11 Emulsification: Vegetable Proteins K A Y H. McWATTERS Department of Food Science, University of Georgia College of Agriculture Experiment Station, Experiment, G A 30212 JOHN P. CHERRY Southern Regional Research Center, Science and Education Administration, Agricultural Research, U.S. Department of Agriculture, P.O. Box 19687, New Orleans, L A 70179
Emulsifiers are classified as a group of surface-active agents that can stabilize a dispersion of two immiscible liquids such as water and oil. In an emulsion, the dispersed droplets are commonly referred to as the disperse, discontinuous, or internal phase, and the medium in which they are suspended is the continuous, or external phase. The most common types of emulsions are oil-in-water (e.g., mayonnaise or milk) and water-in-oil (e.g., butter or margarine) and are classified according to the component that comprises the disperse phase. The terms "oil" and "water" are used in a general manner; almost any highly polar, hydrophilic liquid falls into the "water" category whereas hydrophobic, nonpolar liquids are considered "oils" (1). In the realm of food emulsions, the classic definition has been broadened to include two-phase systems of dispersions of gas in a liquid (e.g., foams) and combinations of liquids, solids, and gases in emulsions or batters (2). Theories of Emulsions and Emulsifier Mechanisms Several d e t a i l e d d i s c u s s i o n s have described the complex theories of emulsion technology (1, 3 , 4 , 5 ). To summarize these t h e o r i e s , e m u l s i f i e r s are e s s e n t i a l f o r emulsion formation and s t a b i l i z a t i o n to o c c u r ; these s u r f a c e - a c t i v e compounds reduce the surface and i n t e r f a c i a l tensions between two immiscible l i q u i d s , but t h i s property accounts f o r only part of the mechanisms a t work i n e m u l s i f i c a t i o n . Three separate mechanisms that appear to be involved i n formation of a s t a b l e emulsion i n c l u d e : 1) reduction of i n t e r f a c i a l t e n s i o n , 2) formation of a r i g i d i n t e r f a c i a l f i l m , and 3) e l e c t r i c a l charges. Molecules i n l i q u i d s e x h i b i t d i f f e r e n t behaviors, depending upon t h e i r l o c a t i o n i n the volume of l i q u i d . Those i n the i n t e r i o r are surrounded by other molecules having balanced a t t r a c t i v e forces. Those a t or near the s u r f a c e , however, are only p a r t i a l l y surrounded by the other molecules of the l i q u i d , and the a t t r a c t i v e forces operating on them are unbalanced. There i s a net
0097-6156/81/0147-0217$06.50/0 © 1981 American Chemical Society
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a t t r a c t i o n toward the main body of the l i q u i d as the l i q u i d tends to reduce i t s surface to a minimum, thus e x h i b i t i n g the behavior o f the s u r f a c e known as surface t e n s i o n . S u r f a c e - a c t i v e agents are important i n t h i s regard because they are p o s i t i v e l y adsorbed at the surface of immiscible components and reduce the surface tension between them. The manner i n which the molecules of s u r f a c e - a c t i v e agents arrange themselves depends upon the d i s t r i b u t i o n of e l e c t r o n s forming the bonds between the atoms of the molecule or p o r t i o n of the molecule. H y d r o p h i l i c (water-loving) molecules have asymmetrically placed e l e c t r o n s and e x h i b i t p o l a r p r o p e r t i e s ; hydrophobic or l i p o p h i l i c molecules have symmetrical arrangements and e x h i b i t nonpolar p r o p e r t i e s . Surface-active agents are compounds that contain both h y d r o p h i l i c and hydrophobic groups and t h e r e f o r e e x h i b i t polar-nonpolar c h a r a c t e r . In e f f e c t , a monolayer that prevents c l o s e contact between d r o p l e t s o f the d i s p e r s e phase i s c r e a t e d ; a s u r f a c e - a c t i v e agent aids i n preventing d r o p l e t coalescence. In a d d i t i o n to lowering surface t e n s i o n , s u r f a c e - a c t i v e agents c o n t r i b u t e to emulsion s t a b i l i t y by o r i e n t e d adsorption at the i n t e r f a c e and by formation of a p r o t e c t i v e f i l m around the droplets. Apparently, the f i r s t molecules of a s u r f a c t a n t i n t r o duced i n t o a two-phase system a c t to form a monolayer; a d d i t i o n a l s u r f a c t a n t molecules tend to a s s o c i a t e with each o t h e r , forming m i c e l l e s , which s t a b i l i z e the system by h y d r o p h i l i c - l i p o p h i l i c arrangements. This behavior has been depicted by Stutz e t a l . (6) and i s shown i n Figures 1 - 5 . Petrowski ( 7 J summarized the e f f e c t s of e l e c t r i c a l charge or balance between a t t r a c t i v e and r e p u l s i v e f o r c e s of p a r t i c l e s on emulsion s t a b i l i t y . The r e p u l s i v e f o r c e s , e l e c t r o s t a t i c i n nature, are s t a b i l i z i n g because they tend to keep d r o p l e t s separated, thereby preventing coalescence. The arrangement of the e l e c t r i c a l charges a t the i n t e r f a c e has been the subject o f some controversy; i t has been described as a double l a y e r of opposite-charged ions or as a s i n g l e , immobile counter-ion l a y e r surrounded by a d i f f u s e l a y e r o f mobile ions extending outward. The r e p u l s i v e i n t e r a c t i o n of d r o p l e t s possessing l i k e e l e c t r i c a l charge seems to c o n t r i b u t e to emulsion s t a b i l i t y . Electrical charges on d r o p l e t s i n emulsions can a r i s e by i o n i z a t i o n , absorpt i o n , or f r i c t i o n a l e l e c t r i c i t y produced by the large shearing f o r c e s required f o r emulsion formation. V i s c o s i t y i s an important physical property of emulsions i n terms of emulsion formation and s t a b i l i t y (1_, 4 J . L i s s a n t [Vj has described several stages o f geometrical d r o p l e t rearrangement and v i s c o s i t y changes as emulsions form. As the amount of i n t e r n a l phase introduced i n t o an emulsion system i n c r e a s e s , the more c l o s e l y crowded the d r o p l e t s become. This crowding of d r o p l e t s reduces t h e i r motion and tendency to s e t t l e while imparting a "creamed" appearance to the system. The apparent v i s c o s i t y continues to i n c r e a s e , and non-Newtonian behavior becomes more marked. Emulsions of high i n t e r n a l - p h a s e r a t i o are a c t u a l l y i n a "supercreamed" s t a t e .
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Food Product Development
Figure 1. Orientation of surfactant molecules at water-air interface (6) \
AIR
Food Product Development
Figure 2. Monolayer coverage (6)
Food Product Development
Figure 3.
Beginning of micelle formation (6)
Figure 4. Micelle cross section (6)
H,O; Food Product Development
Oil
no
Figure 5. Water-in-oil emulsion stabilized by a surfactant Cupper left,) and oilin-water emulsion stabilized by a surfactant (lowerright)(6)
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Proteins c o n s t i t u t e an important group of e m u l s i f i e r s because they behave i n a manner s i m i l a r to that of s u r f a c e - a c t i v e agents by forming strong monolayer f i l m s a t the i n t e r f a c e . Their s t r u c t u r e , and hence t h e i r behavior, i s more a f f e c t e d by such v a r i a b l e s as s a l t c o n c e n t r a t i o n , pH, and temperature than would occur with the use of conventional e m u l s i f i e r s (7). Vegetable p r o t e i n s , along with those derived from d a i r y and meat sources, have been i n t e n s i v e l y studied i n recent years i n e f f o r t s to assess t h e i r p o t e n t i a l as f u n c t i o n a l i n g r e d i e n t s . Because several studies and reviews describe emulsifying properties of soy p r o t e i n in d e t a i l (8-15), the emphasis i n t h i s d i s c u s s i o n w i l l be d i r e c t e d toward vegetable proteins other than soy. Experimental
Procedures
The technique most commonly employed f o r measuring emulsion c a p a c i t y of proteins i s the model system, or m o d i f i c a t i o n s of i t , developed by Swift et a l . (16). This procedure involves the continuous a d d i t i o n of o i l or melted f a t to a p r o t e i n d i s p e r s i o n during high-speed mixing; o i l i s added u n t i l a sudden drop i n emulsion v i s c o s i t y occurs due to separation of o i l and water i n t o two phases. The volume of o i l added u n t i l the "breakpoint" i s reached i s used to express emulsion capacity of a p r o t e i n ; these values may be expressed as t o t a l ml of o i l e m u l s i f i e d , ml of o i l per u n i t weight of sample, or ml of o i l per u n i t of p r o t e i n or nitrogen i n the sample. Comparisons of r e s u l t s obtained from d i f f e r e n t studies i s d i f f i c u l t because small v a r i a t i o n s i n technique, equipment, blender speeds, r a t e of o i l a d d i t i o n , prot e i n source and c o n c e n t r a t i o n , temperature, or type of o i l a f f e c t emulsifying properties of proteins (17, 18, 19). Model systems are useful t o o l s , however, because c o n d i t i o n s and i n gredients can be well defined and c a r e f u l l y c o n t r o l l e d ; they are a l s o simpler and more economical to use than more complicated systems. No standardized t e s t s e x i s t f o r evaluating the emulsifying properties of p r o t e i n s , and i n many cases there seems to be l i t t l e c o r r e l a t i o n between r e s u l t s obtained i n model systems and those obtained i n performance t r i a l s i n food systems. The importance of designing model t e s t systems that simulate actual food-processing c o n d i t i o n s as c l o s e l y as p o s s i b l e has been emphasized by Puski (20) f o r o b t a i n i n g meaningful i n formation regarding emulsifying properties of p r o t e i n s . Emulsifying
Properties of Selected Vegetable Protein Sources
Peanut Seed. Ramanatham et a l . (21_) studied the i n f l u e n c e of such v a r i a b l e s as p r o t e i n c o n c e n t r a t i o n , p a r t i c l e s i z e , speed of mixing, pH, and presence o f sodium c h l o r i d e on e m u l s i f i c a t i o n p r o p e r t i e s of peanut f l o u r (50% p r o t e i n ) and peanut p r o t e i n i s o l a t e (90% p r o t e i n ) . Emulsions were prepared by the blender
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method p r e v i o u s l y described (16); d i s t i l l e d water at pH 6.8 comprised the continuous phase. For those t e s t s i n which f l o u r concentration was held constant, as i n the p a r t i c l e s i z e and mixing speed t e s t s , a l e v e l of 5 g f l o u r dispersed i n 50 ml d i s t i l l e d water at pH 6.8 was used. Data i n Table I show that emulsion capacity of peanut f l o u r decreased with i n c r e a s i n g f l o u r or p r o t e i n concentration while emulsion v i s c o s i t y i n c r e a s e d . This phenomenon was a l s o demons t r a t e d by McWatters and Holmes (22). A decrease i n f l o u r p a r t i c l e s i z e increased emulsion capacity and v i s c o s i t y a p p r e c i a b l y . Increasing the r a t e of mixing, however, decreased emulsion capacity but increased v i s c o s i t y . Increased speeds produce greater shear r a t e , which decreases the s i z e of the o i l d r o p l e t ; thus, there i s an increase i n the surface area of the o i l to be e m u l s i f i e d by the same amount of s o l u b l e p r o t e i n [23 24). Data i n Figure 6 show the e f f e c t of varying the pH and sodium c h l o r i d e concentration on emulsion capacity of peanut p r o t e i n isolate. S h i f t i n g the pH to l e v e l s above or below the i s o e l e c t r i c point improved emulsion capacity of peanut p r o t e i n i s o l a t e i n 0.1M or 0.2M NaCl. S i m i l a r trends were noted when d i s t i l l e d water was used as the continuous phase (data not.shown). At the 0.5M NaCl c o n c e n t r a t i o n , however, l i t t l e d i f f e r e n c e was noted i n emulsion capacity at pH 3 , 4, or 5; appreciable increases occurred when the pH was r a i s e d to 6 and above. At the highest s a l t concentration (1.0M NaCl), a gradual increase i n emulsion capacity occurred when the pH was increased from 3 to 10. An o v e r a l l suppression i n emulsion capacity occurred as s a l t concentration increased except at pH 5 and 6. These emulsion-capacity curves c l o s e l y resemble the p r o t e i n - s o l u b i l i t y curves of peanut p r o t e i n shown i n Figure 7 (25), i n d i c a t i n g that f a c t o r s t y p i c a l l y i n f l u e n c i n g peanut p r o t e i n s o l u b i l i t y a l s o i n f l u e n c e emulsion c a p a c i t y . Thus, a l t e r i n g the e l e c t r o v a l e n t properties by s h i f t s i n pH as well as changing the i o n i c environment of peanut p r o t e i n modifies i t s s o l u b i l i t y and emulsion capacity (21_, 22, 26, 27). Whereas the s a l t - s o l u b l e proteins seem to be the most f u n c t i o n a l i n terms o f emulsion c a p a c i t y of meat proteins ( V 7 ) , those that are s o l u b l e i n water or weak s a l t s o l u t i o n s are the most f u n c t i o n a l i n t h i s respect where peanut proteins are concerned. 9
A t t e n t i o n has been d i r e c t e d toward modifying f u n c t i o n a l properties of peanut proteins by c h e m i c a l , enzymatic, and physical approaches. Chemical m o d i f i c a t i o n has included a c e t y l a t i o n and s u c c i n y l a t i o n treatments (28, 29). Marked improvement i n emulsion c a p a c i t y occurred as a r e s u l t of t h i s t r e a t ment i f the proteins were extracted i n a c i d (28). Beuchat et a l . (30) and Sekul et a l . (31) found that enzymatic h y d r o l y s i s adv e r s e l y a f f e c t e d e m u l s i f i c a t i o n properties of peanut p r o t e i n . However, another study by Beuchat (32J u t i l i z e d a higher conc e n t r a t i o n of peanut f l o u r than reported e a r l i e r (30) and showed that pepsin h y d r o l y s i s at pH 2.0 ( 2 2 ° and 5 0 ° C ) r e s u l t e d i n emulsion c a p a c i t i e s exceeding those of the untreated c o n t r o l ;
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WATER
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Q
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1
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2
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4
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pH OF SUSPENSION Journal of Food Science
Figure 7.
Effect of sodium chloride at different concentrations on the extraction of peanut protein at various pHs (25)
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emulsions formed from pepsin and bromelain hydrolyzates were low v i s c o s i t y , whereas t r y p s i n hydrolyzate emulsions were t h i c k and viscous. Fungal fermentation of peanut f l o u r reportedly decreased emulsion capacity (33), but the e f f e c t s o f using higher concentrat i o n s of peanut f l o u r or continuous phases other than s a l t s o l u t i o n are not known. The a p p l i c a t i o n of moist heat has been employed with c e r t a i n vegetable p r o t e i n s as a physical approach to improving n u t r i t i o n a l and f l a v o r q u a l i t i e s ; consequently, f u n c t i o n a l properties a l s o undergo m o d i f i c a t i o n during h e a t i n g . Emulsion capacity of whole f u l l - f a t peanuts was not adversely a f f e c t e d by heating the seeds i n water (34), although s i g n i f i c a n t changes i n p r o t e i n s o l u b i l i t y and s t r u c t u r a l components occurred at high temperatures (35). Steam heating peanuts i n the form of a f i n e l y ground f l o u r , however, reduced l e v e l s of s o l u b l e nitrogen s l i g h t l y but sharply reduced emulsion c a p a c i t y (36). Thus, processing c o n d i t i o n s and methods can have profound e f f e c t s on prot e i n character and f u n c t i o n a l i t y . Pea and Bean Seeds. McWatters and Cherry (27) i n v e s t i g a t e d the i n f l u e n c e o f pH adjustment on emulsion capacity and v i s c o s i t y of cowpea f l o u r (24.2% p r o t e i n , dry wt b a s i s ) using the procedure of Carpenter and S a f f l e (23). Data i n Table II show that a d j u s t ing the pH from the n a t u r a l l e v e l of 6.4 to 4.0 reduced emulsion c a p a c i t y by about 20%; a d j u s t i n g the pH from the natural l e v e l to 8.2 r e s u l t e d i n a 15% increase i n emulsion c a p a c i t y . A two-step pH adjustment from 6.4+4.0+8.2 produced l i t t l e increase (4%) i n emulsion capacity over the unadjusted sample and was not as e f f e c t i v e as the one-step pH adjustment to 8.2 i n improving emulsion c a p a c i t y . This f l o u r at the natural pH l e v e l produced emulsions that were s i m i l a r i n consistency to a commercial mayonnaise. Flour suspensions adjusted to pH 8.2 were t h i c k e r than the mayonnaise sample while those adjusted to pH 4.0 were thinner. A l l were more viscous than two commercial salad dressings. This e m u l s i f i c a t i o n behavior apparently cannot be a t t r i b u t e d s o l e l y to n i t r o g e n - s o l u b i l i t y patterns (Figure 8 ) , which have been c h a r a c t e r i z e d by Okaka and Potter (37). Though n i t r o g e n s o l u b i l i t y p r o f i l e s of cowpea proteins are i n f l u e n c e d by pH and s a l t concentration changes, these e f f e c t s were not c l e a r l y shown i n emulsion capacity behavior (27). Emulsion-forming and thickening p r o p e r t i e s of cowpeas may a l s o be i n f l u e n c e d by t h e i r high carbohydrate content (about 60%, dry wt b a s i s ) ; the s t a r c h component i s h y d r o p h i l i c and may i n t e r a c t with p r o t e i n and other components i n the system. Sefa-Dedeh and Stanley (38) found that changing the i o n i c strength of the e x t r a c t i n g b u f f e r produced more profound changes i n the nitrogen s o l u b i l i t y of cowpea proteins than heating at temperatures ranging from 25 to 100°C or a t 100°C f o r 90 min. Okaka and Potter (37) found no d i f f e r e n c e i n emulsion c a p a c i t y of cowpea powders prepared from peas that had been soaked i n water at pH 2, 4 , or 6, then steam blanched, and drum d r i e d to reduce beany f l a v o r .
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Table I.
FUNCTIONALITY IN FOODS
Emulsion capacity (EC) and v i s c o s i t y of peanut f l o u r as influenced by f l o u r c o n c e n t r a t i o n , p a r t i c l e s i z e , and rate of mixing.
EC (ml o i l / g f l o u r )
Viscosity (poise)
1 3 5 7 9
50.0 25.0 16.0 12.1 10.5
0.23 1.63 5.83 10.72 19.13
60 100 200
16.0 18.0 21.0
5.83 9.33 10.86
13,000 15,000 18,700
18.0 17.0 16.0
4.89 5.13 5.83
Treatment
F l o u r concentration (g/50 ml)
Particle size (mesh)
Mixing r a t e (rpm)
from Ramanatham e t a l . ( 2 Ï T
Table II.
E f f e c t of pH adjustment on emulsion capacity (EC) and v i s c o s i t y of cowpea f l o u r .
EC ml o i l / g f l o u r
Viscosity cps
6.4 + 4.0
28.9
26,080
6.4
36.5
47,680
6.4 -»• 8.2
42.2
66,240
6.4 •* 4.0 •+ 8.2
38.0
60,160
pH Adjustment
3
For comparison, a commercial mayonnaise had a v i s c o s i t y of 44,640 c p s ; two commercial salad d r e s s i n g s , 6,400 and 16,800 cps. from McWatters and Cherry
(27)
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Sefa-Dedeh and Stanley (39) a t t r i b u t e d decreases i n s o l u b i l i t y c h a r a c t e r i s t i c s of cowpea proteins a t low i o n i c strength to the formation o f i o n i c bonds 1) w i t h i n the p r o t e i n molecule and 2) between adjacent p r o t e i n molecules leading to the formation of aggregates. Other changes i n s o l u b i l i t y at higher i o n i c strength appeared to r e s u l t from " s a l t i n g - i n " and "salting-out" e f f e c t s . Data i n Table III show v a r i a b l e response i n emulsion c a p a c i t y of f i e l d pea and cowpea products with changes i n i o n i c environment. Emulsion c a p a c i t y was higher when the pea products were dispersed i n water than i n any of the s a l t s o l u t i o n s . An inverse r e l a t i o n ship between p r o t e i n concentration and emulsion capacity i s apparent a t each s a l t c o n c e n t r a t i o n . Data i n Table IV show that each pea product formed higher v i s c o s i t y emulsions i n water than i n s a l t s o l u t i o n . The decrease i n emulsion v i s c o s i t y which occurred when s a l t was present i n the continuous phase was greater i n the f i e l d pea products than i n cowpea f l o u r and may have been caused by v a r i a t i o n s i n processing c o n d i t i o n s as well as inherent d i f ferences i n the seeds. Sosulski and Youngs (40) used an a i r - c l a s s i f i c a t i o n process to f r a c t i o n a t e e i g h t legume f l o u r s i n t o f i n e and coarse f r a c t i o n s . The f i n e material was a s s o c i a t e d with the p r o t e i n f r a c t i o n , whereas the coarse material was p r i m a r i l y s t a r c h . A modification of the model system method developed by Inklaar and Fortuin (41) was used to measure o i l e m u l s i f i c a t i o n . This procedure involves s t i r r i n g a p r o t e i n - c o n t a i n i n g material i n t o water contained i n a beaker, adding s a l t , adding a f i x e d amount of o i l from a buret while s t i r r i n g , c e n t r i f u g i n g u n t i l the volume of separated o i l does not change any f u r t h e r , and c a l c u l a t i n g the separated o i l as a percentage of the t o t a l amount of o i l added. Sosulski and Youngs (40) reported o i l - e m u l s i f i c a t i o n c a p a c i t y as the percentage of o i l e m u l s i f i e d . Data i n Table V show that chickpea and Great Northern bean had high emulsion c a p a c i t y . The p r o t e i n f r a c t i o n s had greater emulsifying c a p a c i t y than starch f r a c t i o n s . S a t t e r l e e e t a l . (42) i n v e s t i g a t e d the emulsion c a p a c i t y of f r a c t i o n s of the Great Northern bean by means of a microe m u l s i f i e r apparatus developed by Tsai et a l . ( 4 3 j . It used small q u a n t i t i e s of m a t e r i a l s and provided f o r e x c l u s i o n of a i r and i n j e c t i o n of o i l a t a constant rate u n t i l the emulsion breakpoint was reached. M o d i f i c a t i o n s i n i n i t i a l o i l - w a t e r - p r o t e i n mixing before the beginning of the run and i n blender speed were made (44). Emulsion s t a b i l i t y was determined by preparing sausages from the bean p r o t e i n products and measuring cooking l o s s e s . These authors found that the albumins had greater emulsion c a p a c i t y than the g l o b u l i n s (Table VI). The low emulsion c a p a c i t y of the g l o b u l i n s was r e f l e c t e d i n the low emulsion c a p a c i t y of the bean p r o t e i n concentrate. The g l o b u l i n s showed good s t a b i l i t y i n the heated sausages used i n the emulsion stability test. The high emulsion c a p a c i t y of the albumins was not apparent i n emulsion s t a b i l i t y , i n d i c a t i n g the importance of evaluating both aspects of e m u l s i f i c a t i o n p r o p e r t i e s .
.224 .327 .423
55.6 32.4 21.0
F i e l d pea f l o u r
Cowpea f l o u r
Pea Product
F i e l d pea p r o t e i n concentrate
V To
.208 .206 .247 .363
.178 .262 .414
.391
.279
1.0M NaCl
0.5M NaCl
0.1M NaCl
EC (ml oil/mg p r o t e i n )
Emulsion capacity (EC) of pea p r o t e i n products dispersed i n water and s a l t s o l u t i o n s .
Water
III.
Protei η
Table
11.
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Table IV.
Emulsification: Vegetable Proteins
AND CHERRY
227
V i s c o s i t y of emulsions prepared from pea p r o t e i n products dispersed i n water and s a l t s o l u t i o n s .
Viscosity Pea Product
(cps)
Water
0.1M NaCl
0.5M NaCl
1.0M NaCl
F i e l d pea p r o t e i n concentrate
30,240
10,080
12,640
20,480
F i e l d pea f l o u r
16,480
2,560
1,440
2,560
Cowpea f l o u r
22,880
17,600
16,960
18,880
Table V.
O i l e m u l s i f i c a t i o n capacity of legume f l o u r , p r o t e i n (PF) and s t a r c h (SF) f r a c t i o n s i n percent, dry b a s i s .
% Oil
Emulsified
Flour
PF
SF
Chickpea
94
92
79
Pea bean
64
66
20
Northern bean
92
98
57
Fababean
47
77
24
F i e l d pea
48
48
20
Lima bean
53
70
27
Mung bean
64
64
22
Lentil
56
58
50
from Sosulski and Youngs
(40)
0.8 1.3 1.2
0.03 0.04 0.13
12.4 26.8 15.4
Globulins
Albumins
BPC
from S a t t e r l e e e t a l .
(42)
BPC (bean p r o t e i n concentrate contained 65% g l o b u l i n s and 35% albumins),
a
Water + Suspension S o l i d s
Fat
EC
ES (ml released/10 g e m u l s i f i e d meat)
Emulsion c a p a c i t y (EC) and s t a b i l i t y (ES) of various p r o t e i n f r a c t i o n s from the Great Northern bean.
ml oi1/100 mg p r o t e i n
Table VI.
11.
MCWATTERS
A N D CHERRY
Emulsification: Vegetable Proteins
229
Sunflower Seed. Emulsion capacity of defatted sunflower meal was i n v e s t i g a t e d by Huffman et a l . (45j a t three pH l e v e l s ( 5 . 2 , 7 . 0 , 1 0 . 8 ) , blender speeds (4500, 6500, 9000 rpm), and o i l addition rates (30, 45, 60 ml/min). With low mixing speeds and r a p i d rates of o i l a d d i t i o n , optimum emulsion capacity occurred at pH 7.0. These authors r e l a t e d the observed e m u l s i f i c a t i o n properties to p r o t e i n s o l u b i l i t y , surface area and s i z e of o i l d r o p l e t s , and rate of p r o t e i n f i l m formation. L i n et a l . (46) measured the emulsion c a p a c i t y of defatted sunflower seed products. Data i n Table VII show that sunflower f l o u r was s u p e r i o r i n emulsifying c a p a c i t y to a l l other products tested. The emulsions were i n the form of f i n e foams and were s t a b l e during subsequent heat treatments. The d i f f u s i o n e x t r a c t i o n processes employed to remove phenolic compounds d r a m a t i c a l l y reduced emulsion c a p a c i t y , although i s o l a t i n g the p r o t e i n improved emulsion capacity to some extent. Canella et a l . (47) evaluated the e f f e c t s of s u c c i n y l a t i o n and a c e t y l a t i o n on the emulsion c a p a c i t y of sunflower p r o t e i n conc e n t r a t e (Figure 9 ) . These authors showed that emulsion c a p a c i t y f o r untreated and treated concentrates was low i n the a c i d i c pH r e g i o n , increased u n t i l pH 7 was reached, then decreased a t higher pH l e v e l s . Chemical m o d i f i c a t i o n by s u c c i n y l a t i o n and a c e t y l a t i o n improved emulsion capacity of sunflower p r o t e i n concentrate, with the 10% s u c c i n y l a t i o n treatment producing the g r e a t e s t i n c r e a s e . Emulsion s t a b i l i t y , determined by a heating and c e n t r i f u g a t i o n t e s t , was a l s o improved by these chemical treatments. Improvements i n emulsifying p r o p e r t i e s were a t t r i b u t e d to changes i n p r o t e i n s o l u b i l i t y and e l e c t r i c a l charges. Rapeseed. Methods employed i n processing of rapeseed p r o t e i n products i n f l u e n c e emulsion c a p a c i t i e s (48, 49). Kodagoda et a l . (48) showed that rapeseed p r o t e i n i s o l a t e s from water extracts e m u l s i f i e d m o r e . o i l than i s o l a t e s from a c i d or a l k a l i extracts (Table V I I I ) . Rapeseed i s o l a t e s e m u l s i f i e d more o i l than t h e i r concentrate counterparts. Rapeseed i s o l a t e s and concentrates from a c i d e x t r a c t s were f a r s u p e r i o r i n emulsion s t a b i l i t y to rapeseed p r o t e i n products from water or a l k a l i e x t r a c t s . Sosulski et a l . (49) found that high temperature e x t r a c t i o n procedures used to d e t o x i f y rapeseed p r o t e i n products by removal of g l u c o s i n o l a t e s adversely i n f l u e n c e d emulsion c a p a c i t y (Table IX). An i s o l a t e prepared from a low g l u c o s i n o l a t e c u l t i v a r , however, had higher emulsion c a p a c i t y values than the other d e t o x i f i e d rapeseed products and a soy f l o u r and concentrate a l s o tested. The authors a t t r i b u t e d the low emulsion c a p a c i t y values of the soy products and the Tower rapeseed concentrate FRI (Food Research I n s t i t u t e ) to t h e i r low nitrogen s o l u b i l i t i e s . G i l l berg (50) demonstrated that the s o l u b i l i t y c h a r a c t e r i s t i c s of rapeseed p r o t e i n i s o l a t e s were h i g h l y s e n s i t i v e to and a l t e r e d by small
230
PROTEIN
FUNCTIONALITY
IN
FOODS
100
X 0) " ° 80 £
70
-Q
60
δ "
S Ο Ζ
Ο- deionized H;0 • - 0 05M NaCl • - 0 SOM NaCl a - 1 OOM NaCl
40
30 20 10
5
6
7
10
11 12
PH Journal of Food Science
Figure 8.
Effects of pH and salt concentration on nitrogen solubility index of raw dehulled cowpeas (31)
Ο—0
protein concentrate
ο- -u
2*i.succinylated
ο- -α 10%
α 50% • a 100%
Or
acetylated
υ
ο. υ c 3
Ε
PH
Lebensmittel-Wissenschaft & Technologie
Figure 9. Effect of pH on emulsifying capacity of native and chemically modified (succinylation, acetylation) sunflower seed protein concentrates (41)
11.
M^WATTERS
A N D CHERRY
Table VII.
Emulsification: Vegetable Proteins
Emulsion capacity of defatted seed products.
Defatted Sunflower Seed P r o d u c t
Flour
3
% Protein
sunflower
% Oil
Emulsified
55.5
95.1
Concentrate
(DE-60)
67.9
14.0
Concentrate
(DE-80)
68.3
11.3
Concentrate
(DE-90)
68.0
10.1
90.7
25.6
Isolate
(DE-60)
231
Concentrates were d i f f u s i o n extracted i n tap water (pH 5.5) at 60°C f o r 4 hr, 8 0 ° C f o r 1.5 hr, or 90°C f o r 1.0 hr to remove phenolic compounds. The i s o l a t e was prepared by a d d i t i o n a l washing and pH adjustment of the DE-60 concentrate. from L i n e t a l .
(46)
E x t r a c t i o n Stage-Medium
first-water second-acid third-alkali single-alkali first-water second-acid third-alkali single-alkali
Isolate Isolate Isolate Isolate
Concentrate Concentrate Concentrate Concentrate
mi η
10.0 200.0 8.0 8.5 7.5 200.0 7.0 6.5
45.3 36.3 32.4 37.3 33.2 27.5 25.1 29.5
ES
ml oil/100 mg p r o t e i n
EC
Emulsion capacity (EC) and s t a b i l i t y (ES) of rapeseed p r o t e i n i s o l a t e s and concentrates.
Product
Table V I I I .
11.
McWATTERS
Table IX.
A N D CHERRY
Emulsification: Vegetable Proteins
Emulsion capacity of rapeseed p r o t e i n
Total Glucosinolates mg/g
Rapeseed Product
products.
% Oil Emulsified
Midas
flour
17.0
20
Torch
flour
7.8
29
Tower
flour
1.2
46
Tower meal
1.1
34
Tower concentrate 8 0 °
0.1
17
Tower concentrate
0.1
10
0.0
38
FRI^
Tower i s o l a t e
233
For comparison, a soy f l o u r and soy concentrate e m u l s i f i e d 12 and 10% o i l , r e s p e c t i v e l y . VRI
= Food Research
from Sosulski e t a l .
Institute (49)
234
PROTEIN
FUNCTIONALITY
IN
FOODS
amounts of s a l t s and v a r i a t i o n s i n pH. Because these f a c t o r s a f f e c t i n t e r a c t i o n and i o n i c bonding of molecules i n food systems, f u n c t i o n a l p r o p e r t i e s other than s o l u b i l i t y are a l s o l i k e l y to be affected. Cottonseed. Emulsion c a p a c i t y and v i s c o s i t y c h a r a c t e r i s t i c s of g l a n d l e s s cottonseed f l o u r dispersed i n water are markedly i n fluenced by v a r i a t i o n s i n pH and f l o u r concentration (51_, 52; Figure 10). Emulsion c a p a c i t y was lowest near the i s o e l e c t r i c pH (4.5) and highest a t pH 1 1 . 5 . Suspensions i n the a l k a l i n e pH range e m u l s i f i e d more o i l than those at a c i d i c pH l e v e l s . These authors found that the s o l u b l e f r a c t i o n was c l o s e l y r e l a t e d to the v i s c o s i t y of emulsions formed by g l a n d l e s s cottonseed f l o u r . V i s c o s i t y values were lowest near the i s o e l e c t r i c pH and highest at pH 1 1 . 5 . Emulsions formed by suspensions i n the a l k a l i n e pH range were more viscous than those formed by a c i d i c suspensions. This behavior may be a s s o c i a t e d with the storage g l o b u l i n s o f cottonseed p r o t e i n , most of which are s o l u b l e a t a l k a l i n e pH l e v e l s (5]_). Safflower. Betschart e t a l . (53) studied the i n f l u e n c e of e x t r a c t i o n , i s o l a t i o n , n e u t r a l i z a t i o n , and drying v a r i a b l e s on e m u l s i f i c a t i o n properties ( a c t i v i t y and s t a b i l i t y ) of safflower protein i s o l a t e s . Emulsion a c t i v i t y was determined by measuring the volume of e m u l s i f i e d l a y e r remaining a f t e r c e n t r i f u g a t i o n of a p r o t e i n i s o l a t e - w a t e r - o i l mixture. Emulsion s t a b i l i t y was determined by measuring the amount of e m u l s i f i e d material remaining a f t e r heating a t 7 0 - 7 4 ° C f o r 30 min. Data i n Table X show that n e u t r a l i z a t i o n of safflower p r o t e i n i s o l a t e s to pH 7 was r e quired f o r emulsion formation and s t a b i l i z a t i o n and was more i n f l u e n t i a l than p r e c i p i t a t i o n pH or drying method. The method used to dry safflower p r o t e i n i s o l a t e s had l i t t l e i n f l u e n c e on emulsification properties. The n e u t r a l i z e d safflower p r o t e i n products performed e q u a l l y as well as a soy p r o t e i n concentrate and i s o l a t e a l s o t e s t e d . Thus, pH i s a major c o n t r i b u t i n g f a c t o r to e m u l s i f i c a t i o n p r o p e r t i e s of safflower p r o t e i n i s o l a t e s . Vegetable Proteins as Emulsi f i e r s
i n Food Systems
The general c l a s s of food-grade e m u l s i f i e r s i s broad, but these agents may be grouped according to the f u n c t i o n they perform (2^ 5) i n c l u d i n g : 1)
2)
reduction of surface tension a t o i l - w a t e r i n t e r f a c e s , which promotes emulsion formation, phase e q u i l i b r i a between o i l - w a t e r - e m u l s i f i e r a t the i n t e r f a c e , and emulsion s t a b i l i t y ( e . g . , the a c t i o n of egg y o l k and mustard to emulsify o i l and water i n mayonnaise); i n t e r a c t i o n s with s t a r c h and p r o t e i n components i n foods that modify texture and r h e o l o g i c a l properties
11.
MCWATTERS
Table X.
Emulsification: Vegetable Proteins
A N D CHERRY
E m u l s i f i c a t i o n p r o p e r t i e s of safflower p r o t e i n i s o l a t e s as i n f l u e n c e d by pH and drying method.
Emulsification
Dryi ng pH Adjustment
Method
9
Activity
{%)
Properties Stability
9 + 5
FD
0
0
9 + 5
SD
0
0
9 + 5 + 7
FD
63 ± 1.5
62 ± 1.4
9 + 5 + 7
SD
59 ± 0.8
59 ± 1.0
9 + 6
FD
0
0
9 + 6
SD
0
0
9 + 6 + 7
FD
58 ± 2.8
60 ± 2.3
9 + 6 + 7
SD
52+1.7
51 ± 0.8
a
235
FD = Freeze d r i e d ; SD = Spray d r i e d , from Betschart e t a l .
(53)
{%)
236
PROTEIN
ι
IS
ι
ι ι ι I ι I I S3 55 75 95 pH OF 10% SUSPENSION
FUNCTIONALITY
IN
FOODS
I—L 115
American Chemical Society
Figure 10. Solubility and emulsifying properties of cottonseed proteins at various pH values: (O — O) emulsion capacity; (O O) emulsion viscosity; (% — Φ) soluble protein fraction; (Φ Φ) insoluble protein fraction (5\).
11.
MCWATTERS
3)
A N D CHERRY
Emulsification: Vegetable Proteins
237
( e . g . , the a c t i o n of mono- and d i - g l y c e r i d e s and stearoyl l a c t y l a t e s as dough c o n d i t i o n e r s and a n t i s t a l i n g agents i n bakery p r o d u c t s ) ; control of c r y s t a l l i z a t i o n i n f a t and sugar systems ( e . g . , the a c t i o n of mono- and d i - g l y c e r i d e s i n peanut butter to prevent o i l s e p a r a t i o n , or s o r b i t a n monos t e a r a t e to prevent "bloom" or migration of f a t to the surface of c h o c o l a t e ) .
In the l i t e r a t u r e , reports of using vegetable p r o t e i n s , other than soy, s p e c i f i c a l l y f o r these f u n c t i o n s are sparse. Vegetable proteins have been used i n breads and bakery products, not as emulsifying agents or dough c o n d i t i o n e r s , but f o r p r o t e i n e n r i c h ment (42, 48, 53-62). Instead of c o n d i t i o n i n g the dough and improving bread q u a l i t y , however, the opposite e f f e c t i s produced when vegetable proteins are used as p a r t i a l replacements f o r wheat flour. Because the gluten component i s d i l u t e d i n composite f l o u r mixtures of t h i s type, m o d i f i c a t i o n s i n f o r m u l a t i o n , mixing, or baking procedure are g e n e r a l l y required to o f f s e t adverse e f f e c t s on l o a f volume, crumb s t r u c t u r e , or sensory a t t r i b u t e s . A few reports i n the l i t e r a t u r e d e s c r i b e the e f f e c t s of using peanut p r o t e i n i n comminuted meat systems such as meat loaves (63, 64), f r a n k f u r t e r s (65), and ground beef p a t t i e s (66.). In some i n s t a n c e s , peanut p r o t e i n e i t h e r produced b e n e f i c i a l e f f e c t s ( e . g . , increased tenderness and cohesiveness, 66) or no adverse e f f e c t s from a sensory, p h y s i c a l , or m i c r o b i a l standpoint (65). In other i n s t a n c e s , however, no b e n e f i t s such as reduced shrinkage or increased f a t and water binding were derived by use of peanut p r o t e i n (63, 64). Torgersen and Toledo (64) found that p r o t e i n s o l u b i l i t y had a negative i n f l u e n c e on the d e s i r a b l e p r o p e r t i e s of a comminuted meat system, with the more s o l u b l e p r o t e i n s allowing more f a t r e l e a s e on cooking. These authors suggested that a p r o f i l e of s o l u b i l i t y change with temperature seemed to be a b e t t e r index of f u n c t i o n a l i t y than s o l u b i l i t y at any s i n g l e temp e r a t u r e ; they concluded that the most d e s i r a b l e p r o t e i n s f o r use as a d d i t i v e s i n comminuted meats were those that possessed the combined q u a l i t i e s of good gel s t r e n g t h , high water-absorption c a p a c i t i e s a t 9 0 ° C , and i n c r e a s i n g s o l u b i l i t i e s with i n c r e a s i n g temperatures. The mechanisms involved i n meat systems, t h e r e f o r e , are much more complex than e m u l s i f i c a t i o n alone and seem to be a combination of e m u l s i f i c a t i o n , b i n d i n g , and g e l a t i o n . Vaisey et a l . (67j evaluated the performance of fababean and f i e l d pea concentrates as ground beef extenders. Sensory a t t r i b utes of b r o i l e d meat p a t t i e s containing the p r o t e i n concentrates were more acceptable to t a s t e p a n e l i s t s when these products were added as t e x t u r i z e d f l a k e s than as f l o u r . The extended products had s i g n i f i c a n t l y l e s s d r i p l o s s than a l l - m e a t c o n t r o l s but had s o f t e r textures. T e x t u r i z i n g the concentrates by drum-drying and f l a k i n g reduced t h e i r f a t r e t e n t i o n p r o p e r t i e s but produced p a t t i e s more l i k e the a l l - b e e f c o n t r o l .
238
PROTEIN
FUNCTIONALITY
IN
FOODS
L i n et a l . (68) i n v e s t i g a t e d the e f f e c t s of supplementing wieners with sunflower f l o u r and d i f f u s i o n - e x t r a c t e d p r o t e i n con centrates. The sunflower products showed good emulsion s t a b i l i t y , l e s s shrinkage, and l e s s cooking l o s s than a l l - m e a t c o n t r o l s . Though the i n c l u s i o n of the sunflower seed products i n wieners r e s u l t e d i n s l i g h t l y lower o r g a n o l e p t i c s c o r e s , most of the supplemented products were considered to be a c c e p t a b l e . These authors emphasized that the true value of vegetable proteins can only be determined by i n c o r p o r a t i n g them i n t o actual food products. Conclusions Most of the information p e r t a i n i n g to e m u l s i f i c a t i o n pro p e r t i e s of vegetable p r o t e i n s , other than soy, has been derived by means of model-system s t u d i e s . For the most p a r t , the a p p l i c a t i o n of t h i s information to performance i n actual food systems has y e t to be determined. D i f f i c u l t i e s i n o b t a i n i n g meaningful data regarding e m u l s i f i c a t i o n mechanisms of vegetable proteins stem from the lack of standardized model system t e s t s that can simulate the processing c o n d i t i o n s and i n g r e d i e n t s involved i n complex food systems. The information that has been obtained should not be d i s c o u n t e d , however, but rather should be considered as a p o i n t of departure and as part of a t e c h n o l o g i c a l base con cerning physical and f u n c t i o n a l c h a r a c t e r i s t i c s o f vegetable proteins. The a p p l i c a t i o n of t h i s data base to food products presents a s u b s t a n t i a l challenge to the food s c i e n c e and technology d i s c i p l i n e .
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RECEIVED
September 5, 1980.
