Documenti di Didattica
Documenti di Professioni
Documenti di Cultura
Their Applications
Gerard L. Hasenhuettl
Kraft Foods
Richard W. Hartel
University of Wisconsin
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Cover design: Will T. Comellas, A.G.T. Inc.
t'
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Copyright 1997 Springer Science+Business Media Dordrecht
Originally published by Chapman & Hall in 1997
Softcover reprint of the hardcover 1st edition 1997
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12345678910 XXX 0100999897
Library of Congress Cataloging-in-Publication Data
Food emulsifiers and their applications/edited by Gerard L.
Hasenhuettl and Richard W. Hartel.
p. em.
Includes bibliographical references and index.
ISBN 978-1-4757-2662-6 (eBook)
ISBN 978-1-4757-2664-0
DOI 10.1007/978-1-4757-2662-6
1. Food additives. 2. Dispersing agents. I. Hassenhuettl, Gerard L., 1944II. Hartel, Richard W., 1951TP455.F667 1997
664'.06-dc21
97-1647
CIP
British Library Cataloguing in Publication Data available
Contents
xiii
Contributors
Preface
XV
ONE
Overview of Food Emulsifiers
Gerard L. Hasenhuettl
1.1
1.2
1.3
1.4
Introduction
Emulsifiers as Food Additives
Emulsifier Structure
Emulsifier Functionality
References
1
2
5
7
9
TWO
Synthesis and Composition
of Food-Grade Emulsifiers
11
R. J. Zielinski
2.1
2.2
2.3
2.4
2.5
Introduction
Mono- and Diglycerides
Propylene Glycol Monoesters
Lactylated Esters
Polyglycerol Esters
vii
11
13
15
18
23
viii
26
26
29
31
33
36
36
38
THREE
Analysis of Food Emulsifiers
39
Gerard L. Hasenhuettl
3.1
3.2
3.3
3.4
3.5
3.6
Introduction
Thin-Layer and Column Chromatography
Wet Chemical Analysis
Physical Methods
Instrumental Methods
Setting Specifications for Food Emulsifiers
References
39
40
43
52
54
62
63
FOUR
Carbohydrate/Emulsifier Interactions
67
Lynn B. Deffenbaugh
4.1 Introduction
4.2 Inclusion Complexes of Starch
4.3 Effects of External Lipid Materials on
Starch Properties
4.4 Factors Affecting Complex Formation
4.5 Physical Properties of Starch/Emulsifier Complexes
4.6 Summary
References
67
68
69
81
85
90
90
Contents
ix
FIVE
Protein/Emulsifier Interactions
95
Introduction
Protein Stability
Protein/Surfactant Interactions
Protein/Phospholipid Interactions
Protein/Emulsifier Interactions-Food Applications
References
95
97
98
120
132
137
SIX
Introduction
Surface Activity
Solution Properties of Emulfiers
The Use of Phase Diagrams to
Understand Emulsifier Properties
6.5 Examples of the Relation between
Phase Diagrams and Emulsion Stability
6.6 Some Ways to Classify Emulsifiers
6.7 The Emulsifier Surface
References
147
148
149
153
154
161
167
171
SEVEN
173
Stephen R. Euston
7.1 Introduction
7.2 Ice Cream
173
174
183
187
191
194
197
199
202
203
204
EIGHT
Applications of Emulsifiers
in Baked Foods
211
Frank T. Orthoefer
8.1
8.2
8.3
8.4
8.5
8.6
8. 7
8.8
Introduction
History of Emulsified Shortenings
Emulsifier Function in Baked Goods
Role of the Shortening
Role of the Emulsifier
Emulsifier Interaction with Bakery Components
Applications in Baked Goods
Summary
References
211
211
213
215
216
221
225
233
234
NINE
Emulsifiers in Confectionery
235
Mark Weyland
9.1 Introduction
9.2 Emulsifiers in Chocolate and Compound Coatings
9.3 Antibloom Agents in Chocolate and
Compound Coatings
9.4 Other Emulsifiers Used in Coatings
9.5 Emulsifiers in Nonchocolate Confectionery
235
236
244
248
249
Contents
xi
252
253
TEN
Margarines and Spreads
255
Eric Flack
10.1
10.2
10.3
10.4
10.5
10.6
10.7
10.8
10.9
10.10
10.11
10.12
Introduction
Early Development
Yellow-Fat Consumption
Definitions and Descriptions
Structure and Raw Materials
Fat Crystallization
Emulsifiers
Processing
Reduced- and Low-Fat Spreads
Oil-in-Water Spreads
Liquid Magarine
Summary
References
255
256
257
258
262
263
267
270
274
278
278
279
279
ELEVEN
Emulsifier Trends for the Future
281
Gerard L. Hasenhuettl
11.1
11.2
11.3
11.4
Index
282
282
283
284
285
286
287
Contributors
Thomas Arnebrant
Department of Pharmaceutical Analysis
Ferring AB
Box 30047
S-216 13 Malmo, Sweden
Bjorn Bergenstahl
Institute for Surface Chemistry
P.O. Box 5607
S-114 86 Stockholm, Sweden
Martin Bos
TNO Nutrition & Food Research Institute
P.O. Box 360
3700 AJ Zeist, The Netherlands
David C. Clark
DMV International
NBC-Iaan 80
P.O. Box 13
5460 BA Veghal, The Netherlands
xiii
xiv
Contributors
Lynn B. Deffenbaugh
Hill's Pet Nutrition
Science & Technology Center
P.O. Bo.c 1658
Topeka, KS 66601-1658
Stephen R. Euston
New Zealand Dairy Research Institute
Private Bag 11029
Palmerston North, New Zealand
Eric Flack
Danisco Ingredients, Ltd.
North Way
Bury St. Edmonds
Suffolk, 1P32 6NP, United Kingdom
Gerard L. Hasenhuettl
Kraft Foods, Inc.
801 Waukegan Rd.
Glenview, IL 60025
Tommy Nylander
University of Lund, Chemical Center
Department of Physical Chemistry
P.O. Box 124
S-22100 Lund, Sweden
Frank T. Orthoefer
2004 Me Cracken St.
Stuttgart, AR 72160
Mark Weyland
Quest International
5115 Sedge Blvd.
Hoffman Estates, IL 60192
Richard J. Zielinski
Quest International
Joliet Technical Center
24708 Durke Dr.
Joliet, IL 60410-5249
Preface
Food emulsions have existed since long before people began to process foods for
distribution and consumption. Milk, for example, is a natural emulsion/colloid in
which a nutritional fat is stabilized by a milk-fat-globule membrane. Early
processed foods were developed when people began to explore the art of cuisine.
Butter and gravies were early foods used to enhance flavors and aid in cooking.
By contrast, food emulsifiers have only recently been recognized for their ability to stabilize foods during processing and distribution. As economies of scale
emerged, pressures for higher quality and extension of shelf life prodded the development of food emulsifiers and their adjunct technologies. Natural emulsifiers,
such as egg and milk proteins and phospholipids, were the first to be generally
utilized. Development of technologies for processing oils, such as refining,
bleaching, and hydrogenation, led to the design of synthetic food emulsifiers.
Formulation of food emulsions has, until recently, been practiced more as an
art than a science. The complexity offood systems has been the barrier to fundamental understanding. Scientists have long studied emulsions using pure water,
hydrocarbon, and surfactant, but food systems, by contrast, are typically a complex mixture of carbohydrate, lipid, protein, salts, and acid. Other surface-active
ingredients, such as proteins and phospholipids, can demonstrate either synerXV
xvi
Preface
hope that this effort will stimulate further innovation directed at increasing the
quality and reducing the expense of processed food products.
Acknowledgments
The editors express their apprec1atwn to Barbara Bagnuolo and Julie
Hasenhuettl for typing and proofreading large portions of the manuscript.
Gerard L. Hasenhuettl
Richard W. Hartel
ONE
1.1
Introduction
Food emulsions, colloids, and foams have their origins in the evolution of animal species. Milk has a naturally occurring membrane that allows the dispersion of fat droplets into an aqueous environment. Early food formulations to
produce butter, whipped cream, cheese, and ice cream built upon the natural
emulsifiers present in the system. The development of mayonnaise in France
as a cold sauce utilized the natural egg phospholipids to disperse a liquid oil
into an acidified aqueous phase. The emulsifying power is still very impressive
by today's standards since it allowed up to 80% oil to be dispersed without inversion to an oil-continuous emulsion. The invention of margarine by the
French chemist Hippolyte Mege-Mouries in 1889 utilized the solid fat of tallow to produce a stable oil-continuous emulsion to serve as a low-cost substitute for butter. In this case, the emulsion had to be stable temporarily only
until the product was chilled.
Synthetic emulsifiers have come into common use only in the latter half of
the twentieth century. Their development was driven by the emergence of the
processed food industry, which needed technology to produce and preserve
quality products that could be distributed through mass market channels. A
key technical hurdle was to maintain product stability over an extended shelf
life. With small amounts of emulsifier, for example, salad dressings can be
stored on a shelf for more than a year without visible separation. Now, other
factors such as the development of rancidity limit consumer acceptance of aging products.
Detailed knowledge of the physical chemistry of emulsions is best obtained
when pure oil, water, and surfactants are used. Food emulsions, by contrast, are
extraordinarily complex systems. Commercial fats and oils are complex mixtures
of triglycerides that also contain small amounts of highly surface-active materials (Friberg and Larsson, 1990). Salt content and pH in food emulsions vary
widely enough to have significant effects on their stability. Natural and commercial emulsifiers are often complex mixtures that vary in composition between different manufacturers. Other food ingredients, such as proteins and particulates,
contribute surface activity that may dramatically alter the character of the emulsion. Because of all these complex relationships, the formulation of food emulsions grew up as an art, dominated by individuals having a great deal of
experience. The gradual development of sophisticated techniques such as electron microscopy, nuclear magnetic resonance, and chromatography/mass spectrometry has solidified the art with a scientific dimension.
The subject of food emulsions has been extensively reviewed by Friberg
and Larsson (1990), Krog et al. (1985), and Jaynes (1985). This book will be
oriented to the design, manufacture, and use of food emulsifiers as ingredients
to improve the quality and economy of processed foods.
1.2
2ICFR
No.
Lecithin
Functionalities
Typical
applications
184.1400
Coemulsifier, viscosity
reducer
Margarines, chocolate
products
Monoglycerides
184.1505
Emulsifier, aerator,
crystal stabilizer
Margarines, whipped
toppings, peanut
butter stabilizers
Diacetyltartaric acid
esters of
monoglycerides
184.1101
Baked goods,
confections, dairy
product analogs
Monosodium salt of
analogs, phosphated
monoglycerides
184.1521
Emulsifier, lubricant,
release agent
Dairy products,
soft candy
21CFR
No.
Functionalities
Typical
applications
Emulsifier, plasticizer,
surface-active agent
whipped toppings
Lactylated
monoglycerides
172.850
Acetylated
monoglycerides
172.828
Succinylated
monoglycerides
172.830
Emulsifier, dough
strengthener
Shortenings, bread
Ethoxylated
monoglycerides
172.834
Emulsifier, stabilizer
Cakes, whipped
toppings, frozen
desserts
Sorbitan
monostearate
172.842
Emulsifier, rehydrating
agent
Confectionery
coatings, yeast,
cakes, icings
Polysorbates
172.836
172.838
172.840
Emulsifier, opacifier,
solubilizer, wetting agent
Salad dressings,
coffee whiteners,
gelatins, ice cream
Polyglycerol esters
172.854
Sucrose esters of
fatty acids
172.859
Emulsifier, texturizer,
film former
172.844
172.846
Emulsifier, dough
conditioner, whipping
agent
Bread, coffee
whiteners, icings,
dehydrated
Baked products,
potatoes
Propylene glycol
esters
172.858
Emulsifier, aerator
Cake mixes,
whipped toppings
As with any other totally new food additive, the need to prove safety of the
product in foods at high levels of consumption requires extensive toxicity studies
and enormous documentation. The consequent financial and time commitments
make development of totally new emulsifiers unattractive for emulsifier manufacturers. A somewhat easier development approach is to petition for expanded
use (new applications or higher permitted levels) of emulsifiers that are already
approved. However, even this tactic may require several years of review.
In addition to national regulations, many food processors require their ingredients, including food emulsifiers, to be kosher so that their products are
acceptable to Jewish and many Islamic consumers. For emulsifiers to be
kosher, they must be produced from kosher-certified raw materials. This requirement precludes the use of almost all animal fats. This is not much of a
problem since emulsifiers are easily produced from vegetable fats that can be
blended to give similar fatty acid compositions. The major concern in kosher
certification is to determine in advance whether the customer's rabbinical
council recognizes the Hekhsher (kosher symbol) of the producer's rabbi.
Products labeled as "all natural" must contain ingredients that have not
been chemically processed or modified. Only lecithin or other naturally occurring materials such as proteins would be acceptable for these products.
1.3
Emulsifier Structure
Lipophilic end
Hydrophilic end
-OH
Unsaturated; oleate
Unsaturated; linoleate
Figure 1.1
also contain measurable concentrations of trans (E) fatty acids that have higher
melting points than the cis (Z) fatty acids.
Polar head groups may be present in a variety of functional groups. They
may be incorporated to produce anionic, cationic, amphoteric, or nonionic surfactants. Mono- and diglycerides, which contain an -OH functional group, are
the most widely used nonionic emulsifiers. Lecithin, whose head group is a
mixture of phosphatides, may be visualized as amphoteric or cationic, depending on the pH of the product.
Proteins may also be surface active due to the occurrence of lipophilic
amino acids such as phenylalanine, leucine, and isoleucine. lnterfacially active proteins will fold so that lipophilic groups penetrate into the oil droplet
while hydrophilic portions of the chain extend into the aqueous phase.
Proteins in this configuration may produce a looped structure that provides
steric hindrance to oil-droplet flocculation and coalescence. Charged proteins
may also stabilize emulsions due to repulsion of like charged droplets.
Proteins may also destabilize water-in-oil emulsions, such as reduced fat margarines, by causing the emulsion to invert.
Food emulsifiers may be thought of as designer molecules because the
structure and number of heads and tails may be independently varied. A very
useful conceptual tool is hydrophile/lipophile balance (HLB). The topic has
been extensively reviewed by Becher, so only a brief description will be presented here. The number and relative polarity of polar groups in a surface-ac-
tive molecule determine whether the molecule will be water- or oil-soluble (or
-dispersible). This concept has been quantitated by calculation of an HLB
value to describe a given emulsifier. High-HLB values are associated with
easy water dispersibility. Since conventional practice is to disperse the surfactant into the continuous phase, high-HLB emulsifiers are useful for preparing
and stabilizing oil-in-water (0/W) emulsions. Low-HLB emulsifiers are useful
for formulation of water-in-oil (W/0) emulsions, such as margarine. Extreme
high or low values are not functional as emulsifiers since almost all of the molecule will be solubilized in the continuous phase. They would, however, be
very useful for full solubilization of another ingredient, such as a flavor oil or
vitamin, in the continuous phase. At some intermediate values of HLB, the
molecule may not be stable in either phase and will result in high concentration at the interface.
Example(s)
Whipped toppings
Flavors, vitamins
Bread, buns, rolls
Yeast and sugar manufacturing
Macaroni, pasta
Salad oil
Yeast-raised baked goods
Candy, chewing gum
Margarines, frying shortenings
Frozen toppings and coffee
Gloss enhancer
Cloud formers
Hydrating agents
Encapsulating agents
Dispersion stabilizers
Whiteners
Confectionery coatings
Beverages
Powdered milk drinks
Flavors, aromas
Peanut butter
of solid particles in a continuous fat or oil phase. Chocolate viscosity is controlled by the addition of soy lecithin or polyglycerol ricinoleate (PGPR). Oil
separation in peanut butter is prevented by use of a monoglyceride or highmelting fat. In some cases the secondary effect may be of greater concern than
formation of the emulsion. Strengthening of dough and retardation of staling
are vital considerations to processors who bake bread.
A common practice in the food industry is to use two- or three-component
emulsifier blends to achieve multiple functionalities. In a cake emulsion, for
example, aeration to produce high volume, foam stabilization, softness, and
moisture retention are achieved by using an emulsifier blend. One useful statistical method to optimize emulsifier blends is the full factorial experimental
design using a zero or low level of each emulsifier and a higher level of each
emulsifier. The major advantage of this design is that it will detect two- and
three-factor interactions that are not uncommon in complex food systems.
Small-molecule emulsifiers (e.g., monoglycerides) may exert their effect by
partially or totally displacing proteins from an oil/water interface. This replacement is entropically favored because of the difference in size and mobility of the species. Direct interaction of emulsifiers and proteins may be
visualized through electrostatic and hydrogen bonding, although it is difficult
to observe in a system that contains appreciable amounts of oil. Chapter 5 on
emulsifier/protein interactions will elaborate on these concepts.
Emulsifier suppliers generally employ knowledgeable technical service
professionals to support their customers product development efforts. Their experience in selecting emulsifiers for a functional response is a valuable initial
source of information. However, food processors may want to develop unique
products that have no close relationship to a product currently in commerce. In
this case, the supplier may have some general ideas for emulsifier selection.
However, it may be necessary for product developers to define their own criteria for emulsifiers based on critical functions required in the product.
Statistical experimental design is a very useful tool to optimize food emulsifiers and their concentrations. For example, a full factorial design (Krog et al.,
1985; Jaynes, 1985) may be used to determine the levels of three emulsifiers to
obtain an optimum product. Response surface methodology (RSM) and fractional factorial designs are very useful techniques because they reduce the
number of experiments necessary to obtain optimal concentrations. However,
since synergistic and antisynergistic effects are often observed between emulsifiers, care should be taken to design the experiments so that two-factor inter-
actions are not confounded. Robust design is also highly recommended so that
the food product has minimal sensitivity to uncontrolled noise factors.
The objective of this book is to provide the food industry professional or interested technical professional with an overview of what emulsifiers are, how they
are prepared, and how they are utilized in food products. Although in many
senses food emulsifiers have become commodity ingredients, sophisticated understanding and application in processed foods is likely to continue to advance.
References
Friberg, S., Larsson, K. (eds.) (1990). Food Emulsions, Marcel Dekker, New York.
Jaynes, E. {1985). "Applications in the food industry, II," in Encyclopedia of Emulsion
Technology {ed. P. Becher), Vol. 2, Marcel Dekker, New York, 1985, pp. 367--85.
Krog, N., Rilson, T.H., Larsson, K. (1985). "Applications in the food industry, " in
Encyclopedia of Emulsion Technology (ed. P. Becher), Vol. 2, Marcel Dekker, New
York, pp. 321-66.
TWO
Synthesis and Composition of
Food-Grade Emulsifiers
R.J. Zielinski
2.1
Introduction
The food-grade emulsifiers are generally esters composed of a hydrophilic (water-loving) end and a lipophilic (fat-loving) end. In general, the lipophilic end
is composed of stearic, palmitic, oleic, or linoleic acid or combinations of these
fatty acids. The hydrophilic end is generally composed of hydroxyl or carboxyl
11
12
groups. The melting point of the various esters within each family will be determined by the melting point of the fatty acids used to prepare the emulsifier.
The melting points of the common fatty acids are given in Table 2.1. When
stearic and palmitic acids dominate, the ester will be solid and relatively highTable 2.1 Melting points of fatty acids
C8:0
CIO:O
Cl4:0
Cl6:0
Cl8:0
Cl8:1
Cl8:2
Acid
CMP
Caprylic acid
Capric acid
Lauric acid
Myristic acid
Palmitic acid
Stearic acid
Oleic acid
Linoleic acid
16.3
31.5
44.2
56.4
62.9
69.6
14
-6
eC)
melting; when oleic and linoleic acids dominate, the ester will be low-melting
and could be a liquid at room temperature. The fatty acids present in an emulsifier may be obtained from either a fat or oil or a fatty acid source. All fats and
oils are triglycerides, and the fatty acids can be obtained from the triglycerides
by a hydrolytic process followed by fractional distillation. Generally, either
natural or fully hydrogenated fats and oils are split to obtain the fatty acids.
This process is illustrated in Equation 1.
0
II
CH 2-0-C-R
CH 2-QH
II
CH-o--c-R1
CH 2-0-C-R
3H 20
CH-QH
CH 2-0H
R--cOOH
R1--cOOH
R2--cooH
0
Triglyceride
Glycerine
Fatty acids
13
Kosher oleic acid may be obtained from high-oleic safflower and high-oleic
sunflower seed oils. It may also be obtained from a specially purified tall oil
from pine trees. Commercial stearic acid may be of three types: (i) a mixture of
about 90% stearic acid and 10% palmitic acid, (ii) a mixture of about 70%
stearic acid, 30% palmitic acid, or (iii) a mixture of about 50% palmitic acid
and 50% stearic acid; all are known as stearic acid. Generally, partially hydrogenated fats and oils are used to prepare plastic-type emulsifiers.
2.2
The mono- and diglycerides are the most widely used food-grade emulsifiers.
They may be esters that are solid and high-melting, esters that are liquid at
about room temperature, or a plastic-type ester. There are a variety of monoand diglycerides commercially available at the present time. These are commonly designated as (i) 40% a-monoglycerides, (ii) 50% a-monoglycerides,
and (iii) 90% monoglycerides. Monoglycerides are commonly composed of a
number of components, as indicated in Figure 2.1.
These components are present in different amounts in the commercially
available esters. The various types of commercial compositions are described
in the following discussion.
The mono- and diglycerides may be prepared by either an interesterification process with glycerine and triglycerides or a direct esterification process
with fatty acids. These processes are illustrated in Equation 2. In the latter
case, water of esterification is formed and removed during the process step.
0
CH 2-0H
CH-OH
CH-C-R
CH-OH
II
CH 2-0-C-R
II
CH-0-C-R
II
CH2-0-C-R
II
CH-0-C-R
II
CH 2-0H
CH-0-H
CH 2-0H
CH2-0-C-R
Glycerine
Monoglyceride
Diglyceride
Triglyceride
Figure 2.1
14
II
CH 2-0H
CH 2-0-c-R
CH-QH
II
heat
CH-0-C-R
CH-OH
CH 2-0H
CH-OH
catalyst
II
CH-QH
0
II
CH 2-0H
CH 2-0-C-R
CH 2-0H
CH-Q-C-R
Glycerine
Triglyceride
Glycerine
Monoglyceride
CH-OH + R-cOOH
I
Glycerine
II
CH 2-0H
CH 2-0H
heat
catalyst
CH 2-D-c-R
II
CH-O-c-R
II
II
CH-0-C-R
II
CH 2-0-C-R
CH 2-D-C-R
Diglyceride
Triglyceride
(2)
The catalysts commonly used in these processes are sodium hydroxide or hydrated lime (calcium hydroxide); the temperatures involved range from 200 to
250C. The proportions of each of the products-free glycerine, monoglyceride,
diglyceride, and triglyceride-are purely dependent on the molar ratio of glycerine and oil or glycerine and fatty acid used. The overall composition can be approximated using a random distribution of the free hydroxyl group and fatty acid
groups (Feuge and Bailey, 1946). The various fatty acid groups from the oil or
fatty acid are also assumed to he randomly distributed in the final product.
Once the desired ratio of oil or fatty acid and glycerine has been chosen to
yield the desired monoglyceride content, a number of different types of monoglyceride compositions are still potentially available. For example, if a 40%
monoglyceride composition is desired, then (i) the catalyst may be neutralized,
15
generally by phosphoric acid; (ii) the catalyst may be not neutralized; (iii) the
reaction mixture may be cooled to a determined temperature and the glycerine
that is insoluble in the final composition removed by decantation type process;
and (iv) the excess free glycerine may be removed from the reaction products
by vacuum distillation. A 40% monoglyceride prepared by a decantation-type
process (iii) may contain about 4% free glycerine, while a 40% monoglyceride
prepared by a vacuum-stripping process will typically contain less than 1%
free glycerine. A stripped 40% monoglyceride will typically be composed of
about 46% monoglyceride, 43% diglyceride, 10% triglyceride, and 1% glycerine. Similarly, a 50% monoglyceride may be prepared by decantation from insoluble glycerine or by a vacuum-stripping process. A vacuum-stripped 50%
monoglyceride will typically contain about 55% monoglyceride, 38% diglyceride, 5% triglyceride, and 2% free glycerine.
Under the conditions of high temperature, very low pressure, and an extremely short path, monoglycerides may be distilled and thus concentrated and
purified. This process is known as "molecular distillation." Typically, a 40%
monoglyceride mixture is subjected to molecular distillation to yield monoglycerides of over 90% purity. The nondistilled portion is recycled for an additional interesterification process to yield another 40% monoglyceride-type
composition for use as the feedstock.
As previously mentioned, either nonhydrogenated or partially hydrogenated
triglycerides, or saturated or unsaturated fatty acids, may be used to prepare
solid, plastic, or liquid monoglyceride mixtures.
The mono- and diglycerides have a generally recognized as safe (GRAS)
FDA status.
2.3
16
II
CH 2-QH
CH-OH
CH 2-0-c-R
heat
CH-O-C-R
catalyst
II
CH 3
CH 2-0-c-R
Propylene
glycol
Triglyceride
0
CH 2-QH
CH-QH
II
II
CH 2-o-c-R
CH-OH
CH 2-0-c-R
CH-0-C-R
CH2-0H
CH-OH
CH 3
CH 3
CH 3
CH2-QH
Propylene
glycol
Propylene
glycol
monoester
Propylene
glycol
diester
Glycerine
II
CH 2-0-c-R
CH 2-Q-C-R
CH-QH
II
CH 2-o-c-R
CH-0-C-R
II
CH-o-c-R
CH-QH
CH 2-QH
CH 2-o-c-R
Monoglyceride
Diglyceride
Triglyceride
17
18
CH 2--o-c-R
CH 2-0H
CH-OH
R-cOOH
heat
CH-OH
catalyst
CH 2
CH 3
II
CH 2-0-c-R
CH 2-0-C-R
I
(4)
tripropylene glycol monoesters. These esters are not allowed by the FDA as direct food additives.
The FDA regulations for propylene glycol monoesters allow the use of all
edible oils and edible fatty acids. However, the majority of the propylene glycol monoesters commercially available contain very high percentages of
palmitic acid and stearic acid that are saturated fatty acids. Few unsaturated
propylene glycol monoesters are available at this time.
2.4
Lactylated Esters
19
/R-COOH
OH f--- R-COOH
OH
I 0II
CH 3-c-COOH
f--- R-QH
CH 3-C-c-o-cOOH
f--- R-OH
Lactic acid
CH3
Dilactic acid
Figure2.2
2.4.1
Two broad categories of food-grade emulsifiers that are prepared by the reaction
of the hydroxyl group of lactic acid with either fatty acids or fatty acid chlorides
are commercially available: (i) the lactylic esters of fatty acids and (ii) partial
metal salts of lactylic esters of fatty acids. Each will be discussed in tum.
2.4.1.1
20
acids most probably is prepared by the direct esterification of lactic acid with
fatty acid in about a 1:1 molar ratio (Thompson et al., 1956). A high free polylactic acid content (see Chapter 3 for analytical procedure) indicates that the
product has not been water-washed.
There are no restrictions on the lactic acid content of the lactylic esters of
fatty acids in the FDA regulations (21CFR172). There are no FDA restrictions
on the edible fatty acids that may be used in the preparation of the lactylic esters of fatty acids; however, the stearate esters are most commonly produced.
2.4.1.2
Metallic Salts of Lactylic Esters of Fatty Acids. There are two metallic salts of lactylic esters of fatty acids that are used in the food industry: calcium salt and sodium salt. The calcium salt was commercialized first, as a
dough conditioner for bread, and is known as calcium stearoyl-2-lactylate.
The 2 indicates that 2 moles of lactic acid were used in its preparation. The
sodium salt, which was commercialized later, is known as sodium stearoyl-2lactylate, sodium stearoyllactate, or simply "SSL." It is used extensively in
many food applications.
Both esters can be prepared by the same process, the direct esterification of
the partial salt of lactic acid with a fatty acid in about a 2: l molar ratio of lactic acid to fatty acid (Thompson et al., 1956). Both esters have a monomer content of about 40%, a polymeric content of about 31%, and a polylactic acid
content of about 6%.
Care must be taken to use high-quality starting materials to avoid excessive
darkening during the reaction process. The color of the final product can be reduced by the use of hydrogen peroxide as a bleaching agent.
The amounts of lactic acid and metal ion content of both esters are regulated
by the FDA and are about 34% and 4.5%, respectively. Only the use of specified
grades of stearic acid is permitted by the FDA regulations for the products.
2.4.2
2.4.2.1
can react with the hydroxyl group of other fatty acid-derived compositions to
yield emulsifiers that are useful in many food products. One of the most common
fatty acid compositions used is a reaction product with a monoglyceride to form a
lactylated monoglyceride identified in the FDA regulations as glycerol-lacto esters of fatty acids. The overall simplified reaction is illustrated in Equation 5.
As can be seen from Equation 5, the number of hydroxyl groups remain the
21
OH
I
+ CH -C-COOH
3
I
H
____.
r,--o--c
OH
I I
-c-cH,
+ Hp
CH-OH
I
Monoglyceride
Lactic acid
Lactylated monoglyceride
(5)
22
These glycerol lactates can be removed by water-washing or can be steamstripped from the reaction mixture under reduced pressure.
A distilled monoglyceride could also be used as a reacting monoglyceride
composition. However, during the reaction with lactic acid, the distilled monoglyceride could totally or partially disproportionate to yield free glycerine and
a mixture of mono-, di-, and triglycerides. Thus, the benefit of using a distilled
monoglyceride could be lost.
Other possibilities of starting monoglycerides are an unstripped intermediate monoglyceride with an a-mono content of 28 to 40%, a 40% U-monoglyceride that has been vacuum-stripped to remove the free glycerine, or a 50%
monoglyceride type that has or has not been vacuum-stripped to remove the
free glycerine before reaction with lactic acid. Of course, any diglycerides present in the monoglyceride mixture can also react with lactic acid.
Another variable in the production of lactylated monoglycerides is the
amount of lactic acid reacted with the monoglyceride intermediate. In theory,
a monoglyceride can react with 2 moles of lactic acid to yield the dilactylated monoglyceride ester. Commercially, lactylated monoglycerides are
available that contain either about 15% or 22% lactic acid. There is an analytical constant known as WICLA (see Chapter 3), which stands for water-insoluble combined lactic acid, used by some companies to characterize
lactylated monoglycerides. This value will give an estimation of the amount
of active functional lactylated monoglyceride present in the commercial
product as opposed to a total lactic acid value that is a summation of the
WICLA value and the amount of lactic acid present in a water-soluble form,
such as glycerol lactates.
Because of the wide range of possible starting monoglycerides and the reaction conditions chosen, all commerciallactylated monoglyceride compositions
may not be compositionally the same, and possibly, differences in functionality
can exist between products with a similar lactic acid content.
The FDA regulations for lactylated mono- and diglycerides permit the use
of any edible fat or oil or edible fatty acids to be used in their manufacture.
There are no restrictions on the amount of lactic acid that can be present in the
lactylated mono- and diglycerides.
2.4.2.2
Another
emulsifier, prepared from lactic acid by the reaction of the carboxylic acid
group of the lactic acid with the hydroxyl groups of glycerol and propylene gly-
23
col monoesters, is the lactylated fatty acid esters of glycerol and propylene glycol. In this type of emulsifier, propylene glycol is interesterified with an edible
fat or oil to yield a mixture of mono- and diglycerides and propylene glycol monoesters. The reaction mixture is then reacted with lactic acid. As with the
lactylated mono- and diglycerides, a great variety of propylene glycol monoester and monoglyceride mixtures could be used to react with lactic acid to
yield a final composition. In addition, lactic acid can react with any free propylene glycol and free glycerine initially present or formed by a disproportionation of the starting propylene glycol monoesterlmonoglyceride composition.
These propylene glycol lactates and glycerol lactates are removed by waterwashing or vacuum distillation of the reaction product. Obviously a complex
composition is produced.
It should be noted that only propylene glycol esters prepared by an interesterification process may be used. Propylene glycol mono- and diesters prepared by a direct fatty acid esterification route are not allowed under the
pertinent FDA regulation (21CFR172.850). The FDA regulation further specifies the WICLA content of the final ester composition to be 14 to 18%.
2.5
Polyglycerol Esters
Polyglycerol esters have been commercially available to the food industry for
over 25 years, but they have been known in the nonfood industry for many
more years. The polyglycerol alcohols are most often prepared by the polymerization of glycerine with an alkaline catalyst at elevated temperatures
(Harris, 1941; Babayan and Lehman, 1973). The polymerization is a random
process and a number of different polyglycerols are produced. This is illustrated in Equation 6.
The extent of polymerization is followed by refractive index, viscosity, or
hydroxyl value. When the theoretical hydroxyl value for a diglycerol is obtained, the polyglycerol could be called diglycerol. When the hydroxyl value
for triglycerol is obtained, the polyglycerol could be called a triglycerol, etc.
The hydroxyl values for di- to decaglycerol are summarized in Table 2.2. The
theoretical molecular weight of each polyglycerol is also given in Table 2.2.
In general, no effort is made to separate the various polyglycyerols, and the
entire reaction mixture that contains a distribution of polyglycerols is used to
prepare polyglycerol esters. At lower degrees of polymerization, higher concentrations of lower poyglycerols are present; at higher degrees of polymeriza-
24
heat
catalyst
OH
OH
OH
I I
OH
OH
OH
I I
OH
OH
OH
OH
OH
Diglycerol
OH
I I
Triglycerol
Theoretical
hydroxyl value
Theoretical
molecular weight
1839
1352
1169
92
166
240
314
338
462
536
610
684
758
1072
1012
971
942
920
902
888
25
plex. Since mixtures of fatty acids are used, this makes the polyglycerol esters even more complex. Thus, a triglycerol monostearate polyglycerol ester
is a mixture of the stearate and palmitate esters of glycerine, diglycerol,
triglycerol, tetraglycerol, pentaglycerol, hexaglycerol, heptaglycerol, octaglycerol, nonaglycerol, and decaglycerol, while a decaglycerol tristearate is
a mixture of the stearate and palmitate esters of the same polyglycerols but in
different proportions.
A wide range of polyglycerol esters are commercially available from liquid
esters such as hexaglycerol dioleate to plastic esters like triglycerol monoshortening to solid esters like decaglycerol decastearate.
FDA regulations permit the use of edible hydrogenated or nonhydrogenated nonlauric oils and the edible fatty acids derived from them as well
as oleic acid from tall oil to prepare polyglycerol esters up to and including
decaglycerol.
26
2.6
Sorbitan Esters
Only one sorbitan ester is currently approved in the FDA regulations as a direct food additive in the United States: sorbitan monostearate. However, a
GRAS petition has been filed to permit the use of sorbitan tristearate in confectionery coatings (Federal Register, 1988). The FDA, as of 1996, has not
acted on this petition. The most commonly used process for the production of
sorbitan esters is the direct fatty acid esterification of sorbitol with stearic acid
(Brown, 1943; Japanese Patent, 1974). Generally, the stearic acid used is
about a 50:50 mixture of stearic and palmitic acids. The overall reaction is
shown in Equation 7.
Sorbitan is a monoanhydro sorbitol and sorbide (or isosorbide) is a dianhydro sorbitol, as illustrated in Equation 7. Commercial sorbitan monostearate is
the stearate, palmitate ester of a mixture of about 1 to 12% sorbitol, 65 to 72%
sorbitan, and 16 to 32% isosorbide. It can be demonstrated that the amount of
sorbitan is fairly constant but that the amount of linear sorbitol and isosorbide
can vary among manufacturers.
The use of an acidic catalyst during the esterification of sorbitol promotes
the cyclization of sorbitol to the mono- and dianhydro forms. The use of a basic
catalyst promotes the esterification reaction at the expense of color formation.
It is possible that each commercial supplier has its own blend of acidic and
basic catalysts to generate the ratio of sorbitan esters required to meet tight
FDA specifications for hydroxyl number, saponification number, and acid
number while minimizing color degradation of the product. Hydrogen peroxide
can be used to reduce the color of a sorbitan monostearate.
Stockburger (1981) has patented a process for preparing sorbitan esters in
which sorbitol is first dehydrated with an acid catalyst to the desired ratio of
sorbitol, sorbitan, and sorbide polyols. The mixture of polyols is then reacted
with fatty acids and a basic catalyst to yield the desired sorbitan ester with an
improved light color. The use of sorbitan monostearate in foods is highly regulated. The reader is referred to 21CFR172.842 for further details on where sorbitan monostearate can be used in food products, and for the required
analytical constants.
2.7
Ethoxylated Esters
Four ethoxylated fatty acid esters have FDA approval for use as direct food additives. These are ethoxylated sorbitan monostearate, ethoxylated sorbitan
CH 2-0H
CH 2-DH
CH-OH
CH-OH
CH-OH
27
catalyst
CH-OH
R-COOH
CH-OH
CH-OH
CH-DH
CH-OH
II
CH 2-0H
CH 2-0-C-R
Sorbitol
Linear sorbitol
monoester
J=l
CHI
L<JH
I
r-<JH
I
I
r_j
I
o
CH_j
CH-OH
CH
CH-OH
CH 2-0-C-R
Sorbitan
Sorbide
monoester
monoester
(7)
28
monooleate, ethoxylated sorbitan tristearate, and ethoxylated saturated monoand diglycerides. The base ester that has been ethoxylated is sorbitan monostearate, sorbitan monooleate, sorbitan tristearate, and a 28% monoglyceride,
respectively. For each of these esters the same basic reaction is involved, the
reaction of a hydroxyl group with ethylene oxide. This reaction is illustrated in
Equation 8 for a monoglyceride.
A basic catalyst such as potassium hydroxide is used during the ethoxylation process (Egan and Lampost, 1969). The ethoxylation process is a highly
exothermic reaction and adequate cooling must be available to control the reaction temperature. The ethoxylated esters are highly complex in composition,
and the actual components of the ethoxylated sorbitan or monoglyceride esters
have not been defined. Ethylene oxide itself is a highly explosive compound
under certain conditions. Safety instructions on the handling of ethylene oxide
must be read and understood before an ethoxylation process is undertaken.
Another name for ethoxylated compositions is polyoxyethylene (20) sorbitan
CH 2-0H
I \
CH -CH
CH-QH
CH 2-0-CH 2-CHpH
catalyst
II
CH 2-0-C-R
Monoglyceride
1\
CH 2-CH 2
CH-OH
CH -C-R
2 II
0
Ethylene oxide
CH 2-0-CH 2-CHpH
IH2-0-CH2-CH2 -O-CH2-CH2-0H
CH -O-C-R
2
II
(8)
29
monostearate, for example. Polyoxyethylene refers to the ethylene oxide polymeric chain and the (20) indicates that approximately 20 moles of ethylene oxide
have been reacted with one average molecular weight of the starting fatty acid
ester. Other more common names for these products are polysorbate 60 for
ethoxylated sorbitan monostearate, polysorbate 80 for the oleate, polysorbate 65
for the tristearate, and polyglycerate 60 for the saturated monoglyceride.
The polyoxyethylene group is a hydrophilic, or water-loving, group. By the
introduction of the polyoxyethylene group, the starting fatty acid ester is made
more hydrophilic and the final composition has excellent water solubility or
dispersibility. The melting point of each of the starting esters is decreased and
the ethoxylated esters are liquids or soft pastes at room temperature.
The use of the ethoxylated esters in food products is highly regulated, and
the reader is referred to 21CFR172.834, 21CFR172.836, 21CFR172.832, and
21CFR172.840 for details of where and how much polyglycerate 60, polysorbate 60, polysorbate 65, and polysorbate 80, respectively, may be used.
2.8
Succinylated Esters
Succinylated esters of monoglycerides and propylene glycolesters are permitted as direct food additives. The essential step in the preparation of succinylated esters is the reaction of a hydroxyl group with succinic anhydride. This is
illustrated in Equation 9 for a monoglyceride.
II
CH,--c~
Monoglyceride
II
+
CH -0-C-R
0 -------+ CH-OH
CH--c/
2
II
Succinic anhydride
CH -0-C-R
2
II
Succinylated monoglyceride
(9)
30
2.9
31
There are two fruit acids used to make food-grade emulsifiers: tartaric acid and
citric acid. Tartaric acid-derived esters have been used extensively in Europe,
but they have not been used very much in the United States. One of the foremost uses of tartaric acid-derived esters is a dough conditioner; in the United
States other dough-conditioning agents are used in place of tartaric acid esters.
The tartaric acid esters used are the mono- and diglycerides of diacetyl tartaric
acid (DATEM esters). The synthesis of these esters is shown in Equation 10.
COOH
HO-CH
HO-CH
COOH
Tartaric acid
Acetic anhydride
Diacetyl tartaric
anhydride
Acetic acid
(lOa)
32
CH -0-C-R
2
0
COOH
H3 C-c-O-CH
H3
I
c-c-o-cz o
""IIc-o CH
2
I
CH-QH
DATEM
(lOb)
33
pH; if the food product is basic, the DATEM ester can be converted to the anionic form. The two acetyl groups are somewhat liable to hydrolysis, which
yields acetic acid. These esters should not be used in aqueous systems that are
expected to have a long storage life.
Two different types of citric acid-based emulsifiers are permitted as direct
food additives: stearyl monoglyceride citrate (21CFR172.755) and monoglyceride citrate (21CFR172.832). The monoglyceride citrate is restricted to use in
antioxidant solutions. It is prepared by the reaction of glycerol monooleate
with citric acid. The final product has an acid number of 70 to 100 and has a
14 to 17% total citric acid content.
Stearyl monoglyceride citrate is prepared by the reaction of citric acid,
monoglycerides, and stearyl alcohol to yield a composition with an acid number of 40 to 52, a saponification number of 215 to 255, and a total citric acid
content of 15 to 18%. The expected reaction is shown in Equation 11.
As can be seen, the reaction products are quite complex. Both saturated and
unsaturated monoglycerides may be used. This type of ester is used in shortenings containing other emulsifiers and is not used extensively in the food industry.
2.10
Acetylated Monoglycerides
Another food-grade additive that is formed by the reaction of an acid anhydride with a hydroxyl group is the acetylated monoglycerides. This reaction is
illustrated in Equation 12.
The starting monoglyceride may be saturated or unsaturated and is of molecularly distilled type. 21CFR172.828 permits the use of a non-food-grade catalyst or a food-grade catalyst in the acetylation reaction. The acetic acid
byproduct and any excess acetic anhydride or formed triacetin are removed by
vacuum distillation. Since the acetic anhydride is monofunctional, there is no
danger of polymers being formed.
The acetylated monoglycerides may also be formed by an interesterification
process. In this case, triacetin is interesterified with a suitable mixture of edible fats or oils and glycerol, as illustrated in Equation 13.
In this case, the composition of the final mixture can be calculated using a random distribution (Feuge and Bailey, 1946). The final reaction mixture is separated by means of steam-stripping and molecular distillation. Any unreacted
triacetin is removed by the vacuum steam-stripping process, and the desired
acetylated monoglycerides are concentrated by the molecular distillation process.
34
CH 2-COOH
HQ-C-COOH
CH 2-COOH
Citric acid
CH-QH
CH-QH
II
CH-0-C-R
Monoglyceride
Stearyl alcohol
(ll)
Note that in direct reaction acetylation, the hydrophilic hydroxyl group is replaced by a lipophilic acetyl group; thus, if a fully acetylated ester is prepared,
the product is no longer an emulsifier since it has no hydrophilic groups. In the
interesterification process both fully esterified and partially esterified compositions can also be formed, depending on the amount of glycerine used.
The more fully esterified compositions are useful for their film-forming and
moisture barrier properties.
FDA regulation 21CFR172.828 specifies that the final composition have a
Reichert-Meiss! value of 75 to 200 and an acid value under 6. The Reichert-
------+
CH 3-c-o-c-c-CH 3
CH -0-C-R
2
35
+
CH -0-C-R
Monoglyceride
Acetic anhydride
(12)
0
CH 2-0H
I
CH-OH
CH 2-0-C-cH 3
CH 2-0-c-R
CH-O-C-cH 3
I
CH 2-0H
CH-Q-C-R
CH 2-0-C-cH 3
CH-O-C-CH 3
CH 2-0-C-CH 3
catalyst
CH 2-Q-C-R
CH 2-0-C-cH 3
CH-0-C-R
0
II
CH 2-0-C-R
CH-OH
I
CH -0-C-CH
other
compounds
(13)
36
Meissl number determines the amount of short-chain fatty acids in the final
composition, and this is a measure of the degree of acetylation (see Chapter 3
for methodology). The degree of acetylation in commercially available acetylated monoglycerides ranges from 50 to 90%.
2.11
Phosphated Esters
2.12
Sucrose Esters
The newest family of emulsifiers to obtain FDA approval for use as direct food
additives is the sucrose esters. This occurred in 1982 (47FR5547). The sucrose esters permitted as additives under 21CFR172.859 are the mono-, di-,
and triesters. This regulation does not relate to the fully esterified sucrose esters that have been promoted as nonabsorbable fat and oil replacements. These
esters are a mixture of the octa-, hepta-, and hexaesters of sucrose.
The main method of preparing sucrose monoesters is by means of an interesterification process between sucrose and methyl esters of fatty acids. The basic process is illustrated in Equation 14.
Sucrose and the methyl esters of fatty acids are very insoluble in one another. In addition, sucrose darkens and carmelizes on heating to temperatures
around 150C. This insolubility and darkening of the sucrose has led to the
use of organic solvents such as N,N-dimethylformamide (DMF) or dimethyl
sulfoxide (DMSO) and reduced pressure in the preparation of the sucrose
mono-, di-, and triesters (Haas, 1959). The use of DMSO as a solvent was allowed in 1987 (52FR10883).
37
HOCH 2
HO
H
OH
OH
Sucrose
II
R-C-OCH 3 _______.
Methyl esters
RCOOCH 2
HO
H
Sucrose monoester
OH
OH
Methyl alcohol
(14)
38
produce an emulsion particle size less than one-quarter the wavelength of light,
in contrast to other emulsions in which particle sizes are greater than the wavelength of light. This particle is so small the emulsion appears transparent.
However, the use of the high amount of the metal salts of the fatty acids and unreacted sucrose leads to difficult purification and separation procedures. In general, solvents such as ethyl acetate, methyl ethyl ketone, or isobutyl alcohol are
used during the purification process. FDA rule 21CFII 72.859 specifies the
amount of these solvents that can be in the final sucrose ester. A patent has been
issued regarding the purification of the sucrose ester reaction mixture (Mizutani
et al., 1973), but many other purification processes are possible. Overall, the
production of the sucrose monoesters is a more complicated process than the
production of the other food-grade emulsifiers.
References
Babayan, V.K., Lehman, H. (1973). U.S. Patent 3,637,774, January 25.
Barsky, G. (1950). U.S. Patent 2,509,414, May 30.
Brown, K.B. (1943). U.S. Patent, 2,322,820, June 29.
Egan, R.R., Lampost, S.B. (1969). U.S. Patent, 3,433,645, March 18.
Eng, S. (1972). U.S. Patent 3,636,017, January 18.
Federal Register, 53, 40808, December, 1988.
Feuge, R.O., Bailey, A.E. (1946). Oil and Soap, 23, 259.
Food Chemical Codex (1981). 3d ed., pp. 98-99.
Freund, E.H. (1966). U.S. Patent 3,293,272, December 20.
Harris, B.R. (1939a). U.S. Patent 2,177,983, October 31.
- - (1939b). U.S. Patent 2,177,984, October 31.
- - (1941). U.S. Patent 2,258,892, October 14.
Hass, H.B. (1959). U.S. Patent 2,893,990, July 7.
Japanese Patent Publication (1974). No. 15246, April13.
Martin, J.B. (1968). U.S. Patent 3,375,262, March 26.
Mizutani, N., et al. (1973). U.S. Patent 3,798,324, July 24.
Osipow, L.l., Rosenblatt, W. (1969). U.S. Patent 3,480,616, November 25.
- - (1972). U.S. Patent 3,644,333, February 22.
Stockburger, G.J. (1981). U.S. Patent 4,297,290, October 21.
Thompson, J.B., Buddemeyer, B.D. (1956). U.S. Patent 2,744,825, May 8.
Thompson, J.B., eta!. (1956). U.S. Patent 2,733,252, January 31.
Yamagishi, F., eta!. (1974). U.S. Patent 3,792,041, February 12.
THREE
3.1
Introduction
Analytical methods for food emulsifiers are closely associated with or derived
from methods commonly used for fats and oils (Sonntag, 1982; Karleskind,
1996). Test methods are of several types and are carried out for a variety of
reasons. Analysis ensures that the composition of the emulsifier is correct and
that it has not seriously degraded during processing. Often the composition
and distribution of homologs has implications for the utility of the emulsifier in
the individual food product (see, for example, Halkier, 1980). The final level of
testing is often a measurement of performance in the food system itself.
Specifications are negotiated and agreed upon between the producer and the
customer (usually a processed food manufacturer). Analytical tests are carried
out on the process line or control laboratory of the supplier, who then provides
a certificate of analysis to the customer. The customer may then check the
analysis as part of the receiving procedure and accept or reject the shipment.
Disputes may be settled by submitting a sample to an independent laboratory.
Quality assurance testing is also necessary to ensure that foods conform to
39
40
standards set by government regulatory agencies. The U.S. Food and Drug
Administration (FDA), the European Economic Community (EEC), and the
regulatory agencies of individual countries have limits on the maximum concentration of emulsifiers or other food additives allowed in specific products.
This topic has been reviewed by several authors (Kroeller, 1966, 1968;
Gernert, 1968; Amano, 1979). Professional organizations, such as the
Association of Official Analytical Chemists (AOAC), the American Oil
Chemists' Society (AOCS), International Union of Pure & Applied Chemistry
(IUPAC), Leatherhead Food Research Association, and the National Academy
of Sciences (Food Chemicals Codex), have compiled lists of official methods
that are used for fats, oils, and emulsifiers (Cunniff, 1995; Firestone, 1990;
Paquot and Hauffen, 1987; Slack, 1987; Taylor, 1996).
To determine emulsifiers in intact food products, the fats and emulsifiers must
first be extracted. Fats and oils are soluble in nonpolar solvents, such as hexane
or toluene. However, emulsifiers are amphiphilic and more polar and tend to
be less soluble in cold hexane or petroleum ether, particularly when emulsifier
concentration is high compared to total lipid level. Chloroform or chlorofonn/
methanol mixtures (Flor and Prager, 1980) are sufficiently polar but also sufficiently immiscible to allow for extraction from aqueous food systems. In cases
where total lipid content is high compared to emulsifier concentration,
Halverson and Qvist (1974) have reported extracting total lipids with refluxing
hexane followed by extraction with acetonitrile. Solid foods samples (e.g., cake,
coffee whitener) are conveniently extracted using a Soxhlet extraction apparatus.
Liquid samples (e.g., milk, ice cream mix) are usually separated in a separatory
funnel or countercurrent distribution apparatus. Another factor complicating extraction from foods is that lipids may be tightly complexed by starches or proteins or may simply be encapsulated in a biopolymeric matrix. Jodlbauer (1976)
has demonstrated the use of an amylase enzyme to release lipids from a pasta
product so that they are more easily extracted.
41
42
3.3
43
Wet analysis describes a determination that uses solvents and chemical reagents
to determine an aspect of emulsifier composition. An example would be titration
of free fatty acid with alcoholic potassium hydroxide. Wet methods have been
standard practice for many years. However, a large analysis volume in laboratories poses the problem of disposal or recycling of reagents and/or spent solvents.
For this reason, the current trend is to replace the wet method with instrumental
or physical measurements that supply related or equivalent information. Some of
the most widely used methods for emulsifiers are described next.
3.3.1
a- Monoglyceride t
Monoglycerides may exist as alpha, the most predominant form, which has the
fatty acid esterified in the 1 or 3 positions of the glycerol head group, or beta,
where the substitution is at the 2 position. The alpha isomer has adjacent hydroxyl groups in the 1, 2 or 2, 3 positions. The isomer is determined by titration
with periodic acid resulting in cleavage, as shown in the following equation:
The analysis is carried out by dissolving the sample (extracted lipid or pure
OH
I
OH
----+
HI0 4
II
-c-c
CH
0 -~-~C17 H35
HI04
HI03
Hp
(15)
t
44
3.3.2
Fatty acids may occur during the preparation of many common food emulsifiers. They may be present as residues from direct esterification reactions or
may be due to hydrolysis of glycerides. Fatty acids may act as antiemulsifiers
in some systems. They also make a system sensitive to pH, since alkali transforms fatty acids into soaps, which are powerful anionic surfactants. Attempts
are therefore made to keep free fatty acids in mono/diglycerides as low as possible. Other emulsifiers, such as sodium stearoyl lactylate and lactylated
monoglycerides have high acid values that must fall in a range defined by the
FDA or other regulatory body.
Acid value is determined by dissolving a weighed sample of the emulsifier
in an isopropanol/toluene or ethanol/diethyl ether solvent and titrating with a
standard potassium hydroxide solution to a phenolphthalein endpoint. In cases
where the acid value is used to track the progress of reactions using anhydrides, such as in production of DATEM and acetylated monoglycerides, an
aprotic solvent rather than an alcohol must be used. Alternatively, the sample
solution may be titrated potentiometrically to the equivalence inflection point.
The latter method is particularly useful for dark-colored samples where a visual endpoint is difficult to observe. The acid value is defined as the number of
milligrams of potassium hydroxide necessary to neutralize the free fatty acid
per gram of sample and is determined by the formula
45
(A- B)(N)(56.1)
w
where A
3.3.3
Iodine Valuet
(S- B)(N)(l2.69)
where
s=
w
mL used to titrate sample
w=
t
weight of sample
46
In this test, when reporting the value, it is important to also identify the
method by which the value was determined.
Iodine value may also be calculated from the fatty acid composition (FAC) of
fats, oils, and fatty acids through gas-liquid chromatography (GLC)(Firestone,
1990h). The same method may be used for food emulsifiers, but new conversion
factors would probably need to be developed to translate GLC peaks to iodine
values. Near infrared reflectance (NIR) has also been developed as a rapid
method to determine unsaturation by measurement of the carbon-carbon double
bond absorbance peak. However, a great deal of work is necessary up front to assemble a calibration curve to relate the measured absorbances to traditionally
measured values.
3.3.4
Peroxide Valuet
Peroxides and hydroperoxides may be determined by the reaction of a solution of the sample in acetic acid and chloroform with potassium iodide.
Liberated iodine is then titrated with standardized sodium thiosulfate solution
using a starch indicator. Precautions need to be taken to avoid the presence of
residual oxidizing or reducing agents on glassware. Also, strong ultraviolet
light must be avoided, since it forms peroxides by a photochemically induced
oxygen addition. Peroxide is defined as the number of milliequivalents of peroxide (AOAC uses the term "active oxygen") per 1000 g of sample that oxidize
potassium iodide. The exact value is obtained from the following formula:
(V)(T) (1000)
m
m =
wt
in g of sample
47
Since the practicing formulator needs to minimize the peroxide content of ingredients, the lower the peroxide value, the less is the risk of developing rancidity.
Special surveillance should be maintained on materials that have higher iodine
values (higher unsaturation). Of course, other materials that catalyze oxidation,
such as iron, copper, and lipoxygenase enzymes, must also be considered.
Recently, peroxides have been determined utilizing high-performance liquid chromatography (HPLC) (Yang, 1992; Yang et al., 1991). A sample is separated on a column and fractions then pass through a post-column reaction
chamber where they are mixed with a luminescence reagent. Light emitted
from the chemiluminescence reactions is then measured by a special photochemical detector. The method is fairly rapid and requires only small amounts
of sample. An additional advantage is that different peroxide species may be
measured separately, which may indicate where oxidation is occurring. The
major disadvantage is the capital cost of the equipment and the high cost of the
luminescence reagent.
3.3.5
Saponification Numbert
w=
t
wt of sampling
48
This value is sensitive to both the degree of esterification and the chain
length of the substituent fatty acid. Higher degrees of esterification and shorter
chain lengths are reflected by higher saponification values. Conversely, longer
chain lengths and lower degrees of substitution are reflected in lower saponification values.
3.3.6
Hydroxyl Valuet
_(5_6._1)_(7)_(V._
0 -_V_) + AV
m
where T = normality of standard alkali solution
49
up the determination. Hydroxyl values are used mainly to specify and control
polyglycerols and polyglycerol esters.
3.3.7
Monoglycerides and fatty acids may be reacted with lactic acid to produce
lactylated derivatives that are useful emulsifiers. Lactic acid may be present as
esterified or free lactic acid. In addition, lactic acid has both a carboxyl function and can self-condense to produce small polymers. The total lactic acid
present in a sample encompasses all these species and is indicative of the lactic acid/lipid ratio in the production batch. In this test, a sample is boiled with
alcoholic potassium hydroxide followed by addition of hydrochloric acid solution. Lipid material is separated by exhaustive extraction with diethyl ether.
The aqueous phase is then titrated with standard potassium hydroxide solution
to the phenolphthalein endpoint.
Free lactic acid is determined by dissolving the sample in benzene and extracting the solution with water. Free lactic and poly lactic acids are then quantitated by titration with standard potassium hydroxide. An obvious problem
with this test is the use of benzene, which is a carcinogen.
Water-insoluble combined lactic acid (WICLA) is determined by dissolving a
sample in benzene and extracting the sample as described in the previous test. The
benzene layer (or upper phase from the previous test) is evaporated to dryness. The
residual solid is then heated with alcoholic potassium hydroxide neutralized and
titrated in the same manner as carried out in determination of total lactic acid.
The above methods suffer from the disadvantage of using a toxic, carcinogenic solvent and laborious phase-separation procedures. Franzke and Kroll
(1977) recommend the use of chloroform and petroleum ether for the determination of free and total hydroxy acids, respectively. The authors report the
analysis of lactic, citric, and tartaric acid in emulsifiers. They have also reported the use of an enzymatic method for determination of total lactic acid
(Franzke and Kroll, 1980).
3.3.8
Reichert-Meiss! Valuet
This method is specific for determination of acetic and short-chain fatty acids that
have been esterified to mono- or diglycerides. The method involves (i) cleavage of
Taylor (l996a).
50
the short-chain acid in alkali followed by neutralization with dilute sulfuric acid,
(ii) distillation of the liberated acetic (or other short-chain) acid, and (iii) determination of the condensed acid by titration with standard base to the phenolphthalein
endpoint. The method is therefore equipment intensive (the condensing unit must
be replaced or cleaned between samples) and time-consuming.
3.3.9
Moisture
3.3.10
Color
Color of emulsifier is a reflection of the quality of starting lipid in its manufacture and/or manufacturing conditions. Thermal abuse of fats and emulsifiers
are well known to produce dark colors. In many cases, oxidized tocopherols
are thought to be the chromatic impurities. Dark-colored emulsifiers may detract from the appearance of finished food products where they are used in
high concentrations (e.g., cake icings). However, color evidence that an emulsifier has been abused also suggests that pro-oxidants may be present that may
adversely affect the flavor of the product. Emulsifiers would therefore be
processed to produce the lightest color possible.
Firestone (l990m).
51
3.3.11
Although the official AOCS method has been classified as a surplus method
(obsolete), residual soap remains an important process check in emulsifier production. Many emulsifiers are produced by interesterification using alkaline
catalysts (see Chapter 2). The catalyst is neutralized at the end of the process
prior to removal of excess polyol by distillation. In batch processes, if the alkali is not properly neutralized, disproportionation (reverse reactions) can occur, resulting in emulsifiers that are out of specification. Analysis of soap
(alkali) is used to assess proper neutralization.
The determination is carried out by dissolving an emulsifier sample in a
suitable organic solvent/water mixture and titrating with standard hydrochloric
acid. Bromophenol blue or phenolphthalein may be used as indicators. The
method may also be adapted for potentiometric titration. Another method for
assessing the extent of neutralization is to measure the pH. A 5% solution of
emulsifier in isopropanol/water is equilibrated to room temperature and the pH
is read using a standard electrode. A pH reading of 6.5 to 6.8 indicates that
soap is absent and the emulsifier has been properly neutralized.
3.3.12
52
the lecithin obtained in this way is therefore a major concern. Its chemical
structure is that of a phospholipid where phosphoric acid is esterified to a glycerol molecule and an organic base. Total phosphorus in the sample may be determined by a laborious process in which the sample is saponified, phosphorus
is separated by precipitation with molybdate solution, the precipitate is dissolved in base, and excess base is titrated with standard nitric acid (Firestone,
1990r). Tamanaka and Kudo (1991) have reported a rapid method where phosphorus precipitated by molybdate is determined spectrophotometrically. A
much simpler, albeit less precise, procedure is to determine lecithin content as
acetone-insoluble material based on the principle that neutral lipids are soluble in acetone while phospholipids are not (Firestone, 1990g). This principle
was used by Goldstein (1984) to develop a method for determination of lecithin
in oils. The oil sample was dispersed in acetone and lecithin was quantitated
by measurement of the turbidity.
Individual
phosphatides,
such
as
phosphatidylcholine
and
phos-
3.4
Physical Methods
3.4.1
Melting Point
53
54
3.4.2
Viscosity/Consistency
3.4.3
Refractive Index
3.5
Instrumental Methods
There has been little recent effort toward development of new wet methods for
analysis of food emulsifiers. Studies that are carried out in this area have fo-
55
3.5.1
Gas-Liquid Chromatographyt
Christie (1989).
56
same molecular weight as monomargarine, is commercially available at a reasonable cost and stable to 1,2-acyl shifts (Hasenhuettl, 1990). Cholesteryl acetate has also been utilized as an internal standard (Blum and Koehler, 1970).
Collaborative studies have been conducted with the objective of developing a
uniform method for AOCS, AOAC, and IUPAC (Brueschweiler and
Hautefenne, 1990; Firestone, 1994). Figure 3.1 shows the separation of propylene glycol esters and monoglycerides by GLC.
Polyol distributions of other food emulsifiers can be determined by cleavage of fatty acids, as for fatty acid distribution. The residual polyol can be purified and converted to a volatile trimethylsilyl derivative that is analyzed by
GLC. Use of this technique provides structure information on the underlying
polyol. Percentages of sorbitan, sorbitol, and isosorbide present in sorbitan
mono- or tristearate can be determined using this procedure (Murohy and
Grisley, 1969; Tsuda et al., 1984). Polyols from polysorbates are detectable by
GLC, but under conditions reported by Lundquist and Meloan (1971), they are
eluted as a single peak. Detection of 1,4-dioxane, a carcinogenic byproduct of
ethoxylation reactions, may also be determined in polysorbates by GLC (Birkel
et al., 1979). Polyol distribution from diglycerol through dodecaglycerol are
obtainable for polyglycerol esters (Schuetze, 1977; Sahastrabudhe, 1967).
Polyglycerol polyricinoleate (PGPR), which has been extracted from chocolate
products, has also been determined by gas-liquid chromatography (Dick and
Jmcmal Standard
Internal Standard
PGMS
PGMP
PGMS
GMO&GMS
GMS
12
Time (min)
15
18
21
24
Time (min)
Figure 3.1 GLC separation of monoglycerides and propylene glycol ester emulsifiers.
(a) Commercial emulsifier; (b) in shortening. (Hasenhuettl et al., 1990.)
57
3.5.2
cratic solvent system because gradient elution causes drift of the IR detector.
Recently, an evaporative light-scattering detector (ELSD) has been developed
to overcome these technical problems for lipids (Christie, 1992; Bruns, I 988;
Lee et al., 1993). Figure 3.2 shows a separation of phospholipids using the
ELSD detector. HPLC can also be interfaced with an MS instrument to give
both separation and identification of peaks.
The most commonly reported separations of food emulsifiers under a potpourri of columns, solvents, and detectors have been monoglycerides (Filip
and Kleinova, 1993; Takagi and Ando, 1994; Renger and Wenz, 1989; Tajano
and Kondoh, 1987; Martin et al., 1989; Rilsom and Hoffmyer, 1978;
Brueschweiler, 1977) and phospholipids of soy (Melton, 1992; Sotirhos et al.,
1986; Hurst and Martin, 1984; Huyghebaert and Baert, 1992; Tumanaka and
Fujita, 1990; Rhee and Shin, 1982) and egg (Hsieh et al., 1981; Kaitaranta
and Bessman, 1981). Hurst and Martin (1980) have also employed HPLC to
determine phosphatidylcholine in chocolate. Separation of phospholipids has
58
1. Cholesterol
2. PalmHic acid
3. Phosphatidylethanolamine
4. Phosphatidylserine
5. Phosphatidylcholine
6. Sphingomyelin
12
16
1. Cholesterol
2. Phosphalidylethanolamine
3. Phosphalidylcholine
3 4. Sphingomyelin
12
2DIIin.
Column:
16
2D Min.
Column:
also been reviewed by Christie (1996). Garti and Aserin (1981) reported the
separation of polyglycerol mono- and polyesters by HPLC on a LiChrosorb column. Sorbitan esters of fatty acids could also be separated on the same stationary phase (Garti and Aserin, 1983). Murakami and coworkers (1989)
described the separation of sucrose esters by preparing their 3,5-dinitrobenzoyl derivatives. This technique was used to determine sucrose esters in a
wide range of food products.
The development of facile extraction and rapid sample prepurification
methods as well as autosampling technology will undoubtedly encourage more
widespread use of HPLC.
3.5.3
Spectroscopic Methods
59
lengths. The short-wavelength region is not selective enough to allow determination of lipids in the presence of other organic compounds. However, complexes of emulsifier functional groups have been widely utilized to analyze
emulsifiers colorimetrically. A diagnostic qualitative analysis for ionic surfactants was developed by Lew (1975). Methylene blue was used to analyze for
anionic surfactants at 650 nm while cationic compounds were determined with
Orange 2 at 485 nm.
Several determinations of phospholipids rely on complexation of the choline
moiety of phosphatidylcholine. Molybdenum blue (Hartman et al., 1980),
dipicrylamine (Mueller, 1977), and Reinecke's salt (Moelering and Bergmeyer,
1974) were used to determine lecithins in a variety of food products.
Polyoxyethylene chains in food emulsifiers can complex with cobalt thiocyanate
or Dragondorff's reagent to produce a colored complex. This method has been
used to detect polysorbates in instant noodle soup (Saito et al., 1987; Tonogau et
al., 1987) and in salad dressings (Daniels et al., 1982). Shmidt (1979) reported a
procedure for determination of DATEM esters by mixing saponified lipid with
sodium metavanadate solution and measurement of the absorption at 490 nm.
Infrared spectroscopy is applicable to the functional groups present in the
common food emulsifiers. The emergence of Fourier transform infrared spectroscopy (FTIR) makes rapid detection of functional groups possible. The technique could therefore be used to monitor manufacturing processes. Given
these capabilities, it is somewhat surprising that little work has been done to
measure emulsifiers. Ingber utilized near infrared (NIR) reflectance to determine monoglyceride concentration and determine the hydroxyl value in polyglycerols and polyglycerol esters (N. Ingber, personal communication).
Spectrophotometric analysis has also been utilized to measure impurities in
food emulsifiers. Fujita and Yamanaka (1991) measured the absorbance of iodine liberated from an iodide solution from reaction with lipid peroxides. This
technique yields equivalent information to the peroxide method described previously. Infrared absorption at 1675 cm1 was utilized to determine residual dimethylformamide in sucrose esters (Jakubska, 1977). However, this method is
inadequate to determine the solvent at ppm levels.
3.5.4
60
tain signals. These signals are dependent on the environment of the nuclei and
are therefore useful for determining functional groups and chemical structure.
Relaxation time of the signal is useful to determine the physical state of a molecule. Wide-line NMR is widely used to measure the amount of solid fat or
solid fat content (SFC) in a sample. However, this technique is rarely used for
food emulsifiers and is not included here. Chemical shifts have been utilized
to determine the mesomorphic form of emulsifiers in aqueous systems, which
is likewise beyond the scope of this chapter. The reader is referred to
Lindblom (1996) for a thorough treatment of this subject. Mesomorphic phase
behavior in food emulsions will be discussed in Chapter 6.
Proton (1 H) NMR is the oldest technique. However, because of the large number of similar protons in alkyl chains of emulsifiers, this technique has not shown
widespread utility. Press and coworkers (1981) observed the protons on choline at
3.3 ppm to determine phosphatidylcholine content in lecithin. Sheeley et al.
(1986), in the same laboratory, proposed that the vinylic protons at 5.3 ppm could
serve as an alternative measurement of the iodine value, discussed earlier.
Chemical shifts of carbon (13 C) are useful for determination of the functional environment of the carbon atom being observed. Since lipids have significantly fewer carbons than hydrogen atoms, the problem of spectra
3.5.5
Mass Spectrometry
Mass spectrometry has long been used to elucidate the structures of organic
molecules. The molecular ion is an accurate indicator of the molecular weight
Table 3.1
13 C
61
Emulsifier structure
Gl-1
Gl-2
GL-3
N-CH 2 O-CH 2
Soy phosphatidylcholine
Egg yolk phosphatidylcholine
Soy phosphatidylethanolamine
Egg yolk phosphatidylethanolamine
1-Monoglyceride
1,2-Diglyceride
!-Propylene glycol monoester
2-Propylene glycol monoester
Propylene glycol diester
Monoacetylated monoglyceride:
63.01
62.94
62.81
62.81
65.04
65.04
69.46
65.92
65.42
70.51
70.63
70.59
70.55
70.27
72.25
66.13
63.33
63.78
64.07
64.07
63.47
61.58
19.2 (CH 3)
66.26
66.62
40.69
40.59
71.77
67.98
16.25 (CH 3)
16.5 (CH 3)
62.07
63.00
62.00
72.39
68.19
69.16
61.40
65.26
62.33
a
~
Diacetylated monoglyceride
59.34
59.32
62.08
62.13
62
identified from the spectra. An electrospray technique has also been found to be
useful for analysis of sucrose fatty acid esters (Schuyl and van Platerink, 1994).
This technique showed a series of molecular ions that corresponded to the degree of substitution of fatty acids onto the sucrose molecule.
3.6
63
References
Amano, H., (1979). Tekisuto-Zeminaru, 26th Conference, Tokyo: Nippon Yukagaku
Kyokai, 59-74.
Biacs, 0., et al. (1978). Acta Aliment. Acad. Sci. Hung., 7(3):181-93, CA 89:213672.
Birkel, T., et al. (1979). ]. Assoc. Off. Anal. Chem., 62(4):931--6.
Blum, J., Koehler, W. (1970). Lipids, 5(7):601--6.
Bruemmer, J.M. (1971). Brot Gebaeck, 25(11):217-20.
Brueschweiler, H. (1977). Mitt. Geb. Lebensmittelunters. Hyg., 68(1):46-63.
- - , Dieffenbacher, A. (1991). Pure Appl. Chem., 63(8):1153--62.
- - , Hautefenne, A. (1990). Pure Appl. Chem., 62(4):781-93.
Bruns, A. (1988). Fett Wiss. Technol., 90(8):289-91.
Christie, W.W. (1996). "Separation of phospholipid classes by high performance liquid
chromatography," in Advances in Lipid Methodology-Three (ed. W.W. Christie),
The Oily Press, Ayr, Scotland, pp. 77-108.
- - (1992). "Detectors for high performance liquid chromatography of lipids with
special reference to evaporative light scattering detection," in Advances in Lipid
Methodology-One (ed. W.W. Christie), The Oily Press, Ayr, Scotland, pp. 239-271.
- - (1989). Gas Chromatography and Lipids: A Practical Guide, The Oily Press,
Ayr, Scotland.
Cunniff, P. (ed.) (1995.) Official Methods of Analysis of AOAC International, 16th ed.,
AOAC International, Arlington, VA.
Ibid. (1995a). 41.1.63 Method 969.34.
Ibid. (1995b). 41.1.60 Method 942.19.
Ibid. (l995c). 41.1.20 Method 940.28.
Ibid. (1995d). 41.1.18 Method 920.160.
Ibid. (1995e). 41.1.15 Method 993.20.
Ibid. (19951). 4l.l.l2 Method 965.32.
Ibid. (1995g). 4l.l.l6 Method 965.33.
Daniels, D.H., et al. (1985). ]. Agric. Food Chem., 33(3):368-72.
- - (1982). ]. Assoc. Off. Anal. Chem., 65(1):162-5.
Dawe, R.G., Wright, J.L.C. (1988). Lipids, 23(4):355-8.
Dick, R., Miserez, A. (1976). Mit. Geb. Lebensmittelunters Hyg., 67(4):472-87.
Dieffenbacher, A., et al. (1988). Rev. Fr. Corps. Gras, 35(12):495-9
- - (1989). Rev. Fr. Corps. Gras, 36(2):64.
Duden, R., Fricker, A. (1977). Fette Seifen Anstrichm., 79(12):489-91.
El-Sebaiy, L.A., eta!. (1980). Food Chem., 5(3):217-28.
Erdahl, W.L., et al. (1973). ]. Amer. Oil Chemists Soc., 50(12):513-15.
64
65
66
FOUR
Carbohydrate/Emulsifier
Interactions
Lynn B. Deffenbaugh
4.1
Introduction
Many different types of emulsifiers are used in food products. The inclusion rate is
usually low, < 3%. Composition and characteristics of emulsifiers used in foods
are discussed in other chapters. Emulsifiers often function indirectly in foods by
modifying the properties of major components such as carbohydrates, protein, or
fat. This chapter discusses the impact of emulsifiers on carbohydrates.
Carbohydrates are a large group of diverse food components. Carbohydrates
can be classified into three categories based on degree of polymerization: monoand disaccharides, oligosaccharides, and polysaccharides. Polysaccharides
are further divided into starch and nonstarch fractions. The nonstarch polysaccharides are plant cell wall materials that contribute dietary fiber to foods.
Mono-, di-, and oligosaccharides and nonstarch polysaccharides (fiber) interact minimally with emulsifiers from the perspective of food-product characteristics. Starches, however, form complexes with emulsifiers that significantly
alter the characteristics that the starches provide in foods. Emulsifiers that
complex with starch are notably different in function and usage from proteincomplexing emulsifiers discussed in the next chapter.
This chapter discusses how complexes form between starches and emulsi-
67
68
4.2
lower levels than with amylose (Hahn and Hood, 1987; Lagendijk and
Pennings, 1970; Twillman and White, 1988).
In general, a complexing agent can be any hydrophobic molecule with a rodlike shape and a diameter less than the 4.5 to 6 A-wide internal core of a starch
helix (Rutschmann and Solms, 1989). Complexing agents include iodine (asP-),
fatty acids, hydrocarbon chains of emulsifiers, and various organic molecules
(i.e., n-butyl alcohol, t-butyl alcohol, dimethyl sulfoxide, and a-naphthol).
Organic aroma compounds have been shown to form thermostable inclusion
complexes with amylose (Maier et al., 1987; Schmidt and Maier, 1987).
Amylose alone exists in solution as a random coil. In the presence of complexing agents, energy minimization drives amylose into a helix configuration
(Neszmelyi et al., 1987). Hydrocarbon chains of fatty acids or hydrophobic
complexing adjuncts are attracted into the hydrogen-lined, hydrophobic core
of an amylose helix (Krog, 1971). Once formed, the complex is stabilized by
dipolar interactions (Mikus et al., 1946) and fulfillment of hydrophobic solvation requirements of the helix (Krog, 1971). Computer-derived models of amylose complexes with stearic acid and iodine have been derived on the basis of
the energy minimization concept (Neszmelyi et al., 1987).
Carbohydrate/Emulsifier Interactions
69
The long axis of complexing agents is internal and parallel to the long axis
of the amylose molecule (Mikus et al., 1946; Rundle and French, 1943a, b).
Complexing agents interchange reversibly, competing for the same space in
the amylose helix (Mikus et al., 1946; Schoch and Williams, 1944). The unit
cell packing size and distance between starch helices are not altered by the
type of complexing agent (Raphaelides and Karkalas, 1988).
Lipid chains usually occupy the helix as paired dimers linked by hydrogen
bonds at the carboxyl groups in fatty acids or glycerol in monoglycerides
(Raphaelides and Karkalas, 1988). The hydrocarbon chains in complexes are so
highly ordered that they have been reported to be crystalline (Carlson et al., 1979).
Amylose helices induced by complexation aggregate into partially crystalline structures that exhibit a V-type x-ray diffraction pattern (Szczodrak and
Pomeranz, 1992) and are insoluble. These insoluble complexes consist of
lamellar crystals with the helix perpendicular to the lamella (Raphaelides and
Karkalas, 1988). Amylose and amylopectin complexes can be differentiated by
their physical properties. Amylopectin emulsifier complexes are more soluble
than amylose complexes. The insolubility of the fatty acid/amylose complex
has long been used as a means to selectively precipitate amylose, but not amylopectin, from solution (Schoch and Williams, 1944). Complexes cannot always be assumed to be insoluble, however, since the degree of insolubility
varies with the surfactant (Kim and Robinson, 1979). The complex between
starch molecules and iodine is used to differentiate and quantitate the amylose
fraction, which forms a blue complex, and the amylopectin fraction, which
forms a red-purple complex.
70
4.3.2
Iodine-Binding Capacity
4.3.3
Starch-Pasting
......
......
Sucrose monoesters
GMS
Sucrose ester
GMS, SSL
EMG, polysorbate 60
MG
CTAB, GMS, SLS
Sucrose esters
Sucrose monoesters
GMS, SSL
GMS, SSL
MG
EMG, polysorbate 60
MG
MG
CTAB, GMS, SLS
Sucrose esters
MG
MG,SSL
MG
Wheat
Potato
Maize, potato, tapioca, wheat
Wheat
Potato amylose
Amylose
Tapioca
Amylose
Wheat
Maize, potato, wheat
Wheat
Potato
Potato amylose
Amylose
Tapioca
Tapioca
Amylose
Nonwaxy
Wheat flour
Nonwaxy
Complexing
agent
Starch
type/fraction
Effect of complexation
on starch properties
Table 4.1
Reference(s)
.....
I()
Sucrose ester
Sucrose monoesters
SSL
SSL
DATEM, MG, SSL
POEMS
MG
DATEM, MG, SSL
Wheat
Wheat
Potato
Wheat
Wheat starch
Wheat
Maize, potato, wheat
Wheat
Wheat flour
Potato
Various
Tapioca
Complexing
agent
Starch
type/fraction
continued
Effect of complexation
on starch properties
Table 4.1
Evans 1986
Ghiasi et al., 1982d
Hoover & Hadziyev, 1981
Bourne et a!., 1960
Ebeler & Walker, 1984
Eliasson, 1985
Eliasson, 1986b
Ghiasi et a!., 1982a,b
Pomeranz et a!., 1969
Rilsom eta!., 1984
van Lonkyhuysen &
Blankestijn, 1974
Deffenbaugh, 1990a
Ebeler & Walker, 1984
Eliasson, 1983
Eliasson, 1986b
Evans, 1986
Favor & Johnston, 1947
Krog, 1971
Krog, 1973
Moorthy, 1985
Eliasson, 1986b
Reference(s)
.....
w
CSL, GMS
MG, SSL
POEMS
MG
MG
MG
CTAB, SDS
Wheat
Potato
Potato
Amylose/amylopectin mixtures
POEMS
Sucrose ester
SSL
Sucrose monoesters
MG
MG
GMS, SLS
Pea flour
Wheat flour
Potato
Masa harina flour
Tapioca
....
continued
Complexing
agent
Starch
type/fraction
Rice
Maize
Amylose
Potato amylose
Maize, potato, tapioca, wheat
Potato amylose
Potato amylose
Reference(s)
CSL =calcium stearoyl-lactyl-2-lactylate; CTAB =cetyltrimethylammonium bromide; DATEM =diacetyl tartaric acid esters of monoglylcerides; EMG =ethoxylated
monoglycerides; GMP =glycerol monopalmitate; GMS =glycerol monostearate; MG =monoglycerides; POEMS =polyoxyethylene stearate; SDS =sodium dodecyl
sulfate; SLS =sodium Iaury! sulfate; SSL =sodium stearoyl-2-lactylate.
Decrease amylopectin
recrystallization
Decreased formulation of
resistant starch
Reduced gel breaking strength
Reduced starch extrudate
solubility and retrogradation
Reduced in vitro enzymolysis
with B-amylase
Reduced in vitro amyloglucosidase
digestion
Reduced in vitro a-amylase
digestion
Decreased glucoamylase
digestibility
Slowed rate of in vivo a-amylase
digestion
Effect of complexation
on starch properties
Table 4.1
.......
Ul
Sucrose monostearate
Sucrose esters
MG
DATEM, MG, SSL
POEMS
GMS, SLS
Sucrose monostearate
MG
CSL, MG
Sucrose esters
Potato amylopectin
Waxy maize
Amylopectin
Waxy maize
Waxy maize
Waxy barley
Potato amylopectin
Amylopectin
Waxy maize
Waxy maize
Sucrose esters
Sucrose monostearate
Waxy maize
Potato amylopectin
Deffenbaugh, 1990a
Bourne et al., 1960
MG
Amylopectin
Reference(s)
Complexing
agent
Starch
type/fraction
Effect of complexation
on starch properties
Table 4.2
CTAB, SDS
CTAB, unsat'd MG
CTAB, SDS
Potato amylopectin
CSL = calcium stearoyl-lactyl-2-lactylate; CTAB = cetyltrimethylammonium bromide; DATEM = diacetyl tartaric acid esters of monoglycerides; GMS = glycerol
monostearate; MG = monoglycerides; POEMS= polyoxyethylene stearate; SDS =sodium dodecyl sulfate; SLS =sodium Iaury! sulfate; SSL =sodium stearoyl-2-lactylate.
Deffenbaugh, 1990a
Sucrose esters
Waxy maize
Reference(s)
Complexing
agent
Starch
type/fraction
continued
Effect of complexation
on starch properties
Table 4.2
Carbohydrate/Emulsifier Interactions
.--..
77
1.0
...::::
()
L..
.....0en
0.8
--Hylon VII
o--oWheat
A - - A Maize
c--o Potato
o--oTapioca
v--vWaxy Maize
0>
0.6
IJ")
...._,
c:
0
0
tO
.....
0
en
0.4
0.2
..0
<(
0.0
T--------v-----~v------------------------------v
2
3
% SE (starch wt. basis)
plied. This relative decrease in viscosity after time and application of shear
defines a peak viscosity.
The process whereby the ordered structure of the starch granule is lost is
called "gelatinization." The transition is a first-order water-mediated melting
of crystalline regions in the starch granule (Donovan, 1979; Zobel, 1984).
Maximum solubilization and swelling occur when excess (> 5 times) water is
present, typical of sauces, gravies, and puddings. In lower moisture systems,
such as baked or extruded products, granule swelling can be limited by moisture content. Extremely high viscosities result because diluent is limited.
Uses and functions of starches are multiplied by modifying properties with
emulsifiers. For example, time of addition and/or time for diffusion of fatty adjuncts into starch granules are processing parameters than can be varied to
produce substantially different properties in cooked starch or cereal grain
products (Lund, 1984). If added before starch gelatinization, monoglycerides
penetrate the granule, form molecular complexes, and reduce starch swelling
power. Addition of monoglycerides after gelatinization enhances granule stability (van Lonkhuysen and Blankestijn, 1974). The function of starch-complexing emulsifiers is primarily due to formation of starch/emulsifier
complexes. The magnitude of the effect of starch/emulsifier complexes on
starch cooking properties varies with starch type, emulsifier type, and processing conditions.
78
Emulsifier and starch molecules must have access to each other to interact.
Starch-complexing emulsifiers must be soluble enough or in a phase that
yields monomers (see Section 4.4.4). Starch molecules have limited availability until gelatinization begins. Emulsifiers adhere to the granule surface before
gelatinization, and they begin to form insoluble starch/emulsifier complexes as
soon as the granule begins to swell and amylose begins to solubilize. These insoluble complexes near the surface stabilize the granule. The rate of further
swelling and amylose leaching are slowed. Gelatinization temperature is increased because more energy is required to cook or swell the stabilized granule. Some emulsifiers, such as polysorbate 60, may cover the starch surface
with a film, increase hydrophobicity, and inhibit water transfer into the granule
(Kim and Walker, 1992c).
The effect of emulsifiers on starch gelatinization is detected in many ways
(Table 4.1). When starch pastes were made with SSL or GMS, changes in viscoelastic properties of the paste were coincident with reduced granule swelling
(Eliasson, 1986b). The granules were less deformable (stiffer), as indicated by
an increase in the temperatures where peaks in storage (G') and loss (G") module were reached. Pasting temperature, hot viscosity, and temperature of peak
viscosity of most normal starches are increased in the presence of emulsifiers
that have a fatty adjunct that can form inclusion complexes with starch.
According to Mitchell and Zillman (1951), the increase in starch paste viscosity in the presence of complexing agents is a result of the increased ability of
granules to absorb and hold water without rupturing.
Gelatinization, the melting of crystalline regions in the granule, causes loss
of birefringence and disappearance of a starch x-ray diffraction pattern
(Eliasson, 1986a). When emulsifiers delay gelatinization, the loss of birefringence is delayed (see Table 4.1).
Some emulsifiers with a fatty acid chain adjunct do not form complexes
with starch. Anionic surfactants, such as sodium lauryl sulfate (SLS) and
sodium dodecyl sulfate (SDS), destabilize the granule because of its highly
negative charge (Eliasson, 1986b; Moorthy, 1985). The destabilization leads to
rapid, complete swelling of the granules, and a rapid increase in viscosity
early in the pasting (cooking) cycle. Loss of granule integrity will subsequently
lead to a dramatic loss in viscosity after peak. This is undesirable if the starch
is being utilized for its thickening, texturizing abilities. In the selection of an
emulsifier for product development applications, charge as well as chemical
composition need to be considered when effect on starch is important.
Carbohydrate/Emulsifier Interactions
4.3.4
79
The formation of a composite starch gel is responsible for the texture and
structure of many food systems. A starch gel formed from a paste is a composite of swollen starch granules embedded in and reinforcing the amylose gel
matrix (Ring, 1985). As a starch paste cools, molecules become less soluble
and aggregate, and a three-dimensional gel forms (Osman, 1967). The consistency increases because molecular associations form a cross-linked network
that increases the resistance of the paste to deformation (Zobel, 1984).
Gelation is initiated by rapid precipitation of amylose in solution, while amylopectin gels much slower and requires higher concentrations. Amylose serves
as (i) the chief material for formation of a gel network to entrap unabsorbed water and (ii) a binding material to link intact or fragmented swollen granules.
In starch gels made with emulsifiers, the insoluble complex produced will
form a gel (Conde-Petit and Escher, 1992). The formation of complexes between amylose and emulsifiers accelerates gelation in the first few hours of aging after gelatinization compared to starch gels made without emulsifiers
(Conde-Petit and Escher, 1994). The gelation of maize, potato, tapioca, and
wheat starch during cooling is responsible for the setback viscosity of profiles
shown in Figure 4.2 (Deffenbaugh, 1990a). A sucrose ester emulsifier increased setback viscosity due to complex formation that accelerated gelation.
Gelation can be induced and controlled by use of emulsifiers in starch-containing foods (Conde-Petit and Escher, 1992).
4.3.5
Retrogradation
The primary reasons for using emulsifiers are modification of finished product
characteristics. Retrogradation in starch-containing foods impairs texture and
taste, and emulsifiers inhibit this retrogradation.
Retrogradation is the formation of ordered, partially crystalline sites within
a cooled starch paste or gel. The recrystallization of starch is a long-term
process that occurs during the hours and weeks after pasting and gelation.
Amylose retrogrades so fast that it is often complete before the food is consumed. Retrogradation of amylopectin occurs more slowly and affects the texture and taste offoods throughout shelf life (Miles et al., 1985).
Emulsifiers decrease the rate of retrogradation of starch gels mainly through
emulsifier/starch-complex formation (Miura et al., 1992). The helix structure of
starch in a complex prevents side-by-side stacking of starch molecules and,
80
200
100
100
- MIn
- l S Sit
-2!1. 5I
pp
gao
Sl
];-
'
5
~
1;-
60
40
Ill
-~
40 .
I5>0CI
B
I 0 IZ
Time (n1in)
H.ot to g~0c
I<
16
18
20
100
1:-
8 60
40
...0
40
20
0
~ 20
0~~-+--~+--l~r-- ~--
I50CI
Heat to
10
IZ
Time (min}
95C
I
14
Cool
16
to
18
20
~C
: h
- w,.,.. .
~: ~~
eo
GO
.'-
~il
16
--
IB
20
c~ lo S0C
100
-1"-H
~~--~-+--~~-+_,
120
-Phl
I:IL $(
0
~ 20
~ 20
0
60
/ - - - - - -
;~\:...
~~/----~
;.\\ \..
//1
\~:~. ]!
1 i:
I i:
,_{.~-!:,'
10
12
Timl!!(mln}
Heat to 95C
14
IG
18
Cool lo .50C
Figure 4.2 Rapid Visco Analyzer viscosity profiles of maize, potato, tapioca, and
wheat starches with 0, 1, 2, or 5% (starch wt basis) of a sucrose ester emulsifier. (From
Deffenbaugh, J990a .)
20
Carbohydrate/Emulsifier Interactions
81
A weak, limited interaction between amylopectin and a sucrose ester emulsifier was observed with various methods (Deffenbaugh, 1990a). IBC and glucoamylase digestion of waxy maize starch were slightly reduced by the
emulsifier. Delay in development of viscosity during gelatinization of waxy
maize starch also suggested that the emulsifier interacted with amylopectin.
Conclusive data describing direct and indirect effects of emulsifiers on amylopectin retrogradation are now available (Gudmundsson and Eliasson, 1990).
Emulsifiers were found to be most effective at directly inhibiting amylopectin retrogradation at very high, near 100%, amylopectin levels. Amylopectin/emulsifier
complexes were responsible for this direct effect. In the presence of amylose,
amylopectin will cocrystallize with amylose, which forms nuclei. When amylose
and amylopectin are present together, ligands will preferentially complex with
amylose. The amylose then cannot co-crystallize with amylopectin, and the emulsifier's effect on amylopectin is indirect. In addition, when the amylose component complexes with emulsifiers, less emulsifier is available to interact with
amylopectin, and direct amylopectin/ emulsifier-complex formation is reduced.
4.3.6
Enzymolysis
Complex formation interferes with the ability of enzymes to digest starch molecules
(see Table 4.1). Kinetics of glucoamylase, which cleaves successive glucose units
from the nonreducing end of starch chains, were not directly affected by a sucrose
ester emulsifier (Deffenbaugh, 1990a). Reduced digestibility of normal starches in
the presence of an emulsifier were due to decreased availability of the starch substrate. Precipitation of the complex and reduced steric compatibility of helical
starch with the active site of the enzyme reduced the starch substrate availability.
Glucoamylase digestion of waxy maize starch was not inhibited by a sucrose ester (Deffenbaugh, 1990a). If a complex formed in the outer branches, it
was not stable enough or strong enough to inhibit glucoamylase.
The inhibition of starch enzymolysis by emulsifiers is analytical evidence of
complex formation. It should be noted however, that emulsifier/starch-complex
formation does not have a significant effect on overall extent of digestibility of
starch in vivo as shown in rats (Holm et al., 1933).
4.4
4.4.1
Adjunct Properties
82
has been reported with 18-carbon saturated fatty acids (Hahn and Hood, 1987)
and 14- to 16-carbon fatty acids on monoglycerides (Hoover and Hadziyev,
1981; Krog, 1971; Lagendijk and Pennings, 1970). Geometrical structure as
well as chain length and molecular weight will influence the ability of the ligand to penetrate the starch helix (Miura et al., 1992). If chain length as well as
helix conformation are known, the fatty acid to amylose ratio required to
achieve saturation of the amylose helix is predictable from stoichiometric relationships (Karkalas and Raphaelides, 1986).
Solubility of the adjunct affects the equilibrium between complex formation
and concentration of the adjunct in solution. The lower solubility of fatty acids
versus monoglycerides in aqueous systems drives fatty acids into the lipophilic
core of the amylose helix more than monoglycerides.
Increased unsaturation in free fatty acids or fatty acid chains of monoglycerides reduces the ability of the adjunct to bind with starch (Hahn and Hood,
1987; Krog, 1971; Lagendijk and Pennings, 1970). The 30 bend at cis bonds
inhibits rotation and causes steric hindrance. In addition, lower solubility of saturated fatty acids will favor complex formation with the hydrophobic helix core.
Steric hindrance from bulky groups on emulsifiers and surfactants interferes
with the ability of the adjuncts to form complexes with starch (Gray and Schoch,
1962). Monoglyceride binding is less than binding of fatty acids of the same
chain length because steric hindrance of the glycerol group interferes with complex formation (Hahn and Hood, 1987). Propylene glycol esters of fatty acids,
with a particularly bulky polar group, have lower complexing ability than emulsifiers with smaller polar groups (Krog, 1971). In addition to steric hindrance,
hydrophilic polar groups are not compatible with the hydrophobic helix core.
The higher the proportion of the polar group to the overall molecule, the less
compatible the molecule is with the hydrophobic interior of the helix.
4.4.2
Starch Granule
The starch granules may also sterically hinder starch complexation with adjuncts in solution. Monoglycerides exist in micelles in solution and attach to
the starch granule surface simply by adsorption at low temperatures (< 50C)
(van Lonkhuysen and Blankestijn, 1974). As the temperature increases, the
monoglycerides penetrate swollen granules to complex with amylose. Other
workers measured strong surfactant/starch complexes at temperatures as low
as 60C, where only slight disorganization (gelatinization) of the starch granule
has occurred (Ghiasi et al., 1982c). The amount of the insoluble, possibly crys-
Carbohydrate/Emulsifier Interactions
83
talline complex, increased when held at 60C for 1 hour. These authors concluded that the surfactants do enter the granule and complex with amylose before gelatinization.
4.4.3
Starch Type
The source of starch in part determines its functionality in foods, and different
starches will be affected by emulsifiers in different ways and to varying extents. GMS restricted swelling of potato starch granules more than it did maize
or wheat starch granules (Eliasson, 1986b). Some methods of analysis, such as
iodine-binding capacity and glucoamylase digestion, are less sensitive to the
unique reaction of different starches in the presence of an emulsifier
(Deffenbaugh, 1990a). Other methods, such as viscoelastic properties of starch
gels (Eliasson, 1986b) and viscosity profiles (Deffenbaugh, 1990a) do differentiate the response of different starches in the presence of emulsifiers.
Viscosity parameters of various starches in the presence of a sucrose ester
emulsifier are shown in Table 4.3 (Deffenbaugh, 1990a). Time to peak viscosity was changed more in tapioca than in maize, wheat, and potato starches. The
emulsifier changed peak viscosity most in potato starch, and changed setback
viscosity most in wheat starch. Potato and tapioca granules were stabilized by
complex formation so that granule swelling and disintegration were more gradual. Moorthy (1985) also reported that starch-complexing emulsifiers stabilized the pasting viscosity of tapioca starch. Viscosity profiles are a
convenient, easy way to study complex interactions between components in
food systems.
4.4.4
Environmental Conditions
84
0% SE
1% SE
2% SE
5% SE
Maize
Potato
Tapioca
Wheat
5.431
3.031
3.67 1
7.32 1
5.962
3.642
4.262
8.082
6.72 3
4.083
7.23 3
8.453
7.724
5.154
8.33 4
8.844
Waxy maize
3.45 1
3.54 1
3.862
4.163
Starch
0% SE
1% SE
2%SE
5% SE
Maize
Potato
Tapioca
101.3223
Wheat
57.9 1
256 1
113.2 1
77.2 2
2322
104.92
74.3 2
2263
99.63
65.93
183.64
78.4 1
80.11
81.2 1
81.61
Waxy maize
88.8
101.62
98.02
89.83
Starch
0% SE
I% SE
2% SE
5% SE
Maize
Potato
Tapioca
Wheat
Waxy maize
55.0 1
83.9 1
61.5 1
78.81 1
50.2 1
86.02
83.9 1
68.12
90.92
51.0 1
93.83
110.32
84.83
129.!3
52.61
97.4 3
I, 2, 3, 4
118.04
166.64
51.31
These numbers indicate significant difference (P < 0.05) within starch type.
dimers at lower pH. pH does not affect binding of monoglycerides because the
carbonyl group of the fatty acid is involved in the ester linkage to glycerol.
Amylose-complexing ability of compounds containing hydrocarbon chains
is significantly affected by the phase behavior of the lipid material (Larsson,
1980). The most effective amylose-complexing emulsifiers have a high degree
of molecular freedom in an aqueous phase and exhibit lyotrophic mesomorphism. Micelles or liposomes of emulsifiers are the most effective polymorphs
for providing lipid monomers, and as a result, are more effective complex formers than lipids in other polymorphic states (Riisom et al., 1984; Eliasson,
1986a). Lysolecithin, a native lipid in cereal starches, complexes readily with
Carbohydrate/Emulsifier Interactions
85
amylose because it exists as micelles in solution with a high equilibrium concentration of monomers readily available for interaction (Takahashi and Seib,
1988). Crumb softeners are added to bread dough as J3-crystal monoglycerides
that convert into an a-crystalline state during baking that readily complex with
starch (Krog, 1981; Krog and Nybo-Jensen, 1970). Blends of emulsifiers that
form an emulsion in solution will be less effective starch-complexing agents
than pure monoglycerides.
4.5.1
X-ray diffraction was one of the first methods used to identify and study inclusion complexes with starch (Mikus et al., 1946). X-ray diffraction provides information on the crystallinity of starches. A clathrate (inclusion) complex is
indicated when powder diffractograms of starch have a so-called V-type pattern.
X-ray diffraction has been used to confirm the presence of inclusion complexes
when an aqueous starch system has been heated in the presence of native lipids
or fatty adjuncts (Biliaderis et al., 1986; Deffenbaugh, 1990a; Eliasson, 1988;
Eliasson and Krog, 1985; Hanna and Lelievre, 1975;
Hoove~
and Hadziyev,
1981; Kugimiya et al., 1980; Osman et al., 1961; Rutschmann and Solms, 1990).
The helical conformation of amylose within the complex has also been shown using x-ray diffraction (Rutschmann and Solms, 1990). X-ray diffraction data does
suggest V-type complex specifically between amylopectin and lipid ligands
(Gudmundsson and Eliasson, 1990). It appeared important that "free" amylopectin was used, whereas amylopectin in waxy maize granules did not complex
(Eliasson and Ljunger, 1988; Evans, 1986; Kugimiya et al., 1980)
X-ray diffraction measurements have indicated that the cell dimension of
the helix is essentially the same for complexes with various emulsifiers containing one fatty acid moiety. Emulsifiers with two or more fatty acids are sterically excluded from the helix and complex formation is inhibited (Osman et
al., 1961). Most V complexes have a pitch of approximately 0.8 nm, indicating
86
that the coils of the helix are in contact with each other (French and Murphy,
1977). Jane and Robyt (1984) studied the lamella-like organization of V-type
complexes and concluded that starch chains were folded such that the chain
axes were perpendicular to the surface of the lamella.
4.5.2
Infrared Spectroscopy
Infrared spectroscopy has confirmed the presence of the carboxyl (Bates and
White 1986; Osman et al., 1961), methyl (Bates and White, 1986), or carbonyl
(Hahnel et al., 1995) group of complexing agent inside the starch helix in a
complex. A positive shift in the carbonyl IR absorption peak for GMS in a
starch complex has been measured (Hahnel et al., 1995). The strength of the
carbonyl group double bond increased because of the additional electron density of the starch complex.
4.5.3
Stable free radical spin probes of fatty acids are used to study the interaction between the fatty acid and starch using electron spin resonance (ESR). Changes in
probe motion, reflected as changes of spectral line shape, occur when the probe
is adsorbed or in a highly viscous medium compared to measurements of the
probe in a solvent. The motion of a spin probe of stearic acid was greatly slowed
in the presence of wheat, high amylose maize, and waxy maize starches (Pearce
et al., 1985, 1987a). Binding was weaker in waxy maize than in the other
starches (Pearce et al., 1987b). The interactive moiety of the probe was shown to
be the fatty acid. Results were similar at room temperature or after heating to
95C and cooling back to room temperature. Binding was thought to occur
throughout the interior of the granule since surface absorption could not account
for the amount of probe bound. The presence of water-facilitated binding, presumably by enhancing solvent penetration into the granule (Nolan et al., 1986;
Pearce et al., 1985). Similar results were reported by Nolan et al. (1986) for
probe binding to maize starch at room temperature. Johnson et al. (1990) also
measured spin probe binding by regular and waxy maize starches at room temperature after heating to 95C. Heating was found to decrease the strength of the
complex formed after cooling, most likely because of altered molecular conformation. Spin probe penetration into the granule increased upon heating, due
both to increased granular surface area and increased accessibility of the interior
of the granule to the fatty acid probe.
Additional ESR analysis indicated that the fatty acid spin probe was slowly
Carbohydrate/Emulsifier Interactions
87
4.5.4
13
4.5.5
Differential scanning calorimetry (DSC) data defines the temperature and enthalpy of thermal transitions. Gelatinization of starch is a water-mediated, endothermic melting phenomenon. Complexes between starches and emulsifiers
form exothermally (i.e., crystallize) during heating of starches with the emulsifier
88
and water (Kugimiya et al., 1980). The net enthalpy of starch gelatinization endotherm is reduced in the presence of external lipids due to simultaneous,
exothermic crystallization of starch/lipid complexes (Biliaderis et al., 1986;
Deffenbaugh, 1990a; Eliasson, 1983; Eliasson, 1986a; Eliasson and Ljunder,
1988; Eliasson et al., 1988; Evans, 1986; Kugimiya et al., 1980). Data is given
in Table 4.4 showing the effect of a sucrose ester emulsifier on gelatinization
temperature and enthalpy of various starches (Deffenbaugh, 1990a). Microscopy
and viscosity profile measurements, discussed previously, also detect a delay in
gelatinization of normal starches in the presence of emulsifiers. Depending on
moisture and emulsifier levels, DSC may not be sensitive enough to detect the
delay in starch gelatinization caused by emulsifiers (Deffenbaugh, 1990a; Kim
and Walker, 1992b). See Table 4.4.
Table 4.4 DSC parameters of starch gelatinization endotherm from
thermograms of starch with 0, I, 2 or 5% (starch wt basis) sucrose
ester emulsifier. (From Deffenbaugh, l990a.)
To
eq
Starch
0% SE
1% SE
2% SE
5% SE
Maize
Potato
Tapioca
Wheat
Waxy maize
66.661
59.741
63.54 1
58.701
69.03 1
66.53 1
59.831
63.97 1
59.101
68.40 1
66.491
59.751
64.03 I
58.42 1
68.401
66.42 1
59.611
63.90 1
59.021
68.13 1
Starch
0% SE
1% SE
2% SE
5% SE
Maize
Potato
Tapioca
Wheat
Waxy maize
72.83 1
64.751
70.191
63.691
72.591
64.75 1
70.64 1
63.72 1
72.69 1
64.891
70.82 1
72.661
64.60 1
70.31 1
74.751
74.171
63.301
74.291
63.671
74.24 1
Tpeq
MI (J/g)
Starch
0% SE
1% SE
2% SE
5% SE
Maize
Potato
Tapioca
Wheat
Waxy maize
13.441
16.931
18.191
10.61 1
16.901
11.5()2
16.641
15.282
9.58 12
17.011
10.612
16.2612
13.773
9.33 2
16.961
10.662
15.372
11.834
8.782
16.83 1
12 3 4
Different numbers indicate significant difference (P- 0.05) within starch type.
Carbohydrate/Emulsifier Interactions
89
~and
Eliasson and Krog, 1985; Raphaelides and Karkalas, 1988; Stute and
Konieczny-Janda, 1983). GMS forms a very stable complex with starch and
has some of the most profound effects on gelatinization properties.
Physical properties of starch/emulsifier complexes will depend on conditions
during crystallization. Multiple-melting endotherms for starch/emulsifier complexes or shifting of endotherms upon rescanning indicates the presence of different crystal polymorphs (Kugimiya et al., 1980; Biliaderis and Galloway, 1989;
Eliasson, 1988; Biliaderis et al., 1986; Bulpin et al., 1982; Paton, 1987). At the
onset of gelatinization, association of the amylose chain with a ligand provides
the conformational order (helicity) for nucleation that precedes crystallization.
Complexation during the first heating may be incomplete due to restricted mobility of amylose during gelatinization (Kugimiya and Donovan, 1981).
Small, imperfect crystals formed during gelatinization may have individual
helical segments distributed randomly without crystallographic register. An
annealing process occurs between Tg, amorphous glass transition temperature,
and Tm, crystallite melting temperature. Partial melting of less stable crystals
allows sufficient chain mobility for growth of more stable crystals. The melt-
90
ing/recrystallization process yields a more dense crystal composed of crystallites embedded in and molecularly continuous with disordered chain segments. Annealing of polymorphs as the temperature increases or during
rescans will increase crystallite melting temperature.
Different polymorphs of starch/emulsifier complexes may also exist within a
larger crystal that has folded onto itself (Eliasson, 1988). Complexes in folds or
at the surfaces of crystals would have a lower melting temperature than complexes further inside the crystal.
4.6
Summary
Acknowledgment
The author acknowledges with great appreciation the assistance of Mrs. Teresa
Booher in preparation of the manuscript.
References
Banks, W., Greenwood, C.T. (1975). Starch and Its Components, Edinburgh University
Press, Edinburgh, Scotland.
Batres, L.V., White, P.J. (1986). Interaction of amylopectin with monoglycerides in
model systems,}. Am. Oil Chem. Soc., 63, 1537-40.
Biliaderis, C.G., Galloway, G. (1989). Crystallization behavior of amylose-V complexes:
structure-property relationships, Carbohydrate Res., 189, 31-48.
Carbohydrate/Emulsifier Interactions
91
92
Carbohydrate/Emulsifier Interactions
93
94
FIVE
Protein/Emulsifier Interactions
Martin Bos, Tommy Nylander, Thomas Arnebrant,
and David C. Clark
5.1
Introduction
Many food emulsions are more complex than the traditional definition of an
emulsion: a colloidal dispersion of liquid droplets in another liquid phase.
This is mainly because the dispersed phase is often partially solidified or the
continuous phase may contain crystalline material, as in ice cream. However,
one characteristic that all emulsions have in common is that they are (thermodynamically) unstable. The four main mechanisms that can be identified in the
process of breaking down an emulsion are creaming, flocculation, coalescence,
and Ostwald ripening. There are two ways in which the process of breakdown
of an emulsion can be influenced. First, use of mechanical devices to control
the size of the dispersion droplets and second, the addition of stabilizing
chemical additives like low molecular weight emulsifiers and polymers to keep
it dispersed. The main purpose of the latter is to prevent the emulsion droplets
from fusing together (coalescence), often achieved by repulsive droplet/droplet
interactions. These interparticle interactions are determined mainly by the
droplet surface, which is coated with emulsifiers, often biologically surface-active components like proteins, mono- and diglycerides, fatty acids, or phospholipids. The forces most commonly observed are electrostatic double layer, van
95
96
der Waals, hydration, hydrophobic, and steric forces. They are responsible for
many emulsion properties including their stability.
The complex mechanisms involved in formation, stabilization, and destabilization of emulsions make fundamental studies on applied systems difficult.
One approach has therefore been to clarify the basic physical and chemical
properties of emulsions by the study of simpler model systems. The adsorption
behavior of single-emulsion components like proteins, fatty acids, surfactants,
or phospholipids at liquid/air or liquid/liquid interfaces have given information about surface activity, adsorbed amounts, kinetics, conformation, and surface rheology. The development of experimental techniques has made it
possible to extend these studies to multicomponent systems. This has provided
further information concerning competitive adsorption, displacement, and
complex formation, which can be related to emulsion and foam stability.
For further information concerning the physicochemical factors affecting
the emulsion structure as well as characterization of food emulsion stability,
the reader is referred to the reviews of Dickinson (1987a; 1988), Dickinson
and Stainsby (1982), and Tadros and Vincent (1983), and for the principles of
emulsion formation to the review of Walstra (1983) along with the other chapters in this book. In this chapter we will focus on the molecular interactions
between proteins and other surface-active components present at the interface
of the emulsion droplets that prevent the droplets from fusing together.
Understanding the interaction between these emulsifier components is the key
to increase the emulsion stability as well as to be able to tailor the structure of
these systems. Various surface-active components like lipids, low molecular
weight (LMW) surfactants, and even phospholipids will be regarded as emulsifiers. We will first discuss the stability of the protein in liquid solutions, which
is an important factor for their behavior in emulsion systems. Although the behavior at liquid/liquid and liquid/air interfaces can be best compared with the
situation in an emulsion or foam, we will also discuss some relevant studies
concerning the solid/liquid interface as well as the effect of emulsifiers on the
solution behavior of proteins. A number of techniques can be used to study
protein/emulsifier interactions, including surface film balance, ellipsometry,
Brewster angle microscopy (BAM), circular dichroism (CD), differential scanning calorimetry (DSC), surface rheology, fluorescence spectroscopy, and neutron reflectivity. It is beyond the scope of this chapter to discuss these
techniques in detail, but when necessary a brief explanation will be given.
The link between the molecular interactions between emulsifier compo-
Protein/Emulsifer Interactions
97
nents and the properties of food emulsions will be discussed in the last section
of this chapter.
98
a mean packing density of 0.74, which is comparable with the values found for
crystals and organic molecules (Richards, 1977). In this densely packed environment, short-range forces like van der Waals interactions and hydrogen
bonding play an important role (Privalov, 1978). In addition, the hydrophobic
interactions contribute significantly in stabilizing the structure, as first pointed
out by Kauzmann (1959). Accumulation of hydrophobic residues in the core of
the molecule leads to a strong folding force. In contrast, the polar groups prefer
the outside of the protein molecule due to their solvent affinity.
Due to the delicate balance of forces mentioned above, the protein is marginally stable (Pace, 1981; Privalov, 1988; Dill, 1990). Changing one amino acid
residue might stabilize or destabilize the protein (Matsumura et al., 1988).
Binding of a lipid, surfactant, or another protein molecule can also have the
same effect. For instance, it has been shown that at low surfactant-to-protein ratios, the binding can stabilize the protein against thermally induced unfolding,
as observed for the interaction between fatty acids or sodium dodecylsulfate
(SDS) and bovine serum albumin (Gumpen et al., 1979), as well as SDS and~
lactoglobulin (Hegg, 1980). On the other hand, increasing the surfactant-to-protein ratio above lO moles of SDS per mole serum albumin or 1 mole of SDS per
mole of J3-lactoglobulin monomer causes unfolding of the protein. Creamer
(1995) observed similar stabilization of ~-lactoglobulin against urea induced unfolding at 1:1 molar ratios between the protein and SDS as well as palmitate. In
many emulsions involving proteins and emulsifiers, the interaction could take
place at the interface. The proximity of an interface might disturb the stability
force balance of the protein, resulting in unfolding of the protein. The unfolding
of proteins upon adsorption is entropically favored (Dill, 1990; Norde, 1986) and
might be the driving force for adsorption where the contact between the hydrophobic domains and the aqueous environment are minimized by proper orientation of the molecule (Haynes and Norde, 1994).
5.3
Protein/Surfactant Interactions
5.3.1
Protein/surfactant interactions of solid surfaces have been studied with the aim
of estimating the protein attachment strength to surfaces, for optimizing detergency processes, and for avoiding undesired adsorption in medical applications.
The major part of the work has been carried out with the purpose of characterizing the protein binding to the surface rather than the protein/surfactant interac-
Protein/Emulsifer Interactions
99
tion. Due to this fact, many studies referenced below are concerned with the degree of removal, or elution, of adsorbed protein by surfactant. We have, however,
included these data in this review and tried to evaluate them in connection with
the sparse data that allow understanding on a molecular level. Even if the data
mainly refer to solid surfaces, the basic principles are also valid at liquid interfaces such as those of the emulsion droplet. Since the process of surfactant interaction with proteins at interfaces is determined by the surfactant/protein, the
surfactant/surface and protein/surface interactions, the following brief introduction is intended to provide a background on surfactant association and adsorption, and on surfactant/protein interactions in solution.
The polarized structure of surfactants derives from the hydrophilic head
group and a hydrophobic, usually hydrocarbon, part that promotes interaction
with each other, other molecules, and surfaces by the hydrophilic head as well
as the hydrophobic moieties. These interactions can be expected to be of electrostatic or polar, steric, and hydrophobic type, and the total interaction is
often a sum of or a balance between two or more of these forces. As a consequence of the strong tendency for the hydrophobic chains to avoid contact with
water, self-association will take place both in solution and at interfaces (see
Section 5.4.4). The monomer concentration in solution will be strongly dependent on the association pattern and thus have a pronounced effect on the interfacial behavior (Lindman and Wennerstrom, 1980; Israelachvili, 1992).
The general features of surfactant adsorption are
l. At high surfactant concentrations [around the critical micelle concentra-
100
0.05
"'su
'bn
0.04
0.03
6
0
~ooooosoo8~
~
<
0.02
"a
Q)
Ul
"a
0.01
<:t;
0.00
1800
Time (s)
_Q
3600
iv
ii
c
Figure 5.2 A schematic illustration of an isotherm for adsorption of ionic surfactants
to hydrophilic surfaces (Somasundaran and Kunjappu, 1989). Different stages of the
isotherm are labeled (I) to (IV), and schematic drawings of possible structures that exist
in these regions are presented in Figure 5.3. (Reprinted with kind permission of the
American Chemical Society, Washington, DC.)
Protein/Emulsifer Interactions
101
association behavior of surfactants in the different regions found in the adsorption isotherm, as shown in Fig 5.2. The upper drawings correspond to a hydrophilic surface, the lower to a hydrophobic surface.
5.3.1.1
et al., 1991) in investigations into the adsorption strength of blood plasma proteins, particularly fibrinogen. They introduced the term "elutability" to describe
the degree of removal. The surfactant elutability of proteins is affected by factors
that are known to influence the binding strength of a protein to a surface. Thus,
surfactant elutability has been found to decrease with conditions favoring conformational changes, i. e., decreasing protein concentration (Rapoza and Horbett,
1989, 1990a), increasing temperature (Bohnert and Horbett, 1986; Rapoza and
Horbett, 1990a), time of adsorption or "residence time" (Rapoza and Horbett,
1989; Ertel et al., 1991), and decreasing stability of the protein (McGuire et al.,
1995a, b; Wahlgren et al., 1993). However, surfactant elutability will be influenced not only by protein properties but also by the type of surfactant (WelinKlintstrom et al., 1993; Wahlgren et al., 1993a, b; Wahlgren and Amebrant,
1991) and surface (Wahlgren et al., 1993, 1994; Elwing and Colander, 1990), as
further discussed next.
11-111
Figure 5.3
IVa
ivb
102
Protein/Emulsifer Interactions
103
formation between surfactant and protein is involved in the removal mechanism of proteins from hydrophilic surfaces. In the case of hydrophobic surfaces, the removal processes of protein by the different surfactants, including
non-micelle-forming ones, are in general more similar than for the hydrophilic
surfaces. This might be expected, due to the different orientation of the surfactant, and suggests a replacement mechanism, due to higher surface activity of
the surfactant (Wahlgren, 1992). Blomberg and coworkers used the interferometric surface force technique (lsraelachvili and Adams, 1978) to study the
interaction between lysozyme adsorbed on mica and SDSo (sodium dodecane
sulfonate) and SDS. They found that SDSo, which has a Krafft temperature
above room temperature and hence does not form micelles, had a minor effect
on the interaction between adsorbed lysozyme layers on mica (Tilton et al.,
1993), and from the small change in surface potential, they concluded that few
surfactant molecules were bound to the adsorbed protein. SDS showed a similar low binding to lysozyme on mica at low concentrations (up to 0.5 erne) but
caused a collective desorption of the protein at the erne of the surfactant, indicating that the cac to adsorbed lysozyme is in the range of its self-association
limit in solution (erne) (Blomberg, 1992). These studies show that anionic surfactants bind to an adsorbed layer of lysozyme, which is almost neutral after
binding of the positively net-charged protein to the negative mica surface. The
binding of surfactant thus leads to an increased negative charge of the layer,
which in the case of SDS finally leads to desorption of the protein. It is likely
that this is due to electrostatic repulsion between the negatively charged surface and the protein/surfactant complexes.
Nonionic surfactants are generally found to be ineffective in removing protein from hydrophilic surfaces (Wahlgren, 1992; Elwing et al., 1989). As mentioned above, these surfactants bind to a very low extent to protein in solution
(except when specific binding sites or pockets are present) and to the proteincovered surface. At hydrophobic surfaces, however, they usually have a considerable effect (Elwing et al., 1989; Wannerberger et al., 1994). This was
elegantly demonstrated in a study of surfactant interactions with proteins adsorbed at a surface with a gradient in wettability (Elwing et al., 1989).
The effect of chain length of alkyltrimethylammonium surfactants on the
elutability of fibrinogen at concentrations above their erne was found to be
small at both silica and methylated silica surfaces (Wahlgren, 1992). Rapoza
and Horbett (1988, 1990a) did not find any effects of chain length of sodium
alkyl sulfates on the elutability of fibrinogen and albumin down to a chain
104
length of 6 carbon atoms. However, they found, as expected, that the chain
length did influence the surfactant concentration at which the onset of protein
removal was initiated. The trend was similar to the one observed for the onset
of other cooperative binding events (e.g., micelle formation).
Rapoza and Horhett (1990h) found that surfactants with large head groups
such as Tween 20 gave rise to lower fibrinogen elutahility levels than other surfactants at polyethylene surfaces. Welin-Klintstrom et al. (1993) found that the
elutahility of fibrinogen adsorbed at surfaces with a wettahility gradient decreased with the bulkiness of the hydrophobic part of the surfactant. In this connection it was also found that nonionics showed an increased removal of
fibrinogen into the more hydrophilic region of the gradient surface when the
cloud point (phase separation temperature) was approached (Wahlgren et al.,
1995). These general observations of removal efficiency are in line with the findings of Backstrom and coworkers (Backstrom, 1987; Lindman et al., 1988), who
studied the removal of fat by different surfactants and found a maximum at conditions corresponding to an optimum in the packing of surfactant molecules at a
flat interface and those of Malmsten and Lindman (1989), who investigated,
among other variables, the effect of temperature on cleaning of hard surfaces.
Thus, it may be concluded that at high surfactant concentrations, head group
effects are, as expected, most pronounced at hydrophilic surfaces hut less important at hydrophobic ones. In addition, it appears that principles for detergency in
general, involving the packing efficiency of molecules at interfaces, are applicable to qualitatively describe the removal of proteins from the surface.
Protein/Emulsifer Interactions
105
toglobulin and lysozyme decreased roughly in the order silica > chromium oxide > nickel oxide (Wahlgren and Arnebrant, 1991). The similarity between
the behavior of the two oppositely charged surfactants indicates that the removability of protein in these cases mainly reflects the binding mode of the
protein to the surface.
Elwing et al. (1989, 1990) studied the surfactant elutability of proteins adsorbed to a surface containing a gradient in hydrophobicity and found large
differences in the amounts removed from the hydrophilic and hydrophobic
ends. In the case of a nonionic surfactant (Tween 20), the elutability was
largest at the midpoint of the gradient, which can be attributed to enhanced
conformational changes of the adsorbed protein at the hydrophobic end, in
combination with a lower efficiency of removal by nonionics at hydrophilic
surfaces. At hydrophobic surfaces the removal is generally high (Elwing et al.,
1989; Wannerberger et al., 1994). However, this may not be considered as evidence for weak binding of the proteins to the surface, but rather as an indication of the strong interaction between the surfactants and surface.
Simple models.
can be classified into two types of behavior, a short description of which follows (see Figure 5.4):
l. Complex formation between surfactant and protein that leads to desorption of protein from the surface. In this case the surfactant does not have
to adsorb at the surface, but it has to interact with the adsorbed protein.
2. Replacement of protein by surfactant. This implies that the interaction
between the surfactant and the surface has to be stronger than the interaction between the protein or the surfactant/protein complex and the
surface. Adsorption of the surfactant to the surface is essential in this
connection, but binding of surfactant to protein may not be required.
Of course, the surfactant might bind to the surface and/or to the adsorbed
protein without any net effect on the amount of protein adsorbed. A partial removal according to mechanisms (1) or (2) as illustrated in Figure 5.4 may be
the most common observation and has previously been suggested as one indication of the presence of multiple adsorption states of the protein (Horbett and
Brash, 1987).
It is important to note that surfactant can remove proteins without binding to
the surface [case (1), left side of Figure 5.4]. This could be described as a solubi-
106
Proteins and
Protein/Emulsifer Interactions
107
Jones and Wilkinson, 1976; Coke et al., 1990: Frapin et al., 1990;
O'Neill and Kinsella, 1987; Kresheck et al., 1977; Clark et al., 1992;
Creamer, 1995), serum albumin (Tanford, 1980; Nozaki et al., 1974;
Brown, 1984; Ericsson and Hegg, 1985), and specific lipid-binding proteins such as puroindoline from wheat (Wilde et al., 1993).
2. Cooperative adsorption of the surfactant to the protein without gross conformational changes.
3. Cooperative binding to the protein followed by conformational changes
(Few et al., 1955; Subramanian et al., 1986; Su and Jirgensons, 1977;
Nelson, 1971).
These different interactions can occur in the same system upon increasing the
surfactant concentration. The conformational changes that occur in case (3)
can involve changes in secondary structure (Nozaki et al., 1974; Subramanian
et al., 1986; Su and Jirgensons, 1977). Several models for the protein/surfactant complexes have been suggested, e.g.: rigid rod (Reynolds and Tanford,
1970), pearl and necklace ( Shirahama et al., 1974), and flexible helix model
(Lundahl, 1986). In the cooperative region (2 and 3), above the critical association concentration (cac ), the interaction is mainly one of hydrophobic character (Tanford, 1980; Subramanian et al., 1977; Nelson, 1970).
The adsorption from surfactant/protein mixtures to hydrophobic solid surfaces is to some extent analogous to the adsorption at air/water or oiVwater interfaces, which have been the subject of frequent studies (Courthaudon et al.,
1991a, b; Clark et al., 1994). Competitive adsorption between proteins and
surfactants at these interfaces has recently been reviewed by Dickinson and
Woskett (1989). They conclude that, as expected, small surface-active components above a certain critical concentration will dominate over proteins at
these interfaces, since such components normally have higher surface activity
(superiority in lowering interfacial tension).
Experimentally it is observed that the presence of surfactants in protein solutions may influence the amount of proteins adsorbed to solid surfaces in
three different ways (Wahlgren 1992; Wahlgren and Arnebrant, 1991, 1992;
Wahlgren et al., 1993, 1995):
108
The complete lack of adsorption in case (1) could be explained by complex formation with a surfactant resulting in an entity that has no attraction to the surface
or direct/preferred surfactant adsorption, due to their higher surface activity and
diffusivity, unpending adsorption of proteins, or protein/surfactant complexes. In
cases (2) and (3), the formation of complexes in solution leads to a decrease or increase in the amounts of protein adsorbed, respectively. The presence of surfactant
influences the total amount of protein adsorbed by steric effects or changes in the
electrostatic interaction between complexes as opposed to native protein. In addition, the complex may adsorb in a different orientation than the pure protein, resulting in a positive or negative effect on the adsorbed amount.
Due to the different shapes of the adsorption isotherms of surfactants and proteins, the interaction with the interface is of course, as for protein mixtures,
strongly dependent on the relative concentration of the components. The special
character of the surfactant adsorption isotherms featuring the sharp increase in adsorbed amount in the range of their critical association concentration will influence
these events in a very pronounced way. Studies regarding these surfactant/protein
"Vroman effects" have been reported; for example, adsorption of fibrinogen from
mixtures containing Triton X-100 passes through a maximum (Slack and Horbett,
1988). The adsorption from ~-lactoglobulin/SDS mixtures at different degrees of
dilution was studied by Wahlgren and Amebrant (1992) (see Figure 5.5). At concentrations above the erne for the surfactant, the amount adsorbed corresponded to
a layer of pure surfactant and was found to increase after rinsing. At lower concentrations, the adsorbate prior to rinsing appeared to be a mixture of protein and surfactant, and the total amount adsorbed passes through a maximum. The
composition of the adsorbate after rinsing is most likely pure ~-lactoglobulin, since
interactions between the surface or protein and SDS are reversible.
At high degrees of dilution of the mixture, the absence of surfactant adsorption to the methylated silica, the nonreversible adsorption of ~-lactoglobulin,
and the observed partial desorption of the adsorbate from the mixture imply
that some SDS molecules are bound to ~-lactoglobulin molecules with a higher
affinity than to the surface (see Figure 5.5). The amount of protein adsorbed is
larger, even after rinsing, than for adsorption from pure ~-lactoglobulin solutions, and it can be concluded that SDS binding in this case facilitates the adsorption of protein.
Protein/Emulsifer Interactions
109
0.18
......-: --
..-...: .-- .
0.12
0.06
0
1000
100
Degree of dilution
10
Figure 5.5 The amounts adsorbed to a methylated silica surlace as a function of degree of dilution for a mixture of ~-lactoglobulin and SDS (0.2 w/w), in phosphate
buffered saline pH 7, I= 0 .17. The figure shows the adsorbed amount (!lg/cm2) after 30
minutes of adsorption (e) and 30 minutes after rinsing(+). In addition, the figure shows
the adsorption of pure ~-lactoglobulin, after 30 minutes of adsorption () and 30 minutes after rinsing (x). Finally, the adsorption isotherm of SDS is inserted (0). (From
Wahlgren and Arnebrant, 1992.)
factant surface interactions and surfactant/protein binding. Solubilization requires complex formation between protein and surfactant, and replacement requires adsorption of the surfactant to the surface. As for protein adsorption, one of
the most important protein properties affecting surfactant-induced desorption appears to be conformational stability. Differences between a competitive situation
and addition of surfactant after protein adsorption may derive from the alteration
in surface activity of protein/surfactant complexes formed in solution compared to
pure protein, the difference in diffusivity of surfactants and proteins affecting the
"race for the interface," and time-dependent conformational changes resulting in
"residence time" effects.
The effects observed at low surfactant concentration, below the erne, mvolve the specific binding of surfactant to some proteins and are not fully understood. Further, the exact prerequisites for solubilization versus replacement
as well as detailed information on molecular parameters such as aggregation
numbers are not known.
110
two distinct mechanisms; the one that dominates is dependent upon the molecular composition of the interface (Clark, 1995). Low molecular weight surfactants such as food emulsifiers or polar lipids congregate at the interface and
form a fluid-adsorbed layer at temperatures above their transition temperature
(see Figure 5.6a). When a surfactant-stabilized thin film is stretched, local
thinning can occur in the thin film. This is accompanied by the generation of a
surface-tension gradient across the locally thin region. Surface tension is highest at the thinnest point of the stretched film, due to the local decrease in the
surface concentration of emulsifier in the region of the stretch. Equilibrium
surface tension is restored by adsorption of surfactant from the interlamellar
liquid, which is of very limited volume in a drained thin film. This process is
called the "Gibbs effect." Alternatively, migration of the surfactant by lateral
diffusion in the adsorbed layer toward the region of highest surface tension
Protein/Emulsifer Interactions
111
may also occur (Clark et al. 1990a). Here, the surfactant drags interlamellar
liquid associated with the surfactant head group into the thin region of the film
and contributes to the restoration of equilibrium film thickness. This process is
often referred to as the "Marangoni effect" (Ewers and Sutherland, (1952).
In contrast, the adsorbed layer in protein-stabilized thin films is much
stiffer and often has viscoelastic properties (Castle et al., 1987). These derive
from the protein/protein interactions that form in the adsorbed layer (see
Figure 5.6b). These interactions result in the formation of a gel-like adsorbed
layer, recently referred to as a "protein-skin" (Prins et al., 1995), in which lateral diffusion of molecules in the adsorbed layer is inhibited (Clark et al.,
l990b). Multilayer formation can also occur and serves to further mechanically strengthen the adsorbed layer (Coke et al., 1990). When pure protein
(o) Syrfnctnru-alonc
(b) Pmtein-olpne
h igh mobility
Film sttetchiog
~
diffusion~
~Prot
~ cin
rapid
dcfonnntion
/
no int.emct.i:ons
Pilm stretching
No Protein
defonnation
Slow diffusion
P'lm rui l r e.
Figure 5.6 Schematic diagram showing the possible mechanisms of thin-film stabilization. (a) The Marangoni mechanism in surfactant films; (b) the viscoelastic mechanism in protein-stabilized films; (c) instability in mixed component films. The thin films
are shown in cross section and the aqueous interlamellar phase is shaded. (Reprinted
with kind permission of the Royal Society of Chemistry, London.)
112
films are stretched, the change in interfacial area is dissipated across the film,
due to the cohesive nature of the adsorbed protein layer and possibly the deformability of the adsorbed protein molecules.
Thin-film instability can result in systems that contain mixtures of proteins
and low molecular weight surfactants (Coke et al. 1990; Clark et al., 1991a;
Sarker et al., 1995), as is the case in many foods. The origin of this instability
rests in the incompatibility of the two stabilization mechanisms: the Marangoni
mechanism relying on lateral diffusion, and the viscoelastic mechanism on immobilization of the protein molecules that constitute the adsorbed layer. One
can speculate that in a mixed system, competitive adsorption of low molecular
weight surfactant could weaken or interfere with the formation of protein/protein interactions in the adsorbed layer and destroy the integrity and viscoelastic properties of the adsorbed layer (see Figure 5.6c). This could be a
progressive process, with the presence of small quantities of adsorbed surfactant initially introducing faults or weaknesses in the protein film. Adsorption
of more surfactant could induce the formation of protein "islands" in the adsorbed layer. These structures could be capable of slow lateral diffusion but
would be too large to participate in Marangoni-type stabilization. Indeed, they
could impede surfactant migration in the adsorbed layer. Adsorption of progressively more surfactant would reduce the size of the protein aggregates still
further until the adsorbed protein was in its monomeric form. Ultimately, all
the protein would be displaced from the interface by the surfactant.
Two types of interaction are shown in the schematic diagram of the mixed system. First, there is an interactive process associated with the coadsorption or
competitive adsorption of the two different species at the interface. Second, many
of the functional proteins used in food production have physiological transport activities and therefore possess binding sites, which may allow the formation of
complexes with surfactants. Let us consider each of these processes in tum. The
transition in adsorbed-layer structure at the air/water and oil/water interface
caused by competitive adsorption between protein and emulsifiers has been studied in detail. Oscillatory surface shear (Kragel et al., 1995) and dilation (Clark et
al., 1993: Sarker et al., 1995) measurements have been carried out at the air/water interface and show an emulsifier-induced reduction in surface shear viscosity
and surface dilational elasticity at critical emulsifier/protein ratios. This is consistent with the cartoon depicted in Figure 5.6, where at a specific molar ratio the
emulsifier will break down protein/protein interactions in the adsorbed layer, resulting in a reduction in surface shear viscosity and dilational elasticity. Similar
Protein/Emulsifer Interactions
113
observations have been made at the oil/water interlace under conditions of constant shear in experiments where both components were present in solution when
the interlace was formed (Courthaudon et al., 1991) and when the competing surfactant was added subsequent to formation of the protein-adsorbed layer (Chen
and Dickinson, 1995). Direct measurements of changes in adsorbed-layer rheological properties in foam and emulsion films is possible using the fluorescence
recovery after photobleaching (FRAP) technique (Clark et al., 1990a; Clark,
1995). This method relies on the positioning of a fluorescent reporter molecule in
the adsorbed layer either by covalent labeling of the protein of interest with a fluorescent moiety such as fluorescein isothiocyanate (FITC) or by use of an amphipathic fluorescent molecule [e.g., 5-N-(octadecanoyl)amino fluorescein, ODAF] at
low (submicromolar) concentrations that are insufficient to alter adsorbed-layer
structure. FRAP allows direct measurement of the lateral diffusion coefficient of
the fluorophore in the adsorbed layer of the thin film. The presence of an extensive network of protein/protein interactions in the adsorbed layers arrests lateral
diffusion of the probe molecule, as is the case in all films stabilized by protein
alone (Clark et al., 1990b). This situation persists in the presence of emulsifier
until a critical ratio of emulsifier/protein is achieved. This is marked by the onset
of surlace lateral diffusion in the adsorbed layer. The technique serves as a useful
means of (i) comparing the resistance of different proteins to emulsifier-induced
adsorbed layer disruption, (ii) evaluating the effectiveness of protein modification
strategies at improving the resistance of proteins to competitive displacement,
and (iii) investigating the usefulness of natural food ingredients as crosslinkers of
proteins in the adsorbed layer.
5.3.2.2
114
Tween 20
L-a lysophosphatidylcholine, palmitoyl
Sucrose monolaurate
Sucrose monostearate
Sucrose monooleate
Sodium stearoyl
lactylate, pH 7.0
Sodium stearoyl
lactylate, pH 5.0
Lauric acid
Palmitic acid
Dissociation
constant
Reference
4.6 j.iM
166.J.!M
11.6 j.iM
1.02 j.iM
24.8 j.iM
0.26 j.iM
0.30 j.iM
Clark, unpublished
0.7 j.iM
0.1 j.iM
The general features described earlier are evident with a comparatively low
concentration of protein causing a significant reduction in y. In the absence of
protein, yreduces gradually with increasing Tween 20 concentration. The gradient of the reduction in surface tension reduces at higher Tween 20 concentrations (> 30 J1M) but doesn't become completely flat due to failure to attain
equilibrium y, possibly due to the presence of a mixture of surface-active species
in the Tween-20 sample. In contrast, the curve in the presence of protein maintains a relatively steady surface-tension value of about 50 mN/m up to Tween-20
concentrations of 25 j.iM due to adsorption of the protein. This means that the
curve for the sample containing protein crosses that of Tween 20 alone. This is
strong evidence for complex formation between the two components, since the
curves cross due to a reduction in the concentration of free emulsifier in solution
due to that which interacts with the protein to form the complex.
Thus, great care must be taken when considering the surface properties of
Protein/Emulsifer Interactions
115
80
70
5
~
60
u.;0
~
Q)
~
Q)
<..>
50
$....
;::l
r:n.
40
30
20
40
60
80
100
Figure 5.7 Surface tension isotherm for Tween 20 in the absence (e) and presence
() of 0.2 mg/mL ~-lactoglobulin. The data were recorded after 20 minutes adsorption
and are therefore not at equilibrium.
(l)
[P] [E]
[PE]
(2)
116
[P] = [P101]
[PE]
(3)
[E] = [E101]
[PE]
(4)
where [P101] and [E101] are the total protein and emulsifier m the system.
Substituting (3) and (4) in (2) gives
which can be solved for [PE] and can be used to calculate the relative concentrations of the three components. In addition, the binding data, which may
comprise a change in a parameter (e.g., intrinsic fluorescence) caused by formation of the complex may be fitted using this equation, provided there is a
single active binding site and the titration is carried out to saturation.
Alternatively, it is possible to determine the dissociation constant and number
of binding sites from the Scatchard (1949) equation
[E]
(6)
where v is the fraction of protein with occupied sites (i.e., [PE]/[P101]). If the
Scatchard plot of v against v/[E] gives a straight line, it indicates the presence
of only one class of binding site. The gradient of this line is -l!Kd, and the intercept on the xaxis gives the number of binding sites, n. If the Scatchard plot
does not give a straight line, then the shape of the curve obtained can be used
to identify if the observed binding is positively or negatively cooperative or the
presence of multiple independent sites. In the former case the Hill equation
can be used to determine the Kd and a cooperativity coefficient (Hill, 1910).
Evidence for the formation of a specific complex in solution by direct measurement or by crossovers in surface-tension/concentration isotherms is not
sufficient to allow the conclusion that intact complex adsorbs directly at the
air/water or oil/water interface. There are few studies that provide convincing
data that support such a hypothesis, and that which is provided often only indirectly points toward the presence of adsorbed complex at the interface. One
of the best understood systems is that of Tween 20 and ~-lactoglobulin (Coke
Protein/Emulsifer Interactions
117
et al, 1990; Wilde and Clark, 1993), which are known to interact in solution
to form a 1:1 complex characterized by a Kd = 4.6 ~' which has an increased hydrodynamic radius of 5. 7 nm compared to 3.5 nm for P-lactoglobulin alone (Clark et al., 1991b). Detailed measurements of the properties of
foam films formed from a constant concentration of 0.2 mg/mL mixed native
and fluorescein-labeled P-lactoglobulin as a function of increasing Tween-20
concentration (Wilde and Clark, 1993; Clark, 1995) have been reported. This
study revealed that between molar ratios (R) of Tween 20 to P-lactoglobulin of
0.2 to 0.9, there was a progressive increase in the thickness of the foam films
and a corresponding decrease in the amount of adsorbed protein to an intermediate level of approximately 50% of that which was originally adsorbed
(see Figure 5.8). These changes occurred prior to the onset of surface diffusion of the labeled protein as determined by the FRAP technique at R = 0.9
(Coke et al., 1990). The increase in foam-film thickness was unexpected since
protein displacement by the Tween 20 should have reduced the thickness of
the thin film. One persuasive interpretation of the data is that coadsorption or
trapping of the Tween-20/P-lactoglobulin complex in the adsorbed multilayers could account for adsorbed-layer thickening (Clark et al., 1994), since the
complex is known to have an increased hydrodynamic radius (Clark et al.,
1991b). Calculations reveal that this is a distinct possibility, since 16 to 49%
of the P-lactoglobulin present in solution will be in the complexed form in the
R-value range, 0.2 to 0.9. Further confirmation of this explanation comes from
measurements of foam-film thickness at different P-lactoglobulin concentrations but at constant R value. Film thickness data for P-lactoglobulin at 0.2
and 1.0 mg/mL are also shown in Figure 5.8. The completely different thicknesses observed at the two protein concentrations can be rationalized in terms
of the amount of Tween-20/P-lactoglobulin complex formed. The sharp reduction in film thickness observed in the 0.2 mg/mL P-lactoglobulin sample at R
= 0.9 occurs when there is 5.41-LM complex present in the sample. An equiv-
~-lactoglobulin
118
40
0
0
--,._,
_..,
C l)
;::S
0
30
OJ
43
...
;::S
CIJ
-E
Cl)
Cl)
OJ
20
];a
u
~
4
Figure 5.8 A comparison of foam-film thickness and surface concentration data for ~
lactoglobulin samples as a function of Tween 20 concentration. Surface concentration
of FITC/~-lactoglobulin (0.2 mg/mL) as determined by fluorescence counts (.&); foamfilm thickness for samples containing 0.2 mg/mL () and l.O mg/mL (e) ~-lactoglobu
lin. R refers to the molar ratio of Tween 20: ~-lactoglobulin.
moidal behavior hut was shifted to increased surfactant concentration by approximately 2 orders of magnitude compared to the static measurements.
Inclusion of ~-lactoglobulin (0.4 mg/mL) in the initial solutions caused only a
small reduction in the measured
Ydyn
the presence of Tween 20 up to a concentration of 15 f.!M. Above this concentration a small hut significant further reduction in
Ydyn
fect resulted in a small inflection in the Ydyn curve in the region corresponding
to 15 to 40 J.LM Tween 20. At higher Tween-20 concentrations, the curve for
the mixed system followed that of Tween 20 alone. The inflection in the
Ydyn
Ydyn
isotherm indicates that there is very little difference between the sur-
Protein/Emulsifer Interactions
119
face activity of the three components present in solution, Tween 20, ~-lac
toglobulin, and the complex, since
'Ydyn
ies of the binding of LPC to the a form revealed that the binding became
tighter with increasing acyl chain length, and higher concentrations of the
short-chain-length LPC are needed to achieve optimal foam stability enhancement. One interesting observation was that the micellar form of LPC
was the species that bound to the protein. This finding emerged from the observation that lauryl-LPC showed no interaction with the puroindoline-a until
the levels present exceeded the critical micelle concentration of 400
120
5.4
Protein/Phospholipid Interactions
Introduction. In this section we will discuss the interaction between phospholipids and proteins. Both proteins and phospholipids are major components
of biological membranes, and therefore it is not surprising that a vast number
of publications concerning protein/phospholipid interactions are related to understanding the biomembrane processes. This aspect has recently been reviewed in a book by Watts (1993).
In pharmaceuticals, phospholipids are often used as colloidal drug carriers
in drug delivery systems in various physical forms like liposomes or emulsions. These drug delivery systems are cleared from the circulation (bloodstream) by the reticuloendothelial system. It is believed that this clearance is
triggered by adsorption of serum proteins on the phospholipid interface and
will depend on the properties of both the protein and the phospholipid (Eidem
and Speiser, 1989; Tabata and lkada, 1990; Patel, 1992).
In untreated milk the fat globule is stabilized by a lipid membrane, composed
mainly of sphingo- and phospholipids. Homogenization of milk results in a total
area increase of the fat globules by some 6- to 10-fold, where the increased surface
is stabilized by proteins adsorbed from milk during the process. Phospholipids,
particularly phosphatidyl-choline Oecithin), are added to various processed foods,
where they act as emulsifiers alone or together with proteins. Therefore, proteins
and phospholipids, separately as well as in complexed form, contribute significantly to the physical properties of many systems of technological interest (e.g.,
emulsions and foams). The intention of this section is to emphasize the diverse nature of phospholipid/protein interaction and to point to some implications of this for
the physicochemical as well as the biological properties of phosholipidlprotein systems. It is also our intention to point at some of the major forces involved in phospholipid/protein interactions. We will highlight only some properties of importance
for the understanding of phospholipid/protein interactions. Therefore the classification of interactions made in this section, when the lipid is dispersed in solution,
at different interfaces and with lipid phases, is arbitrary. Assignment according to
polar/nonpolar, anionic/cationic, or soluble/insoluble phospholipids could also
have been possible.
5.4.1
Many proteins have the biological role of transporting molecules with hydrophobic properties, which are bound to a hydrophobic pocket in the protein. ~
Lactoglobulin is thought to transport retinol (Papiz, 1986; Sawyer, 1987; North,
Protein/Emulsifer Interactions
121
1989) but has also been shown to have high affinity for phospholipids, fatty
acids, and triglycerides (Diaz de Villegas, 1987; Creamer, 1995; Sarker et al.,
1995; Kristensen, 1995). An important application of lipid/protein interactions
was reported by Kurihara and Katsuragi (1993), who found that a lipid/protein
complex, formed between ~-lactoglobulin and phosphatidic acid, could mask
bitter taste. This property was suggested to be specific for phosphatidic acid
since no effect was observed for mixtures of ~-lactoglobulin and phosphatidylcholine, triglycerides, and diglycerides. Differential scanning calorimetry measurements confirm the presence of a specific interaction between phosphatidic
acid and ~-lactoglobulin since the presence of distearoylphosphatidic acid
(DSPA) as well as dipalmitoylphosphatidic acid (DPPA) thermally stabilized the
protein, which was not observed when the protein was mixed with phosphatidylcholine, phosphatidylethanolamine, or phosphatidylglycerol (Kristensen et al.,
1995). No interaction could be observed if the lipid contained unsaturated fatty
acid residues or if it was mixed in the gel state with the protein. Thus, the results
show that the interactions between ~-lactoglobulin and phospholipids are
strongly dependent on the acyl chain as well as the head group. This is not simply a question of having a negatively charged head group since no interaction
was observed for phosphatidylglycerol.
The influence of protein structure on lipid/protein interactions has been
demonstrated by Brown et al. (1983), who observed that native ~-lactoglobulin
is unable to bind to phosphatidylcholine vesicles. However, if the protein was
dissolved in a-helix-forming solvent, binding to the phospholipid was observed. Brown suggested that the acyl chains of the lipid interact with the hydrophobic interior of the a helix, while the polar head group is likely to
interact with the hydrophilic exterior of the protein (Brown, 1984). The partially unfolded proteins formed during food processing may give helix structures when interacting with the lipids and these lipid/protein complexes can
improve the emulsification process (Brown, 1984; de Wit, 1989).
5.4.2
A few studies have demonstrated the use of deposited phospholipid layers, deposited via the Langmuir-Blodgett technique or by spincoating, to follow the
interaction between proteins and phospholipids in situ by using ellipsometry
(Malmsten, 1994, 1995; Corselet al., 1986; Kop et al., 1984). This approach
can also be used to analyze the kinetics of interaction (Corsel et al., 1986; Kop
et al., 1984). The work of Malmsten (1994, 1995) showed that the interaction
122
of human serum albumin, lgG, and fibronectin from human plasma with phospholipids spin-coated onto methylated silica surfaces depends on the phospholipid head group. He found no interaction of proteins with phospholipids
that have no net charge or shielded charges like phosphatidylcholine, phosphatidylethanolamine, sphingomyelin, and phosphatidylinositol, whereas interaction was observed with the phospholipid surfaces containing unprotected
charges like phosphatidic acid, diphosphatidylglycerol, and phosphatidylserine. No differences in adsorption behavior were found for spin-coated surfaces
and Langmuir-Blodgett-deposited phospholipid layers.
The consequences of nonspecific interactions with negatively charged dioleoylphosphatidylserine (DOPS) bilayzrs observed for fibrinogen and albumin
in contrast to the specific ones observed for prothrombin have been demonstrated by Corsel et al. (1986). By analyzing their data in terms of intrinsic
binding and transport rate, they found that the initial rate of adsorption of prothrombin was transport-limited, whereas the rate-determining step for the albumin interaction was the binding. The adsorption rate of fibrinogen was either
transport-limited or controlled by the binding, depending on pH and ionic
strength. Since the DOPS bilayer contained biological binding sites for prothrombin, the interaction was completely reversible. However, the interaction
of fibrinogen and albumin with the bilayer probably induced conformational
changes of the proteins and as a consequence the interaction was irreversible.
Kop et al. (1984) found that a high density of DOPS in the bilayer was needed
for the high-affinity binding of prothrombin. Consequently, an introduction of
dioleoylphosphatidylcholine (DOPC) into the bilayer markedly decreased the
affinity for binding.
5.4.3
> ~-lactoglobulin > ~-casein, suggesting that the interaction was of electrostatic nature. The work of Quinn and Dawson (1969a, b) concerning the inter-
Protein/Emulsifer Interactions
123
action between cytochrome c (positive net charge below pH 10) and phospholipids from egg yolk also suggest that it is determined by electrostatics. Their
results show that the limiting pressures for penetration are 20 and 24 mN/m
for phosphatidylcholine and phosphatidylethanolamine, respectively, whereas
penetration into the phosphatidic acid and diphosphatidylglycerol (cardiolipin) monolayers occurred up to pressures (< 40 mN/m) close to the collapse pressure of the film. Furthermore, the presence of sodium chloride
decreased the interaction. Later, Kozarac et al. (1988) confirmed the findings
of Quinn and Dawson with their reflection spectroscopy results. Cornell
(1982) observed a specific interaction between ~-lactoglobulin and egg yolk
phosphatidic acid (e-PA) in spread mixed films at low pH (1.3 and 4) where
~-lactoglobulin carries a positive net charge. No interaction was observed in
the neutral pH range or for egg yolk phosphatidyl choline, (e-PC). ~-lac
toglobulin, adsorbing from solution into a spread monolayers of palmitoyloleoylphosphatidylcholine (POPC) and palmitoyloleoylphosphatidylglycerol
(POPG), was found to interact with the lipids only in the acid pH range
(Cornell and Patterson, 1989). The highest amounts of ~-lactoglobulin
(Cornell and Patterson, 1989), a-lactalbumin, or BSA (Cornell et al., 1990)
bound to mixed monolayers of POPC and POPG were observed below the isoelectric point of the protein, when the lipid layer and the proteins carry an opposite net charge, whereas less was adsorbed around and almost nothing
above the isoelectric point. The interaction was also found to be reduced in
the presence of calcium as well as if sodium chloride was added (Cornell et
al., 1990). Bos and Nylander (1995) used the film balance to study the incorporaffon of ~-lactoglobulin into monolayers of distearoylphosphatidic acid
(DSPA), distearoylphosphatidylcholine (DSPC), and dipalmitoylphosphatidic
acid (DPPA) and some of their results are shown in Figure 5.9. The highest
rate of incorporation was observed for ~-lactoglobulin into the negatively
charged DSPA monolayer. The rate also increases with ionic strength of the
subphase, which probably is due to a decrease of the repulsion within the
phosphatidic acid protein monolayer. The incorporation into the zwitterionic
DSPC monolayers is, as expected, less salt-dependent. The importance of
electrostatic interactions for the incorporation of proteins and peptides in
lipid layers have also been reported for a number of non-food-related systems
like adsorption of SecA (Breukink et al., 1992), actin (Grimard et al., 1993),
IgG (Lu and Wei, 1993), and opioid peptides and opiate drugs (Bourhim et
al., 1993) into phospholipid monolayers.
124
DSPC
. I
''I'
, I
'I
DPPA
-3 [t
(j)
-3.5
~
-4
0_0
-4.5
0
10
15
20
25
30
ri (mN/m)
Figure 5.9 The rate of incorporation of P-lactoglobulin into monolayers of distearoylphosphatidic acid (DSPA), distearoylphosphatidylcholine {DSPC), and dipalmitoylphosphatidic acid (DPPA) versus surface pressure (II). The data were recorded at
constant surface pressure by measuring the area increase of the lipid monolayer
spread on a protein solution containing 1.15 mg/L in 10 11M phosphate buffer pH 7,
with 0 11M (e), 50 11M (), or 150 11M (A) sodium chloride. The rate in mg/m 2 was
calculated from the area increase using the IT-area isotherm of spread monolayers of
P-lactoglobulin. Data adopted from Bos and Nylander (1995), where the experimental
details are also given.
Protein/Emulsifer Interactions
125
5.4.3.1
properties of the protein film (Graham and Philips, 1979; Mitchell, 1986;
Dickinson and Stainsby, 1982). Circular dichroism spectra of ~-lactoglobulin
bound to phospholipid monolayers were found to be similar to those recorded in
solution, indicating that the conformation of the protein did not change significantly when interacting with the lipid monolayer (Cornell and Patterson, 1989;
Cornell et al., 1990). Katona et al. (1978) showed that the~ form of a hydrophobic
myelin protein was more able to interact with sphingomyelin than the <X form, indicating that the phospholipid is able to recognize different conformations of the
protein. Furthermore, the physical state of the phospholipid phase is important for
the interaction between protein and lipid. Fidelio et al. (1986) reported a more favorable interaction between melittin and phospholipids when the lipid film was in
a more liquid expanded state. This might be related to the fact that the protein is
able to penetrate into the phospholipid layer (more hydrophobic interactions).
Interaction (adsorption) of proteins with lipids might also occur at higher surface
pressures, where the lipids are in the condensed or crystalline phase. The protein
then is not able to intercalate the phospholipid monolayer, but might bind to it
forming a second layer, like actin does on negatively charged liposomes through
electrostatic interactions (Grimard et al., 1993).
Another interesting point for discussion is the homogeneity of the mixed
phospholipid/protein film. The homogeneity of mixed protein/lipid films has
been studied by, for instance, electron microscopy of films transferred to a solid
support by Cornell and Carroll (1985). They found that only lipids with the
chains in liquid state, e-PA, dioleoylphosphatidylcholine, and dioleoylphosphatidylethanolamine, formed homogenous films with ~-lactoglobulin, whereas
DPPA and DSPC formed heterogeneous layers. Cornell and Patterson (1989) and
Cornell et al. (1990) observed penetration of ~-lactoglobulin, a-lactalbumin, or
126
BSA into mixed monolayers of POPC and POPG at such high lipid pressure that
they found it unlikely that the proteins could penetrate into a protein layer of
their own at this pressure. Thus, they concluded that the formation of pure protein patches is unlikely and that portions of the protein is suggested to be intercalated in the lipid monolayer. Fluorescence microscopy together with the
surface-film balance technique has also been used to study the structure of
mixed phospholipid cytochrome c and b films (Heckl et al., 1987). It was found
that proteins were located mainly in the fluid membrane phase that coexisted
with solid lipid domains without protein. The penetration in the lipid monolayer
was reduced with increasing pressure. Cytochrome c (positively charged) was
found to interact with dimyristoylphosphatidic acid (DMPA) mono layers but not
with dipalmitoylphosphatidylcholine (DPPC) layers, from which it was concluded that the interaction was largely of electrostatic nature. The effect of different proteins on the organization of lipid monolayer has been demonstrated by
Aynie et al. (1992). Their results indicate that the IX8ccasein in contrast to ~-lac
toglobulin and ~-casein, probably due to the stronger lipid/protein interaction,
induce an ordering of a monolayer of nitroxide fatty acids on the surface of an
emulsion droplet.
5.4.4
As discussed previously, the interaction between surfactants and proteins usually takes place via monomers in the specific binding regime, whereas the cooperative association usually takes place in the range of the erne. However, in
the case of lipids with low aqueous solubility, the association structures are
generally already present when the lipids are mixed with the protein. Thus, the
formation of these structures has a profound impact on protein/lipid interactions. It is therefore important to be familiar with the phase behavior of the
particular lipid under investigation and often seemingly conflicting results can
be derived from differences in the phase structure of the lipids. For instance,
we have observed that the interaction between ~-lactoglobulin and phosphatidic acid occurred only when the lipids were present as a dispersion, but not
when they were mixed in the gel state with the protein (Kristensen et al.,
1995). Hence, we will briefly discuss lipid phase behavior, although a more
detailed description is given elsewhere (see Larsson, 1994).
5.4.4.1
Polar Lipid Phase Behavior. The polar lipids, having large charged or
uncharged polar groups, have an amphiphilic nature and will thus associate in
Protein/Emulsifer Interactions
127
aqueous systems. The common feature for this self-association is the formation
of a polar interface, which separates the hydrocarbon and water regions. An
overview of some of the main types of structures is given in Figure 5.10. The
hydrocarbon chains can exist either in a fluid state, as in liquid crystalline
phases, or in a solid state, as in the lipid gel phases (Larsson, 1994).
Polar lipids can be further divided into two classes on the basis of their interaction with water:
128
Reversed micelles
(L2)
(Solid)
(Melt)
<-Cubic
Reversed hexagonal
(HII)
i=
<1::
a:
f-
w
u
"Water-in-oil"
<-Cubic (Q)
f-
<1::
f-
Lamellar
(La)
I"Mirror plane" I
<1::
LL.
a:
::l
(/)
"Oil-in-water"
<-Cubic
(/)
<1::
w
a:
u
Hexagonal
(Hi)
<Cubic
(H20)
Micelles
(L1)
Figure 5.10 Commonly formed association structures by polar lipids. Phase transitions can be induced by changes in water content, temperature, or interaction with
other solution components, like proteins. The lamellar liquid crystalline phase (LJ ,
can be regarded as the mirror plane, where the aggregates are of the "oil-in-water" type
on the water-rich side and of the "water-in-oil" type on the water-poor side (Fontell,
1992). On both the water-rich and water-poor sides of the L" there are two possible locations for cubic phases. Other "intermediate phases" may also occur.
Rilfors, 1989; Fontell, 1992). One type of cubic structure, consisting of rodlike aggregates, has been proposed by Luzatti et al. (1968). The two polar rodlike networks formed in this way give two continuous and unconnected
systems of fluid hydrocarbon chains. The other main type of cubic phases, observed for water-insoluble lipids, are bicontinuous and based on curved nonintersecting lipid bilayers, which are organized to form two unconnected
continuous systems of water channels (see Lindblom et al., 1979; Larsson,
1989). If an interface is placed in the gap between the methyl end groups of
Protein!Emulsifer Interactions
129
the lipid, it will form a plane that can be described as a minimal smface
(Andersson et al., 1988; Larsson, 1989). A minimal surface has zero average
curvature at any point on the surface; that is, in all points the surface is as concave as it is convex. A minimal surface exhibiting periodicity, as in the cubic
lipid aqueous phase, is termed an "infinite periodic minimal surface" (IPMS),
and three types of IPMS, with different geometry, have been shown to be important in lipid systems (Andersson et al., 1988; Larsson, 1989).
With decreasing water content, the phase behavior of the polar lipids often
follows this sequence: hexagonal phase (HJ ~ lamellar phase (LJ for watersoluble lipids and lamellar phase (LJ ~ reversed hexagonal phase (H1J for
lipids with low water solubility. Cubic liquid crystalline phases (Q) often occur
in between these. Phase transitions can also occur with changes in temperature; with increasing temperature the sequence of thermal transitions is usually the same as with decreased water content.
In nature and in many technical applications the aggregates consist of a
mixture of different lipids, which either exist in a homogenous mixture or separate into domains. As discussed in the review by Raudino (1995), the lateral
distribution in these mixed aggregates is influenced by a number of factors like
ionic strength, presence of polymers/proteins, as well as the composition of the
lipids; thus it is hard to give any general rules to predict when phase separation will occur.
Some Aspects of the Interaction between Proteins and Lipid Structures.
Interactions between proteins and lipid structures are of fundamental importance
in biological membranes, as well as in food emulsions. Our aim is to give some ex-
5.4.4.2
amples of the effects the interaction can have on the lipid phase behavior as well
as on the protein structure. The same type of molecular forces, as discussed in
previous sections, are responsible for the interaction.
Much of the work on lipid phase transitions induced by proteins and peptides has been focused on intrinsic and peripheral membrane proteins. Many
of these proteins carry a positive net charge at neutral pH, and most biological
membranes are neutral, zwitterionic, or negative under physiological conditions (Sankaram and Marsh, 1993). Binding of these proteins generally takes
place via direct electrostatic interaction between basic residues, like lysine
and arginine, and the opposite charges on the lipids. However, other effects
like hydrophobic interaction and steric factors can contribute significantly.
Gramicidin, an uncharged and hydrophobic pentadecapeptide, favors the tran-
130
and
monoolein in the cubic phase shows that the transition of cubic to inverted
hexagonal phase occurred at a much lower temperature at high protein content
than without protein being present (Razumas et al., 1995).
The NMR studies by Spooner and Watts (1991a, b) show that the interaction between cardiolipin bilayers and cytochrome c appears to cause extensive
unfolding of the protein. Similar observations have been made by Heimburg et
al. (1991), who reported that cytochrome c binds to bilayers of phosphatidylglycerol and mixtures between neutral and anionic lipids in two forms:
It was observed that in the fluid state of pure DMPG and DOPG, the bound
protein existed in the more unfolded form (II), whereas in the gel state of
DMPG the native form (I) prevailed. When DOPG, was mixed with nonionic
(DOG) or zwitterionic (DOPC), the bound fraction of the native form (I) increased with the content of neutral lipids.
The work of Minami et al. (1995), in which incorporation of P-lactoglobulin
and P-lactalbumin with negative net charges and lysozyme with positive net
charge at neutral pH, into a sphingomyelin/palmitate gel phase, was studied
found no correlation with the protein net charge. Instead, the amount of protein, which could be dissolved in the thin aqueous layer of the gel phase was
Protein/Emulsifer Interactions
131
suggested to be limited by the dimension of the layer. This is likely to be reduced as a consequence of the osmotic stress exerted by the "outside" solution
phase at high enough protein concentration (Minami et al., 1995).
The lateral phase separation of vesicle bilayers, containing a mixture of
phosphatidic acid and phosphatidylcholine, in the presence of lysozyme has
been followed by differential scanning calorimetry (Raudino and Castelli,
1992). The protein/lipid interaction, which was shown to be dependent on
electrostatic forces, thermally stabilized the protein. As discussed by Raudino,
the lateral phase separation occurring when protein binds to heterogeneous hilayers can eventually lead to the formation of defective lamellar phases, socalled ribbon phases, where the broken lamellae are limited by curved rims
(Raudino, 1995).
It has been shown that a number of proteins can be entrapped in the cubic
monoolein/water phase (Ericsson et al., 1983; Razumas et al., 1994; Razumas
et al., 1995). Not only can enzymes up to 590 kD be entrapped in the cubic
phase but also their activity is retained for a longer time compared to when
they are in bulk solution (Razumas et al., 1994).
How will the structure of the cubic phase be affected by the entrapment of a
protein? It seems that properties like viscoelasticity, microscopic appearance,
and the nature of x-ray diffraction pattern are basically the same as for a pure
monoolein/water phase (Ericsson et al., 1983; Portmann et al., 1991; Landau
and Luisi, 1993; Razumas et al., 1994; Razumas et al., 1995). However, there
are some important differences, indicating that the amount of protein as well
as the amount of water determines the phase behavior (Ericsson, 1986). The
study of Razumas et al. (1995) of the cubic monoolein/cytochrome c aqueous
phase also confirms this and points to the influence of electrolyte present.
Differential scanning calorimetry and enzyme activity measurements show that
lysozyme retains its conformation and activity in the cubic phase (Ericsson et
al., 1983). Similarly, it has been observed by circular dichroism measurements
that bacteriorhodopsin and melittin (Landau and Luisi, 1993) as well as achymokypsin (Portmann et al., 1991) retain their native conformation. On the
other hand, differential calorimetry data, and other observations on cytochrome c entrapped in the cubic phase, suggested some interactions that affect the thermal stability of the protein as well as of the cubic phase (Razumas
et al., 1995). Razumas et al. (1994) used a cubic monoolein-aqueous-phasecontaining enzyme as the biocatalytic layer in amperometric and potentiometric biosensors. They observed large differences in the stability, which
132
decreased in the order lactate oxidase > creatinine deiminase > glucose oxidase > urease, and corresponded to the order of increasing molecular weight.
Further investigations have shown that the introduction oflecithin in the cubic
phase can increase the stability of entrapped glucose oxidase (Nylander et al.,
1995). The mobility of the entrapped enzyme is limited in the cubic phase
compared to bulk solution (1995), whereas the diffusion measurements of
Mattisson et al. (1995) show that a typical substrate molecule, like glucose,
can move relatively freely into the cubic phase. These studies point to an important potential application of cubic lipid phases as a flexible and biocompatible matrix for the immobilization of enzymes for synthesis as well as for
analytical applications. The cubic monoglyceride phases also have the ability
to solubilize lipophilic proteins as well as relatively large amounts of membrane lipids. Larsson and Lindblom (1982) have reported the solubilization of
a-gliadin, which is a hydrophobic wheat protein fraction, into a cubic 1monoolein aqueous phase, with the same characteristics as a binary cubic 1monoolein phase. In this case, the proteins most probably are dispersed in the
lipid bilayer region of the cubic phase.
5.5.1
The action of lipolytic enzymes is of importance in a number of food applications or related areas, ranging from their use in detergents, as tools in modifying lipids for the breakdown of acylglycerides both as unwanted side effects, to
the naturally occurring process in the human intestine. It is well known that lipases work mainly at an interface and thus they are an important example of
Protein/Emulsifer Interactions
133
lipid/protein interactions at interfaces. Excellent reviews regarding lipase action have been written by Verger and Pattus (1982) and Pieroni et al. (1990),
but we will highlight only some aspects in relation to the mechanisms of protein/lipid interactions. There are several types of lipases that act on phospholipids and triglycerides, but we will mainly discuss lipases catalyzing the
hydrolysis of the ester bonds of triacylglycerols. The enzymatic activity is determined by the concentration of lipolytic enzymes associated with the lipid
film and can be inhibited by various proteins (Gargouri et al., 1984).
Experiments carried out with mixed protein/dicaprin films transferred over
pure buffer yielded evidence that inhibition of hydrolysis was caused by proteins bound to the dicaprin film rather than by a direct interaction between
protein and lipase in the bulk phase (Gargouri et al., 1985). Furthermore,
since some lipases were inhibited by adsorption of proteins at the lipid layer,
whereas other lipases were still able to hydrolyze a mixed protein/phospholipid layer, indicating that the inhibition of some lipases cannot be attributed
merely to steric effects hindering accessibility to dicaprin molecules within
the film. Surface concentration measurements of inhibitory proteins showed
that only 5 to 9% of the area of a mixed lipid/protein film was covered by inhibitory proteins, implying that long-range electrostatic forces are likely to be
involved in the inhibition as well as parameters such as surface viscosity and
surface potential. However, similar inhibitory effects caused by melittin (pi >
10) and 13-Iactoglobulin A (pi = 5.2) at pH 8.0 strongly suggest that the nature
134
5.5.2
The formation of the interfacial layer on the emulsion droplet is strongly linked
to the properties of the aqueous environment as well as to the properties of the
oil phase. An example of the former has been provided by Chen and Dickinson
(1995a-c), who showed that the addition of an anionic surfactant, sodium lauryl sulfate (SLES), can introduce flocculation of a gelatin stabilized oil-in-water emulsion at sufficient surfactant concentrations. An increase of the SLES
concentration led to a restahilization of the flocculated emulsion. This could he
related to the hulk behavior, where the anionic surfactant will neutralize the
positively charged gelatin and cause precipitation of the protein. As more surfactant is added, the solubility of the protein increases due to binding of surfactant that increases the protein net charge, and finally the precipitate is
redissolved. Measurements of the composition of the interfacial layer reveal
(Chen and Dickinson, 1995c) that gelatin is initially replaced by SLES, hut as
the surfactant concentration increases, more gelatin can he accommodated at
the interface due to partial neutralization of the gelatin at the interface. This
will eventually cause flocculation to occur (Chen and Dickinson, 1995a). A
further increase of surfactant concentration leads to a decrease in gelatin surface concentration (Chen and Dickinson, 1995c) and a restabilization of the
emulsion (Chen and Dickinson, 1995a). It is noteworthy that an emulsion stabilized by P-lactoglohulin, which was negatively charged under the prevailing
experimental conditions, did not show any signs of flocculation upon addition
of SLES, although a complex was formed in the hulk solution.
Several examples of how the properties of the oil phase composition can affect the structure of the adsorbed layer of protein on the emulsion droplet, and
hence the stability of the emulsion, have been studied. For instance, the work
of Leaver and Dagleish (1992) on the structure of adsorbed layers of ~-casein
on emulsion droplets, where it was found that the cleavage of the protein on
the oil-droplet surface by trypsin gave different products depending on
whether a triglyceride oil or tetradecane, was used. This demonstrates that the
structure of the adsorbed layer depends on the composition of the oil.
Furthermore, it was shown by Heertje et al. (1990) that if a monoglyceride was
dissolved in the oil phase, the displacement of caseinate from the oil/water interface was correlated to the interfacial concentration of monoglyceride. They
also found that at high monoglyceride concentration in the oil phase, the
amount of saturated (monostearoylglycerol) lipid adsorbed at the interface was
Protein/Emulsifer Interactions
135
larger than that of the unsaturated (monooleoylglycerol) one, which also led to
more extensive displacement of caseinate.
We have in the foregoing section mainly been concerned with the binding of
emulsifiers to proteins, but one can also think of cases where there is a repulsion between the lipid and head group and the protein. In this way the structure of the adsorbed protein layer, and hence the properties of the emulsion,
can be changed. This has been demonstrated by Brookbanks et al. (1993), who
found that the thickness of adsorbed layers of milk proteins was significantly
greater when the protein was adsorbed onto negatively charged phosphatidylglycerol (PG) liposomes. The more extended structure was attributed to charge
repulsion between the negatively charged lipid surface and the negatively
charged surface domains of the protein.
5.5.3
Conclusion
136
Protein/Emu1sifer Interactions
137
Acknowledgement
Thomas Arnebrant thanks the Swedish Research Council for Engineering
Sciences (TFR) and Martin Bos acknowledges the EU Human Capital and
Mobility program (ERBCHRXT930322) for their financial support. Tommy
Nylander acknowledges the Swedish Council for Forestry and Agricultural
Research. David Clark acknowledges the support of the Biotechnology and
Biological Sciences Research Council.
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SIX
Physicochemical Aspects of
an Emulsifier Functionality
Bjorn Bergenstahl
6.1
Introduction
2. The lifetimes of bubbles are increased. (Only very pure water displays a
very short lifetime, a few seconds, of bubbles created by shaking.
Normal standard "pure water," double distilled, usually displays a bubble lifetime of about 20 to 30 seconds.)
3. The emulsifiability of oils in water is enhanced. Smaller drops with a
longer lifetime are formed with less stirring.
4. The aggregation rate of dispersed particles is changed. Surface-active
additives may induce or prevent flocculation of disperions.
5. The sediment volume of settling particles is influenced. Surface additives inducing adhesive may create a loose or compact sediment.
6. Crystallization properties are changed. This may include crystallization
rate and crystal shape.
147
148
This chapter aims to discuss the principal physical origin of the various functionalities of typical lipid food emulsifiers. Aspects on the functionality under
very different conditions in various foods will be discussed. I will try to show how
we may select emulsifiers on the basis of their fundamental properties.
6.2
Surface Activity
When an additive is added to a solution, the gain of entropy is very large at low
concentrations. If the additive displays surface activity and adsorbs at an interface, the system loses entropy, which has to be balanced by a gain in free
energy due to the adsorption. At very low concentrations the solubility always
prevails, but when the concentration is increased, more and more of the available surfaces will be covered by the adsorbed molecules. To display surface
activity, the emulsifier needs to have certain properties:
1. It has to form a noncrystalline form t in contact with water.
2. It should have a reduced solubility in water due to a large hydrophobic part.
3. It has to interact with water through polar interactions.
4. It should have a significant molecular weight to reduce the effect of the reduced entropy when it adsorbs.
5. It has to have a reduced solubility in an oil environment due to large size and
the presence of polar groups at the interface.
High-melting emulsifiers do not display surface activity when dispersed in water until a critical temperature, the Krafft temperature, has been reached. At this
temperature the emulsifier solubility in the solution has reached a sufficient concentration to allow for a significant formation of adsorbed layers at the interfaces.
The presence of hydrophobic parts of the molecules increases the energy
gain due to adsorption. In aqueous environments most emulsifiers tend to aggregate in micelles at a critical concentration, erne (critical micelle concentration), or to precipitate as liquid crystals. Above the aggregation concentration
all properties depending on the chemical potential, for instance the adsorption
properties, are more or less constant. The aggregation is mainly driven by the
presence of the hydrophobic parts of the molecules (Tanford, 1973).
t Several lipid emulsifiers are exceptions and are applied in a hydrated gel form (a crystals).
However, this crystal form resembles the liquid crystalline form in terms of interactions with
both phases and spreadability over the interfaces.
149
A polar part of the molecule is necessary to avoid the formation of a separate oil phase. The type of aggregates formed during the adsorption will reflect
the balance between the polar part and the hydrophobic part of the molecule.
The free-energy gain at adsorption is mainly proportional to the molecular
weight, while the entropy loss due to the demixing is independent of molecular
weight. Hence, small molecules, for instance lower alcohols, do not form adsorbed layers at hydrophobic surfaces in contact with water solutions, while
pronounced layers are formed with additives of higher molecular weights, for
instance monoglycerides. Proteins display a much higher surface activity than
protein hydrolysates.
In an oil environment, solvophobic effects are absent and the adsorption
has to be generated by polar interactions between the second phase and the
surface-active molecule.
The interaction between particles is influenced when the particles are covered by an adsorbed layer of an emulsifier. The change in the interaction
strongly influences the macroscopic properties of the dispersions (Table 6.1).
Table 6.1
Interaction
Stability
Sedimentation
Attraction
Repulsion
Flocculation
Stable
The solution properties of emulsifiers are determined for the surface activity of the emulsifiers. In addition, the ability to generate repulsive interactions
is also reflected in the solution properties of emulsifiers.
6.3.
150
reversed
micelles
reversed
hexagonal
lamellar phase
phase
Melting point
M---
hexagonal
phase
,
:micelles
'
'
Krafft
point
Emulsifier
L\Gphase transformation
.L\.Gmixing
where .ilGmixing is negative when changing from large aggregates to small aggregates (micelles and molecular solutions).
is positive and equal to Ahydrocarbonlwater 'Y hydrocarbon/water The hydrophobic effect is the driving force for the aggregation and gives the upper
ilGhydrophobic
molecules:
L l I
next aggregate
LlGpolar group/water
F(l) dl
all neighbors
aggregate
where l is the average distance between the polar groups and F(l) is the interaction.
The area per molecule in the aggregates is given by the balance between
the interfacial tension of the oil/water interface and of the space needed for the
151
polar group itself and the space generated by repulsive interactions between
the emulsifier head groups at the interface.
The area per molecule expands in the series Areversed micelles < Areversed hexagonal
phase~
micellar
solution~
lamellar phase
A2)
enforces
different geometries (Israelachvili et al., 1976, 1977) due to the different ratio
of volume to area of different aggregates, as shown in Table 6.2.
The successive solvation of surfactants in Table 6.2 correspond to a successive change into aggregates that correspond to a more long-range interaction. If there is an upper limit for the repulsion, the solvation series is
terminated at that stage. Hence, the maximum solvated aggregate formed at a
surplus of water is a measure of the ability of the emulsifier to generate repulsive interactions.
The area of the molecule is a measure of the interaction when water is
available, and may be generalized as the hydrophilicity of the molecule. The
spatial requirement of the hydrophobic part of the molecule is of course a measure of the hydrophobicity of the molecule. Consequently, there is a close link
with the classical view of emulsifiers as molecules with a balance between the
hydrophobic and the hydrophilic properties, as they are expressed in the HLB
numbers, proposed by Griffin (1949, 1979).
152
Table 6.2
Area/volume
Sphere (micellar solution)
2
2 1tr
(4/3)1tr
= 3Ao
3 Vhydrophoh
rhydrophob
2r1tl
n. /z
2 vhydrophob
rhydrophob
2A
Vhydrophoh _
rhydrophob
rhydrophob
Ao
A
t The packing constraint is here defined as the necessary cross section of an amphiphilic molecule
in the aggregate at the oil/water interface. This definition is Aofpacking parameter according to
Israelachvili et al. (1992, 1976, 1977).
153
p- Xylene
Micellar solution and
oil phase
lamellar micellar solution
and oil phase
Water
Nonylphenoi-E09
154
talline phase coating the droplet surface reduces the van der Waal's attraction
and that this was an important contribution to the observed effects in the emulsification experiments (Friberg, 1971). However, this explanation is not a useful general explanation since the emulsifier concentration in optimized food
emulsions rarely is high enough to allow for multilayer adsorption (Walstra,
1988; Dickinson, 1986). Obviously, this observation is contradictive to the
need for a separate phase of liquid-crystalline material around the droplet.
However, a correlation between the presence of, or the possibility to form, liquid-crystalline phases and emulsion stability is still experimentally observed
in several systems. To stabilize an dispersion, the emulsifier should:
Stabilizing property
aggregates
Micelles
Bilayers
Reversed
Water-continuous emulsions
Repulsive interactions
Interfacial viscosity
Anchoring
Optimal
Weak Optimal
Too water-soluble
Intermediate
Weak
Optimal
Weak
Acceptable
Oil-continuous emulsions
Repulsive interactions
Interfacial viscosity
Anchoring
Weak
Weak
Acceptable
Intermediate
Optimal
Optimal
Optimal
Weak
Too oil-soluble
155
6.5.2
Lecithins
Lecithin is one of the most commonly used food emulsifiers, and its popularity
can be expected to grow even further due to its natural origin. Technical
'"_
0..
s
.5
";:;'"
"'"0..
:n">
..
c
ccccc
c
c c
"
0..
Lamellar phase 60 C
c Gel phase 23 C
c
00
:;
-1
0
10
15
dw(A)
Figure 6.3 The hydration repulsion between bilayers of monopalmitin in the liquidcrystalline and gel states. (Redrawn from Pezron et al., 1991.)
156
Table 6.4
Liquid-crystalline
phases formed at
Upper swelling
limit (at 25C)
Distilled saturated
C18-16
50%
Krog, 1990
Distilled unsaturated
Cl8:l-2
35%
Krog, 1990
Monoolein
C18:1
40%
Krog, 1990
Emulsifier
Monoglycerides:
Tetraglycerolesters:
Tetraglycerol
monolaurin
C12
55%
Krog, 1990t
C16-l8
Lamellar 45C
55%
Krog, 1990
40%
60%
Krog, 1990
Krog, 1990
Sodium steraoyllactylate:
pHS
pH7
C18
C18
Sorbitan esters:
Polyoxyethylene (20) C18:1
sorbitan monooleate
Polyoxyethylene (20) C18
sorbitan monostearate
Sorbitan stearate
C18
157
properties of the most common phosphatides and then discuss the properties of
various mixtures.
6.5.3
Phosphatidylcholine
6.5.4
Phosphatidylethanolamine
200
Reversed
Hexagonal
~ 150
8 100
~
50
Cubic
Crystalline
phase
Water
Figure 6.4.
Lamellar
phase
Dioleoylphosphatidylcholine
158
90
G
~
c.
E
~
[J
70
[J
50
[J
30
10
-10
10
12
14
16
18
20
22
Chain length
Figure 6.5 The main transition temperature for phosphatidylcholine (PC) () and
phosphatidylethanolamine (PE) (D) as a function of chain length. The sources of data
are given in Table 6.4.
6.5.5
Phosphatidylinositol
The phase diagram of soybean PI and water has been determined by the author
(1991) and by Soderberg (1990). The diagram is characterized by a large
lamellar phase with an unlimited swelling. The liquid-crystalline phase is
formed below room temperature.
6.5.6
Phosphatidic Acid
The phase diagram of the sodium salt of dioleoylphosphatidic acid has been determined by Lindblom et al. (1991). The phase diagram is characterized by a
lamellar phase that transforms to a reversed hexagonal phase at about 30% of
water. This transformation occurs although there is an ionic charge on the molecules and despite the small head group. A possible explanation, supported by
evidence from NMR measurements, is that this is due to ion condensation.
6.5.7
Lysophosphatides
The phase diagrams of a series of different lysophosphatides has been investigated by Arvidsson et al. (1985). Lysophosphatidylcholine has the same hydrophilic polar group as the ordinary PC but only one of the two fatty acids.
This reduces the volume demand of the aggregate, and the packing constraint
allows for the formation of micelles and hexagonal phases.
6.5.8
159
Table 6.5
Phospholipid
Fatty
acids
Liquid-crystalline
phases formed at
Upper swelling
limit (at 25C)
36%
40%
42%
44%
35%
Phosphatidylcholine:
Distearoyl
Dipalmitoyl
Dimyristoyl
Dioleoyl
Egg PC
Soybean PC
CIS
CI6
CI4
CIS:!
CI6-IS:I
CIS:I-2
Small, I9S6t
Insko & Matsui,I97S
Janiak et al., I97S
Bergenstahl & Fortell, I9S7
Small, I9S6
Bergenstahl & Fortell, I9S7
Phosphatidyletanoleamine:
Dipalmitoyl
CI6
Dioleoyl
C1S:l
Soybean PE
CIS
20%
Lamellar phase at 68C
Reversed hexagonal at S4C
Lamellar below 0C
20%
Reversed hexagonal at 5C
l-2+ Reversed hexagonal
30%
above 0C
Caffrey, I9S5
Gawrish et al, 1992
Bergenstahl, 1991
Phosphatidylinositol:
Soybean PI
C1S:I-2+
Lamellar below 0C
Unlimited
Bergenstahl, 1991
Soderberg, 1990
ClS:l
Lamellar below 0C
Unlimited
CI6
Unlimited
Phosphatidic acid:
Dioleoyl
Lyso PC:
Palmi toy!
terrnine the type of liquid-crystalline phase that develops when different phosphatides are allowed to interact together with water. Figure 6.6 shows the
phase diagram of dioleoyl PC and dioleoyl PE in 40% water (Eriksson et al.,
1985). The figure shows that a lamellar phase is formed when the system contains mainly PC, but that about 60% PE nonlamellar phases start to form. This
change is enhanced at high temperatures. Between the hexagonal phase and
the lamellar phase is an area in which a cubic phase appears (above 50C).
160
100
Cubic phase
80
Reversed
hexagonal
phase
e
::l
60
'"<l.l
Lamellar phase
0.,
40
20
100%
DOPE
50150
100%
DOPC
Composition
Figure 6.6
Water
PC
161
Water
PI
Water
Figure 6.7 The phase diagram of soybean PC, soybean PE, and water; of soybean PC,
soybean PI, and water; and of soybean PI, soybean PE, and water. (Redrawn after
Bergenstahl, 1991.) The cubic phase was not included in the original drawing, but it is
a possible interpretation of the x-ray peaks included in the paper. It is also supported
by the data from the study by Eriksson eta!. (1985).
for oil-in-water emulsions are obtained by partial hydrolysis to form
lysolecithins (Emulfluid E). t It is also possible to increase the effective hydrophilicity of the PE by making the polar head group larger through acetylation (Emulfluid A).
6.6
6.6.1
One of the first ideas, proposed by Bancroft (1913) at the beginning of the century, was that the solubility of the emulsifier determines the type of emulsion
that is formed. An oil-soluble emulsifier will create an oil-continuous ernult Emulfluid, Lucas Meyer, Elbdeich 62, Hamburg, Germany
162
sion, and a water-soluble emulsifier will tum the emulsion into a water-continuous one. This is true for low molecular emulsifiers with a high solubility (usually in micellar aggregates), but it is also valid for polymers. However, most
likely, the concept can also, to some extent, be expanded to include emulsifiers
with just a dispersibility in either one of the phases (for instance lecithin).
Experience in this direction is exemplified in Table 6.6. However, the Bancroft
rule provide us just with the first very general directions. To proceed further we
need possibilities to rank emulsifiers quantitatively.
Table 6.6 Emulsifiahility compared with solubility according to the Bancroft
rule. (Ostberg et al., 1995.)
Emulsifier
Soluhility/dispersihility
Type of emulsion
Sorbitan esters
(Span)
Etoxylated sorbitan
esters (Tween)
Hydrophobic lecithin
(normal technical lecithin)
Hydrophiliclecithin
(high LPC or low PE)
Proteins
Fat crystals
Oil-soluble
Oil-continuous
Water-soluble
Water-continuous
Oil-dispersible
Oil-continuous
Water-dispersible
Water-continuous
Water-soluble
Oil-dispersible
Water-continuous
Oil-continuous
163
about 20 to 30C above the final storage temperature [emulsification by the PIT
method (Shinoda and Saito, 1968)].
Shinoda and coworkers (Shinoda and Saito, 1968; Shinoda and Kunieda,
1983; Kunieda and Ishikawa, 1985, reviewed in Shinoda and Friberg, 1986)
have worked according to this concept and characterized a number of different
ethoxylated emulsifiers in combination with various solvents. They then found
that the PIT depends not only on the number of ethoxy groups but also on the oil
phase, indicating the importance of the solubility properties for the stability.
Emulsification experiments performed with a range of different oil-to-water
ratios show that the emulsion type is determined mainly by the emulsifier
properties and is for many systems (pure solvents!) very insensitive to the
phase ratio (Shinoda and Friberg, 1986).
It is obvious that this says a lot about the properties of ethoxylated surfactants
but its applicability to food emulsions is very limited for two main reasons:
1. The concept is based on strongly temperature-dependent properties of
the emulsifiers. This excludes ionic emulsifiers (less important for the
food industry), and it also excludes the most commonly used polyhydroxy and nonionic zwitterionic emulsifiers as they display a very weak
temperature dependence of their hydrophilicity.
2. The solvent properties are important in the PIT concept. However, food
emulsions are made almost solely from triglyceride oils and water that will
behave differently due to the large molecular weight of the oil molecules.
6.6.3
Emulsifiers are molecules with a duality in their properties. The balance between the hydrophobic and hydrophilic properties of the molecules should
then determine the performance, for instance to the type of emulsion formed. If
the emulsifier is changed from being hydrophobic to hydrophilic, the emulsion
formed changes from oil-continuous to water-continuous. The balance of the
emulsifier is recorded as a number, the HLB value. When this concept was introduced by Griffin (1949), the HLB value of unknown emulsifiers was determined by comparing the emulsification properties in a predetermined system
of a mixture of hydrophobic and hydrophilic emulsifiers with a predefined
HLB number.
The important development of the HLB system came when the group contribution system was constructed by Davies (195 7), and it became possible to
164
Group contribution
212
6.8
5.25
2.4
2.1
1.9
1.3
0.33
-0.475
Aggregate
Shape
~>t
..,.,--.
165
Character in solution
HLB
Clear solution
> 13
>2
White dispersion
Milky appearance
7-10
l-2
Lumps of emulsifier
<7
<1
Micelles
Lamellar phase
Reversed aggregates
This result shows that the ability to form liquid-crystalline phases corresponds to the traditional HLB characterization of the emulsifiers.
Dynamic
interfaces
Interfacial
tension
Stabilize the
droplets
Repulsive
surface forces
Diffusion to the
the interfaces
Destabilize the
Diffusion to and
166
The principal role of the intenacial tension is obvious. The presence of emulsifiers lowers the intenacial tension from about 30 mN/m for a triglyceride/water
system to between 1 and lO mN/m. Nonionic emulsifiers close to the PIT create
densely packed intenaces with very low intenacial tensions. However, the effects of the intenacial tension itself are not very large. Walstra (1983) has shown
that the droplet size is only weakly dependent on the intenacial tension.
During the homogenization, new intenaces are formed. The emulsifiers have
to diffuse to the intenaces to lower the intenacial tension during the events when
the droplets are formed. This process must be rapid to be successful, as rapid as
the time scale for the formation of the droplets. For geometrical reasons, the diffusion from the surrounding phase of the droplet is much more rapid than the
diffusion from the internal liquid. This is one important contribution to the validity of the solution rules (Bancroft, PIT, HLB, and phase diagrams).
During the homogenization, the water-soluble substances in the oil phase
diffuse over to the water phase. These types of diffusion across the intenaces
create disturbances that contribute to the emulsification. In many systems, this
effect gives an increased efficiency if the emulsifier is added to the oil phase
before the emulsification. For dispersible emulsifiers (phospholipids) there are
also other reasons why it is more efficient to add the emulsifier to the oil phase
instead of the water phase. During the homogenization, phospholipids tend to
form stable liposomal dispersion in competition to the emulsification of the oil
phase. Westesen has indeed observed that a significant fraction of the phospholipids in a commercial phospholipid emulsion for paranteral use is lost in
liposomal aggregates (Westesen and Wehler, 1992).
Emulsification involves an intensive shear. The shear by itself causes a
high frequency of recoalescence events. If the emulsification is to be successful the formed droplets have to be protected. The repulsive interactions generated by the emulsifiers create a static protection.
The hydrodynamic interaction is crucial for the result of a collision due to
shear. The hydrodynamic interactions depend on the existence of an intenace
with an intenacial viscosity and elasticity. During the collision event, the intenace close to the approaching droplet is depleted of emulsifiers due to the
streaming ofliquid. The sunactant-depleted zone will then have a higher interfacial tension than the surrounding emulsifier-covered areas of the droplets.
This leads to sunace diffusion in the direction opposite to the liquid flow and
ensures the hydrodynamic resistance. If the emulsifier is oil-soluble, emulsi-
167
fier from the internal part of the droplet will diffuse to the depleted area and
thereby reduce the hydrodynamic protection of the droplet.
The discussion in this section has been very qualitative, but an important
point is that the emulsifiers contribute to the emulsification as well as to the
stabilization. The role of the emulsifier for the stabilization is usually difficult
to identify in the simple type of shaking experiments that are the main background to the HLB, the PIT, and the phase diagram concepts. This type of simple, and thereby effficient, experiment provides information about both the
emulsifiability and the stability with a certain emulsifier.
6.7
6.7.1
Competitive Adsorption
168
from a mixture of two emulsifiers, the most hydrophobic emulsifier will have the
strongest affinity to the interface. A consequence is that under competitive adsorption the component with the lowest water solubility will dominate the interface [e.g., the lowest critical micelle concentration (Kronberg, 1983)].
The character of the adsorbed layer, for instance its ability to generate repulsive interactions, is determined by the dominating compound. The structure of the layer depends on the geometrical shape of the molecules and on
lateral interactions between the molecules in the layer. Nonionic surfactants
may form very dense layers due to head-group attraction. Ionic surfactants are
able to form extremely loose layers due to inter-head-group repulsion.
An interesting experimental observation in agreement with this relation is
that the concentration of emulsifier necessary to obtain an emulsion is much
lower for ionic emulsifiers than for nonionic emulsifiers.
In a series of emulsions, we have studied the efficiency of the emulsification
(Ostberg et al., 1995) by droplet size measurements after homogenization. The
results show that for several emulsifiers very small droplets are obtained
(about 0.2 to 0.4 Jlm). The particle size obtained depends on the concentration
of emulsifier. The nonionic emulsifiers leads to a constant particle size down to
a critical concentration below which the ability to form emulsions is strongly
reduced. The critical concentration can be compared with the thickness of the
emulsifier layer on the emulsion droplet. The apparent thickness of the emulsifier layer can be estimated from the particle size and the concentration of
emulsifier (counted on the dispersed phase), if we assume that all emulsifier is
adsorbed to the interface. The apparent thickness gives the upper limit for the
adsorbed layer rather than the correct value:
Thickness of emulsifier layer =
0=
volume of emulsifier
emulsion droplet area
cern Vemulsion droplet
Aemulsion droplet
169
ness. The properties of the ionic emulsifiers are different. These emulsifiers
are able to emulsify the emulsions down to extremely low concentrations corresponding to very low surface concentrations (thin layers).
6.7.3
Layered Adsorption
Adsorption in layers is possible when different classes of surface-active components are present in a mixture. See Table 6.9. The two components must be
very different in character to give a structure with a layered character rather
than a mixed layer. The second component adsorbs to a particle displaying the
characteristic properties of the primary adsorbing emulsifier. This usually
means a more hydrophilic surface, which can be expected to reduce the adsorbed amount. However, in some eases, the presence of certain groups increases the adsorption of specific substances.
The effects of the emulsifiers on protein adsorption is essential in most
emulsifier applications in the food industry.
Ethoxylated surfaetants usually give a strong reduction of protein adsorp-
170
Table 6.9
Emulsifier
cone (%)1
Radius
(Jlm)2
Emulsifier-layer
thickness (1)3
Curve
shape 4
Estimated length
of the emulsifier
(1)5
Dodecylbenzenesulfate
0.1
0.47
1.6
15
10
0.27
90
54
0.20
45
75
lO
0.23
59
93
tion. Courthaudon et al. (1991b) have shown C12E08 totally displace all adsorbed ~-casein from an emulsion system. Similar effects have also been obtained with emulsions formed with polysorbates (Dickinson and Tanai, 1992)
and with monoglycerides (Hall and Pethica, 1967). On the other hand, egg
yolk PC did not reduce the the adsorbed amount of ~-casein more than about
20% (Courthaudon et al., 1991a).
The adsorption of a range of plasma proteins at various phospholipid sur-
171
References
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Bancroft, W.D. (1913).]. Phys. Chem., 17, SOl.
Bergenstahl, B. (1991). In Food Polymers, Gels, and Colloids (ed. E. Dickinson), Royal
Society of Chemistry London, pp. 123-131.
- - - , Claesson, P. M. (1990). In Food Emulsions (eds. K. Larsson, S. Friberg),
Marcel Dekker, New York.
- - , Fontell K. (1983). Prog. Call. Pol. Sci., 68, p. 48-52.
- - , Stenius P.J. (1987). Phys. Chem., 91, 5944-48.
Boyd, J. V., et al. (1976). In Theory and Practice of Emulsion Technology (ed. A. L.
Smith), Academic, London.
Caffrey, M. (1985). Biochemistry, 24, 4826--44.
Courthaudon, J. L., et al. (1991a).]. Agr. Food Chem., 39, 1365.
- - et al. (1991b).]. Colloid Interface Sci., 145, 390.
Darling, D., Birkett, R. J. (1987). In Food Emulsions and Foams (ed. E. Dickinson),
Royal Society of Chemistry, London.
Davies, J. T. (1957). Proc. Intern. Congr. Surf. Activity, 2d, London, 1, 426.
Dickinson, E. (1986). Food Hydrocolloids, 1, 3.
- - - , Tanai S. (1992). Food Hydrocolloids, 6, 163-71.
- - et al. (1991). Food Hydrocolloids, 4, 403-14.
Eriksson, P. 0., et al. (1987). Phys. Chem., 91, 846-63.
- - et al. (1985). Chem. Phys. Lipids, 37, 357-71.
Fontell, K. (1978). Progr. Chem. Fats Other Lipids, 16, 145-62.
- - - et al. (1968). Acta Polytechnica Scandinavica, Chapter 2, Chemistry Series III,
74, 2.
Friberg, S. (1990). In Food Emulsions (eds. K. Larsson, S. Friberg), Marcel Dekker,
New York.
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--,Mandell, L. (1970a). ]. Assoc. Off. Chem. Soc., 47, 149.
- - (1970b). ]. Pharm. Sci., 59, 1001-4.
- - , Rydhag, L. (1971). Kolloid Z. u. Polymere, 244, 233-9.
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Cosm., 85, 27-30.
- - et al. (1969). ]. Colloid Interface Sci., 29, 155-6.
Gawrish, K., et al. (1992). Biochemistry, 31, 2856-64.
Griffin, W. C. (1979). In Kirk-Othmer Encyclopedia of Chemical Technology, Wiley, New
York, Vol. 8.
- - - (1949).]. Soc. Cosmetic Chemists, 311-26
172
Hall, D.G., Pethica, D.A. (1967). In Nonionic Surfactants (ed. M.J. Schick), Marcel
Dekker, New York, p. 516.
lnoko, Y., Mitsui, T.J. (1978). Phys Soc. ]ap., 44, 1918.
lsraelachvili, J. (1992). Intermolecular and Surface Forces, Academic, London.
--eta!. (1977). Biochim. Biophys. Acta, 470, 185-201.
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Krog, N. (1990). In Food Emulsion (eds. K. Larsson, S. Friberg), Marcel Dekker, New
York, p. 127.
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Kunieda, H., Ishikawa, N. (1985). ]. Colloid Interface Sci., 107, 122-28.
- - , Shinoda, K. (1985). ]. Colloid Interface Sci., 107, 107-21.
Larsson, K., Krog, N. (1973). Chem. Phys. Lipids, 10, 177.
Lindblom, G., et al. (1991). Biochemistry, 30, 10938-48.
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Ostberg, G., et a!. (1995). Colloid Surfaces A, Physichemical Engineering Asp., 94,
161-71.
Pezron, 1., eta!. (1991).]. Colloid Interface Sci., 144, 449-57.
Rydhag, L. (1979). Fette Seifen Anstrichm., 81, 168-73.
--,Wilton, I. (1981). ]. Assoc. Off. Chem. Soc., 58, 830-7.
Shinoda, K., Friberg, S. (1986). Emulsions and Solubilization, Wiley, New York.
- - - , Kunieda H. (1983). In Encyclopedia of Emulsion Technology, Vol. 1 (ed. P
Becher), Marcel Dekker, New York.
---,Saito, H. (1968). ]. Colloid Interface Sci., 30, 258-63.
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Plenum, New York.
Soderberg, I. (1990). Structural Properties of Monoglycerides, Phospholipids and Fats
in Aqueous Systems, PhD Thesis, University of Lund, Sweden.
Tanford, C. (1973). In The Hydrophobic Effect, Wiley, New York.
Walstra, P. (1983). In Encyclopedia of Emulsion Technology, Vol. 1 (ed. P. Becher),
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Westesen, K., Wehler, T. (1992). ]. Pharm. Sci., 81, 777.
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emulsions,]. Assoc. Off. Chem. Soc., 48, 771-4.
SEVEN
Emulsifiers in Dairy Products
and Dairy Substitutes
Stephen R. Euston
7.1
Introduction
Bovine milk has been an important source of food for human beings for thousands of years. Not only is milk a very nutritious food in its own right, but it is
also a very versatile starting point for many other dairy products.
Milk is a complex food emulsion and colloidal sol. Table 7.1 gives the composition of whole cow's milk. The emulsion component is composed of fat droplets
dispersed in an aqueous phase containing protein. The protein is in the form of
both casein micelles, which are themselves colloidal particles, and free in solution as whey protein. A considerable reserve of knowledge has been assembled
on the structure and properties of the milk proteins (Swaisgood, 1992). The fat
droplets are stabilized by an adsorbed layer of protein and phospholipid called
the "milk fat globule membrane" (MFGM), which is distinct from the aqueous
phase protein (Walstra and Jenness, 1984). The average composition of the
MFGM has been estimated to be about 48% protein, 33% phospholipid, and
ll% water, with the remainder made up of other minor lipid components
(Walstra and Jenness, 1984). The phospholipid fraction of the membrane is composed of lecithin, phosphatidyl ethanolamine, phosphatidyl serine, phosphatidyl
inositide, plasmalogens, and sphingomyelin. Phospholipids are important food
173
174
Table 7.1 Approximate composition of bovine milk. (From Walstra & Jenness, 1984,
Dairy Chemistry and Physics. Copyright 1984, Wiley-lnterscience. Reprinted by permission of John Wiley & Sons, Inc.)
Component
Average content
(wt%)
Water
Solids, nonfat
Fat in dry matter
Lactose
Fat
Protein
Casein
Mineral substances
Organic acids
Miscellaneous
87.3
8.8
32.0
4.6
3.9
3.25
2.6
0.65
0.18
0.14
Range (wt%)
85.5-88.7
7.9-10.0
21.(}.-38.0
3.8--5.3
2.4--5.5
2.3-4.4
1.7-3.5
0.53-0.80
0.13-0.22
Average dry
matter (%)
69.0
36.0
31.0
26.0
20.0
5.1
1.4
l.l
emulsifiers in their own right. The contribution that they make to the stability of
the milk fat globule is not well understood, but their use as food-grade emulsifiers has been the subject of extensive fundamental research (Courthaudon et al.,
1991; Dickinson et al., 1993a; Dickinson and Iveson 1993).
Until this century the conversion of milk to butter, cream, ice cream, and
various types of cheese had been more of a craft than a science. It is only relatively recently that an albeit incomplete understanding of these processes has
been available. It is now understood that the formation of these milk-based
products is a consequence of either the destabilization of the dispersed-phase
fat droplets (as in butter, ice cream, and whipped cream) or of the dispersed
aqueous-phase proteins (as in cheese). Emulsifiers play an important role in
the formation of structure in many dairy products.
In the following pages, the major emulsifier-containing dairy and imitation
dairy products will be reviewed. A brief description of their production will be
given where relevant, with emphasis on the role that emulsifiers play in the
formation and stability of the product.
175
phase whose freezing point is depressed by freeze concentration of salts, sugars, and polysaccharide stabilizers. Despite its obvious complexity, ice cream
has been widely studied, and much is known about the formation of its structure and the role that low molecular weight emulsifiers play in this (Covin and
Leeder, 1971; Lin and Leeder, 1974; Goff, 1988; Krog and Barfod, 1990;
Barfod et al., 1991). A typical ice cream formulation is shown in Table 7.2.
Constituent
Fat
Milk solid, nonfat
Sugar
Stabilizer
Emulsifier
Water
10.0
11.0
13.0
0.2
0.5
65.3
176
and Barfod, 1990; Barfod et al., 1991). Before describing the role of emulsifiers in ice cream structure formation, it is pertinent to consider the structure
of the adsorbed layer formed around the fat droplet in the ice cream premix
emulsion. During the homogenization stage the ice cream mix is subject to
high shear. This results in the disruption of the fat phase into small droplets.
Surface-active components of the mix will adsorb onto the nascent-oiVwater
interface, lowering the interfacial tension and thus stabilizing the emulsion
droplets. The exact composition of the interfacial layer will depend on the proportions of each type of surface-active component and their relative surface
activities. Recent research has thrown light on the competitive adsorption between different milk proteins (Dickinson, 1986; Dickinson et al., 1988b;
Dickinson et al., 1989a; Dickinson et al., 1989c; Euston, 1989; Dickinson et
al, 1990b; Dickinson, 1992), milk proteins and emulsifiers (Dickinson and
Woskett, 1988a; Dickinson et al., 1990a; Dickinson, 1992; Euston et al.,
l995a; Euston et al., 1995b), and on the cooperative adsorption between proteins and polysaccharide stabilizers (Dickinson and Euston, 1990; Dickinson,
1992; Dickinson, 1993). These studies have identified that, in general, the
more surface-active component will predominate at the fat-droplet interface.
Thus the more surface-active protein in a mixture will dominate the adsorbed
layer initially, low molecular weight emulsifiers will, generally, displace protein from the surface with time, and under certain conditions polysaccharides
can interact with proteins and/or emulsifiers to contribute to the structure of
the adsorbed layer (Bergenstahl et al., 1992). The adsorbed layer of the ice
cream emulsion will be a composite of these functional ingredients.
The key to the formation of ice cream structure is the formation of a stable
foamed product. In ice cream this is achieved in two ways. The foam in ice
cream is not a typical protein-stabilized foam, where air bubble stabilization is
achieved by protein adsorption at the air/water interface. The initial stabilization of the foam network may indeed proceed by this mechanism, but prior to
freezing, the foam structure is stabilized primarily by the partial coalescence
of emulsion fat droplets at the air bubble interface. Figure 7.1 is a cryo-SEM
micrograph of ice cream. Adsorption of fat globules at the air bubble surface is
clearly visible. For this to be able to occur, the emulsion must be relatively unstable to the shearing forces exerted on the mix during air incorporation.
Several workers have attempted to demonstrate the link between emulsion coalescence stability and the mechanical or viscoelastic properties of the adsorbed protein layer (Doxastakis and Sherman, 1984; Rivas and Sherman,
177
1984; Dickinson and Stainsby, 1988; Dickinson et al., 1988a). Dickinson et al.
(1988a) have shown a correlation between the surface shear viscosity of adsorbed layers of various proteins at the planar oil/water interface, and the coalescence stability of emulsions made with these proteins. The higher the
interfacial viscosity, the more stable the resultant emulsion under perikinetic
conditions. It has also been demonstrated that the orthokinetic stability of protein-stabilized emulsions (stability under turbulent or shearing conditions) is
reduced by the presence of low molecular weight emulsifiers (Chen et al.,
1993; Dickinson et al., 1993; Dickinson and Williams, 1994). The explanation
for this lies in the ability of the low molecular weight emulsifier to displace
protein from the fat-droplet smface, thus reducing the mechanical strength of
the adsorbed layer. Emulsifiers present at concentrations too low to cause significant protein displacement can interfere with interprotein interactions
within the adsorbed layer and reduce the interfacial viscosity in this way
(Dickinson et al., 1990a).
Figure 7.1 Cryo scanning electron micrograph of ice cream. Note the partial coverage
of the air bubble (AB) surface by fat globule (FG) and the partially coalesced fat globules (PC) in the foam lamellae. (Courtesy of A.B. McKenna.)
178
The temperature dependence of emulsifier surface activity can be explained in terms of the phase behavior in aqueous solution. In the bulk phase,
monoglycerides exhibit a phase behavior similar to that of triglycerides (Krog,
1990). That is, they can exist in two polymorphic forms, the a. and ~forms,
that differ in the way the molecules pack in the crystal structure. When cooled
from a random molten state, a metastable crystalline structure termed the a.'
state is formed. Further cooling leads to a transition to the ~ state. The most
stable ~-crystalline structure will form if the a.' state is stored at ambient temperature. Lutton et al. (1969) proposed that the temperature dependence of
GMS surface activity can be explained by either the formation of a condensed
crystalline monolayer at the oiVwater interface at 5C or by micelle formation
close to the surface. The possibility that both mechanisms contribute to the increased surface activity on cooling cannot be ruled out.
Monoglycerides, and indeed other emulsifiers, have also been shown to exhibit a complex liquid-crystalline phase behavior in aqueous solution (Krog,
1990). Under certain conditions, the ~-crystal form of a monoglyceride will interact with water to form a lamellar liquid crystal. On cooling, this can transform to a so-called a.-gel phase (Krog, 1990). It has been hypothesized that
179
30
Q)
rJ)
cc
..r::
c.
.....cc
20
c
c
Q)
0....
c.
eftQ)
>
;:;
10
cc
Q)
a:
24
Aging time at 5C
Figure 7.2 Changes in amount of protein adsorbed to fat globules in an ice cream mix
during the aging period at 5C. The amount of protein bound in the fat phase is calculated relative to the total protein in the mix. (From Krog, N., Barfod, N.M., AIChE
Symposium Series, 86, 1-6. Reproduced with permission of the American Institute of
Chemical Engineers. 1990 A!ChE. All rights reserved.)
180
believe that in ice cream insufficient emulsifier is present in the system for liquid-crystalline phases to form. For this to occur the emulsifier must be adsorbed on the fat droplet surface at a concentration far greater than that
required for monolayer surface coverage. This is not the case in ice cream
emulsions (Darling and Birkett, 1987).
Once the emulsion has been destabilized in the freezer, partial coalescence of the fat droplets has to occur at the air/water interface to partially
stabilize the foam structure prior to freezing. Research (Boode, 1992; Boode
and Walstra, 1993) has indicated that the crystal structure of the fat in emulsion droplets is important in determining their susceptibility to partial coalescence. Van Boekel (1980) has shown that when fat crystals form at the
surface of emulsion droplets, and are large enough to penetrate the adsorbed
layer, a lipid bridge can form between two droplets that are in contact with
each other. The proportion of solid fat in emulsion droplets is important in
determining instability (Walstra, 1987). If the majority of the fat is solid, coalescence, or partial coalescence will not occur, and the droplets will be stable. Similarly, if the droplets contain a very low proportion of solid fat,
coalescence can occur. If the solid fat content is in the approximate range 10
to 50%, partial coalescence is possible. A proportion of the emulsion droplet
is required to be in the form of liquid fat for partial coalescence to occur.
Walstra (1987) envisages a partial coagulum offat droplets as being held together by necks of liquid oil.
When fat crystallizes in dispersed emulsion droplets, considerable supercooling can be observed. Crystallization of fats occurs at nucleation points that
already exist in the fat phase. These nucleation points occur relatively infrequently in emulsion droplets where the fat is dispersed into a large number of
small droplets. Consequently, in dispersed systems the triglyceride needs to be
cooled below its bulk phase freezing point before crystallization is initiated.
Emulsifiers in adsorbed monolayers can act as templates for the surface crystallization of triglycerides. Emulsifiers containing saturated hydrocarbon
chains have been shown to be good initiators of fat crystallization, whereas
those with unsaturated hydrocarbon chains are not as good (Berger, 1990;
Barfod et al., 1991). Figure 7.3 gives the solid fat content (SFC) as a function
of time for model ice cream emulsions stored at 5C. Both saturated (GMS) and
unsaturated (GMO) emulsifiers initiate crystallization compared to control
emulsions with no emulsifier, but the SFC of GMS-containing emulsions is always greater than for those containing GMO.
181
100
.....
c:
Q)
.....
c:
80
.....
.....co
60
0Cll
40
"0
20
0
24
Hours at 5C
Figure 7.3 Recrystallization of fat phase of ice cream emulsions without emulsifier
(control) or with saturated (GMS) or unsaturated (GMO) monoglycerides after cooling to
5C and aging. (From Barfod et al., 1991, and reproduced with permission.)
Darling and Birkett (1987) point out that in a mixed triglyceride system,
such as is found in milk fat, single discrete crystals are unlikely to form under
the rapid cooling conditions used in ice cream manufacture. They have shown
that in a cooled vegetable oil emulsion, concentric layers of triglyceride crystals are formed at the surface of oil droplets. These contain imperfections that
may be due to dislocations or recrystallization processes, and these are likely
to cause the destabilizing effect.
The ability of triglyceride crystals to penetrate the adsorbed layer depends
on a number of factors. The surface tension between crystal and oil, crystal
and water, and oil and water will determine how far into the oil phase the crystal will penetrate (i.e., if it is preferentially wetted by the oil or water phase).
Emulsifiers, by way of their surface activity, will alter these surface tensions,
and this may result in the crystals being able to penetrate further into the
aqueous phase. This would lead to a decrease in emulsion stability. The desorption of protein by emulsifier also aids destabilization by reducing the thickness of the layer through which fat crystals have to penetrate. The polymorphic
form of the fat crystal will also play a role in fat-droplet instability.
Triglycerides can exist in three general polymorphic forms, the a, ~. and W
polymorphs. When cooled from a melt, triglycerides will generally form a-type
182
crystals. These are not stable (Larrson and Dejmek, 1990) and will transform
formed at lower temperature, a crystals contain triglyceride in a more disordered liquid-like arrangement. These disordered crystals are softer and are
able to deform and follow the contours of the fat droplet more easily.
Consequently, they are less likely to penetrate the adsorbed layer. The ~-crys
talline structure is more solid-like, with the triglyceride molecules arranged in
ordered arrays. The ~ crystals have a greater mechanical strength and are unable to deform to the shape of the fat droplets. This leads to their bursting out
of the droplet into the aqueous phase (Darling, 1982).
In practice, two types of emulsifier are commonly used in ice cream: monoand diglycerides and polyoxyethylene derivatives of glycol or glycol esters, for
example polysorbates (Keeney, 1982). Sucrose esters have also been evaluated
and have been found to be suitable as ice cream emulsifiers (Buck et al.,
1986). Mono- and diglycerides and polysorbates are usually all found in current ice cream emulsifier blends. The explanation for this lies in the relative
abilities of polysorbates and mono- and diglycerides as emulsion destabilizers,
or as foam-forming agents. Polysorbates are far more efficient at displacing
protein from the oiVwater interface than are mono- and diglycerides and thus
are better emulsion destabilizers (Keeney, 1982). Mono- and diglycerides are
better foaming agents and thus are able to aid the formation of the initial foam
prior to fat-droplet agglomeration at the air/water interface (Keeney, 1982). A
second factor is the differing abilities of emulsifiers to influence fat crystallization. Figures 7.2 and 7.3 show that whereas GMO is able to displace more protein from the fat globule surface during aging than does GMS, GMS initiates
more fat crystallization than GMO. Use of a mixed emulsifier system would
also allow optimum protein displacement combined with optimum fat crystallization.
In summary, nonprotein emulsifiers are important in ice cream in several
respects:
l. They promote protein desorption from the surface of fat droplets, both by
their higher relative surface activity and the possible formation of liquid-crystalline mesophases.
2. They can act as nucleation points for surface crystallization of triglycerides.
3. They may promote fat crystal penetration of the adsorbed protein layer by
alteration of the surface tension between various phases.
183
4. They may help in the initial formation and stabilization of the ice cream
foam prior to partial fat-globule coalescence and freezing. Monoglycerides
are particularly good at this function.
184
Figure 7.4 Cryo scanning electron micrograph of whipped cream. Note the greater coverage of the air bubble (AB) surface by adsorbed fat globules (FG) than occurs in ice
cream (Figure 7.1). (Courtesy ofA.B. McKenna.)
Nonhomogenized cream separated from milk will whip satisfactorily without addition of emulsifiers. If the cream is homogenized prior to whipping and
the mean fat-globule size is reduced, emulsifiers have to be added to aid destabilization. Anderson and Brooker (1988) have attributed the differences in
whipping ability of homogenized and nonhomogenized cream to the differences in interfacial composition of the emulsion droplets in these systems.
Nonhomogenized cream contains fat droplets stabilized by native MFGM. The
composition of this has been described earlier (Section 7.1). After homogenization, the particle size is reduced, interfacial area is increased, and consequently, native MFGM is insufficient on its own to stabilize the newly formed
interface. A combination of increased interfacial area and competitive adsorption between MFGM material and cream serum proteins means that proteins
from the aqueous phase (caseins and whey proteins) contribute to the interfa-
185
ciallayer. The adsorbed layer in homogenized cream has been found to consist
mainly of caseins, with smaller amounts of ~-lactoglobulin and ~-lactalbumin
(Anderson et al., 1977; Darling and Butcher, 1978; McPherson et al., 1984).
This is consistent with the available data on competitive adsorption between
casein and whey proteins in model systems (Euston, 1989; Dickinson et al.,
1990b). In the early stages of whipping, before fat-globule adsorption and partial coalescence occurs to any great extent, the air bubbles are stabilized by
adsorbed milk serum proteins. Since this air/water interface is composed of the
same proteins that surround the fat/water interface in homogenized cream, the
difference in interfacial tension between the two interfaces is not very great
(Anderson and Brooker, 1988). Interfacial tension differences have already
been put forward as a driving force for fat-globule adsorption at the air/water
interface in ice cream (Section 7.2). Because of the small interfacial tension
differences between fat globule and air bubble in homogenized cream, the driving force for fat-globule adsorption is low. In nonhomogenized milk the interfacial tension differences between a fat globule stabilized by MFGM and an air
bubble stabilized by cream serum proteins are sufficient to act as the driving
force for fat-globule adsorption at the surface of air bubbles. Of course, this is
also aided by the presence of fat crystals at the fat-globule surface and by the
shearing forces introduced during whipping.
The importance of the fat phase manifests itself in two ways. As in ice
cream (Section 7.2), fat crystals are known to be important in the shear-induced coalescence of the fat globules (Darling, 1982), and the presence of a
certain amount of liquid fat is a prerequisite for good whipping properties.
Bucheim (1986) put forward the idea that the interfacial layer surrounding the
fat droplets ruptures when they collide during agitation. The subsequent
spreading of liquid fat is the first stage in destabilization by aggregation of adjacent droplets. The importance of the solid fat content of the fat globules has
been demonstrated by Darling (1982), who observes a direct correlation between the SFC and whipping time in natural cream.
Recombining technologies are becoming an increasingly important process
for making whipping cream bases. These would encounter the same problems
as homogenized cream (i.e., the similarity in composition and interfacial tension between air/water and oil/water interfaces), if they were formulated with
milk protein as the only surface-active material. For this reason, recombined
whipping creams, and indeed homogenized natural creams contain added low
molecular weight emulsifiers. These will alter the composition of the fat-
186
187
l. They destabilize the cream through their ability to displace protein from
the oil/water interface. This changes the adsorbed layer composition
and interfacial tension of the fat droplet.
2. They may destabilize the emulsion through their ability to form lyotropic
liquid-crystalline mesophases and the subsequent phase transformations that occur to form stable crystalline forms.
3. They may participate in the initial foam stabilization.
4. They aid in the formation of fat crystals at the fat-droplet surface, which
crystals are essential for fat-globule partial coalescence.
This last point is reinforced by electron microscopic studies of emulsifier
containing whipping creams. Figure 7.5 is a cryo-TEM micrograph of a whipping cream containing Tween 80. The large needle-like crystals that emanate
from the fat droplet are, apparently, typical of those formed in Tween-containing systems (McKenna, 1995 personal communication). The bridging of two fat
droplets by these needle-like crystals is clearly visible.
Composition (%)
Hardened coconut or
palm kernel oil
(melting point 3l-36C)
Maltodextrin
Sodium caseinate
Emulsifiers
52.0
32.0
8.0
8.0
188
Figure 7.5 Cryo transmission electron micrograph of whipped cream containing Tween
80 as emulsifier. A partial agglomerate of fat globules is shown. The spikes emanating
from the fat globule (FG) are fat crystals (FC) and can clearly be seen to bridge between
two fat droplets (B). These spikes are, apparently, features of Tween-containing systems. (Courtesy of A.B. McKenna.)
be important in the destabilization of the emulsion, while the protein is important in giving initial stability to the oil-in-water emulsion. The mechanism by
which emulsion destabilization is achieved, however, is different. Whereas
whipped dairy creams and whipped imitation creams are stabilized by partially coalesced, relatively intact fat globules adsorbed at the air/water interface, whipped toppings are stabilized by crystalline fat at the air bubble
surface. Krog and coworkers (Barfod and Krog, 1987; Bucheim et al., 1985;
Krog et al., 1986) have carried out extensive studies of the factors that effect
structure formation in whipped toppings. They have shown (Barfod and Krog,
1987) that part of the fat in spray-dried topping powders is in a supercooled
state. When these topping powders are reconstituted in water at low temperature, they show large structural changes that determine whipping characteris-
189
tics and foam structure. The emulsion becomes unstable due to spontaneous
recrystallization of the supercooled fat. The destabilization of the emulsion is
probably promoted by the temperature-dependent desorption of protein from
the surface, followed by coalescence. This makes crystallization of the supercooled fat more likely, due to the increased probability of nucleation sites
(Bucheim et al., 198S).
Scanning electron microscopy studies (Bucheim et al., 198S) show that the
final structure of the aerated whipped topping is stabilized by a layer of crystalline fat of about 0.1jlm thickness. The aqueous phase lamellae between air
bubbles also contains large proportions of crystalline fat, with smaller proportions of relatively intact fat globules.
The kinetics of fat crystallization and emulsion destabilization depend on
the type of emulsifier used in the formulation. Bucheim et al. (198S) have investigated the effect of distilled propylene glycol monostearate (PGMS), distilled unsaturated monoglycerides (glycerol monooleate, GMO), and distilled
saturated monoglycerides (glycerol monostearate, GMS) on the structure of
whipped toppings. Only PGMS is typically used in commercial formulations.
Figure 7.6 shows that all the emulsifiers promote fat crystallization when compared to toppings without added emulsifier. The effect of PGMS and GMO,
however, was greater than for GMS. The increased emulsion destabilization
caused by the enhanced fat recrystallization led to PGMS- and GMO-containing whipped toppings being more stable than those made from GMS-containing emulsions (Bucheim et al., 198S). The GMS-containing reconstituted
topping emulsion is too stable to allow consequent stabilization of incorporated
air (whipping).
The importance of protein desorption on the whipped topping structure and
stability has been demonstrated by Krog and coworkers (Krog et al. 1986;
Barfod and Krog, 1987). Table 7.4 gives the percentage protein contents of the
soc and
soc in the ab-
sence of emulsifier, almost one-quarter of the protein is associated with the fat
phase. This falls to 1.3% when PGMS is present and 7.7% with GMS added.
The temperature dependence of protein displacement is also evident. At 30C,
over 40% of the protein is in the fat phase when emulsifier is absent, but this
drops to about 33.7% in the presence of PGMS and 12.3% when GMS is included in the formulation (Krog et al., 1986). The GMS-containing reconstituted toppings have a significant proportion of protein still associated with the
190
40
~
.....
c
Q)
.....
c
0
.....Ctl
30
......
"0
(/)
20
...
...
...
10
0
10
20
30
Figure 7.6 Crystallization of supercooled lipid fractions in topping emulsions with different surfactants, reconstituted (I :3) in deuterated water (D20), measured by pulsedNMR at l5C. e GMO (Dimodan 0), PGMS (Promodan SP), o GMS (Dimodan PV),
A no surfactant added. [From Bucheim et al. (1985). Reprinted by permission of
Scanning Microscopy International.]
Table 7.4 Distribution of protein between the fat cream phase and the water
phase of centrifuged topping emulsion. (From Barfod and Krog, 1987. Reprinted by
permission of the American Oil Chemists Society.)
Surfactant
After I hour
at SC
%protein in
fat phase
After I hour
at SC
%protein in
water phase
After I hour
at 30C
%protein in
fat phase
After I hour
at 30C
%protein in
water phase
l0%PGMS
l0%GMS
None
1.3
7.7
24.0
98.7
92.3
76.0
33.7
12.3
41.7
66.3
87.7
58.3
fat-droplet surface, even after aging of the emulsions at 5C. This is obviously
enough to form an adsorbed layer strong enough, in combination with the lower
degree of fat recrystallization, to prevent stabilization of the incorporated air
bubbles. This is in contrast to the situation in ice cream and whipping cream,
where GMS is capable of destabilizing the fat emulsions to a degree that partial coalescence can occur. Darling and Birkett (1987) point out that the level
191
of emulsifiers in whipped toppings is sufficient to allow the formation of emulsifier adsorbed layers of far greater than monolayer coverage. They suggest that
the formation of liquid-crystalline mesophases, and the a.-gel phase of the adsorbed emulsifier, is a possibility. Westerbeek et al. (1991) have shown that a
common emulsifier used in whipped toppings, glycerollactopalmitate (GLP),
is capable of forming the a.-gel phase at oil/water interfaces, and this may contribute to emulsion destabilization.
The present understanding of the formation of structure in whipped toppings suggests the following mechanism for emulsion destabilization and formation of the foam structure:
1. The powdered, dried topping emulsion is stable.
2. When reconstituted at low temperature, protein desorption is initiated and
is aided by added emulsifiers such as PGMS. The mechanism of protein
desorption will be similar to that already described for ice cream. There is
7.5
Cream Liqueurs
Cream liqueurs are dairy emulsions of high added value. The combination of
milk protein-stabilized cream emulsion and high alcohol content make cream
liqueurs unique among dairy emulsions. Table 7.5 gives a typical range of
compositions for cream liqueur. In practice many commercial formulations
also have small amounts of GMS added.
The production of cream liqueurs is governed by the relative poorness of
the alcoholic aqueous phase as a solvent for proteins and sugars. Two commercial processes are in common use (Banks and Muir, 1988), namely, the single-
192
Table 7.5 Range of compositions of a standard cream liqueur. (Reprinted from Banks
Component
Composition (wt%)
Milk fat
Added sugars
Sodium caseinate
Nonfat milk solids
Ethanol
Water
12-16
15-20
2.6-3.5
l.0-3.5
14
46-51
stage process and the two-stage process. Figure 7. 7 presents flowcharts for
both processes. The main difference between the two processes lies in the
stage at which the alcohol is added. In the single-stage process this is prior to
homogenization, whereas in the two-stage process it is after homogenization.
Banks and Muir (1988) found that homogenization in the presence of alcohol
leads to the formation of fewer large fat globules, and as such is preferable in
terms of emulsion stability. A characteristic of cream liqueur production is the
harsh homogenization conditions used (two passes at 300 bar). This results in
a product in which more than 97% of the fat droplets have a diameter less than
0.8 f..Lm. A second factor favoring formation of smaller fat droplets is the significant lowering of interfacial tension observed at the oil/water interface when
alcohol is added to the aqueous phase (Bullin et al., 1988; Dickinson and
Woskett, 1988; Burgaud and Dickinson, 1990). As a result of the very fine
droplet size, the protein in the added cream has to be supplemented by sodium
caseinate (to a fat-to-caseinate ratio of approximately 5:1) to provide adequate
coverage of the newly formed fat surface by protein (Banks et al., 1981). The
fine particle size of the dispersed fat droplets gives the product an excellent
stability with respect to creaming. Banks et al. (1981) have noted no signs of
creaming in liqueurs with a composition within the range quoted in Table 7.5
after 12 months storage. The high level of added sodium caseinate, however,
leads to cream liqueur emulsions being unstable in acid environments. This
means that they are not suitable for combination with acidic beverage mixers
such as lemonade. A cream liqueur that is stable in an acid environment can
be made by replacing the sodium caseinate with GMS. The emulsifier replaces
milk protein as the primary emulsion stabilizer at the oil/water interface, and
the nonadsorbed protein is unable to aggregate the fat droplets when exposed
193
Single-Stage Process
Water
Sugar
Caseinate
Cream
Cream Base
Citrate
Alcohol
Product
Homogenize
2 x 300 bar, 55"C
Two-Stage Process
Cream Base
Homogenize
2 x 300 bar, 55"C
coolto<21rC
Homogenized Base
Gentle mixing
Alcohol
Product
Figure 7.7 Flow diagrams for the process of manufacture of a cream liqueur in (a) a
single stage and (b) two stages. (From Banks and Muir, 1988. Reprinted by permission
to acidic surroundings (Banks and Muir, 1988). Acid stability in this type of
product is gained at the expense of emulsion stability and shelf life. In practice, legal limits in some countries set the concentration of GMS at no more
than 0.4 wt%, and so total replacement of caseinate by GMS is not feasible.
Many manufacturers add low concentrations of GMS as well as sodium caseinate to cream liqueur formulations. Dickinson and coworkers (1989b) have
shown that, in addition to displacing some, but not all of the milk protein from
the fat droplet surface, which presumably infers some acid stability on the
product, GMS also improves the creaming stability of a model cream liqueur.
When model cream liqueurs were stored at room temperature for 12 weeks, no
creaming was observed with added GMS concentrations above 0.5 wt%. Below
this level of added GMS a reduced degree of creaming was observed compared
to control samples with no emulsifier (Dickinson et al., 1989b). The increased
creaming stability was associated with rheological changes in the emulsifier
aqueous phase. At low CMS concentrations the emulsions exhibit Newtonian
behavior, whereas above 0.5 wt%, a clear yield stress is found. Dickinson et al.
194
150 . ------------------------------,
Figure 7.8 Effect of GMS on the shelf life of simulated cream liqueurs on storage at
45C. The time for serum separation to first become visible is plotted against the GMS
concentration. Different symbols refer to separate experiments.(Based on Dickinson et
al., 1989. Reprinted by permission of the Institute of Food Technologists.)
195
syneresis when stored for any length of time. This leads to separation of the
aqueous phase and formation of a distinct, clear serum layer at the bottom of
the sample container. Whether this syneresis will occur at room temperature is
not certain, and Dickinson et al. (1989b) stress that a correlation between the
shelf life at 45C and that at room temperature may not follow. Cream liqueurs
stored under ambient conditions can have shelf lives of several years. Clearly
these are likely to be consumed before serum separation becomes evident.
Since the legal limits on the amount of emulsifier that can be added are set at
about 0.4 wt%, the problem of gel syneresis is unlikely to be encountered. At a
level of 0.4 wt% added GMS, creaming under gravity would not be eliminated
completely, but would be reduced to a level acceptable to the consumer
(Dickinson et al., 1989b).
7.6
Cream products containing 10 to 20% fat have been popular as coffee creamers for over 50 years (Abrahamson et al., 1988). Coffee creamers and whiteners
perform several functions: they give coffee a white color, reduce bitter taste by
complexation of the tannic acids with milk proteins, give the coffee a creamlike flavor, and give body to the coffee (Sims, 1989).
Traditionally, coffee cream is produced by simple concentration of milk up
to the required fat content. The cream is usually heat treated using a UHT
process, homogenized either before or after heating, and packed aseptically to
give a long shelf life. Emulsifiers are not usually added to this product. More
recently, with the advent of recombining technology and with the preference of
some consumers for vegetable oil-based products over those containing milk
fat, new products have appeared that require the addition of emulsifiers to
their formulations. If a recombined coffee cream is produced, the formulation
is more complex than for coffee cream. Table 7.6 gives a typical formulation
for a recombined coffee cream containing 19% milk fat, as suggested by
Zadow (1982).
The recombining process involves a two-stage homogenization with an 18MPa first stage and a 3-to-4-MPa second stage. Presumably the emulsifiers are
added to aid in the homogenization process by reducing the energy required to
form the fat/water interface. It may also be assumed that since in natural coffee
creams the milk fat is already in a dispersed state, the energy required to reduce the particle size is less, and so emulsifiers are not needed. However,
196
Table 7.6 Typical formulation for a recombined coffee cream. (From Zadow, 1982. Reprinted by permission of the International Dairy
Federation.)
Ingredient
Composition (wt%)
3.0
4.5
19.0
0.03
0.05
0.1
73.32
Zadow (1982) states that emulsifiers and stabilizers are required only if the
product is to be given a high heat treatment (a UHT process or steam injection). This may indicate a role for the emulsifiers in protecting the protein or
emulsified fat from heat damage. Evidence exists to support this hypothesis
and will be dealt with in more detail in Section 7.8.
Whereas coffee cream and recombined cream are used in a liquid form, coffee whiteners based on vegetable fat are also popular in dry powder form. Typical
formulations for liquid and dry powder coffee whiteners are given in Table 7.7.
Table 7.7 Typical formulations for liquid and powdered coffee whiteners. (From Si, 1991. Reprinted by permission of the
Society of Dairy Technology.)
Ingredient
Composition (wt%)
Liqnid
Powder
Fat
10.0
Sodium caseinate
Maltodextrin (DE28)
Monoglycerides
Tartaric acid esters
of monoglycerides
Carrageenan
Sodium alginate
K2HP04
Flavor
Water
l.O
30.0
4.0
62.0
-1.5
0.5
10.0
0.2
0.2
0.05
0.2
300 ppm
to 100%
0.05
1.5
lOOOppm
197
7.7
Processed Cheese
198
199
7.8
Recombined and concentrated milk products are produced for economic reasons. The cost effectiveness of transporting milk products that have been concentrated by removal of a proportion of the water phase, and the associated
changes in shelf life, make milk concentration a viable process. Similarly, it is
cost effective to transport dehydrated ingredients for recombination into milk.
The function of emulsifiers in these products is, primarily, to aid in the formation and stabilization of the emulsions. A secondary function, which is claimed
by many manufacturers of emulsifiers, is the effect that emulsifiers have on the
heat stability of milks and milk proteins.
200
7.8.1
Recombined Milk
Recombination of dairy ingredients into milk products is a popular and viable alternative to the export/import of fresh dairy products. It is particularly important
in countries where, for various reasons (e.g., transport delays, high temperatures),
the shelf life of fresh products prohibits their importation or local production.
In such cases, dried dairy ingredients are recombined close to the point of
sale, so as to reduce these problems.
Two approaches to recombining of whole milk can be used:
201
during homogenization. Recent research by Mayhill and Newstead (1992), however, suggests that little benefit in terms of emulsion formation and stability is
gained by their addition. In the case of mono-/diglyceride emulsifiers, it appears
that tradition dictates their presence in the formulation. It is possible that any reduction in creaming due to reduced fat-droplet size in the presence of emulsifier
is cancelled out by reduced emulsion stability caused by protein displacement.
7.8.2
Evaporated and concentrated milks are produced by removal of water from natural or recombined milks. The technology used to make these products includes
evaporation under reduced pressure, reverse osmosis, ultrafiltration, and freeze
concentration (Knipschildt and Anderson, 1994; Varnan and Sutherland, 1994).
These concentrated milk products are more susceptible to heat coagulation,
when UHT processed or sterilized, than are normal concentration milks.
It has been known for some time that the heat stability of skim milk can be
altered by surfactant molecules (Singh and Creamer, 1992). Anionic surfactants such as SDS have been shown to shift the maximum in the heat stability/pH profile of skim milk to more acidic values and to give a marked increase
in maximum heat stability (Fox and Hearn, 1978). Cationic surfaetants such as
CTAB move the maximum heat stability to more alkaline values and give only
a slight improvement in heat stability at the maximum (Pearce, 1978; Shalabi
and Fox, 1982). The mechanism by which these changes occur is not known
for certain. It has been suggested that binding of the surfactant to casein micelles alters the surface charge, which leads to changes in heat stability (Fox
and Hearn, 1978; Pearce, 1978; Shalabi and Fox, 1982). This view is supported by the fact that nonionie surfaetants such as Triton X and Tween 80
have no effect on the heat stability of skim milk (Fox and Hearn, 1978).
Research into the effect of addition of SDS and CTAB on the heat stability of
milk proteins is useful only in helping to understand the process of heat coagulation. These surfactants cannot be added to milk products. In addition to
this, in most concentrated milk products and in whole milk, the fat globules
play a role in heat stability. Surfaetants would interact with both the fat-droplet
surface and the milk proteins. This makes the process of heat coagulation in
fat-containing milks more complicated than in skim milk.
The milk fat globule membrane is known to play a role in the heat stability of
milk. In nonhomogenized whole milk the fat globules have little effect on heat
stability (Singh and Creamer, 1992). However, after homogenization the heat eo-
202
7.9
Emulsifiers have been added to other dairy products to exploit functional properties not normally associated with such emulsifiers. In recombined butter,
phospholipids are added as antispitting agents, to prevent fat spitting during
heating, and monoglycerides have been claimed to provide better "stand-up"
properties during storage (Kieseker, 1983).
Both sucrose esters and glycerol esters of fatty acids (monoglycerides) are
finding a wide range of novel uses. In addition to being good emulsifiers for use
in ice cream (Bucket al., 1986), they are known to improve the mouthfeel in
yoghurt (Farooq and Haque, 1992), inhibit microbial growth (Conley and
Kabara, 1973; Kato and Shibasaki, 1975; Shibasaki, 1979; Beuchat, 1980;
Kabara, 1983; Tsuchido et al., 1987), enhance the thermal death rate of bacte-
203
ria and bacterial spores (Tsuchido et al., 1981; Tsuchido et al. 1983), and increase the heat stability of bovine serum albumin (Makino and Moriyama,
1991 ). It appears that these functions are a result of their ability to bind to proteins (Clark et al., 1992; Fontecha and Swaisgood, 1994).
7.10
Summary
Emulsifiers are very versatile food additives. They can be used as aids to
emulsion formation (e.g., in coffee whiteners/creamers and recombined products), or in contrast, as emulsion destabilizers as in ice cream, whipping
cream, and whipped toppings. These two functions rely on the classical ability
of emulsifiers to act as surface-active agents. In this way they can influence the
formation and stabilization of the fat-droplet adsorbed layer and the composition of this layer. This ability of emulsifiers to displace protein from the fatdroplet surface also, probably, accounts for the increase in heat stability of
concentrated milks when phospholipids are added.
In a similar vein, displacement of adsorbed caseinate by GMS in cream
liqueurs can be used to give increased acid stability to these products. A secondary function of the GMS in cream liqueurs is its ability to interact with proteins, thereby forming a weak gel in the aqueous phase. The associated increase
in viscosity gives improved creaming stability. The ability of the emulsifier SSL
to interact with the proteins in caseinate is also exploited in coffee whiteners.
204
lipids, and milk fat-derived mono- and diglycerides. The future may see a large
increase in the use of products such as BMP, which is rich in natural milk phospholipids as well as protein, and milk fat that has been enriched in mono- and
diglycerides by processes such as controlled glycerolysis of triglycerides.
Acknowledgments
I would like to thank the following people: Drs. Jeremy Hill, Peter Munro, and
Mike Boland (NZDRI) and Dr. Susan Euston (Massey University) for proofreading this manuscript; Dr. David Newstead (Milk Powder Technology
Section, NZDRI) for valuable discussion and comments on UHT fouling and
heat stability of concentrated milks; Dr. Siew-Kim Lee (Cheese Technology
Section, NZDRI) for comments on and suggestions for the section on processed
cheese; Mr. Bing Soo (Food Systems Section, NZDRI) for supplying information on coffee whitener/creamer; Mr. Tony McKenna (Food Science Section,
NZDRI) for supplying the electron micrographs used in Figures 7.1, 7.4, and
7.5; Vanessa Ellery (Graphics Unit, Information Centre, NZDRI) for preparation of Figures 7.2, 7.3, 7.6, and 7.7. Finally, I would like to thank the New
Zealand Dairy Board for permission to publish this work.
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---,Jenness, R. (1984). Dairy Chemistry and Physics, Wiley, New York.
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210
EIGHT
Applications of Emulsifers
in Baked Foods
Frank T. Orthoefer
8.1
Introduction
8.2
Historically, animal fats were used for bakery products because of their natural
plasticity and flavor (O'Brien, 1996). Lard was the preferred animal fat for its
pleasing flavor. Faced with excess supplies of cottonseeds and cottonseed oil,
vegetable shortenings were invented by the cottonseed industry early in this
211
212
century. Initially, cottonseed oil was blended with lard as a "lard compound"
or simply compound shortening. Hydrogenation as a process was invented in
1910. This new process allowed the development of substitutes for the semisolid (plastic) animal fats and permitted the creation of products with improved functional properties.
Along with this modification technique came improved methods for processing the oil, including refining, bleaching, and deodorization. The products
possessed improved oxidative stability, uniformity, and enhanced performance. With the development of lipid chemistry came methods for alcoholysis,
esterification, interesterification, and isomerization. These advances in lipid
chemistry brought new emulsifiers and shortening formulations. High-ratio
shortenings were introduced about 1933. These contained mono- and diglycerides that brought about finer dispersion of fat particles in the dough, giving
strengthened cake batters. These finer particle dispersions allowed higher
sugar levels, with increased water added, to produce sweeter tasting and more
tender cakes. High-ratio shortenings possessed excellent creaming properties.
Moist, higher volume, fine-grained, even-extured cakes were produced. Icings
were improved as well (Hartnett, 1977).
Emulsifier development also proceeded as well in the 1930s (Stauffer, 1996).
Specialty shortenings were the result. Commercial layer cakes, pound cakes,
cake mixes, creme fillings, icings, whipped toppings, bread, and sweet dough
shortenings were created. The development of specialty products resulted in improvements in processing and improved product performance for the retail, food
service, and food-processing industries. In addition to the traditional plastic
shortenings, liquid shortenings, fluid shortenings, and powdered products were
produced. All these shortenings involved formulation with emulsifiers.
The terms emulsifiers and emulsifying agents, surfactants and surface-active agents are used interchangeably in the literature. The terms "emulsifier"
and "emulsifying agents" are, strictly speaking, chemicals or compounds capable of promoting emulsification or stabilization of emulsions by their effect
on interfacial tension. Surfactants for foods may include not only emulsifiers
but also compounds with other functions such as protein or starch interaction.
In most cases, the terms are used interchangeably.
The role of the emulsifier and that of the shortening are intimately bound in
bakery products. Generally, the food emulsifiers for bakery products supplement and improve the functionality of a properly developed shortening.
Emulsifiers act as lubricants, emulsify fat in batters, build structure, aerate,
213
improve eating quality, extend shelf life, modify crystallization, prevent sticking, and retain moisture. A list of emulsifiers commonly used in shortenings is
given in Table 8.1. The selection of or addition of an emulsifier to a shortening
base may significantly change the application of the shortening (Table 8.2).
Table 8.1 Emulsifiers used in shortenings
Mono- and diglyceride
Lecithin
Lactylaled monoglyceride
Calcium stearyllactylate
Sodium stearoyllactylale
Propylene glycol monoesters
Diacetyl tartaric monoglycerides
Ethoxylaled monoglycerides
Sorbilan monostearate
Polysorbate 60
Polyglycerol esters
Succinylated monoglycerides
Sodium stearoyl fumarale
Sucrose esters
Stearoyllactylate
Emulsified
All-purpose
Puff pastry
Pie crust
Cookie
Danish roll-in
Donut fry
8.3
Baked goods without emulsifiers have been described as tough, dry, stale,
leathery, or tasteless (Brandt, 1996). Current processing, distribution, and
storage of baked goods requires the use of additives that maintain quality and
freshness. Emulsifiers are commonly used in many food products. These supplementary materials or food additives are used to
214
Fraction
Percent
Starch
Protein
Pentosan
Lipid
Crude fiber
Ash
70.0-75.0
11.5-12.5
2.0-2.5
l.0-1.5
0.2
05
Table 8.3
Emulsifier
Monoglycerides and diglycerides (GRAS)
Succinyl monoglyceride
Lactylated monoglyceride
Acetylated monoglyceride
Monoglyceride citrate
Monoglyceride phosphate (GRAS)
Stearyl monoglyceride citrate
Diacetyl-tartrate ester of monoglyceride (GRAS)
Polyoxyethylene monoglyceride
Propylene glycol monoester
Lactylated propylene glycol monoester
Sorbitan monostearate
Polysorbate 60
Polysorbate 65
Polysorbate 80
Calcium stearoyllactylate
Sodium stearoyllactylate
Stearoyllactylic acid
Stearyl tartrate
Sodium stearyl fumarate
Sodium Iaury! sulfate
Dioctyl sodium sulfosuccinate
Polyglycerol esters
Sucrose esters
Sucrose glycerides
Lecithin (GRAS)
Hydroxylated lecithin
Triethyl citrate (GRAS)
8.4
215
U.S. FDA
(2ICFR)
EEC
nlllllber
182.4505
172.830
172.852
172.828
172.832
182.4521
172.755
182.4101
172.834
17.854
172.850
172.842
172.836
172.838
172.840
172.844
172.846
172.848
E47l
E472
E 472
E472
E472
E472
E477
E491
E435
E 436
E433
E 482
E 481
E483
172.826
172.822
172.810
172.854
172.859
184.1400
172.814
182.1911
E 475
E 173
E474
E322
E322
A shortening when mixed into a dough interupts the development of the gluten
structure. Literally, the structure is "shortened" and the baked product is tender. Overall, the shortening also contributes to the quality of the finished product by imparting a creamy texture and rich flavor, tenderness, and uniform
aeration for moisture retention and size expansion. Fat-based products are formulated and processed for plasticity to allow spreadability and to disperse
216
8.5
217
8.5.1
Monoglycerides are prepared by reacting glycerine with edible fats and oils or
fatty acids in the presence of a catalyst (Henry, 1995). The important characteristics are melting point and monoglyceride content. The monoglyceride content varies from 40 to 95%. Two crystalline forms are generally present: alpha
and beta. The alpha form is the most functional in bakery products. The major
variables involved in the production of monoglycerides are source of the fat,
monoglyceride content desired, iodine value or degree of unsaturation, and
fatty acid composition. Approximately 40 million pounds (18 million kg) of
monoglycerides are used in the United States in yeast-raised bakery products
(WH. Knightly, 1988). An equal amount was believed to be used in cakes, icings, and other applications. In addition to the improved aeration and sugarholding capacity of cakes formulated with monoglycerides, breads were found
to have an improved shelf life due to retarded staling rate. Various techniques
were applied to improve monoglyceride formulations including other emulsifier types. Today, plastic, hydrated, powdered, and distilled monoglycerides
are used in bakery applications.
In addition to their antistaling effects, monoglycerides in bakery products
result in
Increased aeration
218
When R equals
Emnlsifier
HO-
CH CH-C00-
31
OH
CH 3
-coo-
OOCCH
-QOC-cH-CH-COQ-
monoglyceride (DATEM)
OOCH 3
Polyoxyethylene monoglyceride or
ethyoxylated monoglyceride
(EMG)
H-
Figure 8.1
219
8.5.2.
Sorbitan Emulsifiers
8.5.3
Anionic Emusifier
The anionic emulsifiers include SMG, DATEM (see (Figure 8.1), and other lactic acid derivatives (Figure 8.3). Sodium stearoyl lactylate (SSL) is the most
widely used in the United States. The calcium version, insoluble in water, is
also widely used. Both are dough strengtheners.
SSL may be added as a stabilizer to hydrated monoglyceride preparations.
Other benefits from the lactic acid emulsifiers are antistaling, aeration, and
starch and protein complexing.
8.5.4
Polyhydric Emulsifiers
The polyhydric emulsifiers include the polyglycerol esters and sucrose esters
(Figure 8.4). Both have multiple applications as emulsifiers for foods in general and in bakery products, particularly the sucrose esters. The sucrose esters
are common food emulsifiers in Japan. They provide emulsifying, stabilizing,
220
Where
Sorbitan monostearate
R1 = H (CH 2 CH 2 0)n-
R2 = H
(Polysorbate 60)
R3 = H
Figure 8.2.
and conditioning effects in baked goods. Eight hydroxyl groups that may be esterified are present in sucrose. As expected, the degree of esterification affects
the hydrophylic lipophilic balance (HLB) of the sucrose ester (Table 8.4). The
product is considered a noncaloric fat substitute when six or more of the hydroxyls are sterified (Olestra by Proctor & Gamble).
8.5.5
Lecithin
Lecithin is a coproduct of soybean oil and, to a limited extent, of other vegetable oil refining (com oil and rapeseed oil). It is obtained by water-washing
crude vegetable oil and separating and drying the hydrated gums. The crude
material is very dark colored and viscous, consisting of a variety of phospholipids (Figure 8.5). In addition to the phospholipids, these are also present
221
CH 3
II
II
triglycerides, tocopherols, and glycolipids. Various purified grades are produced by bleaching and fractionating as well as by chemical modification
(Schmidt and Orthoefer, 1985). Commercial lecithin products are specified
based on their acetone-insoluble fraction (a measure of the phospholipid content), viscosity, and color. Lecithin is present in other sources such as egg yolk,
butter, beans, nuts, etc. Lecithin is usually the least expensive emulsifier.
8.6
222
OH
CH 2-CH-cH 3
OH
0-H C-HC-CH
2
OH
Polyglycerol monostearate
II
CH 2-0-C-(CH 3 ) 16CH 3
0
H
HO
H
OH
OH
Sucrose diester
Figure 8.4
8.6.1
Starch
Starch exists in a helical, coiled configuration with about six residues per tum.
This structure is basically a hollow cyclinder that has a hydrophyllic outer surface and a hydrophobic inner core. The inner space is about 45 nm in diameter. Straight-chain, alkyl molecules such as stearic acid will fit in the inner
Table 8.4
223
Percent
monoester
Percent
diester
71
61
50
46
42
33
24
30
36
39
42
49
Percent
triester
Percent
tetraester
0
1
2
2
2
2
12
13
14
16
HLB
15
13
ll
9.5
8
CH 2-0-R 1
r~~
CH -0-P-Q-R
2
lb
Then
Figure 8.5.
Lecithin.
space. The n-alkyl portion of emulsifiers such as present in GMS form a complex with the helical regions of the starch. It is this complex that is thought to
retard starch crystallization in bread, often called "retrogradation," slowing
the staling process.
The influence of emulsifiers on starch consists of (i) affect on the rate of
gelatinization, the gelatinization temperature, the peak viscosity, and the gel
224
8.6.2
Protein
The wheat flour proteins, gliadin and glutenin, form a viscous, colloidal complex known as "gluten" when mixed into a dough. Lipids are also involved in
the formation of the gluten complex. The properties of the gluten are influenced by the lipids and the emulsifiers present. Lipophilic parts of surfactants
interact with hydrophobic regions of proteins, contributing to unfolding or denaturation of the protein. Generally, surfactants contribute to protein denaturation, enhancing interfacial absorption and emulsion stabilization.
Most dough strengtheners are anionic surfactants. The association of the
lipophilic portion to the hydrophobic protein portion incorporates the negative
charge into the complex, promoting aggregation in the dough. The overall effect is aggregation of the gluten protein and an increase in dough strength.
The ionic surfactants induce protein insolubilization, resulting in increased
viscosity and elasticity of the dough. Nonionic surfactants disrupt the hydrophobic portion of the protein, leading to reduced dough viscosity and elasticity and increased protein extractability. A blend of emulsifiers generally
provides the best dispersibility and functionality.
8.6.3
225
Lipids
Wheat flour contains 1.4 to 2.0% lipids divided into free (0.8 to 1.0%) and
bound (0.6 to 1.0%) forms. They may be further divided into nonpolar (50.9%)
and polar (49.1 %) forms. The bound lipids exist as starch inclusion complexes. The nonstarch lipids, about 85% of the total, participate in the chemical, physical, and biochemical processes important for the preparation of
baked goods. The nonstarch lipids consist of glycolipids, phospholipids, and
stearyl esters. Interaction between nonstarch lipids and emulsifiers is limited.
Likely, added lipids merely influence or substitute for each other.
Non-polar-lipid addition to untreated flour gives a deterioration of baking
properties (Schuster and Adams, 1984). Addition of polar lipids to untreated
flour increases loaf volume in breadmaking. The improvement is likely based
on the effects of galactolipids and phospholipids. Emulsifiers may interact with
the water phase of the dough, forming associated lipid-water structures with
free polar flour lipids (Krog, 1981). Emulsifiers may compete with naturally
occurring lipids in wheat flour for the reactive groups of the wheat flour dough.
Their effect on protein components was modulated as a consequence.
8.7
8.7.1.1
tioners. The term is commonly used to describe adjuncts that aid in the development of less tacky, more extensible doughs that are processed through
machinery without tearing or sticking or that result in a product of finer crumb
structure and improved volume and symmetry. These may be summarized as
226
Improved gas retention giving lower yeast requirements, shorter proof times,
and greater finished product volumes
The dough conditioners recognized as highly functional dough strengtheners are calcium stearoyl lactylate, ethoxylated monoglycerides (EOM), polyoxyethylene sorbitan monostearate (PS 60), succinylated monoglycerides
(SMG), and sodium stearoyl lactylate (SSL) (Tenny,l978). Comparative loaf
volumes found for the various conditioners are shown in Figure 8.6, for fully
proofed dough shocked to mimic abuse in production.
Dough conditioners are available in plastic, powdered, and liquid forms
that may he added to the sponge, brew, or dough. The use levels are normally
in the range of 0.25 to 0.5% of flour weight. Often the dough conditioners are
combined with crumb softeners. For example, EOM, PS 60, and SMG are
blended with monoglycerides, a crumb softener.
8.7.1.2
Crumb-Softening.
227
2900
2800
1-
2700
2600
_fl
2500 -
1-
r-----
f-----
- :--
f-
'.:
- :'I!
r--: :-
f-
1--
f-
~l ~
r---:
1---
00
NO
ADD
CSL
EOM
PS-60
SMG
SSL
%Flour, wt
228
170
160
150
140
o_
12
11
130
NO
ADD
r---. ; r---
~- ~
.10 .25 .50
CSL
r---
10.25.50
EOM
~!I_
Jl
_Jl ~Jr---
r-~ r-J
JJ
10.25 .50
SMG
Mo-Di
%Flour, wt
Figure 8.7
8.7.1.3
Emulsifier Blends.
goods longer than any other emulsifier. It was shown to give higher ductility
through interaction with the gluten. Other activity from the lecithin is delayed
staling and shortening reduction. A synergistic effect also occurs with lecithin
and monoglycerides. The monoglycerides produce better crumb grain, softer
bread, and higher loaf volumes. Ethoxylated monoglycerides with monoglyceride is an effective dough conditioner. The negative effects of liquid oils in
place of shortening in bread preparation is overcome with the combination.
DATEM also acts as a dough conditioner, shortening saver, and antistaling
agent in combination with glycerol monostearate. Others include SMG, sucrose esters, polysorbate 60, SSL, and CSL. The SSL and CSL can form complexes with gluten acting as a dough strengthener.
The direct and indirect action of the emulsifier begins with dough preparation and ends with oven baking and storage (Figure 8.8). The first stage begins
Improvement of
wettability
Stabilization of
distributed phases
Mixing
Fermentation
Scaling, kneading,
moulding
Baking
Storage
229
Improvement of gas-retaining
properties
Shorter fermentation
Greater shock-tolerance
Improvement of gas-retaining
properties
Improved loaf volume
Better texture
Better crumb grain
Better uniformity
Decrease of water loss
Figure 8.7 Influence of emulsifiers on production and quality of baked products. (From
Schuster arul Adams, 1984.)
with wetting, and dispersing activity then follows with interactions with flour
components during mixing and in the baking process itself.
8.7.2
8.7.2.1
Cakes. The role of emulsifiers in layer cakes or snacks cakes is diverse. They include aeration, emulsification, and crumb-softening.
The aerated structure of batters depend on whipped-in-air and carbon diox-
230
ide from the leavening agent. The use of emulsifiers lowers the surface tension
of the aqueous phase, improving the amount of air that can be whipped into the
batter. Large amounts of finely divided air cells are important for development
of uniform grain (Randleman et al.,1961). The dissolved C0 2 evolves at air
cell sites and does not spontaneously form bubbles. If the original batter contains many small air cells, the final cake will have a larger volume and fine
(close) grain. The creaming of the sugar and shortening have a major influence
on air incorporation. The incorporation of monoglycerides in the plastic shortening (2 to 5% alpha-monoglycerides) ensures numerous small air cells being
created during beating or creaming.
Cake batter is, of course, an aerated emulsion. The integrity of the air cells
determines cake volume and uniformity. Shortening is an antifoam that disrupts
foam cells. Emulsifiers, however, coat the exterior of the fat particles protecting
the integrity of the air cell (Wooten, 1967). Proper emulsifier selection has permitted the use of liquid oil where only shortening could previously be used.
Light, tender, moist cakes are preferred by the consumer. Emulsifiers provide the desired aeration, emulsification, and crumb-softening. Crumb-softening in cakes is a function of moisture retention, shortening activity, and
starch-complexing. The starch-complexing is the same as for breads. The
emulsifiers complex with the starch to soften the product.
Several types of emulsifiers are used in cakes. Propylene glycol monoester
(PGME) is utilized at 10 to 15% of the shortening. Monoglycerides and mixtures of lactated monoglycerides with PGME are also used in cake mixes.
In bakers' cakes, a large number of emulsifiers are used, depending on the formula, production equipment, and labeling requirements. With soybean oil as the
shortening, a hydrated blend of emulsifiers such as PS 60, SSL, sorbitan monostearate, and distilled monoglycerides work well. Fluid shortenings are produced
containing lactated monoglycerides. The traditional bakers'-cake system is a plastic shortening with 5 to 10% monoglycerides (3% alpha-monoglyceride content).
Packaged cake mixes often use emulsified PGME at 10 to 15% of the oil. The
cakes are unusually tender and are not suitable for commercial cake production.
Emulsified cake shortenings are also used for cake donuts. The degree of air
entrapment during the creaming phase determines the grain in the final donut.
8.7.2.2
extent in cookies and crackers, they play a significant role in (i) controlling
spread, (ii) improving cutting and appearance, and (iii) improving texture. The
231
limited use probably stems from the reluctance to change the successful formulas in this segment of the baking industry. New products utilizing emulsifiers are receiving greater acceptance since they have been shown to play
functional roles.
Certain emulsifiers have the ability to change the degree of spreading of
cookie dough as it is baked (Table 8.5). The effect likely occurs because of altering of the viscosity of the dough. Cookie dough with SSL shows increased
spread compared to a nonemulsified control (Rusch, 1981). The SSL may interact with the starch granule, delaying hydration of the granule and subsequent gelatinization (Tsen et al., 1973).
Lecithin may be used to produce a drier dough that machines better and
that improves release from the rotary die surface. Use is from 0.25 to 0.7% of
the flour. Part of the effect may simply be reduction of available water because
of lecithin hydration.
Lecithin that is highly fluidized with other oils or fatty acids is widely used
as release agents in cookie baking due to improved release from rotary dies.
Heat-resistant lecithins such as those modified with acetic anhydride are especially adaptable to the application. Lecithin is used in cookie and cracker formulations at 0.25 to 1.0% of flour weight. It may be added with the shortening
at the creaming stage or simply combined with the shortening when rotated.
Antistaling is of less significance in cookies and crackers since they are of
Spread
ratio
8.3
8.8
10.4
10.0
9.8
9.6
9.7
9.2
9.3
9.2
232
8.7.3
Extruded Snacks/Cereals
8.7.4
Cream Icings
Cream icings are prepared by creaming sugar with fat, then adding flavor, egg
white, and perhaps a small amount of water. The emulsified shortening generally contains 2 to 3% alpha-monoglyceride. Polyoxyethylene sorbitan monostearate at 0.5% is included in some icings to assist in air incorporation.
Propylene glycol esters when incorporated into the shortening produce icings
with excellent gloss and gloss retention.
8.7.5
The fat-free or low-fat foods are marketed in almost every segment of the food
industry. In most instances, there is no one solution for removal of fats from the
formulation. Skillful formulation using fat replacers, emulsifiers, bulking
agents, flavors, and other ingredients has been applied to fat replacement.
Low-fat and fat-free cakes have been produced using additional emulsifiers
in conjunction with starch-based fat replacers and gums or hydrocolloids for
233
moisture retention and functionality. PGME and DATEM products have assisted in formula development.
Emulsifiers are not generally regarded as fat substitutes or replacers.
Emulsifiers affect the texture and mouthfeel by their effect on surface interactions. The caloric value of emulsifiers varies depending on their actual composition and digestibility. They tend to have fat-like properties through their
hydration, water-binding, and dispersing effects in processed foods. The general functions of emulsifiers in low-fat or no-fat applications are to
Mono- and diglycerides are the most used emulsifiers. Distilled monoglycerides have a caloric advantage over the lower mono-containing preparations.
Other emulsifiers in reduced fat products include the polysorbates, polyglycerol esters, and sorbitan monoesters.
Many noncaloric fats have been synthesized. Sucrose polyesters (OlestraTM
from Proctor & Gamble) is probably the best known. Emulsifier use in products utilizing the noncaloric, synthesized fat replacers is very likely similar to
that utilizing the traditional caloric version.
8.7.6
Release Agents
A separate application of emulsifiers in bakery products, although not incorporated into the dough, is release agents or pan sprays. Lecithin is the primary
emulsifier used. Often the pan sprays are formulated with an oil in combination with mold inhibitors and lecithin. One to 6% lecithin is added. Modified
lecithins that exhibit improved heat stability may used. The pan spray may
simply be brushed or sprayed to achieve a thin film promoting easy release of
baked products from pans or belts.
8.8
Summary
234
References
Brandt, L. (1996). Emulsifiers in baked goods, Food Product Design, Feb., pp. 64-76.
Chawla, P., deMan, J.M. (1990). ]. Am. Oil Chem. Soc., 67, 329.
Hahn, D.E., Hood, L.F. (1987). Factors influencing com starch lipid complexing, Ger.
Chem., 64,81-85.
Handleman, A.R., et al. (1961). Bubble mechanisms in thick foams and their effect on
cake quality, Ger. Chem., 38, 294.
Hartnett, D.I. (1977). ]. Am. Oil Chem. Soc., 54, 557.
Henry, C. (1995). Monoglycerides: the universal emulsifier, Ger. Foods World, 40(10),
734-738.
Knightly, W:H. (1988). Ger. Foods World, 33,405-412.
Krog, N. (1981). Theoretical aspects of surfactants in relation to their use in breadmaking, Ger. Chem., 58, 158--164.
Lagendijk, J., Pennings, H.J. (1970), Ger. Sci. Today, 15, 354-356, 365.
O'Brien, R.D. (1996). Shortening: types and formulations, in Bailey's Industrial Oil and
Fat Products (ed. Y.H. Hui), 5th ed., Vol. 3, pp. 161-193, Wiley, New York.
Rusch, D.T. (1981). Emulsifiers: uses in cereal and bakery foods, Ger. Foods World,
26(3), 1ll-ll5.
Schmidt, J.C., Orthoefer, F. (1985). Modified lecithins, in Lecithins (eds. B.F. Szuhaj
and G.List), Chap.lO, pp. 203-213, Am. Oil Chern. Soc., Champaign, IL.
Schuster, G., Adams, W.F. (1984). Emulsifiers as additives in bread and fine baked
products, in Advances in Cereal Science and Technology (ed. Y. Pomeranz), Chap.
4, pp. 139-242, Am. Assoc. Cer, Chern. Inc., St. Paul, MN.
Stauffer, C.E. (1996). Emulsifiers for the food industry, in Bailey's Industrial Oil and
Fat Products, 5th ed., Vol. 3, pp. 483-516, Wiley, New York.
Tenney, R.J. (1978). Dough conditioners/bread softeners, Baker's Digest, 52 (4), 24.
Tsen, C.C., et al. (1973). High protein cookies I. Effect of soy fortification and surfactants, Baker's Digest, 47(4); 34-39.
Weiss, T.J. (1983). Food Oils and Their Uses, AVI, Westport, CT.
Wooten, J.C., et al. (1967). The role of emulsifiers in the incorporation of air into layer
cake batter systems, Ger. Chem., 44, 333.
NINE
Emulsifiers in Confectionery
Mark Weyland
9.1
Introduction
235
236
sumer the product has an acceptable taste, appearance, and texture. These
qualities in modern food products are often critically dependent on the type
and level of emulsifiers used in the product.
The second function of an emulsifier is often to act as a surfactant. In these
cases the role of the emulsifier is to modify the behavior of the continuous
phase of a food product so as to bring about a specific effect or benefit. The
most common example of this in confectionery is the use of lecithin in chocolate to reduce the viscosity of the product and improve the ease of handling
and processability.
Many of the classes of emulsifiers described in this book have also found
their way into confectionery products. These include lecithin and modified
lecithins such as YN and phosphated monoglycerides, glycerol monostearate,
polyglycerol esters including polyglycerol polyricinoleate (PGPR), sorbitan esters, polysorbates, lactic acid and tartaric acid derivatives of monoglycerides,
acetylated monoglycerides, sucrose esters, and propylene glycol monoesters.
All these compounds have a common feature that makes them suitable as
emulsifying agents: they are ambiphilic, possessing both lipophilic and hydrophilic properties. The nature of this property is often expressed as hydrophilic/lipophilic balance or HLB. The HLB number is an indication of the
properties of an emulsifier and is usually on a scale of 0 to 20. An emulsifier
with a low HLB will tend to be more oil-like and will therefore have a greater
affinity for the oil phase of a confectionery product. Lecithin, for example, has
an HLB of 4 and has a great affinity for the oil phase in chocolate. Polysorbate
60, by contrast, has an HLB of 15 and is quite soluble in water and therefore
has an affinity for the syrup phase in toffees and caramels.
It is often the case in food products, and in confectionery too, that a combination of two emulsifiers in a recipe or formula containing two distinct phases
will result in the longer lasting and more uniform product. In these cases, combinations oflow- and high-HLB emulsifiers give the best results.
In the remainder of this chapter, a number of the more common confectionery types using emulsifiers are described, along with a review of the available knowledge relating to the most optimal emulsifier types and their benefits.
9.2
The use of emulsifiers in chocolate and compound coatings is perhaps the best
documented in the literature of any of the applications of these ingredients in
Emulsifiers in Confectionery
237
9.2.1
Lecithin
Chocolate and compound coatings are dispersions of solid particles in a continuous fat phase. The solid particles are composed of sugar granules, milk
solids, and cocoa solids. In chocolate the fat is cocoa butter and comes directly
from the crushing of cocoa nibs, but in compound coatings the fat comes from
vegetable oils added to the formula to give the same fat content as chocolate
overall. Moisture is also present in the coating, introduced indirectly via the
sugar or other solid ingredients. It is the presence of these solid particles and
moisture that causes chocolate and compound coatings to deviate from true
Newtonian viscosity behavior. The solid particles tend to abrade each other,
and the resultant internal friction causes the viscosity of the material to vary
according to the applied shear stress.
Viscosity is very important when considering how chocolate and compound
coatings are used, because they always have to flow to either fill a mold without
defects or air bubbles or cover a candy piece with a thin, even coat. These
properties are controlled almost completely by the rheological behavior of the
coating, which in turn is dependent on the nature of the continuous liquid
phase, that is, the fat and fat-soluble ingredients. Considering the flow properties of molten chocolate, the two most important parameters are plastic viscosity and yield value. "Plastic viscosity" is defined as the force required to keep
liquid chocolate flowing once it has started moving, whereas "yield value" is
the force required to start the mass of liquid chocolate moving. Both plastic
viscosity and yield value can be decreased by the use of specific surfactants,
and this enables the chocolate manufacturer to have greater control of cocoa
butter levels. Plastic viscosity and yield value are often combined in a value
called "apparent viscosity," but it is important to understand that chocolates
with equal apparent viscosities can have different yield values and different
plastic viscosities.
238
Coatings can always be made more fluid for better control by adding more
cocoa butter or vegetable fat to the mix, but since these are the more costly ingredients in coatings, this is often an unattractive solution. Better by far is to
add a surfactant like lecithin to reduce coating viscosity. It is possible to add
0.5% lecithin to a coating to have the same viscosity reduction effect as the
addition of 5% cocoa butter or vegetable fat (Minifie, 1980). Lecithin allows
coating users to operate efficiently at much lower fat contents than would otherwise be the case (Figure 9-1).
Lecithin is commercially extracted from the soybean by solvent extraction
and precipitation. It is a light brown fluid that contains approximately 65%
acetone-insoluble phosphatides and 35% soybean oil. An oil-free product is
also available that is in the form of granules. The chemical composition of soy
lecithin is as follows (Minifie, 1980):
Soybean oil
Phosphatidyl choline
Phosphatidyl ethanolamine
Phosphatidyl inositol
Other phosphatides and polar lipids
Carbohydrates, e.g., sterols
35%
18
15
11
9
12
39
38
L.
Q)
~
::I
.0
co
37
36
(.)
35
'#-
34
(.)
33
32
1\
""
0.1
0.2
..........
0.3
['...
0.4
0.5
0.6
0.7
% lecithin (soya)
Figure 9.1 Effect of addition of lecithin on the fat content in dark enrobing chocolate.
Curve represents chocolate formulas of equal viscosity. (Minifie, 1980.)
Emulsifiers in Confectionery
239
CH 2-o-c-R
?!
CH-0-C-R
lipophile
II
I
hydrophilic
Figure 9.2
II
OH
240
100
[',_
1'---
90
80
r--__
c---
..
50
>"'
40
30
20
10
1---
--
!\
--: 60
~
>.
---- - -
~--
70
"'utJ
~Cocoa/Fat
\\
\l
""
0 .1
'
- - .........
t---- -
Chocolate
I'-.
1'--.:_ugor I Fat
0 .2
0 .3
0.4
0.5
0 .6
0 .7
process in the fat phase. Levels of over l% in typical 30% fat coatings should
not be used.
Emulsifiers in Confectionery
241
a neutral flavor and can be used at higher dosage levels than natural lecithin
without the negative impact on viscosity (Bonekamp-Nasser, 1992; Klienert,
1976; Nakanishi, 1971). See Figure 9.4.
YN is also claimed to reduce the thickening of chocolate and compound
coatings due to moisture and overheating (Bradford, 1976). A comparison between lecithin at 0.3%, YN at 0.3%, and cocoa butter added at 5% to chocolate gave the same similar overall viscosity readings (see Figure 9.5), but a
calculation of the Casson yield values showed that YN produced significantly
lower values than with the other systems. The viscosity reducing effect of YN
is reportedly less in milk-free coatings than with milk coatings (Klienert,
1976; Hogenbirk, 1989). Milk coatings have generally higher viscosities than
milk-free coatings due to the effect of milk solids/fat/emulsifier interactions.
150
140
130
120
~ 1 RP.M.
90
tl
ao
0"'
\\
\\
100
--
\"''
""
'
''
60
50
'-..5
0
0
40
L.
cD
--- -
..............
.....
..........
20
20 R.P.M.
...
0
0.1
0.2
0
/o
--.. _
... -r--
~--
10
Figure 9.4
---
r--
..........
R.P.M.
'<"..... r::--..........
30
r--------
'~
--
?- 70
"'u0
>"'
"0
u
Soya lecithin
---YN
~\
110
....
~\
0.3
.... --
--- 1----------04
0.5
Lee ith 1 n
242
700
c:;-
5% cocoa butter
E
..s?.
Ul
600
0.3% lecithin
Q)
c:
>
500
~
Ul
Ul
Q)
--+-
0.3% YN
__.._
400
....
....
+'
Ul
300
ctl
Q)
..c:
(/)
200
Figure 9.5
10
20
30
40
50
Viscosity plot comparing lecithin, YN, and cocoa butter. (Bradford, 1976.)
9.2.3
where R= H or a fatty acyl group derived from poly condensed ricinoleic acid
and n is the degree of polymerization of glycerol.
Emulsifiers in Confectionery
243
A number of studies have been published that compare the effects of PGPR
with lecithin and YN. Most conclude that PGPR, when added to chocolate or
compound coatings at 0.5% or less, can reduce the coating yield value to almost zero (Application Notes Admul WOL, Quest International; Bamford,
1970). The practical benefit of such a feature is that in a chocolate bar molding
operation, PGPR addition would allow the chocolate to flow easily into even
complicated mold shapes without entrapping air bubbles and also to flow
around inclusions. Furthermore, the opportunity exists to reduce the fat content of the chocolate as well as the coat of chocolate formulations.
A typical comparison of lecithin and PGPR additions to a milk and to a
dark chocolate using a Rotovisco viscometer is given in Tables 9.1 and 9.2
(Application Notes Admul WOL, Quest International).
The values in Tables 9.1 and 9.2 indicate that in milk chocolate it is possible to reduce the yield value to almost zero. The presence of lecithin in the
emulsifier mix allows the plastic viscosity to be minimized, and this blend pro-
Table 9.1
2.0
4.0
5.0
45
29.8
26.5
16.3
15.3
97
62
58
58
Lecithin
0.05
0.1
0.2
0.4
30.0
26.7
20.0
15.6
79
54
40
37
PGPR
0.075
0.175
0.3
0.5
0.6
30.0
29.2
26.8
30.5
32.0
86
38.5
22
2.5
2.0
Lecithin + PGPR
0.1
0.2
0.3
14.1
13.4
12.7
34
32
29
Addition
Amount
Cocoa butter
0.0
l.O
llO
244
Table 9.2 Casson plastic viscosities and yield values of a dark chocolate when cocoa butter, lecithin and PGPR are added
Addition
Amount
Casson plastic
viscosity (poise)
Casson yield
value (dynes/cm2)
Lecithin
0.3
0.7
0.97
1.3
18.5
17.1
14.4
12.4
155
221
297
285
PGPR
0.0
0.1
0.2
0.5
1.0
12.9
12.5
14.8
14.9
15.9
199
151
82
13
0
vides the optimum solution. In dark or semisweet chocolate the effect of PGPR
on plastic viscosity is small, while it can reduce yield values to very low values
at 0.5% addition. The mechanism by which PGPR affects yield value is not
understood but its practical benefits are widely reported.
PGPR is also claimed to be advantageously used in ice cream coatings since
it allows low apparent viscosities in the presence of low levels of moisture
(Bamford, 1970). Also claimed is PGPR's beneficial effect on fat-phase crystallization leading to easier tempering, improved texture, and longer shelflife of
coatings (Application Notes Admul WOL, Quest International.). The author has
found that the viscosity-reducing properties of PGPR does lead to significantly
reduced viscosity at temper and a level of temper, as measured by a tempermeter, which is easier to maintain over long periods in an enrober without significant recirculation of chocolate via melt-out and retempering circuits.
PGPR's most recognized benefit remains that of fat reduction, and manufacturers claim that a blend of 0.5% lecithin and 0.2% PGPR allows cocoa butter
reductions of approximately 8%.
Emulsifiers in Confectionery
245
ter-based chocolate is to ensure that the cocoa butter crystallizes in the correct crystal form or polymorph. Cocoa butter has several different polymorphic
forms that have melting points ranging from 17 to 35C. The forms are represented by the greek letters y, a, pt, and~ As the polymorphic form increases
in stability it also increases in melting point. To make chocolate in the familiar
glossy, fast-melting form with good snap, it is necessary to crystallize the cocoa
butter in the highest melting polymorph, which is ~ This form of cocoa butter
is also needed to ensure good contraction in molded products and the long
bloom-free shelf life expected for good quality chocolate goods. Bloom is the
phenomenon in which liquid fat is forced to the surface of a product and spontaneously crystallizes there as a grayish layer that looks to the uninformed consumer as if the product has gone moldy. The liquid fat is forced out to the
surface in this way by the following mechanisms:
l. Chocolate is not preconditioned (tempered) correctly such that insufficient concentration of seeds in the ~ form is present in the crystallizing
chocolate mass. This leads to a higher level of less stable Wforms in the
chocolate mass that later transform to the more stable ~ form. This
transformation causes the chocolate coating or bar to contract and
squeeze liquid fat to the surface. Chocolate contains liquid fat even at
room temperature, where cocoa butter attains a maximum solid fat content of approximately 85%. This liquid fat at the surface crystallizes in
an uncontrolled fashion and is a mixture of ~' W, and even possibly
some a forms.
2. Chocolate is tempered correctly, but in storage or distribution the product is subjected to wide temperature variations, resulting in partial
melting and resolidification of the chocolate. Under these conditions
uncontrolled recrystallization takes place and extensive bloom can occur. This kind of change is often referred to as "heat damage" and the
product is classified as not heat resistant.
3. In molded bars that contain peanuts or other nutmeats as solid inclusions, or in enrobed products that have centers containing quantities of
soft vegetable oil or dairy butter oil, this oil can "migrate" from the center to the chocolate shell. The soft oil will cause the chocolate to become
soft and cocoa butter will dissolve. This will cause severe damage to the
product due to physical handling prior to consumption or due to discoloration and bloom of the chocolate shell, which will now be far more
heat-sensitive.
246
9.3.1
the~ form thus deterring bloom (Anon., l99la). In this way STS behaves as a
crystal modifier.
The situation is further complicated, however, because there exists not one
hut two ~ forms of cocoa butter, usually referred to as forms V and VI. In this
classification system the Wform is called form IV and transitions from form IV
to form V, the stable form, occur via the liquid phase only. The transformation
of form V to form VI takes place only via the solid phase and can take many
months to occur. Also, identification of form VI is difficult and requires specialized x-ray diffraction techniques. However, this ~transformation is also as-
Emulsifiers in Confectionery
24 7
sociated with bloom in solid chocolate that has been well tempered. Work done
by Garti et al. (1986) has indicated that STS is particularly effective at blocking this V to VI transformation and, hence, preventing bloom even after extensive temperature cycling between 20 and 30C.
Other emulsifiers studied by Garti included sorbitan monostearate and
Polysorbate 60, but these were only half as effective as STS. STS is a highmelting-point emulsifier (approximately 35C} whose structure is more closely
related to cocoa butter triglycerides than to most other emulsifier types. It is
speculated that it is due to this similarity that STS co-crystallizes with cocoa
butter from the melt and, due to its rigid structure, binds the lattice in form V.
Other more liquid or less triglyceride-like emulsifiers tend to depress the melt
point of crystallized cocoa butter, increasing liquidity and promoting form IV
to V transformations in preference.
This is presumably why STS is a more effective antibloom agent in solid
chocolate than in enrobed chocolate items, where soft center oils can soften
cocoa butter crystals via migration and lead to type IV-V bloom occurring (see
type 3 bloom previously mentioned). Krog (1987), however, claims that STS
locks fats in the less stable Wform and prevents the transformation to ~
Berger (1990) also claims that STS performs well as a bloom inhibitor or gloss
enhancer in palm kernel oil-based compound coatings used to enrobe cakes
by stabilizing the Wform of the vegetable fat, a situation also observed by the
author in several practical cases using lauric coating fats but with much less
reliability when using domestic fats such as soybean or cottonseed-based coating fats. Such products tend to have longer bloom-free shelf lives in many
cases so that the need for antibloom additives is not so imperative.
The use of STS in food in the United States is limited to chocolate and compound coatings for which there is a petition pending for acceptance by FDA as
a food additive. This uncertain status has not deterred United States food companies from enjoying the benefits that STS can bring to coating appearance.
STS is, however, more widely accepted as an additive in EC countries.
9.3.2
SMS and polysorbate 60 [also known as polyoxyethylene (Player, 1986) sorbitan monostearate] are also used as antibloom agents, especially in compound
coatings based on vegetable butters. They are not as effective as STS, but they
have the advantage of being already accepted by FDA as food-grade emulsifiers. They are usually used in combination, where the SMS acts as a crystal
248
modifier and the polysorbates act as hydrophilic agents to improve emulsification with saliva and aid flavor release (Dziezak, 1988; Lees, 1975). SMS can
also be used at high levels in coatings to increase heat resistance due to its
high melt point, 54C; unfortunately, the addition of SMS also causes the coating to become waxy.
Up to 1% of this combination can be added to coatings to improve initial
gloss and bloom resistance. The optimum ratio of SMS to polysorbate 60 is
60:40 (Woods, 1976). These emulsifiers are claimed to function by forming
monomolecular layers of emulsifier on the surface of sugar and cocoa particles,
thereby inhibiting the capillary action that causes liquid fat to migrate to the
surface and cause bloom. Lecithin is still needed in these systems to control
coating viscosity and reduce fat content.
In another nomenclature system, SMS is also called Span 60 and polysorbate 60 is called Tween 60 (Lang, 1974). Spans and Tweens are also claimed to
reduce the rate of fat crystallization; therefore, to develop proper crystal size a
suitable tempering system needs to be employed. They are employed in both
chocolate and compound coatings and may be used with advantage iffast crystallization of the coating would be disadvantageous.
9.4
Mono- and diglycerides are also used as additives to chocolate and compound
coatings, often as their purified or distilled forms. They can act as seeding
agents especially when in high-melting-point forms such as glycerol monostearate (GMS). They are more commonly used as antibloom agents in laurictype palm kernel oil compound coating to extend useful shelf life. A typical
usage level would be 0.5%. Berger (1990) claims good results in hydrogenated
palm kernel oil coatings when using glyceryllacto palmitate at 1 to 5% as a
gloss improver; the application was as a coating for a baked product. Moran
(1969) found that a polyglycerol ester of stearic acid reduced the viscosity of
fat sugar systems better than lecithin as well as retarded crystallization, improved gloss, and better demolding.
Lactic acid esters of monoglycerides have also been used to control gloss in
compound coatings (Hogenbirk, 1989; Dziezak, 1988) and to improve demolding performance (Anon., 1991b). Woods (1976) describes the use of triglycerol
monooleate in compound coating chocolate to improve initial gloss and gloss
retention, and triglycerol monostearate as a whipping agent to aerate coatings
Emulsifiers in Confectionery
249
giving them a lighter texture for filling applications. Herzing (1982) describes
in detail the types of polyglycerol esters needed to optimize the glossy properties of lauric and nonlauric compound coatings: triglycerol monostearate, octaglycerol monostearate, and octaglycerol monooleate. These emulsifiers are
added to the coating fat at up to 6% by weight.
Polyglycerol esters have also been claimed to speed up the setting time of
chocolate panning coatings when used at levels of 0.4 to 0.6% (Player, 1986).
Hogenbirk (1989) has found advantages of viscosity reduction to some degree
with examples of mono- and diglycerides, diacetyl tartaric acid esters of monoglycerides (DATEM), acetylated monoglycerides, and propylene glycol monoesters. Musser (1980) has published results showing the benefits of adding
up to 1.5% DATEM to chocolate and compound coatings to modify viscosity
properties and to improve the rate of crystallization of coating fat phases. In a
series of experiments, Musser found that the addition of DATEM to fully
lecithinated milk and dark chocolates, and dark sweet coatings, could further
reduce the viscosity of the coatings as measured by a Brookfield viscometer. At
the same time Musser found that the DATEM was acting as a seeding agent in
chocolate systems, improving the speed of crystallization and resulting in a
finer grain and better gloss in molded bars. Musser's conclusions relative to the
effect of DATEM on viscosity is supported by the author's own published study
on chocolate viscosity and emulsifiers (Weyland, 1994).
9.5
250
9.5.1
Chewing Gum
Chewing gums contain fats and emulsifiers that act as softening agents or plasticizers to the gum base. In this role, emulsifiers can also act as carriers for
colors and flavor aiding in the dispersion of these important ingredients within
the gum base. Up to 1% lecithin can be used to soften chewing gum to the desired consistency (Patel et al., 1989) and can be hydrated or mixed with a vegetable oil or suitable fatty emulsifier such as mono- and diglycerides, to aid in
dispersion within the chewing gums. Chewing gums prepared in this way have
the desirable soft, chewy properties popular in today's top products.
Other emulsifiers are also used in chewing gum to provide suitable textural
and antistick properties to the chewing gum base; and these include monoand diglycerides, glyceryl lacto palmitate, sorbitan monostearate, triglycerol
monostearate, triglycerol monoshortening, and polysorbates 60, 65, and 80.
Lecithin is also used to provide a protective coating to chewing gum pieces
prior to a hard panning process (Dave et al., 1991). Normally, only hard chewing
gums can be hard panned in this way, but by using a hydrated lecithin coating it
is possible to candy coat the gum and then allow the lecithin to soften the chewing gum in storage prior to consumption. By this technique the emulsifier coating, when set, forms a suitable base for syrup-based candy coatings.
9.5.2
Emulsifiers in Confectionery
251
9.5.3
252
and emulsifiers creates a water-insoluble helical complex and creates an irreversible textural effect. This interaction was quantified by Krog (1977) using
the amylose-complexing index or ACI. The ACI is defined as the percentage of
amylose precipitated at 60C after l hour and after reacting 5 mg of the emulsifier with 100 mg of amylose in solution. See Table 9.3.
Table 9.3
87
35
42
49
18
16
32
0
9.6
Processing Aids
Emulsifiers are sometimes used in small amounts in confectionery products either to control aeration or to prevent product sticking to machinery and packaging. They can also be used to displace starch from starch-molded jellies and
gums and provide a shiny attractive appearance as well as a barrier to degradation from atmospheric oxygen and moisture.
Aeration of protein systems containing small amounts of fat, such as
nougats, can be facilitated by the addition of triglycerol monostearate. Liquid
fat or lipophilic emulsifiers such as GMS or acetylated monoglycerides usually
tend to destabilize forms and cause deaeration.
Emulsifiers are also useful release agents providing barrier properties between product and molds, tables, metal, conveyor belts, utensils, and machin-
Emulsifiers in Confectionery
253
References
Anon. (199la). Confectionery Production, 57(2), 136-7, 140.
- - - (l991b). Confectionery Production, 57(6), 451-2.
Bamford, H.F., eta!. (1970). Rev. Int. Choc. (RIC), 25, 6.
Berger, K.G. (1990). World Conference on Oleochemicals into the 21st century, AOCS,
Champaign, IL, pp. 288-91.
Bonekamp-Nasser, A. (1992). Confectionery Production, 58(1), 66, 68.
Bradford, L. (l976).Int. Flavors Food Addit., 7(4), 177-9.
Dave, ].C., eta!. (1991). U.S. Patent 5,135,761, March 28.
Dziezak, J.D. (1988). Food Techno!., 42(10), 171-86.
Garti, N., eta!. (1986). ]. Assoc. Off. Chem. Soc., 63(2) 230-236.
Gregory, D.H. (1982). Confectionery Production, 48(10), 437-9.
Harris, T.L. (1968). Surface Active Lipids in Foods, Monograph No. 32, Society of
Chemical Industry, England.
Herzing, A.G., eta!. (1982). U.S. Patent 4,464,411, November 5.
Hogenbirk, G. (1989). Confectionery Production, 55(1), 82-3.
Jeffrey, M.S. (1991). Manufacturing Confectioner, 71(6), 76-82.
Klienert, J. (1976). Rheol. Texture Food Qual., pp. 445-73, AVI, Westport, CT.
Krog, N. (1977). ]. Assoc. Off. Chem. Soc., 54(3), 124--31.
Lang, M. (1974). Confectionery Manufacturing and Marketing, 11(2), 3-5, 13.
Lees, R. (1975). Confectionery Production, 41(6), 296,298, 304.
254
TEN
Margarines and Spreads
Eric Flack
10.1
Introduction
255
256
tificial gastric juices to facilitate separation of tissue from pure fat and then,
following crystallization at 25 to 30C, extracting under pressure about 60% of
a soft semifluid fraction-oleomargarine-and about 40% of a hard white
fat-oleostearine. The softer fraction had a melting point similar to butter fat,
a pleasant flavor not unlike melted butter fat, a pale yellow color, and could
easily be plasticized, and although different in fatty acid composition from butter fat, it offered a useful basis for the production of a substitute. He assumed,
therefore, that this new butter fat substitute consisted of glycerides of margaric
and oleic acids (the former then considered to be a C17 homolog of the saturated fatty acids, although now known to be a eutectic mixture of palmitic and
stearic acids) that solidified in crystals having a pearly luster. The name margarine was derived from the Greek word "margarites," meaning pearl and thus
is pronounced with a hard "g" according to its derivation.
Mege's process was to take a proportion of the soft fat with appropriate
quantities of milk and water, into which a small amount of udder extract was
stirred. Following agitation, a stable emulsion similar to thick butter cream
formed that, on further churning, thickened to resemble butter. While this
process may now sound rather rudimentary, it nonetheless comprised all the
elements of margarine production as we know them today.
10.2
Early Development
Mege took out patents in France and England, but the next major development
took place in Holland when, in 1871 in the absence of patent law in Holland at
that time, he sold his invention to the butter merchants Johannes and Anton
Jurgens in the village of Oss. Also in the same locality was another family of
butter merchants-Simon and Henry Van den Bergh-who were to commence
production shortly afterward and who later were to join forces with the Jurgens
and eventually become the largest margarine manufacturers in the world
(Schwitzer, 1956).
Production followed in most countries in Europe-Austria (1873/4), Italy
(1874), Germany (1874), Norway (1876), and Denmark (1883), which was later
to become the largest per capita consumer of margarine. Mege applied for a
United States patent that was granted in 1874.
Many variations were developed in the following years and patents on new
formulations and processes were taken out. At the same time there was strong
opposition to the introduction of margarine into the market by the farming
257
Year
Butter
Margarinet
1930
1935
1940
1945
1950
1955
1960
1965
1970
1975
1980
1985
1990
1991
1992
1993
1994
17.8
17.6
17.0
10.9
10.7
9.0
7.5
6.5
5.3
4.7
4.5
4.9
4.4
4.2
4.2
4.5
4.8
2.8
3.0
2.4
4.1
6.1
8.2
9.4
9.9
11.0
11.0
11.2
10.8
10.9
10.6
11.0
10.8
9.9
258
A similar development has been seen in Europe (Table 10.2), where in recent years (and also forecast to continue) the trend is for a reduction in yellowfat consumption generally, with small reductions in both butter and margarine
in the order of 4 to 5% and an increase in spreads of 17 to 19% by 1997.
Table 10.2 Europe-Average consumption of butter, margarine, and spreads
(kg per capita)
1987
Country
Belgium/
Luxembourg
Denmark
France
Germany
Greece
Ireland
Italy
Netherlands
Portugal
Spain
UK
Total EC
1992 (Est.)
Butter
Margarine
Spreads
Butter
Margarine Spreads
8.3
10.2
8.8
8.3
0.9
5.8
2.3
4.1
0.8
0.5
4.9
5.1
12.8
16.9
3.9
7.7
1.4
4.5
1.2
13.7
3.4
1.6
7.5
5.4
3.2
3.5
0.4
0.3
0.1
3.4
7.4
9.0
8.2
6.7
1.0
3.1
1.9
3.4
1.0
0.5
3.9
4.5
11.7
14.4
3.4
8.0
1.4
4.2
1.3
11.9
3.0
3.3
0.1
0.8
0.6
0.7
2.2
6.1
5.1
5.1
6.2
0.9
0.9
0.2
6.8
4.0
0.3
1.6
2.4
1.4
Source: Food Source '92 (1992), Frost & Sullivan, New York.
10.4
259
be similar to butter. However, with the new developments during more recent
years involving different components and varying combinations of vegetable
oils, animal fats, and milk fat, a range of new descriptions have come into being such as low-fat spreads, low-calorie spreads, and yellow-fat spreads. In
1993, Moran listed the varieties then available, as shown in Table 10.3.
Table 10.3
High-fat spreads
Low-fat spreads
Butter
Margarine: Packet
Soft
Polyunsaturated fatty acids
(PUFA) spreads
Vegetable/butter-fatblended spreads
Vegetable fat spreads
80
80
80
80
80
70
60
40
Vegetable/butter-fatblended spreads
Butter-fat spreads
Very low-fat spreads
Water-continuous spreads
70
40
40
20-30
15
9
5
Source: Moran (1993).
10
l. Butter
A. Milk fats
Products derived exclusively from milk
and/or certain milk products for which fat
is the essential constituent of value
l. Blend
2. Three-quarter-fat blend
3. Half-fat blend
4. Blended spread xo/o
but not more than
l. Margarine
2. Three-quarter-fat
margarine
3. Half-fat margarine
4. Fat spreads xo/o
B. Fats
Products derived from solid and/or liquid
vegetable and/or animal fats suitable
for human consumption, with a milk-fat
content more than 3% of the fat content
2. Three-quarter-fat butter
3. Half-fat butter
4. Dairy spread xo/o
Sales description
Fat-group definitions
Table 10.4
Milk-fat content not less than 80% but less than 90%.
Max. water 16%; max. dry nonfat milk materials 2%
Milk-fat content not less than 60% but not more than 62%
Milk-fat content not less than 39% but not more than 41%
Products with milk-fat content:
Less than 39%
More than 41 %; less than 60%
More than 62%; less than 80%
Vegetable and/or animal fats not less than 80%; less than 90%
Not less than 60%; less than 62%
Product categories
(additional description with
indication of% fat content by weight)
261
Whether the listed descriptions will gain greater prominence other than as
the required secondary descriptions is doubtful since consumers, presumably,
will prefer to favor the brand names that have been applied to the broad range
of products now available.
Fat spreads falling outside these standards, for instance those with fat contents below 10% and concentrated products with fat contents of 90% or more,
will not be classed as spreadable fats.
EC legislation does not specify vitamin fortification, which is left for decision at the national level. However, in the UK the requirement will remain as
previously specified in the Margarine Regulations (Sl 1867) 1967. That is,
each 100 g of margarine shall contain 800 to 1000 mg of vitamin A and 7.05 to
8.82 mg of vitamin D. This does not apply to other spreads, so that vitamin fortification in these cases is a matter for individual decision by the manufacturers and/or retailers.
Legislation in the United States and Canada is currently less detailed than
in Europe, but both require a minimum of 80% fat. In the United States, FDA
156.110 describes margarine (or oleomargarine) as "food in plastic form or
liquid emulsion" containing not less than 80% fat and describes a method for
fat determination. It specifies vitamin A fortification to be not less than 15,000
I.U. per pound but leaves vitamin D as an optional ingredient while stating not
less than 1500 I.U. per pound. While spreads are not specified in the legislation, low-fat spreads were stated in 1990 as the fastest growing category, with
spreads from 20 to 72% fat on the market (Borwanker and Buliga, 1989).
The National Association of Margarine Manufacturers of the United States
describes spreads as having fat contents ranging from 18 to 79%. Details of
market shares in the United States are shown in Table 10.5.
Stick margarine
Soft margarine
Liquid margarine
Low-fat vegetable oil spreads
Total retail market
1980
1990
65
19
1
15
44
100
100
l3
2
41
262
263
10.5.1
Oils and fats, otherwise known as triglycerides, are esters of glycerol and fatty
acids having a basic formula
where R 1, R2 , and R3 are the fatty acid residues, which may be the same but,
invariably, are different according to the source of the fat. Examples of the
variations in fatty acid compositions of different oils and fats are shown in
Table. 10.6.
Modification of the fat by, for instance, hydrogenation, whereby some of the
unsaturated links (double bonds) become saturated (for instance oleic C18:1 to
stearic CIS) offer the opportunity of considerable variation in the melting
points of fats, especially the liquid vegetable oils. The exact physical character
of a fat depends upon that of the constituent fatty acids that can vary widely in
melting point, as illustrated in Table 10.7.
10.6
Fat Crystallization
""'
0.2
1.6
tr
25-30
2.5-3.5
15-25
35-45
7-12
0.5-1
1-4
59-70
0.2
3-4
0.5-1
26-28
2-3
23-27
30-35
1-1.5
0-1
45-57
4-8
Lard
9.2-13.9
tr
2.2-4.4
36.6-65.3
15.6-40.7
tr
2-8
84-105
7.7-9.7
2.3-3.2
5.4-7.4
1.3-2.1
tr
tr
7.5-10
tr
13-17
Groundnut
45.9-50.3
16.8-19.0
Coconut
Fatty acid
Beef
tallow
tr
0.8-1.3
tr
43.1-46.3
tr
4.0-5.5
36.6-65.3
9.4-11.9
0.1-0.4
[xx]1
50-54
Palm
1.9-3.0
11.9-18.5
1.4-3.3
tr
[xx]1
16-19
7.2-10.0
43.6-51.4
15.3 17.2
Palm
kernel
3.4-6
0.2-0.6
l.l-2.5
52-65.7
16.9-24.8
6.5-14.1
1-6
109-126
tr
Lowerucic
rapeseed
9.9-12.2
tr
3.6-5.4
17.7-25.5
50.5-56.8
55-95
[xx]l.5
125-136
tr
tr
Soya
5.6-7.4
tr
3.0-6.3
14.0-34.0
55.5-73.9
tr
[xx]l.5
85
tr
Sunflower
265
C12lauric
C14 myristic
C 14: 1 myristoleic
C16 palmitic
C16:1 palmitoleic
C18 stearic
C18:1 oleic
C18:2 linoleic
C18:3linolenic
C20 arachidic
M.P. eC)
44.2
54.4
Liquid
62.9
Liquid
69.6
16.2
Liquid
Liquid
74.4
1
1
1
2
Table 10.8
Fat ( 0 C)
Time (min)
Temp.
Coconut
Coconut/palm 1:1
Coconut/palm, interesterified
Palm-hardened
Palm-hardened/palm 2:1
Lard
Lard/palm 1:1
Palm
Sheafat
3
4
5
5
8
14
15
27
45
20
15
18
17
13
lO
10
lO
10
mediate, and the most stable ~ form the highest values. Transition from one form
to another is from
The polymorphism of crystalline fats may cause problems with the consistency
of margarine and spreads. During production, the fats initially crystallize in the a
form and normally will rapidly transform to the Wform. This is the desirable form
for spread production since the small needle-shaped
p form (> 20 Jlm) they will give the spread a grainy consistency known as
266
(a)
(b)
Figure 10.1 Fat crystals in (a) normal margarine with good consistency and (b) a margarine with large ~crystals causing a "sandy" texture. (Magnification 200x.) (Courtesy
Concurrently, the specific area of the crystal surface will decrease, allowing
the liquid oil to penetrate to the surface of the margarine, which may then lead
to oiling out, especially if the margarine comes under pressure. Some vegetable oils such as partially hardened sunflower or low erucic rapeseed are
particularly prone to form ~ crystals and thus can cause sandiness in spreads.
In this case, sorbitan tristearate at 0.3% of the fat has been found to inhibit the
26 7
be adsorbed at the oil/water interface and that only lipophilic emulsifiers with
high solubility in the oil phase can perform as crystal inhibitors in emulsions.
Varying the level of saturated fat affects the degree of crystallinity of the oil
phase (Borwanker and Buliga, 1989). Spreads with 40% fat were made using
oil phases of varying crystallinity by blending either liquid soyabean oil or
stick margarine oil with soft margarine oil. The textures of the spreads made
with these blends were visibly very different, increasing in softness in the
same direction as the oil itself. As a consequence of increased firmness, the
stability of the resulting spread was expected to increase with increasing crystalinity in the oil phase. A centrifugation procedure was used to measure the
stability of the emulsions (Figure 10.2). The higher amount of water and/or oil
(especially water) released corresponds to a less stable emulsion.
10.7
Emulsifiers
60
"0
+-'Cil
Ul
Cll co
u Cll
Cll Cll
....
.s-.:
>.0
..~I...
==o
..c.._
co Cll
+-'+J
CJ)co
3:
""' '"--
50
40
30
-a...___
20
O Oil
"-~
10
O Water
0
4.5
6.3
8.6
10.4
13.0
50 Soft
50 liquid
75
25
Soft
75
25
50 Soft
50 Stick
Figure 10.2 Effect of the degree of crystallinity on the emulsion stability of 40% fat
spreads as measured by ultracentrifugation. (From Borovanker and Buliga, 1989.)
268
They are either oil-in-water (0/W) emulsions, where the continuous phase is
water (such as in milk or ice cream) or water-in-oil (W/0), where the continuous phase is oil (as in butter and margarine).
The stability of two-liquid phase emulsions is kinetic stability, i.e., the system
is not thermodynamically stable (Friberg et al., 1990). Thermodynamically stable emulsions would spontaneously reform following separation by, for instance,
centrifugation, while experience shows that an emulsion that has separated remains so unless mixed by some external action. In reality, the two separated
phases are in the most stable state to which all emulsions will tend. Thus, a stable emulsion is one in which this inevitable trend has been retarded so that it is
not noticeable during the normal life of the product--even if it be several years.
Margarine is a water-in-oil emulsion in principle only because, in reality, it is
a dispersion of water droplets in a semisolid fat phase, containing fat crystals
and liquid oil (Krog et al., 1983). Preparation of the emulsion requires considerable energy to reduce the droplet size of the disperse phase, thereby creating an
increase in the surface area between the two immiscible phases.
The initial water-in-oil emulsion is prepared in mixing tanks with vertical or
horizontal stirrers to ensure emulsification but without incorporating too much air.
It is rather coarse in water-droplet size and fairly unstable if not kept agitated.
In modern continuous production, the emulsion exists for only a brief period before passing into the chilling unit where final emulsification and crystallization of the fat phase take place. Thus, the emulsion need not be very
stable against coalescence as the water droplets become fixed within the semisolid phase. However, the droplet size is important when considering flavor release and microbiological spoilage. Thus, emulsifiers are used to lower the
interfacial tension between the oil and water phases, which will generally result in a smaller water-droplet size. Distribution of the water droplets in the
range 2 to 4 11m will give better stability against deterioration such as mold
growth. However, it is still desirable for some of the water droplets to be larger
in size-10 to 20 11m-to give a better flavor release in the mouth.
For these purposes, therefore, lipophilic emulsifiers such as mono-/diglycerides of long-chain fatty acids (C16-C18) are used at levels of 0.1 to 0.3%,
often in combination with 0.05 to 0.1% refined soya lecithin.
Comparisons of the water-droplet distribution in margarine emulsion and in
finished margarine are shown in Figure 10.3. Droplet size in the emulsion is
further reduced during the cooling and kneading processes in the tube chiller.
The water droplets in finished margarine are stabilized by adsorbed fat
(a)
269
(b)
Figure 10.3 Water-droplet size distribution in (a) margarine emulsion and (b) the finished margarine. (Magnification 200x.) (Courtesy of Danisco Ingredients, Denmark. )
270
Figure 10.4 Freeze fracture electron micrograph showing the microstructure of margarine. Water droplets (w) are covered with fat crystals. (Bar: l!lm.) (Courtesy of Dr. W.
Buchheim, Keil.)
limited effect in reducing spattering in low-salt margarine, there are other
emulsifiers that, either alone or with lecithin, will help prevent coalescence of
water droplets during frying. In this respect, citric acid esters of mono- and
diglycerides (Citrem), polyglycerol esters, and thermally oxidized soybean oil
interacted with mono- and diglycerides used at 0.3 to 0.4%, together with soya
lecithin can be very effective.
10.8 Processing
Typical modern processing involves the separate processing of the oil and water phases. The oils and fats are kept in storage tanks at temperatures just adequate to maintain them at the required fluidity. Preferably, they should be
bottom fed to avoid splashing and aeration. It is essential that the plant, i.e.,
tanks, pipes, and pumps, be entirely free from copper or copper alloys to avoid
the high risk of oxidation.
The final oil blend together with oil-soluble components such as emulsifiers, colors, flavors, and vitamins, is prepared separately from the water phase
271
that may contain milk components such as whey powder, skim milk powder, or
sodium caseinate and also salt, gelatin, or thickener, water-phase flavors where
appropriate, and preservatives such as potassium sorbate for low-fat products.
Modern methods involve the feeding of the phases by computer-controlled
load cells, which ensures fast throughput and constant composition.
A second emulsion tank is used for maintaining quantities of finished emulsion for feeding the cooling and kneading equipment. Cooling and kneading
can be carried out by either of two different methods: (i) tube chiller or (ii)
chilling drum-complector.
In the former, cooling and kneading take place in a closed system and in a
single process. In the latter, the cooling and kneading are carried out separately-cooling on the chilling drum and kneading in the complector. The advantage of the chilling drum-complector method is that it allows the product to
rest between cooling and kneading, which is important with formulas based on
slower crystallizing fats, for instance, puff pastry margarine. By comparison,
the tube chiller is relatively more compact considering throughput, is easy to
operate, and reduces the possibility of spoilage.
Most types of margarine can be made very satisfactorily using the tube
chiller method, although in some cases extra working units (pinning machines)
may prove advantageous. However, the chilling drum-complector has been the
preferred method for puff pastry margarine, although tube chillers are now
widely used, and is perfectly satisfactory for normal table margarine and for
cake margarine. The suitability of both methods is summarized in Table 10.9.
Table 10.9
Type
Tube chiller
complector
Block/stick margarine
Soft table margarine
Low-fat spread
Cake and creaming margarine
Puff pastry margarine
Shortening
W/0 emulsion with high water
content
XXX
XXX
XXX
XXX
XXX
XXX
XX
XXX
XXX
XXX
XXX
= Excellent;
XX
= acceptable; X = poor.
272
The comparative merits of the two methods were investigated using four different fat combinations (see Table 10.10) in puff pastry margarines that were then
evaluated by various parameters including a "finger" judgement (see Table
10.11) of plasticity after one week (Madsen, TPlOl).
Table 10.10
Dilatation, iodine value, and melting points of fat blends used for
comparison of puff pastry margarines
Solid fat index
Fat blend
so
10
15
20
25
Iodine
value
l.
2.
3.
4.
37
40
43
37
37
40
42
35
34
36
37
31
27
31
29
25
22
24
23
19
67
7l
66
74
l8Ct animal
18Ct vegetable
12Ct animal
soct animal
M.P.
eq
37
37
41
37
tThe figures indicate storage and rolling temperature of the puff pastry margarine.
Madsen (fPlOl).
Source:
Table 10.11
Fat blend
Method
Plasticity judgment
Score
l. l8C animal
Chilling drum
Tube chiller
5
0
2. 18C vegetable
Chilling drum
Fine plasticity
Lumpy/gritty, a little firm,
becoming slack and very
greasy when worked
Good plasticity, tending to
become greasy
Gritty, firm, becoming slack
and greasy when worked
Fine plasticity
Tube chiller
3. 12C animal
Chilling
Drum
Tube chiller
4. soc animal
Source:
Madsen, TPlOl.
Chilling drum
Tube chiller
4
1
5
0
5
0
273
Similar results were found when the same products were judged after 3
weeks' storage.
This rather critical evaluation did not fully take into account the differences that might result from the use of varying techniques possible with tube
chillers, which was investigated separately.
Table 10.12 Plasticity judgment by means of the finger method in puff pastry
margarines made on tube chiller with varying production techniques
Production
technique
Fat blend
l. Normal
Animal
Vegetable
2. Intermediate
crystallizer
Animal
Vegetable
3. Pinning machine
Animal
Vegetable
4. Resting tube +
pinning machine
5. Low capacity
Animal
Vegetable
Animal
Vegetable
Plasticity judgment
after I week at l8C
Points
l
3
l
1
2
l
2
0
The important issue relates to the postcrystallization of the fats, which was
further investigated (Madsen, 1981 ). A good plastic puff pastry margarine can
be bent without breaking and the plasticity evaluated by repeated handworking to check stability, firmness, and greasiness (Figure 10.5).
In the case of cake margarine, the most important points are that the fat
blend is soft and easy to incorporate into the batter and has good creaming
properties. This, therefore, suggests the use of lauric fats (coconut, palm kernel) that crystallize quickly and, thereby, facilitate creaming.
274
275
of the fat blend, the type and level of emulsifier, and the addition of thickeners
such as gelatin, sodium alginate, pectin, and carrageenan to the aqueous phase.
Low levels of whey protein may be used to improve flavor release with the further
advantage that the pH of the aqueous phase can be lowered to improve keeping
properties because, unlike casein, whey proteins do not precipitate at low pH.
The speed of processing, i.e., the rate of throughput and the emulsion temperature, are also important factors in the stability of the spread.
Examples of the effect of various types of emulsifiers on the stability of lowfat spreads with fat contents of 40% and 20% are shown in Tables 10.13 and
10.14, respectively.
Table10.13
spread
Water separation % after
Levels
of use
Types of emulsifier
5 min
0.6%
1.6
2.6
9.5
15.8
l.O
2.6
6.3
0.6%
plus
0.2%
0.4%
plus
0.2%
IV = Iodine value.
Formulation of spread:
Water phase
56.4% water
0.5% whey powder
1.5% salt
0.1% potassium sorbate
Fat phase
276
Table 10.14 Effects of types and blends of emulsifiers on the stability of a 20% fat spread
Levels and types of emulsifiers
Results
IV = Iodine value.
Formulation of spread:
Water phase
69.9% water
4.0% skim milk powder
pH6.8
3.0% gelatin
Source:
Madsen (1989).
The trial batches in both cases were made on a three-tube Perfector pilot
plant using a fat blend composed of
60 parts soya oil
30 parts partially hydrogenated soya oil
lO parts coconut oil
Both examples indicate the potential instability of these emulsions and the
variation in the stabilizing effects of different emulsifier blends even when used
at low dosage levels. These results suggest that emulsifier structure at the interface plays a critical role in retarding water-droplet flocculation and coalescence.
While the evaluation of spreads by measuring water separation provides
quantifiable results, a quicker and more practical evaluation of the stability of
277
It is rather more difficult to produce low-fat butter spread from butter oil
due to the relative hardness of the fat at low temperatures. Nonetheless, spread
with a butter fat content of about 40% has been produced using 5% sodium caseinate plus 2% sodium alginate in the water phase and 0.5% distilled monoglyceride, IV approximately 55, in the fat phase.
However, in this case it is not possible to lower the pH of the aqueous phase
without precipitating the caseinate, which will reduce its emulsifying properties, and therefore, the keeping properties of the spread will be limited.
However, a satisfactory low-fat butter spread can be produced from dairy
cream, using a distilled monoglyceride with high iodine value (80 to 105) in
the cream and a thickener such as sodium alginate. In this case, phase inversion from 0/W to W/0 can be achieved in the tube chiller, using normal or
slightly reduced cooling and operating at 40 to 50% of the usual capacity. To
obtain a satisfactory working effect, a high rotor speed in the tube chiller cooling cylinder would be preferable (Madsen, 1989).
Factors that may affect the efficiency of phase inversion in low-fat emulsions are listed in Table 10.15.
278
Table 10.15
Most of these factors concern the increase in rate of collision of oil droplets
upon which the rate of coalescence is dependent (Moran, 1993). The emulsifier(s) likely provide steric hindrance to keep droplets separated.
Against this, however, one major drawback is that unless products are prepared at comparatively low pH, then ultra-high-temperature processing and
possibly an aseptic filling procedure must be followed if shelf lives comparable
to conventional spreads are required.
Oil-in-water spreads remain a relatively unexplored area of the spreadable
fats market, possibly due to the problem of microbiological deterioration, but
may offer a potential for future expansion in markets where adequate levels of
preservatives are permitted. In contrast to W/0 emulsions, these systems
would utilize high-HLB emulsifiers to stabilize the water-continuous emulsion.
279
In markets such as the UK that have traditionally used solid fats like lard or
cooking fat or, more recently, liquid vegetable oils, there is less interest in using liquid margarine.
A typical composition has a fat phase of about 82% based on soya or sunflower oil, in which emulsifiers perform two functions. First, an emulsifier such
as citric acid ester of monoglyceride (Citrem) at, say, 0.4% plus 0.2% soya
lecithin produces a stable water dispersion with limited spattering during frying; and second, a selected mono- and diglyceride blend prevents oiling out on
storage and thus gives a homogenous product with low viscosity. If the margarine is not intended for frying, the Citrem can be replaced with 0.2% distilled monoglyceride. The water phase may contain proportions of skimmed
milk powder and salt as well as a preservative such as potassium sorbate.
Flavoring may be added to both phases.
The aqueous phase should be adjusted to 4.5 to 6.5 pH and pasteurized at
80C. The fat phase should be tempered to about 60C and the emulsifiers
melted into a small amount of the liquid oil to a temperature of 65 to 70C,
which is then stirred into the main part of the fat phase. The water phase is
added to the fat phase under continuous agitation before cooling through the
tube chiller with an outlet temperature of approximately 5 to 7C.
The emulsion should be rested for 2 hours to allow proper fat crystal formation and then stirred vigorously for 15 minutes before topping off for packaging.
10.12
Summary
Standard margarines containing 80% fat are, under normal conditions, relatively stable, requiring minimal quantities of lecithin and/or mono- and diglycerides over and above the quantities of milk proteins usually present.
However, where the margarine is required for special purposes, i.e., cakemaking or frying and particularly in products with reduced fat levels, emulsifiers specific to the unique functional requirements of the application should
be selected (see Chapter 8 for applications of emulsifiers in baking products).
References
Andersen, A.J.C., Williams, P.N. (1954). Margarine, 2d ed., Pergamon, Oxford.
Borwanker, R.P., Buliga, G.S (1989). Emulsion properties of margarines and low fat
spreads, in Proc. Food Emulsions and Foams-Theory and Practice (eds. P.G. Wan
eta!.), San Francisco, pp. 44-52.
280
Flack, E. (1992). The role of emulsifiers in reduced fat and fat free foods, in Food
Technology International Europe (ed. A. Turner), Sterling Publications, London, pp.
179-181.
Friberg, S.E. et al. (1990). Emulsion stability, in Food Emulsions (eds. K. Larrson and
S.E. Friberg), Marcel Dekker, New York, pp. 1-40.
Garti, N., et al. (1982).]. Am. Oil Chem. Soc., 59, 181.
Krog, N., et al. (1988). Applications in the food industry, in Encyclopaedia of Emulsion
Technology, Vol. 2: Applications of Emulsions (ed. P. Becher), Marcel Dekker, New
York, p. 321.
Madsen, J. Puff Pastry Margarine: A Comparison of the Chilling Drum and Tube Chiller
Methods of Production, TP101, Grinds ted, Denmark.
- - - (1981). Post-Crystallisation in Puff Pastry Margarine, at 11th Scandinavian
Symposium on Lipids, Bergen, Norway.
- - - (1983). Product Formulation and Processing of Margarine and Yellow Fat
Spreads, at Margarine and Yellow Fat Seminar, Coventry, England.
- - - (1989). Low Calorie Spread and Melange Production in Europe, at World
Conference on Edible Oils and Fats Processing, Maastricht, Holland.
- - - , Als, G. (1968). Sandiness in Table Margarine and the Influence of Various
Blends of Triglycerides and Emulsifiers Thereon, at /Xth Int. Soc. of Fat Research
Congress, Rotterdam.
Moran, D.P.J. (1993). Reduced calorie spreads, in Porim Technology, No. 15., Feb '93,
Palm Oil Research Institute of Malaysia.
Schwitzer, M.K. (1956). Margarine and Other Food Fats, Leonard Hill, London.
ELEVEN
Emulsifier Trends
for the Future
Gerard L. Hasenhuettl
As in many other industries, forecasting the future is usually done by examining the trends of the past and extrapolating the changes for several years into
the future. While this works well for continuous functions, discontinuous
events often produce dramatic changes in market structures. Forecasting technology is essentially an observation of technical events, extrapolating the
trends, and trying to estimate which technologies are about to jump to the next
S curve. Food technology abhors radical change. Development of new crops is
contingent on being able to diffuse the crop into the massive agronomic supply
chain. Issues such as cross-pollination and identity preservation may pose significant problems. When a new product is brought to market, it needs to be
thoroughly tested or examined for its safety. Programs like HACCP are aimed
at institutionalizing safety into the industry. Even if the government is assured
of a product's safety, consumer acceptance is by no means assured.
Controversies around genetically engineered and irradiated foods are testimony to this principle.
As we stated in Chapter l, development of totally new emulsifiers is unlikely because of the time and cost involved with regulatory approval.
However, this by no means sounds a death rattle for innovations in the field of
281
282
11.1
283
part of the physical structural or primary functionality of food. For example, the
development of high liquid-oil margarines will require an emulsifier that can stabilize a W/0 emulsion where the continuous phase is liquid. Adjunct technologies, such as the development of molecular networks to thicken the oil phase can
make a significant contribution to providing structure to the processed food.
Solid fat is implicated in such functions as aeration of whipped toppings and formation of evenly dispersed air cells in cakes. Replacement with liquid oil places
more of a functional burden on the emulsifier system.
Health authorities have recommended that total dietary fat should not exceed 30% of calories. Some consensus is also building that some forms of cancer may correlate to total caloric consumption. Since fat represents 9 cal/gram,
cutting fat consumption will also reduce total calories (if the same weight of
food is consumed). If consumers respond to these recommendations, pressures
for availability of reduced-fat and fat-free foods will continue or even increase.
Elimination of fats from products normally containing high levels creates enormous challenges to maintain texture and flavor. Flack (1992) has speculated
that emulsifiers can play an important role in reclaiming some of these qualities. Structured emulsifier phases have been patented as the basis for formation of a pseudo-fat phase (Heertje, et al., 1994; Kleinherenhrink et al., 1995).
This trend toward replacement of fat functionality with emulsifiers will undoubtedly continue.
284
very efficient surfactants for application such as enhanced oil recovery and oilspill control. A logical extension of these technologies would be to develop
food-grade organisms that produce natural surfactants. However, extensive
evaluation would still be essential to ensure the safety of these products when
consumed in processed foods.
Interfacial
(hydrophobic)
attraction
Volume v
-._ \
Area a 0
Figure 11.1
Table 11.1
285
Lipid type
Critical
packing
shape
Critical
packing
parameter (VIaJ.)
Structures
formed
Single-chain lipids
with large head
group areas
Single-chain lipids
with small head
group areas
Double-chain lipids
with large head
group areas
Double-chain lipids
with small head
group areas
Double-chain lipids
with small head
group areas
< l/3
Cone
Spherical micelles
l/3-l/2
Truncated cone
Cylindrical micelle
(hexagonal)
l/2-l
Truncated cone
Flexible bilayers,
vesicles
-l
Cylinder
Planar bilayers
(lamellar)
>l
Inverted truncated
cone
Inverted micelles
the functional role of a mesophase is defined, rational design of head and tail
groups can he accomplished and the emulsifier synthesized by techniques described in Chapter 2.
Emulsifiers may he viewed as one class of functional food ingredients. As
Chapters 4 and 5 have pointed out, these ingredients may interact with carbohydrates and proteins to produce a number of technical effects. The area of ingredient interactions is the subject of a recent book by Gaonkar (1995). New
experimental techniques such as scanning tunneling microscopy and interfacial rheology have the potential to provide greater understanding of these interactions. Emulsifiers can then be designed to form more selective interactive
structures that may result in improved or entirely new functionalities.
11.5
Production of emulsifiers involves contacting lipid and polar phases that are
mutually insoluble. To achieve homogeneity, temperatures as high as 500F
are applied and held until reaction is complete.
286
catalyzed reactions in monolayers and microemulsions. The major disadvantages of enzyme-catalyzed interesterification are high enzyme cost (relative to
sodium or calcium hydroxide catalysts), longer reaction times, and the tendency of enzymes to denature. However, the problems are surmountable and
the use of enzyme technology is expected to increase.
References
Arcos, J.A., Otero, C. (1996). ]. Am. Oil Chem. Soc., 73:673-82.
Flack, E. (1992). Food Techrwlogy International-Europe, 79-81.
Gaonkar, A. (ed.) (1995). Ingredient Interactions: Effects on Food Quality, Marcel
Dekker, New York.
Heertje, 1., et al. (1994). European Patent 0 558 523 Bl, July 13.
lsraelachvili, J.N. (1985). Intermolecular and Surface Forces: With Applications to
Colloidal and Biological Systems, Academic, New York, pp. 246--62.
Kleinherenbrink, F.A., et al. (1995). International Patent Application WO 95/35035,
December 28.
Larsson, K. (1994). Lipids-Molecular Organization. Physical Functions and Technical
Applications, The Oily Press, Ayr, Scotland, pp. 156--62.
Shah, D.O., et al. (1994). ]. Am. Oil Chem. Soc., 71:1405-9.
Vulfsen, E.N. (1993). Trends Food Sci. & Techno!., 4:209.
Index
Index
acetylated diglycerides, 33
acetylated monoglycerides, 33, 44,
60,253
preparation of, 33-36
acetylated monoglyceride citrate,
285
33-36
289
290
Index
B
baked products:
cream icings, 232
crumb-softening, 226
emulsifiers, 216
applications, 225
extruded snacks/cereals, 232
fat-free products, 232
dough conditioning, 225
monoglycerides in, 217, 218
production influences, 229
shortening, 215
yeast in, 217
Bancroft rule, 162, 166
birefringence, loss of, 78
bovine milk, 173
composition of, 174
bovine serum albumin 98 102 113
' '
'
'
126
bread dough, 85
c
cake batters, 212, 230
calcium stearoyl-2-lactylate, 20, 57
calcium hydroxide, 17
capillary melting point, 53
Index
291
0
dairy applications, emulsifiers, 202
decaglycerol decastearate, 25
decaglycerol tristearate, 25
derivatives, bakery products, 217
diacetyl tartaric acid esters of:
monoglycerides (DATEM)
292
Index
233,249,266
differential scanning calorimetry
(DSC),53,87,96,131
diglycerides, 6, 15, 23, 25, 33, 40,
41,49,55,60,182,198,
200,204,212,227,240,
249,250,268,274,279
hydroxyl values, 23, 24
interesterification process, 15, 17
diacetyl tartaric acid, 31
dimyristoylphosphatidic acid, 126
dioleoyl PC, PE, 160
dioleoylphosphatidylserine, DOPS, 122
dioleoylphosphatdylcholine, 125
dioleoylphosphatidylethanolamine, 125
dipalmitoylphosphosphatidic acid, 121
diphosphatidylglycerol, 122
dipalmitoylphosphatidic acid, DPPA,
123
dipalmitoylphosphatdylcholine, 126
distearoylphosphatidic acid, 121, 123
DMPG, 130
dodecyl sulfate, SDS, 78
DOPG, 130
DTAB, adsorption, 100
E
egg yolk, 123
electron microscopy, 2, 266, 269, 270
ellipsometry, 121, 171
emulsifiers:
anionic, 219
analytical methods for, 39
baked foods, 211-216
bakery products, role of, 212-214
breakdown, process of, 95
bilayers, 154
binary mixture, liquid-crystalline,
150
carbohydrate interactions, 67
l3C chemical shifts, 61
chocolate confectionery, 235, 236
classified as, l l
classification, 161
coalescence, 6, 176
color of, 50
compound coatings confectionery,
236
consistency, 54
crumb softeners in bread, 70
dairy applications, other, 202
determination of, 40--52
destabilization, whipped toppings,
189
dispersion, stabilize, 154
emulsifier/starch-complex formation, 69, 79
enzyme synthesis, 286
fat, crystal structure of, 180
flocculation, 6
food additives, 2-5
functionality, 7-9
future trends, 281-286
hexagonal phase, 149
high-melting, 148
ice cream, 175
instrumental methods, 54--62
interaction, ingredients, 284--285
interfacial viscosity, 154
lactoglobulin, 114
lamellar phase, 149
margarine, function of, 267-270
melting point of, 52-54
mesophases, 284--285
micelles, 149, 154
Index
163-164
layers for, 170
phase inversion, 162
solubility, 161
emulsifier interaction, bakery components, 221
emulsifier surface, 167-169
emulsion stability, 154--155
emulsion phase diagrams, 154--155
entropy, gain of, 148
enzymatic synthesis, food, 285
293
enzymolysis in starches, 81
ethoxylated esters, 11, 26, 169, 266
sorbitan monostearate, 26
sorbitan monoolate, 27-28
sorbitan tristearate, 28
mono- and diglycerides, 28
surfactants, 169
preparation of, 28--29
polyoxyethlene (20) sorbitan, 28
ethoxylated fatty alcohols, 266
ethoxylated monoglycerides, 228
ethoxylated nonyl-phenol and xylene,
153
evaporated milk, 201
European Economic Community
(EEC), 40
baked goods in, 214
extruded snacks/cereals, 232
F
fat content, solid, SFC, 180
crystallization:
margerine, 263-267
polymorphic, 263-265
whipped toppings, 189
fat:
crystals of:
bakery products, 217
margarine, 266
effect of emulsifiers on,
266-267
crystallization:
confectionery, 239
chocolate, 248
margarine, 265
fat, yellow, consumption, 257
fat-free baked products, 232
294
Index
32
hydroxyl, 26
propylene glycol, 18
sorbitan monostearate, 26
succinylated monoglycerides,
30
Reichert-Meiss! value, 34
form stability, bakery products, 217
Fourier transform infrared spectrosopy, FTIR, 59
food emulsifiers, solution properities,
155-156
free fat acids, 60
measuring, in olive oil, 60
freeze fracture, margarine, 270
fruit acid esters, 11, 31-33
tartaric acid-derived esters, 31
fudge, 251-252
G
gas-liquid chromatography GLC), 46,
55-57
Gibbs effect, 110
performed on lipids, 55
GLC, 56
gliadin, 132
glyceryl monostearate (GMS), 86,
197
gelatin, 134
gelatinization, 78, 89
gelatinization endotherm of starches,
88
globalization, food industry, 282
globular proteins, interior, 97
hydrogen bonding, 98
hydrophobic interreaction of, 98
van der Waals interaction, 98
gluten structure, 215
180-18,189,236,248,
250,251,252
glycerol monooleate, 189
GRAS, 2-3, 15, 26
monosodium phosphate, 36
status of, 32
gums, 251
H
helix starch, structure of, 79
hexaglycerol dioleate, 25
hydroxyl value, 48
high-performance liquid chromatography (HPLC), 47, 52,57-58
detectors of, 58
Index
separations of phospholipids, 58
homogenization, 166
hydrated distilled monoglycerides, 227
hydrodynamic interactions, 166
hydrophile/lipophile balance (HLB),
6-8,151,163-164,169,
220,236
hydrophilic, 5, 11, 100
hydrated lime, 17
hydration:
force, emulsion droplets, 155
repulsion, monopalmitin, 155
295
J
jellies, 251
ACI values, 252
K
kosher, 5, 13
Krafft temperature, 148
296
Index
173,199,202,220,223,
228,231,233,236,
238-239,248,250,253,
268,274
phosphorus, procedures for, 51-52
phosphatidyl, 171
phosphatidylcholine content in,
60,155
soy lecithin, 51, 239
synthetic, 240
viscosity, 54
linear molecules, starches, 68
lipid emuisifiers, liquid-crystalline,
156
lipid fractions, topping emulsions,
190
lipid materials, effects of 69, 96
lipid/protein interaction, 122
lipid structures, 129, 133
lipids in baked foods, 225
lipophilic, 5, ll
lipolytic emzymes, 133
liposomes, 84
liquid chromatography, 42
liquid-crystaline mesophases,
whipped toppings, 191
liquid margarine, 278-279
low-fat spreads, 274-278
low molecular weight (LMW), 96
lysolecithin, 84
lysophosphatidyl cholines (LPC),
119,159,283
lysophosphatides (LPC), (LPE), 156,
158
lysozyme, 102, 104
M
Marangoni effect, Ill
margarine, l, 255, 258-279
emulsifiers, 267-270
liquid margarine, 278
low-fat spreads, 274-276
market share, United States, 261
oil-in-water (0/W) spreads, 7, 278
phase inversion, 277-278
processing, 270-27 4
puff pastry margannes, 272-274
plasticity, 272-427
reduced fat spreads, 6, 274
spreadability, 277
structure, 262
fat crystallization, 263, 265-266
fat melting points, 272
polymorphism, 263-265
oils and fats, 263-426
melting points, 265
raw materials, 262
water-droplet distribution,
268-269
water-in-oil (y//0) emulsion, 7,
268,274,277-278
mass spectrometry, 60
mayonnaise, l
melting point, 12, 52-53
melittin interactions, 125
Mettler dropping point, 53
micelles, 84, 148, 165
microemulsion process, 37
milk, l, 173
composition of bovine milk, 174
concentrated, 199-202
fat globule membrane (MFGM),
173,184,185
proteins, 122, 135
recombined, 199-201
Index
21-22,25,40-44,55-57,
59--61, 134, 149,
154--156, 178, 182,
198--200, 202, 212,
217-219,227,230--1240,
248--250,252,268--270,
274,279
acetic acid esters of, 187, 236,
249,252-253
acetylated, 33, 44, 60
preparation of, 33-36
297
N
near infrared reflectance (NIR), 46,
48,59
nougat, 250
nuclear magnetic resonance (NMR),
2,59,60
266,270,279
diacetyltartaric acid esters of
(DATEM), 3, 31-33, 41,
44,59,228,249,252,266
distilled, 22, 30, 233
distilled saturated, 156
distilled unsaturated, 156
ethoxylated, 231, 250
lactic acid esters of, 20--22, 44,
187,230,236,248
phase behavior, 178
phases of:
cubic phase, 156
lamellar phase, 156
reversed hexagonal, 156
phosphated, 3, 11, 236
propylene glycol esters of, 60, 187
stearyl monoglyceride citrate, 33
succinylated (SMG), 30, 213, 227,
278,231
tartaric acid esters of, 197, 236
0
oil-continuous emulsions, 154
oleic acids, 11-13, 256
oleomargarine (see margarine)
oleostearine, 256
organic acid esters, 156
diacetyl tartaric acid, 156
monoglyceride ester, 156
orthokinetic stability, 177
Oswald ripening, 95
olalbumin, 102
p
packing parameter, 165
palmitic acids, 11-13, 256
palmitoyloleoylphosphatidylcholine,
123
298
Index
palmitoyloleoylphosphatidylglycerol,
123
para-K-casein, 124
peanut butter, 7
penetrometer, 54
pentosan, 214
peptide chain, 97
peroxide value, 46, 4 7
phase diagrams, 153, 166
emulsion stability, 154
lamellar liquid crystalline phase,
153
water and dioleoylphosphatidylcholine, 157
phase inversion, concept, 162
phase inversion temperature (PIT),
162
phosphated esters, 36
monosodium phosphate, 36
preparation of, 36
phosphatides, mixtures of, 158
phosphatidic acids, 122, 158, 240
dioleoyl, 159
lysophosphatidylcholine (lyso PC),
159
phosphatidylglycerol, 135
phosphatidylcholine (PC), 52, 57-61,
122,130,155,157-159
dimyristoyl, 159
dioleoyl, 159
dipalmitoyl, 159
distearoyl, 159
egg, 159
soybean, 159
transition temperature for, 158
phosohatidylserine, 122
phosphatidylethanolamine (PE), 52,
dipalmitoyl, 159
soybean, 159
phosphatidylinositide, 173
phosphatidylinositol (PI), 122, 156,
158,159
soybean, 159
phosphatidyl serine (PS), 156, 173
phospholipids, 41, 42, 52, 57-61,
159,166,171,173,186,
200,202
liquid-crystalline phases, 159
phosphoric acid, 15, 17
phosphorus pentoxide, 240
photobleaching, fluorescence
recovery, 113
polar lipid:
classes, 127
cubic phase, 127
lamellar phase (LJ, 127
liquid crystalline phase, 127
hexagonal phase, 127
hydrophobic interaction, 127
micelle concentration (cnc ), 127
phase behavior, 126
surfactants, 127
structure, 128, 129
plasmalogens, 173
plasticity, 5
polyglycerate 60, 251
polyglycerol, 59
polyglycerol esters, 3, 11, 23, 54, 58,
59,213,233,249,266,
270
of stearic acid, 248
preparation of, 23-25
polyglycerol polyricinoleate (PGPR),
8,56,236,242-244
polyhydric emulsifiers, 219
polyglycerol esters, 219
sucrose esters, 219
Index
polylactic acid, 20
polymerization, 242
polymorphic form, 245, 246
polyoxyethylene (20), sorbitan monostearate, 28, 29, 114, 115,
118,156,182,220,232
polyoyl, 56
polyphosphates, 198
polysorbates, 3, 29, 59, 61, 178, 182,
219,236,250
polysorbate 60, 29, 213, 220, 227,
230,233,236,247-248,
250,252
processed cheese, 197-199
manufacture of, 198
programmed temperature x-ray diffraction, 53
propylene glycol, 17
propylene glycol esters, 15, 17, 23,
55,232
propylene glycol lactates, 23
propylene glycol monoesters
224
protein/emulsifier:
interactions, 95-135
food applications, 132
lipolytic enzymes, 132, 133
lipid structure, 133
protein stability, adsorption, 97
299
protein, lipid:
interactions, 126
monolayers, 125
protein hydrolysates, 149
protein/phospholipid interactions, 120
dispersed systems, 120, 121
solid surface interactions, 121
protein stability, 97
protein/surfactant interactions, 98--119
adsorption:
binding to, 107
competive, 111
cooperative, 107
isotherm, 99
mixtures of, 106
adsorbed:
surface properties, 103
complex formation, 105
replacement of, 105
binding, protein and emulsifiers,
113
coadsorption, 112
dissociation constant (Kd),113
emulsifier properties, 134
foam, emulsion stabilization, 110
hydrophobic surfaces, 103
hydrophilic surfaces, 103
Krafft temperature, 103
liquid air, 110
liquid/liquid interfaces, 110
lactoglobulin, 117
electrostate repulsion, 103
monolayer of, 99
nonionic surfactants, 106
interactions, solid interfaces, 106
ionic surfactants, 99, 106
solid surfaces, 98
solution influence, 107
micelles, critical concentration of,
99
300
Index
122
protein/protein interactions, ll2
puddings, 77
puroindolines, ll9
R
refractive index, 54
regulatory agencies, 40
Reichert-Meiss! value, 34, 49
reverse osmosis, 201
reversed aggregates, 165
s
saponification number, 26, 4 7
sauces, 77
Scatchard equation, ll6
shelf life, 200
milk, 200
salad dressings, 2
shortening, 2ll-216
emulsified, 2ll-213
fluid, 230
melting point, 216
nonemulsified, 213
plasticity, 216
solid-fat index, 216
sodium caseinate, 192
sodium dodecylsulfate, SDS, 98
sodium hydroxide, 17
sodium lauryl sulfate (SLS), 78, 134
sodium hydroxide, 17
221,231
sodium stearoyllactylate (SSL), 20,
41,42,44,156,197,203,
213,219,221,224,227,
228,230-232,266,283
softening point, 53
solubility, 161
Bancroft rule, 162
sorbide, 26
polyoyls, 26
sorbitan, 26
polyoxylated (20), 231
monostearate (SMS), 26, 56, 219,
230,231,247,248,250,
252
tristearate (STS), 56, 246, 24 7,
266
sorbitan esters, ll, 26, 162, 164,
169,220,233,266
ethoxylated, 26, 28, 266
production, 26
polyxyethylene (20), 156
sorbitan monooleate, 156
sorbitan monostearate, 156, 247
sorbitan stearate, 156
soy lecithin, 8
soybean phospholipids, 161
phase diagram, 161
spectroscopy, 59
sphingomyelin, 125, 173
spread ratios of cookie dough, 231
spreadable fats, 259, 260
starch:
complexes, in baked foods, 222
complexing, 77, 214
differential scanning calorimetry, 85
Index
83,85-90
electron spin responce, 85
infrared spectroscopy, 85
pH effects, 83
temperature, 83
paste gelation, 79
unsaturated fatty acid monoglycerides, 224
waxy, table of, 75
starch/emulsion complexes:
V-type x-ray diffraction pattern, 69
statistical experimental design, 8
fractional factorial, 8
301
213,227,236
preparation of, 36--38
sucrose monopalmitate, 231
sucrose monostearate, 231
sucrose distearate, 231
sucrose monostearate, 266
sucrose polyesters, 233
Olestra, 233
surface activity, 148, 167
components, 167
polar interactions, 148
surfactants, 5
surface tension, emulsifiers, 147,
227
surfactants, 168, 199, 201, 212
anionic, 199, 201, 224
cationic, 199
cetyl-trimethyl ammonium
bromide (CTAB), 199
ionic, 99, 100, 168
interaction of, 101
lecithin, 199
molecules, arraingments of, 101
nonionic, 168, 201, 224
properties, 102
sodium dodecyl sulphate (SDS),
199,201
solidlaqueos interfaces, 99
sucrose ester, 223
zwitterionic, 199
302
Index
T
tetraglycerolesters, 156
monolaurin, 156
tetraglycerol, 156
thin film stabilization, 111, 112
tocopherols, 50
toffee, 250
triglycerides, 2, 12-5, 17, 204, 240,
263
glycerolysis of, 204
triglycerol, 23
triglycerol monooleate, 248
triglycerol monoshortening, 250
triglycerol monostearate, 248, 252
triglycerol monoshortening, 25
triglycerol monostearate polyglycerol
ester, 25
tripropylene glycol monoesters, 18
u
ultrafiltration, 201
w
water-continuous emulsions, 154
water-insoluble combined lactic acid
(WICLA), 49
waxy starch table, 75
wet chemical analysis, 43-50
titration, 43-49
wheat flours, 214
whipped cream, whipping cream,
183,185
electron micrograph, 188
recombined, 185, 186
whipped toppings, 187
emulsion destabilization, 189
fat crystallization, 189
powder composition, 187
protein desorption, 189
Wiley melting point, 53
v
van der Waals interaction, 98
vesicle bilayers, 131
viscosity, 8, 54,177,237,241-244 ,
248
apparent, 237
interfacial, 177
plastic, 237, 243, 244