297,315,322,335 6-Amino-o-cresoI80
Alkanoic acids 32 2-Amino-p-cresoI80
s-Alkyl diamidophosphorothiolates Aminocresols 80,81, 195
141 2-AminophenoI79,81
N-Alkyldichlorornaleimides 66-68 4-Aminophenol 79, 81
Alkyldithiocarbamates 52 Aminophenols 81
Alkylpbosphoric triamides 349 N-2-Aminophenyl-N' -allylthiourea 51
Alkylphosphoroamides 348 N-(4-Aminophenyl)phosphoric
Alkylphosphorodiamidates 141, 199, triamide 143
201 4-Aminophenylphosphorodiamidate
N-Alkyl-2-pyrrolidone 244 113
Allium cepa 315 8-[(4-Aminophenyl)sulfonyl]amino-2-
Allylphosphorodiamidate 112 naphthalenyl phosphorodiamidate
Allylthiourea 51 356
Aluminium 19, 20 [[( 4-Aminophenyl)sulfonyl]amino]
- chloride 19,20 phenylphosphorodiamidates 356
- dimethyldithiocarbarnate 55 2-Aminopyridine 222
- nitrate 20 5-Amino-I,3,4-thiadiazole-2-thiol 358
- oxide 2 3-Amino-l,2,4-triazole 221,223,225
- salts 111, 112 4-Amino-l,2,4-triazole hydrochloride
- sulfate 2, 19,20,247-249,286 221,223-225,227,231,238,268,
Amidase 322 289,291,292
Amidophosphoric acid 105 Amitrole 225, 226
- - monophenyl ester Ill, 112 Ammonia 1,26,28,37,47,49,53,58,
- - - - ammonium salt III 63, '66,67,69, 73, 97, 98, 110, 113,
- - - - heavy metal salts 111 114, 120, 123, 124,26, 129, 130,
132, 135, 137, 147, 149, 162, 164
396
Ammonia (continued) 173-175, 178, Ammonium (continued)

275,276,315,352-354,355,357 - thiocyanate 220
- loss/volatilization 1,3-6,9, 12, 15, - thiophosphorodiamidate 213-215
18-21,23,26-33,37-41,43,44, - thiosulfate 36-42, 183, 184, 188,
46-48,51-57,61-64,66-73,75, 192,198,204,235,243,244,252,
77,84-86,88,93,94,97,98,107, 253,262-264,296,298,299,306,
108, 110-129, 133, 135, 146-154, 307,313,334,347,352
158,159,162-172,174,175,177, - trithiocarbonate 36, 223
179-181,183-185,187-189,192, Amylammonium salts 73, 74
195-206,208,210,211,217-219, Amylase 321,324,325
221,224,225,230-233,235,236, Anacardium occidentale 176
242,244-247,249-251,257-260, Aniline 104, 105
265,269,270,274,275,277,279, p-Anisidine 178
286,288,290,292,293,297-299, Anisole 178
307,308,311,318,335,340,352 Anthraquinone 83, 91, 95, 96
Ammoniated lignosulfonate 241 Anthraquinone-2,6-disulfonate 91
- nitrohumate 166 Anthraquinone-I-sulfonate 91
Ammonification 20 Antimetabolites 162-165, 261
Ammonium 1,4,53,90,94,98,115, Antimicrobial agents 70, 71, 135
116,119-123,125-127,130,134, Antiseptic/antiseptics 2, 35, 79
135, 148, 153, 154, 164, 165, Apatite 356
167-169,172-176,204,209,211, Arginine deamination 336
266,267,282,343,347,348 Arrhenius plot 100, 213
- carbonate 1,23,26,29 Arsenic chloride 34
- chloride 26, 133, 134, 159,241 - compounds 34
- N-p-chlorophenyldithiocarbamate - pentoxide 34
54 - trioxide 34
- dihydrogenphosphate 254 Arylsulfatase 321,322,325,328,330,
- fluoride 188 334,339
- humate 167,328 Aspergillus ochraceus 177
- hydroxide 33, 167 - oryzae 355
-lignosulfonate 168, 169, 184, 185, Asphalt 2, 5, 6, 30, 33, 46, 47
261,262,279,328,332,333,335, Avena sativa 292
341 Azadirachta indica 171
- molybdate II, 15, 187,246 Azadirachtin 171, 172
- nitrate 10, 17,28,38,49, 115, 148, Azide 221
153, 154, 183, 184, 198, 199,201, Azotobacter 329, 330, 332
214,243,244,252-260,263,265, - chroococcum 332
269-275,289,290,292-301,303-
305,308-310,313,316,317,348- B
350,352 Bacillus pasteurii 216, 355
- nitrohumate 166, 167 Bacteria 329-333, 354-358
- polyphosphate 37-40, 244, 253, 262, -, ammonifying 330-332
299 -, autotrophic ammonium-oxidizing
- salts 72, 111, 112, 181 227
- sulfate 29,130,131,167,199,224, -, butyric acid 330
226,235,237-241,263,276 -, cellulolytic 333
397
Bacteria (continued) Bioinsecticide 171

-, copiotrophic 330 Bis(acetylvinyl) sulfide 178
-, denitrifying 331 N,N-Bis-(2-chloroethyl)phosphoric
-, heterotrophic 329, 332, 333 triamide 142
-, nitrifying 330, 331, 333 N,N' -Bis-(diaminophosphinyl)-1,6-
-, N 2-fixing 330, 331 diaminohexane 139
-, oligotrophic 330 N,N'-Bis-(diaminophospbinyl)-1,8-
-, phytopathogenic 65, 66, 70 diamino-p-menthane 139
-, proteolytic 333 N,N' -Bis-(diaminophosphinyl) pi-
- tolerant to Cu and Cd 330 perazine 139
-, ureolytic 331 Bisdiethylpbospboric triamide 349, 350
Banana plantation 231 Bisdimetbylphospboric triamide 349,
Barium 179 350
- chloride 21, 24 Bismuth nitrate 12
Barley 10, 16, 19,49,212,275,276, Bis-(2-oximinophenylacetonitrile) ester
288-293,295,328,339,348 of cyclohexylthiophosphonic acid
Bassia latifolia 176 160
Beet 16 Bis-(2-oximinophenylacetonitrile) ester
3-Benzamido-3-carbostyrylpropanoic of phenylphosphonic acid 160
acids 356 Bisthiourea 51, 52
Benzene 178 Bitumen Ill, 114,289,293,294,304
y-Benzene hexachloride 163, 164 Biuret 48,49, 101,223,276,347,348
2-Benzenesulfonamido-2-thiono-5,5- Bluegrass 21, 146,204,210,297,298,
dimethyl-1,3,2-dioxaphosphorinane 306,307
161 Borate 45, 134
Benzohydroxamic acid 60, 61, 63, 64, Borax 30, 32, 33, 179, 180, 188,280,
178,265,314 292,298
o-Benzoquinone 83 Boric acid 2,31-33, 134, 180, 182-184,
p-Benzoquinone 59, 82-96,98-101, 347,353
172,177,178,180,191-193,237, Boron 179, 188,349
266,267,275,278,301,312-314, - oxide 31
324-326,334,335,337,338,353 Botriochloa caucasica 297
Benzoyl crysean 77, 78 Brassica napus 315
3-Benzyl-5-hydroxyethyltetrahydro- Brij 92 32
1,3,5-thiadiazine-2-thione 71 Broadbean 16, 313, 329
N-Benzyl-N-methylphosphoric Bromegrass 297,299,309
triamide 196, 197,200,201,216, Bromoacetohydroxamic acid 64
257,336 I-Bromo-3-chloro-4,4,5,5,-tetramethyl-
3-Benzyl-5-methyltetrahydro-l,3,5- 2-imidazolidinone 103,239
thiadiazine-2-thione 71 3-Bromo-4,4-dimethyl-2-oxazolidinone
Benzylphosphorodiamidate 112, 113 103
3-Benzylrhodanine-5-acetic acid 76 2-Bromoethylphosphorodiamidate 112
Bermudagrass 53, 297-300 2-Bromo-2-nitrobutyl-N-methylcarba-
Betanal317 mate 70
Betanil 70 317 Bromo-nitro compounds 70, 265, 300
Beta vulgaris cv. altissima 315 2-Bromo-2-nitropropane-l,3-diol 70
BIN No.7 3 2-Bromo-2-nitropropanoI70
398
2-Bromo-2-nitropropyl-N-methyl- Calcium ammonium nitrate 150, 152,

carbamate 70, 265, 300 308,310
4-Bromophenylphosphorodiamidate - carbide 182, 183,244,245,249,280
110 - carbonate 21-24,26,27, 72, 90, 151,
N-(3-Bromopropyl)phosphoric tri- 234,298
amide 142 - chloride 20-30,38, 188, 189,211,
Bromus inermis 297 234,251,297-299,316
n-Butylamine 31 - cyanamide 229, 232
2-(t-Butylamino)-4-(ethylamino)-6- - dibydrogenphosphate 29, 234, 247,
(methylthio)-s-triazine 182,249 248,281,348,350,351
4-t-ButylcatechoI91, 92, 267 - fluoride 33
S-Butyldiamidophosphorothiolates 142 - hydroxide 218
S-(iso-Butyl)diamidophosphorothiolate - ions 22,53
141 -lignosulfonate 168
S-(n-Butyl)diamidophosphorothiolate - sulfate 22, 29, 30, 234
141 - sulfide 35, 191
S-(sec- Butyl)diamidophosphorothiolate Camellia sinensis 169
141 Campylobacter pylori 356
N-n-Butyldichloromaleimide 66, 67 - pyloridis 356
2-n-Butyl-3,5-di-n-propylpyridine 228 Canavalia ensiformis 25
N-(n-Butyl)phosphoric triamide 143, Caprohydroxamic acid 60
182,199,215-220,249,250,288, Caprylohydroxamic acid 60, 61, 64
336 Carbamates 56
N-(sec-Butyl)phosphoric triamide 143, l-Carbamoyl-3-methylpyrazole 227
199 Carbohydrate content 310
n-Butylphosphorodiamidate 348 Carbon disulfide 35, 36, 59, 223
3-n-Butylrhodanine-5-acetic acid 76 Carboxymethylthio-p-benzoquinone 84
N-(n-Butyl)thiophosphoric triamide Cardonite 24, 25
143,144,146-154,167,177,182- Carrot 16
184, 195-213,215-220,238-241, Cashew 176
244-246,248,249,256-260,263, Castor-oil 177
275-280,285,286,288,291,296, Catalase 322, 323, 326-328
297,306-311,318,319,332,335- Catechin 356
338,355,359 Catechol 3, 82-86, 89-91, 94-96, 98-
n-Butylurea 47, 48 101, l36, l37, 177, 180, 191-193,
t-Butylurea 47,48 237,266-268,275,278,314,324-
Butyric acid 353 326,337-339
Caucasian bluestem 297,309
C Cellulase 321
Cadmium 15,313,322,336 Cellulose decomposition 335
- chloride l3, 14, 17 - xanthate 58, 236
- ions 9, 13, 17,233 Cereals 271,272,289,290,304,305
- nitrate 9,15-17 Chlorides 7, 9, 13,26
- salts 76 3-Chloroacetanilide 222
- sulfate 11, 13, 14, 19,322,330,342 Chloroacetohydroxamic acid 64
Caffeine 178, 192 o-Chloro-p-aminobenzoic acid 163,
164,261
399
2-Chloro-4-aminothiophenol, zinc Chromium 322, 330, 334, 339

mercaptide of 81 - chloride 7, 13, 14,233,321
m-Chloroaniline 222 - ions 9, 13, 16
o-Chloroaniline 222 - oxide 15,321
p-Chloroaniline 222 - sulfate 13, 15
1-Chloroanthraquinone 91, 325 Citraconic acid 178
Chloro-p-benzoquinone 84 CitraI311,312
4-Chloro-N-( diaminophosphinyl) benz- Citric acid 178
amide 193, 278 Citrus nobilis 187
3-C hloro-4 ,4-dimethyl-2 -oxazo lidinone CL-1580 221, 222
103 Clay 2,52,244
N-(2-Chloroethyl)phosphoric triamide Coal 165
142,199 Coal tar 173, 176, 225, 232
2-Chloroethylphosphorodiamidate 112 Cobalt 8, 20
Chloroform 97 - salts 76, 112
Chlorohydroquinones 95 Cobaltous chloride 7, 13
Chloromaleimide 67, 68 - sulfate 12, 13, 15, 187
p-Chloromercuribenzoate/-benzoic acid Cocoa palm 173
43-45,190,191,235,337-339 Cocos nucifera 173
4-Chloro-2-nitrophenol 79 Coefficient of nitrogen utilization by
4-Chlorophenol 79 plants 269, 280, 301-304, 312, 313,
4-Chlorophenyl-N,N' -di-n-butylphos- 350
phorodiamidate 106 Common bean 313, 314, 348, 350, 351
4-Chlorophenyl-N,N' -diisopropyl- Copper 15, 76,179,322,330,336,349
phosphorodiamidate 106 - chelate 248, 249, 285, 286
4-Chlorophenyl-N,N' -dimethylphos- - salts Ill, 112
phorodiamidate 106 Corynebacterium renale 356
4-Chlorophenyl-N,N' -diphenylphos- Cotton 224,231,242,315,318,355
phorodiamidate 106 m-Cresol78
4-Chlorophenyl-N,N' -di-n-propyl- o-Cresol78
phosphorodiamidate 106 p-Cresol78
2-Chlorophenylphosphorodiamidate Cresols 78
106,110 m-Cresyl-N,N' -dimethylphosphorodi-
3-Chlorophenylphosphorodiamidate amidate 106
110 o-Cresylphosphorodiamidate 109
4-Chlorophcnylphosphorodiamidate Critical temperature 100
106, 107, 109, 110, 114, 181,278, Crotonylidene diurea 2
315 Crysean 77, 78
3-Chloropropylphosphorodiamidate Cucumber 227,315,316
112, 113 Cucumis sativus 315
2-Chloropyridine 222 Cucurbita pepo 315
6-C hloro-6-( trichl oro methyl )pyridine Cupric acetate 8
222 - carbonate 15
Chromatography, gel 166 - chloride 7, 12-15, 17, 18, 233, 321
-, high-performance liquid 110, 132, - diethyldithiocarbamate 52
133,215-218 - hydroxide 15
-, soil thin-layer 45, 100, 219
400
Cupric (continued) D
- ions 5, 8, 9, 11, 13, 14,20,53, 192, Dactylis glomerata 28, 201, 297
233 Dadap 170
- nitrate 8, 11, 15 DAM 390 41, 115, 159,261
- oxide 6 Dazomet 71, 72, 128, 181, 195,236,
- sulfate 5-10,12-15,17,30,33,177, 316,324,331
179,180,187,188,190,242,247, Dehydrogenase 17,321-326,328,329,
248,250,288,318,330,343 342
Cuprous chloride 7 -, actual 326, 327
- sulfate 12 -, potential 326, 327
Cutrine Plus 248 Denitrification 120, 121, 127, 149, 168,
Cyanamide 229, 230 274,337-340
Cyanobacteria 247 Desthiobiotin 163-165
4-Cyano-N-(diaminophosphinyl) benz- Desulfuration 216, 217
amide 193, 278 Dhaincha 175
Cyanoguanidine 229 Dialkyldithiocarbamates 52
Cyanuric acid 276, 354, 355 3,5-DiaUyltetrahydro-l ,3,5-thiadiazine-
S-Cycloalkyldiamidophosphoro- 2-thione 71
thiolates 141 Diamidophosphoric acid 105
N-Cyclohexylchioromaleimide 68 - - phenyl ester 105
S-Cyclohexyldiamidophosphoro- Diamidophosphorothiolate compounds
thiolate 141 141, 142
N-Cyclohexyldichloromaleimide 68 Diamidothiophosphoric acid 105
N-Cyclohexylmaleimide 68 Diamidothiophosphorothiolate com-
N-Cyclohexylphosphoric triamide 196, pounds 141
199-202,217-220,249,250,257, N-(Diaminophosphinyl)arylcarbox-
275,279,288,336,352,353 amides 356
Cyclohexylphosphorodiamidate 112, N-(Diaminophosphinyl)benzamide
113 182, 193, 195, 196,275,278,337,
3-Cyclohexylrhodanine-5-acetic acid 338
76 N-(Diaminophosphinyl)benzene-
N-Cyclohexylthiophosphoric triamide acetamide 193,278,337,338
143,217-220 N-(Diaminophosphinyl)benzenesulfon-
Cyclophosphazane compounds 350 amide 141
Cyclophosphazanic acids, potassium N-(Diaminophosphinyl)-2-chloroacet-
salts of350 amide 143,240
Cyclotetraphosphazatetraene deri- O-Diaminophosphinyl derivatives of
vatives 159 oximes 140
Cyclotriphosphazatriene derivatives N-(Diaminophosphinyl)-2,2-dichloro-
155-160,220,260,278,291,297, acetamide 143,240
315,318,332,335 N-(Diaminophosphinyl)-4-fluorobenz-
Cymbopogam confertiflorus 170 amide 357
Cymbopogon flexuosus 297 N-(Diaminophosphinyl)-4-( l' -male-
- - var. flexuosus 311 imido)benzamide 144
- winterianus 231 N-(Diaminophosphinyl)-4-methoxy-
Cynodon dactylon 297 benzamide 240
401
O-(Diaminophosphinyl)-2-propanone 2,4-Di-t-butylphenol 79, 325

oxime 140 4,6-Di-t-butylpyrogallol 92, 325
N-(Diaminophosphinyl)-3-pyridine- 4,6-Di-t-butylresorcinol 92, 325
carboxamide 193, 278 Dicalcium phosphate 293
N-(Diaminophosphinyl)sulfamides 140 2,5-Dichloro-p-benzoquinone 84-87,
N-(Diaminophosphinyl)sulfinamides 237,278,314,337,338
140 2,6-Dichloro-p-benzoquinone 84-87,
N-(Diaminophosphinyl)sulfonamides 89,237,337,338
140 2,3-Dichloro-5,6-dicyano-p-benzo-
N-(Diaminophosphinyl)-p-toluene- quinone 87
sulfonamide 141 2,5-Dichloro-3,6-dihydroxy-p-benzo-
N-(Diaminophosphinyl)-2,2,2-tri- quinone 87
chloroacetamide 143,240 (l,~-Dichloro-formyl-acrylic acid 69
N-(Diaminophosphinyl)-2,2,2-tri- 2,3-Dichlorohydroquinone 91,92,267
fluoroacetamide 143 2,5-Dichlorohydroquinone 89
3,5-Diamino-l,2,4-thiadiazole 75 Dichloromaleimide 67, 68
2,2' -Di(5-amino-l,3,4-thiadiazole) - alkyl derivatives 66, 67
disulfide 74, 75 - aryl derivatives 66, 67
O-Diaminothiophosphinyl derivatives - cyc10alkyl derivatives 66, 67
of oximes 140 2,5-DichlorophenoI79
N-(Diaminothiophosphinyl)sulfamides Di-( 4-Chlorophenyl)phosphoroamidate
140 106
N-(Diaminothiophosphinyl)sulfin- 1,3-Dichloro-2-propylphosphorodi-
amides 140 amidate 112, 113
N-(Diaminothiophosphinyl)sulfon- 2,6-Dichloroquinone-4-chloroimide 83,
amides 140 89,99,100
2,4-Diamino-6-trichloromethyl-s-tri- Di-o-cresylphosphoroamidate 109
azine 222 Di-p-cresylphosphoroamidate 109
Diammonium monoamidothiophos- Dicyandiamide 75, 221, 229-234, 236,
phate 348 239-241,243-246,262,263,273,
- phosphate 168, 169,239,332,348- 274,279,280,287,288,313,318,
350 319,336,340
- thiophosphate 348 Dicyandiamide-urea 262
- thiophosphoroamidate 213, 214 Didin 229
Dibenzylthiuram disulfide 57 2,5-Diethoxy-p-benzoquinone 84, 85
2,6-Dibromo-p-benzoquinone 87 2,5-Diethyl-p-benzoquinone 84
2,5-Dibromo-3,6-dihydroxy-p-benzo- Diethyldithiocarbamate 53, 55, 312
quinone 87 Diethylphosphoric triamide 196, 197,
2,6-Oi bromoquinone-4-chloroimide 199,201,254-257,336,349,350
100,326 Diethylphosphoroamidate 349
1,3-Dibromo-4,4,5,5-tetramethyl-2- Diethylphosphorodiamidate 308
imidazolidinone 103 3,5-Diethyltetrahydro-l,3,5-thiadiazo-
4,6-Di-t-butyl-o-benzoquinone 91, 92 line-2-thione 71
2,5-Di-t-butyl-p-benzoquinone 87 N,N' -Dihalo-2-imidazolidinones 103
2,6-Di-t-butyl-p-benzoquinone 87 Oihydric phenols 82, 136, 192
4,6-Di-t-butylcatechol 91, 92, 267,325 1,2-Dihydro-3,6-pyridazinedione 69
2,5-Di-t-butylhydroquinone 91, 325 Dihydroxamic acids 60, 65, 66, 352
402
1,2-Dihydroxyanthraquinone-3-sulfo- Dinonyl-N,N' -diethylphosphoro-

nate 91 amidate 106
2,5-Dihydroxy-p-benzoquinone 85, 87, 2,4-Diphenoxy-2,4,6,6-tetraamino-
88 cyclotriphosphazatriene 155-158,
6-(3' ,4' -Dihydroxy-6' -fluorophenyl) 278
hexanehydroxamic acid 65 2,4-Diphenoxy-2,4,6,6-tetrachloro-
3,6-Dihydroxy-2-isopropyl-5-methyl- cyclotriphosphazatriene 156
p-benzoquinone 87 2,5-Diphenyl-p-benzoquinone 87, 88
2,5-Dihydroxymethyl-p-benzoquinone Diphenyl-N-methylphosphoroamidate
84 106
3,6-Dihydroxypyridazine 69 Diphenylphosphoroamidate 106, 107,
DiisobutyIthiuram disulfide 57 109, 181
Diisopropylthiuram disulfide 57 Diphenylphosphorodiamidate 114, 133,
Dill 16 134
2,5-Dimercapto-l,3,4-thiadiazole 73- 5,5' -Di(3-phenyl-l,3,4-thiadiazoline-2-
75,358 thione) disulfide 74, 75
-, amylammonium salt of 73, 74 Diphosphorodiamides 139, 140
2,5-Dimethoxy-p-benzoquinone 87 DipiperidyIthiuram disulfide 57
2,6-Dimethoxy-p-benzoquinone 87 Dipiperidylthiuram sulfide 57
p-Dimethylaminobenzaldehyde 35 Di-n-propylthiuram disulfide 57
p-Dimethylaminopropionohydroxamic 2,2' -Dipyridil 10 1, 102
acid 64 Disodium anthraquinone-I ,5-disulfo-
Dimethylammonium dimethyldithio- nate 91
carbamate 53, 55 - ethylene-l ,2-bisdithiocarbamate 54
Dimethyl-p-benzoquinone 344 2,2' -Dithiobis-5-nitropyridine 101, 102
2,3-Dimethyl-p-benzoquinone 87, 88, 2,2'-Dithiobispyridine 101, 102
237 - N-oxide 101, 102
2,5-Dimethyl-p-benzoquinone 84-90, Dithiocarbamates 52-56, 180,264,300,
180,193,237,266,267,277,278, 316,323,330,334,340,342
300,314,324,325,337,338 Dithiophosphorodiamides 139
2,6-Dimethyl-p-benzoquinone 87-89, Dolomite 24, 299
237,238,337,338 Duameen T 31
Dimethyldithiocarbamate 53 Dwell 226
3-( I ',1' -Dimethylethyl)-4-hydroxy-
phenylphosphorodiamidate 113, E
138,239 Ebrotidine 357
N,N-Dimethylphosphoric triamide 196, Ecobet357
201,257,336,349,350 Ecological dose-50% 14, 16, 19,322
Dimethylphosphoroamidate 348 Energy of activation 99, 100
3,5-DimethyItetrahydro-I,3,5-thia- - barriers 98
diazine-2-thione 71, 72, 324, 331 Enthalpy of activation 99
DimethyIthiophosphoroamidate 349 Entropy of activation 99, 100
DimethyIthiuram disulfide 57 Epicatechin 356
N,N-Dimethylurea 47,48 3-(1' ,2' -epoxypropyl)-5,6-dihydro-5-
Dinitrogen fixation 314, 340, 341 hydroxy-6-methylpyran-2-one 177
4,6-Dinitro-2-methoxyphenol 79 Eragrostis curvula 297
2,4-Dinitrophenol 79, 189 Erysiphe graminis 347
403
Erythrina lithosperma 170 Fertilization/fertilizers (continued)

Ethanol 3, 35, 110, 353 199,201,211,212,214,215,217,
5-Ethoxy-3-trichloromethyl-l,2,4-thia- 224,229-232,235,242-246,251-
diazole 226 263,269-277,279-281,283,284,
P-Ethylaminopropionohydroxamic acid 286-296,298,300,301,303-306,
64 309-313,315-318,340,347-351,
N-Ethyldichloromaleimide 66-68 353,359
Ethyl-N,N' -diphenylphosphorodiami- Fescue, meadow 125
date 106 -, tall 297, 298, 355
Ethylene 340 Festuca arundinacea 297
Ethylene-l,2-bisdithiocarbamates 53 Fiber digestibility 355
Ethylenediaminetetraacetate 209, 210 Fluorides 33, 34, 234, 251, 263, 293,
Ethylenediaminetetraacetic acid 313,316,323,330,340
disodium salt 178 4-Fluoro-N-(diaminophosphinyl)benz-
2-Ethyl-n-hexylphosphorodiamidate amide 193,275,278,336-338
348 Flurofamide 357
N-Ethylmaleimide 66, 67, 69 Fodders 353-355
Ethylphosphorodiamidate 109,348, Foliar burn 306
349 Formaldehyde 41, 46, 47, 179, 188
N-Ethylphosphoric triamide 199 Free energy of activation 99, 100
3-Ethylrhodanine-5-acetic acid 76 Fumigant 36
2-Ethynylpyridine 226, 288 Fungal biomass 333
Etridiazolc 226, 239 Fungi 329-333, 355
-, cellulose-decomposing 330
F -, glucophilic 330
Fatty acids 31, 32, 163 -, phytophathogenic 65, 66, 70
- -, volatile 354, 355 -, thermophilic/thermotolerant 330
Ferbam 55, 181 - tolerant to Cu and Cd 330
Ferric chloride 7, 9, 12, 187 Fungicides 53, 55, 56
- dimethyldithiocarbamate 55 Furan 176
- hydroxide 2, 9, 37 Furanoflavone 175
- ions 53 Fusaria 333
- nitrate 9, 17, 20, 288
- oxide 9 G
- sulfate 9, 12 p-Galactosidase 355
Ferrous chloride 13 Gallic acid 82, 85, 95, 96
- ions 37, 233 Geranio1311,312
- sulfate 2, 10, 12, 13, 187, 189,246 Geranium 315,318
- sulfide 18 p-Glucosidase 321, 323
Fertilization/fertilizers 1-3,9,10,17, y-L-Glutamyl 2-methoxy-p-nitroanilide
20,24,27-30,32,33,36-41,46,49, 104
50,52,56,64-66,70,72,92,95, y-L-Glutamyl m-nitroanilide 104
101,102,110,111,113,115,119, y-L-Glutamyl p-nitroanilide 104
123, 126, 129, 130, 133, 135, 139, y-L-Glutamyl nitroanilides 104, 105
140 142, 144, 145, 150-153, 157- Glycine max 225, 313
159,165,167,169,170,172,174- Glycolic acid, xanthate of 60
177,179,180,182-184,188,197- Glycols 144
404
Glycols (continued) Hexabromoethane 53, 265

-, derivatives of 144 2,2,4,4,6,6-Hexachlorocyclotriphos-
G1yco1uril301 phazatriene 156
Glycosylation 253,314,317 cis-l,2,4,5-trans-3,6-Hexachloro-
Goethite 18 cyclohexane 163
Gold chloride 7 2,2,4,4,6,6-Hexa(methylamino)cyclo-
Gossypium hirsutum 315 triphosphazatriene 156
Gramineous plants 45, 251-313 Hexamethylenetetramine 47, 180, 347
Grape 356 Hexamethylphosphoric triamide 349,
Grasses 27,125,151,297-313 350
Growing season 49,151,152,232, S-( n- Hexyl)diamidophosphorothiolate
253,254,263,264,268,271,273, 141, 142
279,289,293,298,305,309,316, N-(n-Hexyl)thiophosphoric triamide
351 143
Growth/vegetation period 10,34,281, Hordeum vulgare 288
293,316,323,336,350 Humic acids 165, 166
Guaiacol 79 - substances 1, 166, 170, 183, 184,
Guanidine 229,230 241,318,328,332,341
Guanylthiourea 52, 75, 195,229,236, Hybrid sorghum-sudan grass 297
243,293 Hydrazoic acid 229
Guanylurea 48, 229, 230 N-Hydrocarbylthiophosphoric
- sulfate 47 triamides 144
Guatemala grass 170 Hydrochloric acid 19,38,343
Gypsum 30, 174, 176 Hydrogen peroxide 73, 216
Hydrohumate 318
H Hydroquinone 3,82-86,88-101, 104,
N-Halamine compounds 103,238,239 105,136,137,160-162,164,177,
Halogenated alkanes 53, 180 180,181,189-195,201-205,212,
N-Halo-2-oxazolidinones 103 213,237,238,244-246,253,254,
Heavy metal compounds/salts 2,5-20, 266,268,269,275,278,280,281,
75,76, 179, 18~233,246,288, 286,288,291,292,300-304,308,
292,313,315,321,322,329,334, 312-314,316,317,325-327,329,
336, 339-342 334,336-340,344,359
- - chlorides 10 Hydroxamic acids 60-66, 180,265,314
- - hydroxides 2 - , derivatives of 356, 357
- - ions 233 4-Hydroxyaminobenzeneboronic acid
Helicobacter pylori 356, 357 46
Hematite 18 Hydroxybenzene 78
Herb extracts 357 3-Hydroxybenzeneboronic acid 46
Herbicides 71,197,225,247,317 4-Hydroxybenzoic acid 79
Heterocyclic sulfur compounds 71-78, 3-Hydroxy-2,5-dimethyl-p-benzo-
236,316,324,331 quinone 87, 88
Hevea brasiliensis 93 2-Hydroxy-3,6-diphenyl-p-benzo-
2,2,4,4,6,6-Hexaaminocyclotriphos- quinone 87
phazatriene 155-160, 220, 260, 261, Hydroxyethy1 cellulose 31
332,335,350,351
-, hydrolysis of 159
405
5-(2-Hydroxyethyl)-3-(4-hydroxy-2- 3-Isopropylrhodanine-5-acetic acid 76

methyl-5-pyrimidinylmethyl)-4-
methylthiazolium chloride 163 J
3-Hydroxy-2-isopropyl-5-methyl-p- Jackbean 25,36,37,41,49,65, 1l3,
benzoquinone 87 143,144,177,216,217,220
6-Hydroxy-2-isopropyl-5-methyl-p- Japanese mint 315,318
benzoquinone 87 Java citronella 231
Hydroxylamine 353
- reductase 322 K
p- Hydroxymercuribenzoate/-benzoic Kaolin 2, 163
acid 43, 44, 190, 344 Kaolinite 135
5-Hydroxy-p-naphthoquinone 84 Karanja 175,176,241
N-( 4-Hydroxyphenyl)glycine 81 Karanjin 175,176
Hydroxyurea 48, 49, 220, 357 Kerosene 173, 176, 232
Klebsiella 356
I Krylon 32
Illite l35
Imidodiphosphoric acid diphenyl ester L
111,112 Lactuca sativa 315
- - - -, aluminium salt of III, 112 Lansoprazole 357
- - - -, calcium salt of 111, 182 Lead 233,313,322
- - - -, diammonium salt of 111, 182 - acetate 5,6, 9, 11, 12, 15,30,33
- - - -, heavy metal salts of 111, 112 - chloride 7, 13, 14
lmidodiphosphoric tetraamide 349 - ions 9, l3, 16, 17
Inhibition constant 9, 45, 100, 326 - nitrate 7, 9, 11, 15-17
Inknut 169,170 - oxide 9
Inorganic boron compounds 30-33, - salts 76
188,280,292,298,347 - sulfate 13
- sulfur compounds 35-42, 189,235, - sulfide 35
252,263,295,298,334,347 Leaf tip necrosis/scorch 276,277,297,
Insecticides 163, 171 . 309-311,314,315
Invertase 321-328 Leguminous plants 45, 313-315
Iodine 219 Lemongrass 297, 311
Iodoacetamide 178 Leonardite 183
Iodoacetic acid 178 Lettuce 246, 315, 318
Ipomoea batatas 315 Light metal compounds/salts 2, 19-20
Iron 9,20, 38,41, 76, 179 Lignins 166,241,311
- dimethyldithiocarbamate 55 Lignite 92
- ore 9 Lignosulfonates 166, 168, 184,241,
- oxides 18 261,279,328,332,335,341
- salts Ill, 112 Lime ammonium nitrate 271, 272, 289,
Isobutylidene diurea 2 290,294,295,304,305,317
3-Isobutylrhodanine-5-acetic acid 76 - nitrogen 229
Isocyanic acid 49 Lindane 163
Isopropanol 130 Lipase 353
2-Isopropyl-5-methyI-p-benzoquinone Lithium sulfate 25
87,91 Livestock wastes 352, 353
406
Lolium multiflorum 297 Margosan-O 171

- - var. westerwoldicum 303 Medicago sativa 313
- perenne 150,297 Mentha arvensis 315
Lovegrass 297,301 2-Mercapto-5-amino-l ,3,4-thiadiazole
Lupinus luteus 313 73, 75
2-Mercaptobenzothiazole 77
M 2-Mercapto-5-benzylmercapto-l ,3,4-
Magnesium 19,24 thiadiazole 74
- carbonate 22, 26, 234, 298 2-Mercapto-5-ethylmercapto-l ,3,4-
- chloride 20, 23, 26, 38, 204, 234, thiadiazole 74
306,307,316,322,335 2-Mercapto-5-isopropylmercapto-
- hydroxide 26 I,3,5-thiadiazole 74
- oxide 9 2-Mercapto-l-methylimidazole 74, 75
- stearate 52 2-Mercapto-5-methylmercapto-l,3,5-
- sulfate 19,22,24,27,28,234 thiadiazole 73, 74
Mahua 176, 177,241 5-Mercapto-3-phenyl-l,3,4-thiadiazo-
Maize28,33,41, 117, 118, 124, 137, line-2-thione potassium salt 73-75
148,153,154,160,171,197,201, 5-Mercapto-5-n-propylmercapto-l,3,4-
205,210,216,227,232,244,251- thiadiazole 74
264,274,276-278,280,347,348, 2-Mercaptopyridine 101, 102
351,352,355 - N-oxide 101, 102
Maleic acid 178 2-Mercapto-3-pyridinol 101, 102
- hydrazide 69, 181 2-Mercaptopyrimidine 101, 102
- - ammonium salt 181 2-Mercapto-1,3,4-thiadiazoles 73, 74
- - diol form 69 Mercuric azide 229
- - dione form 69 - chloride 3, 5-8, 10, 12, 13,321,329,
- - sodium salt 181 334, 340-343
Maleimides 66-69 - ions 5, 7, 9, 13, 16
4-( I' -Male imido )butanehydroxamic - nitrate 11
acid 65 - sulfate 7, 12, 13
Mana grass· 170 Mercury 43, 233
Mandarin 187 - salts 76, Ill, 112
Maneb 53-56, 323, 324, 330, 331, 334, Metal sulfides 358
340-342 Methane emission 336
Manganese 8, 11,38,41, 179,349 - oxidation 336
- dioxide 37 Methanol 45, 80,110,195,215,216,
- ethylene-l,2-bisdithiocarbamate 54, 219,352
55,323,330,334,340 2-(p- Methox y)-benzenesulfonamido-2-
- salts 112 thiono-5,5-dimethyl-l,3,2-dioxa-
Manganous chloride 7, 10, 13, 189 phosphorinane 161
- ions 9, 11, 13,37,53 Methoxy-p-benzoquinone 87
- sulfate 11, 13, 187, 189,246 2-Methoxy-3,5-dimethyl-p-benzo-
Manure, farmyard 190,280,287,329 quinone 87
-, green 154, 175 2-Methoxy-3, 6-dimethyl-p-benzo-
-, pet 353 quinone 87
-, poultry 171, 352 3-Methoxy furano-2',3',7,8-flavone
Marcol 72 31, 48, 179 175
407
2-Methoxyphenol 79 Methylphosphorodiamidate 112, 141,

2-[ 4-(3-Methoxypropoxy)-3-methyl- 142,348
pyridin-2-yl]methylsulfinyl-l H- 3-Methylpyrazole 241
benzimidazole 356 3-Methylpyrazole-l-carboxamide 227,
2-Methoxy-trimethyl-p-benzoquinone 292
87 N-Methyl-2-pyrrolidone 244
S-Methyl-N-acetamidodithiocarbamate 3-Methylrhodanine-5-acetic acid 75-77
54 Methylthiocarbamate 53
4-(N-Methylamino )phenol 81 Methylurea 47, 48
~-Methylaminopropionohydroxamic Microbial activity 98, 169, 171, 188,
acid 64 204
2-Methylanthraquinone 91 - biomass 167,212,321,329,330,
2-(p-Methyl)-benzenesulfonamido-2- 332,339
thiono-5,5-dimethyl-l,3,2-dioxa- - counts 321,329,332
phosphorinane 161 - products 177
Methyl-p-benzoquinone 84, 87, 89 Microbiota 354, 355
2-Methyl-3-butyn-2-o1222 Microelements 349
3-Methylcatechol91 Micronutrients 32, 33, 279, 347
4-MethylcatechoI91, 99-101, 326 Microorganisms 1,2, 10, 15,22,26,
2-Methyl-6-chlorophenylp hosphoro- 62,79,118,162,163,165,170,
diamidate 109 191,331-334,337,354
2-Methyl-6-chlorophenylthiophos- -, ammonifying 329, 332
phorodiamidate 109 -, cellulolytic 329, 330, 332
S-Methyldiamidophosphorothiolate -, proteolytic 329, 332
141,142 Mildew disease 347
N-Methyldichloromaleimide 66-68 Mineralization, alanine 337, 338
2-Methyl-5-ethyl-p-benzoquinone 84 -, biuret 348
Methylethylphosphoroamidate 348 -, hexamethylenetetramine 347
2-Methyl-5-ethylpyridine 228 -, nitrogen 267, 336-338, 352
N-Methyl-N-( 4-hydroxyphenyl) Molybdenum 8,9, 349
phosphoric triamide 142, 143 Monoammonium phosphate 187
N-Methyl-N' -hydroxyurea 48,49 Monohydric phenols 78-82, 236, 324,
N-Methylmaleimide 300 331
N-Methyl-N-(4-methoxyphenyl) Monohydroxamic acids 60-65
phosphoric triamide 142 Montmorillonite 135
Methylnaphthol 80 Marganella 356
2-Methyl-p-naphthoquinone 91, 92, 99- Mucochloric acid 69,70, 178, 181
101,267,326 Mucoraceae 333
N-Methyl-N-( 4-nitrophenyl)phosphoric Muriate of potash 151, 152, 174
triamide 142, 199 Mustard 349,350
N-Methyl-N-nitrosoaniline 222
5-Methyl-2-oxo-4-imidazolidine- N
caproic acid 163 Nabam 53
Methylphenols 78 a-Naphthaldehyde 31, 180
4-Methylphenylphosphorodiamidate a-Naphthol 78, 79
110 a-Naphthoquinone 83, 85, 86, 90, 266,
314
408
p-Naphthoquinone 83, 84,91,92,99, Nitroanilides 104, 105

100,267,326 m-Nitroaniline 104, 222
w-(Naphthoxy )alkanohydroxam ic acids o-Nitroaniline 104,222
64,65,352 p-Nitroaniline 104,222
N-~-Naphthylchloromaleimide 68 Nitroanilines 104, 105
N-~-Naphthyldichloromaleimide 68 Nitrobacter 203
a-Naphthyl ester ofphosphorodiamidic 7-[N-(2' -Nitro-2' -bromovinyl)-N-
acid 108, 109 ethylamino]heptanehydroxamic
~-Naphthyl ester ofphosphorodiamidic acid 65
acid 108, 109 Nitrogen immobilization 129,211-213,
N-~-Naphthylmaleimide 68 270
Natrosol 250 31 - loss 284,285,287,292
Natural products 163-177,359 - recovery 150,212,213,250,267,
Necton 37 31 282-286,289,307,309,310
Neem cake 171-177,232,233,241, - uptake by plants 152, 175, 251-253,
262,279,280,287,311,312,318, 255,256,258,261,262,264,265,
351 267-275,279-289,291-295,297-
- extracts 171, 262, 287 304,306,308,309,311-313,318,
- kernel extract 190, 191,329 348, 350-352
- oil 171-175, 241, 311, 312, 318 - use efficiency 252, 253, 258, 273,
Nematicides 71,316 281,290,298,299,303,305,312,
N-HIB Ca 316 316
Nickel 15, 233, 315, 336 Nitrogenase 321
- ions 9, 16,53 Nitrohumic acid 166, 167
Nickelous chloride 7,10, 12-14 Nitromagnesia 300
- nitrate II Nitrophenol 82, 324, 331
- salts 76, Ill, 112 2-Nitrophenol 79, 236
- sulfate II 4-Nitrophenol79
Nicotiana tabacum 170, 315 Nitophenols 236
Nicotine 178 N-( 4-Nitrophenyl)phosphoric triamide
Nitrapyrin 221-227, 229, 230, 237-239, 142,193,278
268,289,291,292,312,313,318, 4-Nitrophenylphosphorodiamidate 110
336 N-Nitrosodimethylamine 222
Nitrate reductase 323, 326, 329 Nitrous oxide 2, 3, 5, 18,29,43, 120,
Nitric acid 2, 166 233,244,245,337,339,340
Nitrification 4,36,37,39,41,59, 75, Nodulation 225, 313, 314
91,92,103,104, 1I5, 121, 127,
138,143-145,149,167-170,172,
174-176,182,184,221,223-231, o
233-246,249,250,262,279,280, Oak 18
287,288,291,292,312,318,336, Oats 19, 23, 49, 117, 130, 224, 272,
337,340,347,348,351,352,359 275,292-294,348-350
Nitrilotriacetic acid trisodium salt 178 Octaaminocyclotetraphosphazatetraene
Nitrite 228, 229 159,350
- accumulation 202, 203, 235, 240 Octadecylamine 30, 31,179,188,298
- reductase 321,322 N-(n-Octyl)phosphoric triamide 199
m-Nitroacetanilide 222 Odor emission 352, 353
409
Oil, cashew nutshell 176 p

-, essential 311, 312 Paraformaldehyde 47,69
-, linseed 311 Pea 313,314
-, mineral 111, 114,289,293,294, Peat 165,166,187,318
304 Pelargonium graveolens 315
-, naphthenic 31 Penicillia 333
-, olive 31 1,2,4,5,8-Pentahydroxyanthraquinone
-, paraffinJparaffmic 31, 13 7, 197, 91
211,257 Peptides 253, 314, 317, 357
-, rapeseed 316, 317 Permissible ecological value 281
-, silicone 31 - healthy value 281
-, vegetable 31 Pesticides 1,4,321,340,341
-, white 31 Phaseolus vulgaris 313
-, see also Neem oil Phenanthrenequinone 83, 95, 96
Oils 2 Phenol 2, 3, 78-80, 110, 132-134, 136,
Oil-seed rape 94, 177,231,315-317 192, 195
Oleates 31 Phenoxyacetohydroxamic acid 64
Oleyl ether of polyethylene glycol 32 2-Phenoxy.-2,4,4,6,6-pentaaminocyclo-
Omeprazole 357 triphosphazatriene 155-158, 278
Onion 34,49,167,315,316 2-Phenoxy-2,4,4,6,6-pentachlorocyclo-
Orchardgrass 28, 201, 297, 302-304, triphosphazatriene 156
308 Phenylacetohydroxamic acid 60, 61
Orchex 792 31 Phenylacetylene 226, 250, 288
Organic mercury compounds 43-45, Phenyl-p-benzoquinone 84, 87
178,190,235,253,264,278,280, N-Phenyl-N' -t-butylthiourea 51
288,296,313,323,339 N-Phenylchloromaleimide 68
Organo boron acid compounds 45, 46 N-Phenyl-f}-chloropropionohydrox-
Organosilicone 31 amic acid 64
Oryza sativa 280 N-Phenyl-a-cyclohexylaminoaceto-
Osmocote 313 hydroxamic acid 64
Oxalic acid 2, 178 Phenyl-N,N' -dibenzylphosphorodi-
Oxidation, abiotically catalyzed 75 amidate 106
-, bacterial 1 N-Phenyl-N,N' -dibenzylthiourea 51,
-, biological 358 52
-, chemical 358 Phenyl-N,N' -di-n-butylphosphorodi-
-, sulfur 341 amidate 106
Oxidized diaminophosphinyl sulfur N-Phenyldichloromaleimide 68
derivatives 140, 141 Phenyl-N,N' -diisopropylphosphoro-
Oximated O-diaminophosphinyl diamidate 106
derivatives 140 Phenyl-N,N' -dimethylphosphoro-
2-(Oximinophenylacetonitrile)-2-oxo- diamidate 106
5,5-dimethyl-l,3,2-dioxaphosphori- Phenyl dimethylpolysiloxane 31, 32
nane 160 2-Phenyl-3,5-dimethylpyridine 228
Oxyhumate 318 Phenyl-N,N' -di-n-propylphosphoro-
Oxythiamine chloride 163-165,261 diamidate 106
p-Phenylenediamine 178
410
N-Phenyl-N' -ethyl-N' -benzyl thiourea Phosphoric triamides 142-144, 154,

51 199-201,213,239,275
N-Phenyl-N' -ethyl-N' -cyc1ohexyl- - -, N-acy1143
thiourea 51 - -, N-aliphatic 142
N-Phenyl-N' -isopropylthiourea 51 - -, N-aryl 142, 143
N-Phenylmaleimide 68 Phosphoroamidates 105, 109,336,348,
Phenylmercuric acetate 7, 43-45, 100, 354
190-193,235,264-266,275,278, Phosphoroamides 105-108, 113, 196,
280,313,314,323,329,343,344 200,239,256,257,262,274,336-
- borate 43, 45, 344 338,340,348,354
Phenylphosphoric acid 130 Phosphoroamidic acid 105
N-Phenylphosphoric triamide 142, 193, Phosphorodiamidates 105, 106, 109,
275, 278, 336-338 112,113,256,348,354
Phenylphosphoroamidate 132 Phosphorodiamides 239, 254, 269, 278,
-, hydrolysis of 132 281,289,293,296,303,314,317,
Phenylphosphorodiamidate 10 I-I 03, 328,331,335
105-107,109-137,144,145,155- Phosphorodiamidic acid 42, 105, 107-
158,181-185,189,192-197,199- 109,130,133,136-138,193,278
213,216,217,238,241,247-249, - - alkyl esters 107-109
254-257,269-279,281-286,288- - - 4-(2-amine-2-oxide-I,3,2-benzodi-
291,293-297,303-308,315,317, oxaphosphole) ester 138
328,331,332,335-338,340,344, - - 5-(2-amine-2-oxide-l,3,2-benzodi-
345,349,352-354,359 oxaphosphole) ester 138
-, decomposition of 132-138 - - diphenyl esters 108, 109
-, hydrolysis of 130, 132-134,200, - - esters 138
249 - - (l-methylethylidene)-di-4, I-phe-
-, stability of 130, 134, 137 nylene ester 139
-, 4-substituted derivatives of 357 - - naphthyl esters 108, 109
N-Phenyl-a-pyrrolidinoacetohydrox- - - 1,3-phenylene ester 139
amic acid 64 - - 1,4-phenylene ester 139, 140
Phenylthiophosphorodiamidate 109 - - phenyl esters 105, 107-109
N-Phenylthiourea 51 - - - -, N-alkyl derivatives of 108
Phenylurea 47-49 Phosphorotriamides 216, 336, 356
Phleum pratense 121 Phosphorus 143
Phloroglucinol 82, 85 - oxychloride 11 0
Phosphatase 321-328,330,334,339 - pentasulfide 35, 191
-, acid 322, 328 Phosphorylated 2-oximinophenyl-
-, alkaline 321, 332, 328 acetonitrile compounds 160, 161
-, neutral 322 Phosphoryl triamide 42, 105, 144, 145,
a-Phosphate 134, 135, 159,209 154-158,178,193-195,213,216,
Phosphogypsum 29, 189, 198, 199 217,220,249,275,278,332,335,
Phospholipase 321 337,338,348,350
Phosphonitrilic hexamide 42, 155,350 - -, linear thermal polymers of 145
Phosphoric acid 2,35, 180, 188 Photodegradation 92
Phosphoric triamide compounds 193, Photosensitive compounds 92
198,256,274,285,291,294-296, Photosynthesis 248
306,315,318,328,332,335,349 Phthalimide 178
411
Phytotoxic/phytotoxicity 17,49,259, Potassium (continued)

261,266,267,277,287,292,309, - fluoride 20, 33
313, 315, 348-350 - hydrogenphosphate 19,234,269
Picric acid 79 - hydroxide 134
Pine 9, 32, 224, 311, 353, 356 - isopropoxyethyl xanthate 59
Pinus cemhra 9 - isopropyl xanthate 59
Pisum sativum 313 - 2-methoxyethyl xanthate 59
Plant extracts 169 - methyl xanthate 59
- growth 19, 56,175,251,253,259, - nitrate 18, 130, 131, 274
263,264,267,275-277,279,287, - oxide 29, 38
293,300,301,308,309,313-316, - permanganate 42
351 - phosphoroamidate 194
- materials 170, 177, 187, 192,241, - salts 74
311,318,329,351 - sulfate 20, 22, 153, 167,234,246,
Plastics 2 248,276,281,293,309,311
Poa pratensis 146,204,297, Potato 315, 318
Poisoning 353, 354 Primary amines 30, 32
Pollution, environmental 1,43,45, Proanthocyanidins 356
133,281,358 Propanol 353
Polyhydric phenols 82-101,178,180, Propionate 355
191,237,238,253,266,278,280, Propionic acid 353
289,296,300,314,316,324,325, Propionohydroxamic acid 60
334,339 N-n-Propyldichloromaleimide 66-68
Polyphenol oxidase 326, 328 2-n-Propyl-3,5-diethylpyridine 228
Po1yphenols 99, 169, 170,241 n-Propylphosphorodiamidate 112, 113,
Polyphosphates 130 141
Polyphosphorodiamides 139 3-n-Propylrhodanine-5-acetic acid 76
Pongamia glabra 175 Protease 321, 325, 326,
Poplar 45 -, NIl-benzoyl-L-argininamide-hydro-
-, white 166 lyzing 322, 323
Populus alba 166 -, casein-hydrolyzing 322
Potassium acetate 134 Proteins 300, 310, 312, 316-318
- allyl xanthate 59 Proteus mirabilis 356
- azide 228, 229 - morganii 356
- carbonate 22, 134,234 Protocatechuic acid 82, 85
- chloride 8, 10, 19-22,26, 28-30, 38, Proton pump inhibitors 357
II~ 121, 122, 14~ 152, 182, 18~ Providencia 356
188,198,199,204,212,234,250, Pumpkin 315-317
251,253,261,269,273,274,291, Pyrane 177
297-299,306,307,311,315-317 Pyridine 102
- dichromate 16, \9,292,330 - derivatives 227, 228
- dihydrogenphosphate 20, 309 - N-oxide 10 1, 102
- 2-dimethylaminomethyl xanthate 59 - 3-sulfonic acid 163-165, 261
- ethylene glycol xanthate 59 Pyrimidine 102
- ethyl xanthate 58-60, 192,236,242, Pyrite 18, 19, 198, 199,233,234,241,
300318 315
- ferricyanide 42 Pyrogallol 82, 83, 85, 89, 95
412
Q Secale cereale 294

Quinhydrone 83, 88,89,95,96, 177, Secondary amines 30, 32
180, 192,269,280,317 Seed germination 1, 10, 16,251,253,
Quinones 82-10 1, 178, 180, 191, 192, 255,264-268,272,275-278,288,
237,238,253,266,278,280,289, 290,295,297, 300, 313-315, 317
296,300,314,316,324,325,334, Seedling emergence 278, 293, 294
339 - growth 251,253,275,277,279,310
Selenious acid 34
R N-Serve 222, 225
Rabeprazole 357 Sesbania aculeata 175
Radish 49,315,316,348 Shell-lac 173-175,232,233
Rapeseed 347 Silicon dioxide 2
Raphanus sativus 315 Silver chloride 13
Rebamipide 356 - ions 5, 7, 9, II, 13
Red clover 56,313,314 - nitrate 7, 8, 12, 344
Reductases 18 - sulfate 7, 8, 12, 13,343,344
Residues, crop/plant 23,39,40, 124, Simazine 247
125,137,149,150,169,197,198, Sodium acetate 21, 247, 248
205,241,244,252,256,257,259, - arsenate 34
263,322,335 - arsenite 34
Resins 31,176,311 - azide 228
Resorcinol 82, 85, 89, 95, 96 - benzene alkyl sulfonate 31
Respiration 321,325,327,329,334, - bisulfite 35
335,339 - borate 21, 246
Rfvalue 100, 101,220 - n-butyldithiocarbamate 55
Rhizosphere 56 - carbonate 21, 22, 234
Rhodanine-5-acetic acid and deri- - chloride 21-26, 30, 234, 322, 323,
vatives 75-77 334,335
Rice 4, 43, 94, 119, 121-123, 125, 126, - diamidothiophosphate 348
135,148,149,152,172,175,177, - diethyldithiocarbamate 55
182, 187, 190, 195, 196,205-207, - dihydrogenphosphate 276
210,211,217,232,248-250,280- - di-n-propyldithiocarbamate 55
288,323,334,336,340 - fluoride 33, 34,179,234,251,263,
Rubber plantation 93 293,313,316,323,330,340
Ruminants 353-355 - humate 167,332,341
Rye 16,271,275,276,294,295,289, - hydrogencarbonate 24, 26, 30
290 - hydroxide 19, 166
Ryegrass 94, 226, 231, 299-303 - ions 53
-, Italian 297, 300, 301, 312, 349-351 - methoxymethyl xanthate 59
-, perennial 27, 150, 152,297,302- - 2-nitrilo-2-propyl xanthate 59
304, 308-311 - nitrite 238
s - pyrophosphate 166
Saccharum officina rum 315 - selenite 34
Saint Augustine grass 297, 298 - silicate 21
Salicylohydroxamic acid 60, 63 - sulfate 21-26,30,234,253,322,
Saponins 171,262 323,334
Sarsaponin 171,262,352,355 - sulfide 36
413
Sodium (continued) Soils, clay/clayey (continued)

- sulfite 35, 38, 253 -,198,205,211,225,230,246,249,
- tetraborate 166 261,264,296,298,313,314,318,
- tetrathionate 41, 189,253,264 322,325,330,350,351
- thiophosphorodiamidate 194 -, -loam 6,7,10-13, 15, 16,20,23,24,
- thiosulfate 37, 38, 41, 42, 189,253, 27,29,30,33,34,36,37,41,43,
264,352 44,50,55,56,58,59,61,63,79,
- trithiocarbonate 36, 58, 59, 223, 242 85-87,92,94, 128, 148, 159, 166,
- tungstate 34 170,221,224,226,231,233,235,
Soils 237,251,252,277,278,285,287,
-, acid/acidic 8, 11,45,60,61,64,98- 291,309,310,321,323,324,328,
101,121,162,165,167,170,192, 329,331,334,336,337,340,342,
197,218,227,234.238,349,350 347
-, alfisol182, 195, 196 -, coarse textured 273
-, alkali/alkaline 45,51,93,99-101, -, fine loamy 146,204,210,306
122,123,154,165,167,172,173, -, - sandy 5, 19,30,188
192,197,202,204,234,264,283 -, - -loam 5, 23, 24, 82, 194, 198,
-, alluvial 9, II, 15,22,35,45,47,51, 241,291,298,328,336
69, 77, 78, 81,92,95, 100, 104, -, fine-textured sandy 298
160-162,175,178,181,188,192, -, forest 8, 11,24, 78, 192,227,232,
223,225,234,237,325-327,334, 233,301
347,350,351 -, grey 8, 9, 192,208,212,224,227,
-, base-saturated 125 232,264,313,314
-, black 16, 64, 79, 80, 91, 92, 192, -, grey-brown 41, 115
225,237,267.278,317,325,333 -, heath 227,349
-, brown 11, 16, 18,21,24,52, 78-80, -, heavy-textured 10, 15,44,45,56,
91,92,96,97, 129, 193, 195, 196, 63,67, 72, 86, 88, 92, 93, 95, 104,
232,233,248,253,264,268,273, 113, 114, 128, 160, 181, 195,202,
279,281,291,293,301,313,325, 239,303,304,318
326 -, laterite 173,237
-, calcareous 1,9,21,22,25-27,29, -, light-textured 1, 10, 15,44,45,63,
33,41,51,85,97,166,176,188, 72, 86, 88, 92, 95, 104, 106, 113,
198,223,227,234,292,298,322, 114,128,160,178,181,195,202,
349 237,239,273,303,311,318
-, chemozemlchemozernic 8, 9, 11, -, loarnlloamy 5,8, 16, 18,36,37,40,
15, 16,21,22,24,35,45,47,58, 43,50,56,63,79,82,85,93,94,
69, 77, 78, 81,92-95,97, 100, 121,127-129,131,159,174,202,
104,160-162,165,178,181,187, 205,212,226,231,238,243,245,
192,198,224.227,236,238,268, 273-275,277,290,302,303,318,
272,278,292,301,312,322, 325,338,349
325-327,334,335,350 -, loamy clay 223
-, chestnut 15,25,34. 167, 192,272, -, - sand 6, 11-13, 16, 19,31,38-41,
328,340 55, 56, 60, 61, 63, 72, 80, 85, 98,
-, clay/clayey 8, 12, 14, 17,22,29,39, 99, 114, 115, 129, 148, 162, 163,
41,45,82,83,93,95,99, 119, 188,195,206,208,224,226,227,
122,126-128,137,149,150,153, 230,251,269,271,273,291-293,
166,168,169,174,182,184,188, 298-300,302-304,317,324,352
414
Soils (continued) Soils (continued)

-, loess 18,27,64, 192, 195,225,232, -, sierozem 93, 94, 302-304
233,237,248,293,317,349 -, silt 223
-, meadow II, 15,21,97,279,326, -, silty clay 20,38-40,51, 177,205,
328,333,347 206,211,243,246,261,277,287
-, medium-textured 106, 113, 178 -, - -loam 6,20,21,27-29,34-36,38,
-, muck 19 43, 50, 58, 63, 79, 80, 83, 85, 94,
-,neutral 218, 234,349,350 95,99, 121, 122, 131, 137, 166,
-, organic 10,56,58,93,224,321, 168, 172, 174, 175, 191, 192, 194,
323-325, 340-342 195,200,205,210,217,230-232,
-, paddy 5,6, 14, 15,41,42,80,95, 257,296
178,232 -, silt/silty loam 8, 14, 16,23-25,28,
-, pararendzina 224 41,43,49,56,60,64,74,75,79,
-, peat 9, 14, 33, 190 80,89,97, 102, 103, 112, 115,
-, podzol/podzolic/podzolized 8, 41, 117, 123, 124, 128, 130, 135, 137,
115,166,170,196,208,209,211, 139-141, 143, 145, 148-150, 152,
212,279,301-304,350,351 153, 156, 158, 173, 189, 194, 196,
-, pseudogleyic 301 199,201,205,206,210,216,218,
-, red 173,187,225,231,237,311 225,239,240,247,248,251,252,
-, red-brown 178, 208, 209 254-259,263,273-275,281,286,
-, reddish-brown 166 288, 296, 325, 326, J48
-, red-yellow 8, 170 -, soddy-podzol/podzolic 8-11, 15, 16,
-, rendzina 348, 332, 335 21,34,49,56,93,97,227,291,
-, saline/salinized 25,45,94,99-101, 292,318,323,347
174,177 -, solonetz 279,328,333,341
-, sand/sandy 14, 15, 17, 19,27,28, -, solonetz-solonchak 21
41,44,52,63,64,67,86,88,90, -, solonized 196,264,279,313
93,94, 114, 115, 125, 164, 167, -, vertisol182, 195, 196,216
173,183,184.192,195, i23, 231, -, volcanic ash 6, 43
234,266,271,290,293,294,296, -, yellow 208,209,211,212
302-304,318,322,329,330,333, N-So141
342,349 Solanum tuberosum 315
-, sandy clay 14, 80, 153,246 Sorghum 275-278, 295-297
-, - -loam 11,18,23,29,44,49,172, Sorghum bicolor 295-297
178,183,184,189,190,195,205, - sudanese 297
217,221,225,234,235,237,241, Soybean 64,66,147,225,260,313-
326,329,336 315,350
-, sandy loam 6, 8,10,11,13-17,19, Spectroscopy, infrared 216
20,22-24,26, 30, 32, 33, 44-47, -, mass 216
50,55,56,58,61,63,77,85,93- -, 31p nuclear magnetic resonance 216,
95, 112, 114, 115, 122, 125, 142, 217
154,163,167,172,173,176,199, Spruce 8,121,135,331,335
205,210,217,221-224,230-234, Stannium salts 76
236,244-246,253,262,264,279, Stannous chloride 12
280,283,287,294,297,311,321, - ions 9
323-325,329,330,334,336,340- Staphylococcus aureus 356
342
415
Starch 22 2,2,6,6-Tetrachlorocyc1ohexylphos-
Stearamine 31 phorodiamidate 112, 113
Stearic acid 2, 83 1,1,2,2-Tetrachloro-l ,2-dibromoethane
Stenotaphrum secundatum 297 53,54,256
Strontium chloride 24 2,2,4,4-Tetrachloro-6,6-di(dimethyl-
Struvite 356 amino )cyc1otriphosphazatriene 156
Subtropical plants 231 Tetrachlorohydroquinone 89
Succinic acid 2 Tetrachloroquinhydrone 89
Succinomonohydroxamic acid 66 2,2,4,4-Tetra(dimethylamino)-6,6-di-
Sugarbeet 198, 199,315,317,318 aminocyc1otriphosphazatriene 156
Sugarcane 30,315,316,318 Tetrafluoro-p-benzoquinone 87
Sulfanilamide 162, 163, 180 Tetrahydro-l,3,5-thiadiazine-2-thiones
Sulfanilic acid, amide of 162 71, 72, 236
Sulfates 7, 9, 13, 26, 37, 38, 358 1,2,5,8-Tetrahydroxyanthraquinone 91
Sulfathiazole 222, 225, 237, 312, 313 Tetrahydroxy-p-benzoquinone 87, 88
Sulfide 37 Tetramethoxy-p-benzoquinone 87
- minerals 358 Tetramethyl-p-benzoquinone 87, 89
Sulfite 37 Tetramethylthiuram disulfide 56,57,
Sulfur 2, 143, 173, 176, 252, 263, 318 88,180,236,314,324,331,341
- Cote 313 Tetranortriterpenoid isomers 171
Sulfuric acid 18, 72 Tetraphosphorodiamide 139
Supergranules 129, 173-175,232,233, Tetrathionate 41
296 - anion 37
SuperN 258 Thermodynamic parameters/values 99,
Superphosphate 10,29,32,49, 121, 100
122,148,151-153,167,174,182, 1,3,4-Thiadiazoline-2-thiones 72, 73,
187,234,246,250,264,273,274, 358
283,291,311,347,348,350 -, thiol form of 72, 73
Super Urea 258 -, thione form of 72, 73
Sweet potato 315, 317 Thioacetamide 191
Synergism/synergistic 59, 84, Ill, 163, Thiobacillus ferrooxidans 358
180-182,210,242,244 - thioparus 358
2-Thiocarboxamido-5-aminothiazole
T 77
Tailings 358 2-Thiocarboxamido-5-benzamido-
Tamarind 177 thiazole 77
Tamarindus indica 177 2-Thiocarboxamidothiazoles 77
Tannic acid 353 Thione 72, 128, 195
Tannins 166, 169 2-Thiono-5,6-dimethyl-l ,3,2-dioxa-
Tartaric acid 178 phosphorinane compounds 161,
Tea 169, 170, 177,241,353 162
Temperature coefficient 99 Thiophosphoric triamide compounds
Terbutryn 182, 183,249,250,288 143,198,213,256,274,285,291,
Terminalia chebula 169 294-296,306,315,318,328,332,
Terradiazole 226 335
Tetrachloro-o-benzoquinone 89 Thiophosphoric triamides 142, 144
Tetrachloro-p-benzoquinone 87, 95 - -, N-acy1143
416
Thiophosphoric triamides (continued) Toxic/Toxicity 110, 135, 275, 289,

- -, N-aliphatic 142 317,348,349,353
- -, N,N-aliphatic 142, 143 Trace element 32
- -, N-alkyl 144 Triazine 312
- -, aryl 143 Trichloroacetate 354
Thiophosphorodiamidates 105, 112, Trichloro-p-benzoquinone 87
113,354 Trichloroethylphosphorodiamidate
Thiophosphorodiamidic acid 42, 105, 112, 137, 196, 197,201,239,254-
138 257,275,308,336
- - esters 138 S-Trichloromethylthiodithiocarbamate
- - phenyl esters 108, 109 54
Thiophosphorotriamides 278, 336 Triethylphosphate 349
Thiophosphoryl triamide 42, 105, 144, 2,2,2-Trifluoroethylphosphorodi-
145,155,178,194,195,199-201, amidate 112
213-217,220,240,241,249,278, 3-Trifluoromethyl-N-(diaminophos-
348 phinyl)benzamide 193
- -, hydrolysis of214, 215 N-(3-Trifluoromethylphenyl)phos-
- -, linear thermal polymers of 145 phoric triamide 193, 278
- -, pyrolysis products of 349 Trifolium pratense 313
Thiopyridine-N -oxides 101, 102 Trimethylamine 353
Thiopyridincs 10 I, 102 Trimethyl-p-benzoquinone 87
Thiopyrimidines 10 I, 102 2,4,6-Trimethylpyridine 228
Thiourea 47-52,188-192,220,235, 2,4,6-Trinitrophenol 79
236,243,246,287,289,293,299, 2,4,6-Triphenoxy-2,4,6-triaminocyclo-
312,316,323,330,344,347,348, triphosphazatriene 155-158
353 2,4,6-Triphenoxy-2,4,6-trichlorocyclo-
- derivatives 51. 52 triphosphazatriene 156
Thiram 56-58,88, 181,227,236,324, Triphosphorodiamides 139
331,341,342 Tripsacum laxum 170
Thiuram disulfides 56,57,236,314, Triterpenoides 171
324,331,341,342 Trithiophosphorodiamides 139
Thiuram sulfides 56, 57 Triticum aestivum 263
Thomas potash 300 - durum 273
Thymoquinonc 87 Triurea phosphate 350
Till/tillage, conventional 196, 197, 210,
211,232,257,297,328,337
-, no- 196-198,210,211,251-253, U
257-260,262,263,295,297 Urea 1-9, 11-75, 77-86, 88-137,139,
-, ridge- 252, 259 140,142-160,162-170,172-221,
-, zero 256,328,332,337 223-320,325,326,328,329,331-
Timothy 121, 125,254,255 333,335-337,339,340,343-357,
Titanium chloride 15 359
Tobacco 53, 72, 170, 177,315,316 - adducts 2
p-Tolualdehyde 31,180 - carboxylase 209
Toluene 32, 35 - derivatives 47-52, 191,235,289,
Tomatoes 34, 167 293,299,316,323,330,347
417
Urea (continued) W
- hydrolysis 1-3,5,6,8,9, 12 13, 16, Waste rocks 358
20-26,29,35-45,47,50,52,56, Waxes 2, 5, 6,30, 33,46,47, 182
58-61,63,64,66,67,69, 71, 72, Wheat 54, 98, 117, 129, 149, 150, 154,
74,77,79,81,83,84,86,88,90- 198,199,230,243,245,246,253,
95,97-100,105,109,115-119, 257,259,262-280,291,295,310,
121,122,124-127,129-131,134, 313,328,332,347,348
135,137,146-150,152-154,156,
160,163-165,167-170,172-178, X
181, 182, 184, 188-192, 195-212, Xanthates 58-60, 236, 250
216-219,221,223,225-236,238, -, branched-chain 59
244,245,247-249,252,257,259, -, straight-chain 59
260,264,276,277,279,282,285, Xylanase 321
315,326,331,347,352-354,356
- phosphate 126,284 Y
Urea-ammonium nitrate 17, 36-41, Yellow lupine 313
115, 145, 150, 153, 154, 158, 183, Yields, crop/plant 17, 19,23, 144, 150,
184,198,201,214,243,244,252- 152,153,246,248,251-275,277,
260,263,265,273-275,296,298, 279-309,311-318,347-351
299,309,352 Yucca schidigera 171,262,355
Urea-calcium nitrate 298
Urea-formaldehyde 41 Z
Ureaforms 2 Zea mays 33,117,124,137,148,171,
Ureaplasma urealyticum 356, 357 251
Urease 1-26,29,32-39,41-49.51-83. Zinc 8, 9, 11. 13, 15-17, 179, 188,233.
85-93,95-107,110,112-114,117, 313,322,329,330,336,349
118.121,127-130,134-148,152- - acetate 8
167,169-172,174,177,179-183, - chloride 7, 10, 12-14
185,187.189-201.205,208,210, - d~methyldithiocarbamate 55
211.213,215-243,245-251,258, - ethylene-l ,2-bisdithiocarbamate 54,
261-267,275-277,280,281.287, 55
288,291-293,297,300,310-313, - ions 53, 233
315,316,318,321-331,333-343, - oxide 16
347,351-359 - salts 76. 112
Usnic acid 3 - stearate 52
- sulfate 10, 11, 13-17, 187, 246, 264,
V 274,283,313
Vanadium 15,336 Zineb 53-56, 265
Vanadium (II) ions 9 Ziram 55
Vanillic acid 178
Vegetable crops 347
Veratro1178
Verdure density 298
Vetch 23,224
Viciafaba 313

S. Kiss, M. Simihăian (Auth.) - Improving Efficiency of Urea Fertilizers by Inhibition of Soil Urease Activity-Springer Netherlands (2002) PDF

Caricato da

Informazioni sul documento

Titolo originale

Copyright

Formati disponibili

Condividi questo documento

Condividi o incorpora il documento

Opzioni di condivisione

Hai trovato utile questo documento?

Questo contenuto è inappropriato?

Copyright:

Formati disponibili

S. Kiss, M. Simihăian (Auth.) - Improving Efficiency of Urea Fertilizers by Inhibition of Soil Urease Activity-Springer Netherlands (2002) PDF

Caricato da

Copyright:

Formati disponibili

IMPROVING EFFICIENCY OF UREA FERTILIZERS

BY INHIBITION OF SOIL UREASE ACTIVITY

SPRINGER-SCIENCE+BUSINESS MEDIA, B.V.

ISBN 978-90-481-5966-6 ISBN 978-94-017-1843-1 (eBook)

Printed on acid-free paper

AII Rights Reserved

Chapter 2. Organic Compounds Tested for Evaluation of Their

2.14.3. Rhodanine-5-acetic Acid and Its Derivatives 75

2.31.3.1. Residues and Extracts Containing Polyphenols 169

Chapter 3. Combined Use of Inhibitors of Soil Urease Activity 179

Chapter 4. Comparative Studies on the Efficiency of Different

4.9. COMPARISON OF PHOSPHORIC TRIAMIDE (PTA) AND

Chapter 5. Compounds Tested for Evaluation of Their Inhibiting

5.2.2.3. Thiuram Disulfides 236

Chapter 6. Soil Urease Inhibitors Used in Combination with

Chapter 7. Effect of Soil Urease Inhibitors on Germination,

7.2.10. Effect of Lignosulfonates 279

7.8.4. Effect of Urea Derivatives 299

Chapter 8. Effect of Urease Inhibitors on Other Enzyme Activities,

8.1.3. Effect of Fluorides 323

8.2.4.2. Effect of Phosphoroamides 336

Chapter 9. Use of Urease Inhibitors in the Analysis of Urea

Chapter 10. Urease Inhibitors Used with Another Purpose

Chapter 1. Inorganic Compounds Tested for Evaluation of Their Inhibiting Effect

1.1. HEAVY METAL COMPOUNDS

urea solution containing 10 mg of urea-N (1,000 ppm N on soil basis), and 5 m1 of

TABLE I. Effect of heavy metal compounds on soil urease activity"

Inhibition of urease activity (%)

Mercuric chloride Hg2+ 42 38

Gold chloride (HAue!.,) Au·l + 18 20

Zinc chloride Z,n 2~ () ()

"From Bremner and Douglas (1971). by permission of Pergamon Press PLC.

'Ouration of incubation is not specilied in the paper.

TABLE 2. Inhibition of soil urease activity by heavy metal salts"

TABLE 3. Effect of inorganic salts on urease activity in a periodically water-logged

In the laboratory experiment described by Todorov et al. (1987), surface samples of

1.2. LIGHT METAL COMPOUNDS

1.3. SALTS OF ALKALI METALSANDALKALINEEARTHMETALS

o or 243.5 mg of urea or 400 mg of Cardonitell 00 g soil, were incubated at 37°C for 2

TABLE 5. Effect of sodium chloride on activity of soil urease and of purified

determined. Table 5 shows that at concentrations of 0.1-4 M, NaCl inhibited urease

Addition of KCl and P fertilizers to urea solution is recommended for inhibiting

1.4. BORON COMPOUNDS

formula H2N-{CH2h-NH-R (where R is a C, - C20, preferably a C IX fatty acid group) may

polysiloxane), 2% Brij 92 (oleyl ether of polyethylene glycol) or 1% Krylon (a 6% acrylic

1.6. ARSENIC COMPOUNDS

1.7. SULFUR COMPOUNDS

"The reagent is prepared as follows: I g ofp-dimethylaminobenzaldehyde is dissolved in 30 mI of absolute ethanol,

Sodium trithiocarbonate (Na2CS3) is known as a nitrification inhibitor and fumigant. In

2Fe(OHh + 2S 20/- + 6H+ -----:>~ 2Fe 2+ + S40t + 6H zO

1.8. OTHER INORGANIC COMPOUNDS

Chapter 2. Organic Compounds Tested for Evaluation of Their Inhibiting Effect on

2.1. ORGANIC MERCURY COMPOUNDS

p-Chloromercuribenzoic acid Phenylmercuric acetate

HO-H9-Q-COOH Q-Hg- B02

2.2. ORGANO BORON ACID COMPOUNDS

4-Aminobenzeneboronic acid hydrochloride (Ill) 3-Hydroxybenzeneboronic acid (IV)

The most preferred rate of addition of these inhibitors to urea-based fertilizer

Hexamethylenetetramine (HMTA; Figure 4) as an inhibitor of soil urease activity was

Figure 4. Structure of hexamethylenetetramine.

2.5. UREA DERIVATIVES

Figure 5. Urea derivatives tested as inhibitors of soil urease activity.

In an experiment described by Sor et al. (1971), volatilization of NH3 from pellets