EFFECT OF SUGAR INDUSTRIES EFFLUENTS ON CHANGES OF GROWTH AND CHLOROPHYLL CONTENTS OF MUNG PLANTS

PROJECT REPORT SUBMITTED

TO

NEW ARTS COMMERCE AND SCIENCE COLLEGE, AHMEDNAGAR, UNIVERSITY OF PUNE IN PARTIAL FULLMENT OF REQIRMENT OF THE DEGREE
OF

MASTEROF SCIENCE
IN

ENVIRONMENTAL SCIENCE
BY

HULAGE ASHISH DATTATRAYA UNDER THE GUIDANCE OF DR. S.D KULKARNI POST GRADUATE DEPARTMENT OF ENVIRONMENTAL SCIENCES NEW ARTS COMMERCE AND SCIENCE COLLEGE, AHMEDNAGAR (2011-2012)

Ahmednagar Jilha Maratha Vidya Prasarak Samajs

Ahmednagar --------------------------------------------

CERTIFICATE
This is to certify that, Mr. ASHIAH DATTATRAYA HULAGE student of M.Sc. Environmental Science has completed the project entitled Effect of Sugar Industry effluent on changes of Growth and Chlorophyll contents of Mung plants. for the partial fulfillment of the syllabus of University of Pune, for the award of Post Graduation in Environmental Science in the academic year 20011-2012. He has completed his work satisfactorily and sincerely.

Project Guide

External

Co-ordinator Dept. of Environment Science

AKNOWLEDGEMENT

It is indeed a great pleasure to submit this project report entitled Effect of sugar industry effluent on changes of growth and chlorophyll contents of Mung(vigna radiata) Plants. I dedicate This project to the Almighty without whose blessing this work would not have been possible. I feel great pleasure to honor and thank my project guide Dr. S.D. Kulkarni H.O.D. Environmental Science Department of New Arts, Commerce and Science College Ahmednagar. I am indeed very much thankful to our Principal and Vice-Principal for providing necessary facilities for the project. I will feel satisfied if I acknowledge the blessing of all my teachers, parents as well as supported help of my friends who have always boosted my skills and encouraged me to complete this work. I am greatful to Prof. P.L. Wagh for providing the necessary information regarding my project. Last but not the least I am thankful to all the staff of the Department.

INDEX

Sr.No. 1 2 3 4 INTRODUCTION OBJECTIVES REVIEW OF LITRATURE

Title

Page no.

METHODS AND MATERIALS 1) COLLECTION OF EFFLUENTS 2) COLLECTION OF SEEED MATERIALS 3) COLLECTION OF SOIL 4) CHARACTERISTICS OF EFFLUENT 5) CHARACTERISTICS OF SOIL 6) GERMINATION PERCENTAGE 7) ROOT LENGTH AND SHOOT LENGHT 8) ESTIMATION OF CHLOROPHYLL

5) 6) 7)

RESULT AND DISCUSSION CONCLUSION REFERENCES.

INTRODUCTION Industrialization is an important tool for the development of any nation. Consequently, the industrial activity has expanded so much all over the world. Today, it has become a matter of major concern in the deterioration of the environment. With the rapid growth of industries (sugar , paper, tannery, textile dairy, dye industries) in the country, pollution of natural water by industrial waste water has increased tremendously. Among them, sugar industry plays a major role in producing a higher amount of water pollution because they contain large quantities of chemical elements. Sugar industries effluent or waste water is purely of plant origin. It contain large quantity of soluble organic matter & plant nutrients. They contain higher amounts of BOD, COD, electrical conductivity and colour. A common practice is to discharge the untreated industrial effluents in nearby water bodies or agricultural land. This has rendered some water bodies polluted to the extent where the water is not safe for human or livestock consumption.(IUCN 1991). Discharge of Raw spent wash into open land or nearby water bodies is resulting in a number of environmental problems of water and soil pollution including threat to plant and animal lives. The effluents not only affect the plant growth but also deteriorate the soil properties when used for irrigation. In arid and semi-arid regions of our country where shortage of water becomes limiting factor in agriculture. The effluent mixed polluted water is used for irrigational purposes by farmers. As this polluted water is being used for irrigation to cultivate the crops, it is necessary to conduct experiments to find out the impact of these effluents on agricultural crops before they are recommended for irrigation. Earlier reports revealed that inadequately diluted effluents discharged drum industries have potential of inhibiting seed germination, seedling growth, chlorophlly, protein (Pandey G.C, Neraliya S. 1999) As the effluent contains some major and secondary elements useful for plant growth, so it can be used in agriculture up to some extent with proper treatment or precautions. Experimental cropMung

The mung bean also known as mung bean, mung, green gram or golden gram is the seed of vigna. Classification- Kingdom- plantae Family-fabaceae Species- V. radiate Binomial Name- vigna radiate Green gram or mung (vigna radiata) is a leguminous pulse crop, growth all over India a protein rich dal. Many botanists agree that it originated in India, since its closely related species like vigna radiate grows wild in India and it has been grown in this region since prehistoric period. Green gram is cultivated in India, Burma, Ceylon, Pakistan, China and Africa. In India the important states growing this crop with a total area as about 30 lakh hectares are Orissa, Maharashtra, Andhra Pradesh, Madhya Pradesh, Gujarat, Rajasthan and Bihar. Its grains are used as Dal, soup and feed for animals. Its straw is used as fodder and fuel. Green gram grows in a wide range of climatic condition. A warm humid ultimate with temperature range of 25 C to 35 c with moderate rains of 80-100cm, well distributed during growing period of 100 days, It is cultivated during warm and wet kharif season and in south India it is cultivated in mild winter rabbi season. Chlorolphyll is photosynthetic pigment which directly affect to photosynthetic productivity. The chlorophyll are the essential components for photosynthesis and occur in chloroplasts as green in all photosynthetic plant tissues. Chlorophyll gives leaves their green colour and absorbs light that is used in photosynthesis. Chlorophyll is found in high concentration in chloroplasts of plant cells. Chlorophyll is a green pigment found in almost all plants, algae and cyanobacteria. They are bound loosely to protein but are readily extracted in organic solvents such as acetone or other. There are at least five types of chlorophyll in plants. Chlorophyll a & b occur in higher plants, fern & mosses, chlorophyll c, d, e are only bound in algae and in certain bacteria. The experiment was conducted to study the change in germination % and seedling growth of Mung (Vigna radiata) after the exposure of sugar industry effluent. The change in chlorophyll content in mung plants in different concentration of effluent.

OBJECTIVES

1) To examine germination percentage & root & shoot growth of Mung after the exposure to various concentration of effluents.

2) To determine properties of effluents used for experiment.

3) To examine a, b, and total chlorophyll content of mung after the exposure to various concentration of effluents.

Review of Literature

The work carried out in India & Abroad on beneficial effects or otherwise & their usages in agriculture and environment have been reviewed in this chapter under-

Sahai and et al. (1985) reported distillery effluents application upto 5 percent effluent increased root and shoot length, plant biomass, net primary productivity, chlorophyll content and seed output compared to the control. But about 5% effluent all the values decreased, so it was concluded that reuse of distillery effluent in irrigation of increased crop yield.

Mukherjee and sahai (1988) studied The effect of distillery waste 1-100% concentration on seed germination, seeding seedling establishment and early seedling growth of Cajannus cajan L. (car. 15-16).The values for percentage germination, rate of germination, root length and seedling were highest in 5% effluent while shoot length was maximum at 2.5 percent effluent.

Parmila Rani and Sanjeev Kumar (Jan 2010) worked on Soil irrigation effect of sugarcane industrial effluent on changes and level of chlorophyll, growth and yield of Triticum aestivum CV. P B W -226. For the study, Plants were grown in pot and irrigated with selected concentration of sugarcane industrial effluent with control ( i.e. from 10% to 100%) upto development of yield. The effluent reflected promontory effect to 20% concentration of effluent on chlorophyll level, growth & yield of plant. The experiment suggested that effluent can be used as fertilizer after dilution.

Salunke K.J. Gopal Reddy and Salve A.W. (2010) worked on effect of sugar factory effluent on seed germination & early seeding growth in groundnut. (Archis hypogea) verities. For study used different concentration of effluent as 25%, 50%, 75% and 100%. The effluent had most beneficial effects on percentage of seed germination & seeding development at lower (25%) concentration. Increase in effluent concentration showed decrease in seed germination & vigour index. The treated effluent can be used as a liquid fertilizer after dilution to a suitable concentration.

Sahai et al (1987) reported that rice seeds treated with different concentration of spent wash ( 0.5,10, 25, 50, 75, & 100%). At higher concentration (25% above) both the speed of germination & seedling growth were retarded because it contains excess amount of cations and anions it injurious to plant growth. At 5% concentration overall growth of seedling was better than in control and suggested that by diluting the spent wash to 5% the spent wash can be used as substitute for chemical fertilizer. The chlorophyll a and b decreased with increase in the effluent concentration. The cartoeniod continued to increase up to 5% spent wash concentration.

Rajaram et al (1988) soaked black gram seed in different concentration of distillery spent wash (1, 2.5, 5.0, 10, 25, 50& 100%) using water soaked as control. After 8 days of germination period it was noticed that spent wash treatment upto 2.5% concentration, spent wash treatment promoted seeding growth, but at higher concentrations, the spent wash has salinity due to dissolve solid accumulation. At higher concentration, trace element like zinc, hinder the plant growth by binding enzyme protein.

Shrivastava and Sahai(1987) suggested that the early seedling growth of cowpea was promoted by sugar factory effluent up to 10% concentration. The effluent at concentration of more that 10% suppressed the seeding growth. They expressed the nutrients and organic pollutant inhibit seedling growth by altering the osmotic relations & the distillery effluent could be used as a liquid fertilizer only for certain crops after proper dilution with water.

Pandey G. C. et al (1999) worked on phyto-toxic effect of sugar and distillery factory effluent on seed germination & protein contents of T. aestivum (k-68). These effluents exhibited stimulating (l50% concentration) and inhibitory (>50% concentration) effects on the seed germination, and the level of protein contents of T. aestivum(k-68) because of high pollution in these effluents.

Dhanam S.(2009)-Investigation was carried out To study seed germination ,seedling growth and certain biochemical parameters in paddy variety ADT38. paddy seeds were raised in petriplates irrigated with various concentrations of dairy effluent ( 0,5,10,25,50,75,100%). At lower dilutions, the paddy showed favorable effect on seed germination, seedling growth & dry matter production amino acid, starch pigment content over control. Among them 100% concentration of effluent caused inhibitory effect.

Rani and Shrivastava (1990) studied The impact of various concentrations (10, 25, 50, 75 & 100%) of distillery effluent on growth of peas. Found that shoot length, leaf number per plant ,leaf area, chlorophyll content & biomass exhibited a gradual increase upto 25%. However, the effluent at all concentration inhibited root growth. The number of pod per plant, seeds per pod & seed per plant in peas increased upto 25% concentration.

Singh et al (2006) revealed The effect of fertilizer effluent on seedling growth and chlorophyll content of gram Cicer aeritenum at different concentration of the effluent & time intervals. The effluent was alkaline in nature with strong ammonia odour. The seedling growth & chlorophyll content which showed a gradual decline with increase effluent concentration. An evaluation of seedling at 21 days at 25% concentration recorded and decrease in root & shoot length & chlorophyll content. Thus the study suggested that effluent can be used safely for gram (Cicer aeritenum) cultivation after proper dilution.

Babu et al (1996) worked on foliar application of distillery spent was liquid fertilizer for betterment of growth of sorghum Vulgare and Cajanus cajan . found that application of spent wash without dilution did not result in growth deformities or any alternation in plant metabolism& agricultural increase in plant growth attribute were observed. Further they noticed increased total chlorophyll, phenol, pigments, and protein, amino acid, starch and nitrogen contents in the leaves in treated plants.

Material & Methods

The materials used and methods followed during the course of investigation are described in this chapter under the following headlines. 1) Collection of effluentSugar mill effluent samples were collected from the point of discharge of Ashok sahakari Sakhar Karkhana Ltd; Ashoknagar (Assk), Tal.-Shrirampur, Dist-Anagar. The effluent sample was collected in plastic containers & stored in refrigerator. 2) Collection of soil sampleThe soil sample was collected from agricultural field up to 0-30 cm. height for experiment.

3) Collection of seed MaterialsThe certified seeds of Mung (Vigna angularis ) were purchased from Mahatma Phule Krishi Vidyapeeth Rahuri (M.P.K.V.) Dist:- Anagar. Seeds with uniform size, colour and weight were chosen for the experimental purpose.

4) Characteristics

of effluent-

Parameters

Methods used

References

1) pH

pH meter (potentiometer)

Jackson (1973)

2) EC(ds/m)

3) BOD (mg/L)

Digital Jackson (1973) conductivity meter (conductometric) Dissolved oxygen Franson (1985) method Open Reflux method Franson (1985)

4) COD(mg/L)

5) Characteristic of Soil-

No.

Parameter

Method used

References

1)

Soil pH

pH meter (potentiometer) Digital conductivity meter (conductometric) Walkey &Black Oxidation method

Jackson (1973) Jackson (1973) Nelson & Sommer (1982)

2)

EC(ds/m)

3)

Organic Carbon(kg/ha) Available Nitrogen (kg/ha)

4)

Alkaline paramagnet Subbain & method Asija (1956) Olsen et al (1982)

5)

6)

Available Phosphorus 0.5M N2HCO3 (kg/ha) (Ascorbic acid) using Spectrophotometer Available Potassium N NH4OAC (kg/ha) (Neutral N ammonium acetate solution

Jackson (1973)

6) Germination Percentage (%)A seeds of Mung ( Vigna angularis) were sterilized with 0.1% mercuric chloride solution for 5 minutes to remove microbes and then washed three times with sterile distilled water. A series of six sterilized Petri dishes with Whatman no.1 filter paper. Plant seeds were spread on each sterilized Petri dish lined with blotting paper and then irrigated with 5ml of the different concentration (0%, 1%, 5%, 10%, 15%, 20%) diluted sugar industry effluent .Each Petri dish containing 40 seeds of Mung. Observation were recorded at 24 hrs intervals, The germination seeds were counted on 96 hrs after sowing & germination percentage was calculated by using formula-

Germination % = No. of seeds germinated x 100 Total no. of seeds sown

7) Root length and Shoot lengthFive seedling were taken randomly from each treatment & their root & shoot length ( cm\seeding) were measured by using a scale and these values are recorded.

8) Estimation of ChlorophyllFor the estimation of chlorophyll from treated & controlled fresh green leaves were plucked randomly from the crops at 7 days interval at morning hrs.

Chlorophyll content was measured according to Arnon (1949) method for this, 1 gm of fresh leaves were homogenate in 20 ml of 80% acetone in dark using mortar & pestle. The homogenate was centrifuge at 3000 rpm for 15 min & transfer the supernatant. To 100ml volumetric flask. Repeat the procedure until residue is colourless. Make the final volume 100 ml with 80% acetone. The absorbance of solution was recorded at 645 nm. 652nm&663nmagainst the solvent ( 80% acetone) blank using visible spectrophotometer. The amount of chlorophyll a, b & total chlorophyll was calculated according to following formula.

Chlorophyll a mg\g = 12.7(A 663) - 2.69 (A 645) x

V 1000 x W

Chlorophyll b mg\g = 22.9 (A663) - 4.68 (A652) x

V 1000 x W

Total Chlorophyll mg\g = 20.9 (A 645) + 8.02 (A 663) x

V 1000 x W

Where, A Absorbance at specific wavelength V Final volume of chlorophyll extract in 80% acetone. W Fresh weight of tissue extracted.

Result and Discussion

Characteristics of Sugar Industry Effluents-

Sr.no. 1) 2) 3) 4) 5) 6)

Parameters Colour Odour pH EC (ds/m) COD(mg/L) BOD(mg/L)

Values Dark Brown Unpleasant smell 4.18 42.3 2500 1250

The physic-chemical parameters of the sugar industry effluent are given in table. The analysis of sugar mill effluent showed that it is acidic in nature with dark brown in colour. The pH was relatively low (4.18) soil becomes acidic resulting in poor crop growth and yield. The electrical conductivity (EC 42.3 ds/m)being very high indicate presence of high concentration of dissolved salts.

The Chemical Oxygen Demand (COD) valve of the effluent was 2500 mg/L. While the recommended level set by BIS is 250 mg/L; the measured COD indicates the high organic load.

It also contained higher amount of Biological Oxygen Demand (BOD) 1250 mg/L. Similar findings of Chandrasekar et al (1998), Rathore et al (2000) and Borole and Patil (2004)

Characteristics of Soil-

Sr.no. Parameters
1) 2) 3)

Values 7.2 0.69 0.25

pH EC (ds/m) Organic Carbon(%) Major Nutrients (kg/ha) N P K

126 76 98

Effect of sugar industry effluent on seed germination of mung (Vigna angularis)

Effluent concentration

Germination percentage (%) hrs 24hrs 48 hrs 87.5 97.5 92.5 97.5 85 72.5 72hrs 97.5 97.5 100 97.5 90 82.5 96hrs 97.5 100 100 97.5 90 87.5

Control 1% 5% 10% 15% 20%

80 87.5 90 95 75 60

Graph shows germination % of mung at different concentration. of effluent at different time interval. The germination % was increased at low concentration. (1%, 5%) resulting stimulating after out of all promontory doses. 1% & 5% dose show maximum prvonotion in comparison to other doses. Lower concentration of effluent was shown to support 100% seed germination. The most drastic reduction in seed germination percentage as the effluent concentration increased from 10% to 20%. Thus the germination percentage decreased with increased in effluent concentration.

Root Length and Shoot Length (in cm)

5th day

15th day

21th day

Effluent concentration Root

Shoot

Root

Shoot

Root

Shoot

Control

1.1

2.3

2.3

4.3

3.8

7.4

1%

1.2

2.7

2.9

5.8

4.3

8.9

5%

1.4

3.1

3.3

6.9

4.9

10.7

10%

1.6

3.6

3.7

8.2

5.3

12.2

15%

1.3

3.2

2.2

4.1

3.2

5.1

20%

2.4

1.9

3.8

2.7

4.5

Values are arithmetic mean (cm)

Data of root & shoot length at different concentration of effluent presented in table of graph. The higher shoot & root lengths were recorded at 10% effluent concentration The root & shoot length were adversely affected by higher concentration of sugar industry effluent treatment. A gradual decrease in these growth parameters were observed. The reduction in shoot & root growth at higher concentration of effluent. The higher toxicity were more pronounced in seedlings at higher conc. Effluent.

Change in Chlorophyll a content (mg/g) of Mung at different concentration of effluent

Sr.no.

Concentration 7 days (%) Control 1% 5% 10% 15% 20% 0.042 0.044 0.045 0.050 0.046 0.043

14days

21days

1. 2. 3. 4. 5. 6.

0.345 0.364 0.433 0.441 0.210 0.303

0. 456 0.720 0.723 0.741 0.362 0.423

Values are arithmetic mean and % change of chlorophyll a(mg/g)

Change in Chlorophyll b content (mg/g) of Mung at different concentration of effluent

Sr.no. 1. 2. 3. 4. 5. 6.

Concentration (%) Control 1% 5% 10% 15% 20%

7 days 0.043 0.046 0.055 0.058 0.063 0.057

14days 0.461 0.511 0.589 0.596 0.248 0.318

21days 0.649 0.701 0.851 0.889 0.585 0.529

Values are arithmetic mean and % change of Chlorophyll b (mg/g)

Change in Total Chlorophyll content (mg/g) of Mung at different concentration of effluent

Sr.no. 1. 2. 3. 4. 5. 6.

Concentration (%) Control 1% 5% 10% 15% 20%

7 days 0.082 0.090 0.112 0.119 0.128 0.102

14days 1.06 1.28 1.40 1.57 0.609 0.458

21days 1.106 1.456 1.637 1.647 1.001 0.990

Values are arithmetic mean and % change of Total Chlorophyll (mg/g)

Data of chlorophyll content at different duration of exposure and concentration of effluent represented in tables and shows in graphs. At 10% effluent conc. Chlorophyll a, b, and total chlorophyll content increased as 0.741, 0.889,1.647 mg/g respectively upto 21 days. At 15% and 20% effluent concentration overall decrease in chlorophyll content was recorded at all intervals as compared to control. A change in total chlorophyll synthesizing capacity of crop has diminished affecting the overall photosynthetic process. Increase in chlorophyllase enzyme. Nag (1981) had suggested increase in chlorophyllase activity by chemical agents. Another reason in due to the presence of Mg ++ ions in effluent which are required for Synthesis of different chlorophyll molecules.