Food Technology. Vol. 38, Num. 5. May 1984. pp. 67-96.

Symposium: Gelation in Food Protein Systems

Quantitative Analysis of Gelation in Egg Protein Systems

P.W. Gossett, S.S.H. Rizvi, and R.C. Baker

Authors Rizvi and Baker are, respectively, Associate Professor, Dept. of Food Science, and Professor and Chairmen, Dept. of Poultry and Avian Sciences, Cornell University Ithaca, NY 14853. Author Gossett formerly with Cornell, is a Scientist with The Pillsbury Co. 311 Second St. S E., Minneapolis, MN 55414
Based on a paper presented during the symposium, "Gelation in Food Protein Systems," at the 43rd Annual Meeting of the Institute of Food Technologists, New Orleans. La., June 19-22, 1984.

An important function of proteins in food systems is gelation. This phenomenon involves the formation of a three-dimensional matrix mainly through inter-protein hydrogen bonding and allows the immobilization of water within the gel structure. The coagulation or gelation of proteins—in particular, the irreversible, heat-induced coagulation or gelation of egg proteins—often controls the success of certain conked food products. It is of interest to the food scientist to be able to quantitatively monitor the gelation process to better predict end-product characteristics, as well as understand the mechanism of network formation. A better understanding of the gelation process will permit manipulation of variables to obtain a gel of desired textural characteristics and functional properties.

Although the coagulation, i.e., denaturation of proteins can be brought about by heat, high pressure, salts, acids, alkalies, alcohols, or denaturing agents such as urea (Mirsky and Pauling 1936), this article will concentrate on the quantitative analysis of heat-induced coagulation or gelation of egg-albumen proteins (Table 1). Factors affecting the kinetic parameters of gelation and the rheological properties of the gel will be discussed, as well as methods used to measure gelation.

Table 1-PHYSICAL AND CHEMICAL PROPERTIES of egg albumen proteinsa
Protein % in albumen pI Molecular weight Intrinsic viscosity theta (dL/g) Characteristics
Ovalbumin 54 45-4.6 45,000 0.043b Phosphoglycoprotein; denatures easily; has four sulfhydryls
Ovotransferin 12-13 6.1-6.6 76,000-80,000 0084c Glycoprotein, complexes iron and other metals
Ovomucoid 11 3.9-4.3 28.000 0.055d Glycoprotein, trypsin inhibitor
Ovomucin 3.5 4.5-5.0 110,000 2.10e Glycoprotein, fibrous, viscous
Lysozyme 3.4-3.5 10.7 14,300-14,600 0.027c Spherical protein; four disulfide bonds; lytic action
Ovoinhibitor 1.5 5.1-5.2 44,000-49,000 - Inhibits trypsin and chymotrypsin
Ovoglycoprotein 1.0 3.9 24,000.24,400 - Glycoprotein
Ovoflavoprotein 0.8 4.0-4.1 32.000-35,000 - Binds riboflavin
Ovomacroglobulin 0.5 4.5-4.7 760,000-900,000 0.0651d Glycoprotein
Avidin 0.5 9.5-10 55,000-68,300 - Binds biotin

a Adapted from Osuga and Feeney (1977) and Powrie (1977); data refer to albumen from chicken eggs
b From Yang (1961)
c From Kuntz and Kauzmann (1974)
d From Vadehra and Nath (1973); ovomucoid pH 3.8-4.6; ovomacroglobulin: pH 6-9
e From Fasman (1976)


A discussion of gelation necessitates defining some commonly used terms associated with the phenomenon. At first glance, some of these terms, may appear to overlap in meaning. However, each has its particular role in this discussion.

Theory of Gelation

Attempts to describe the mechanism and theory of gel network formation are numerous. The classic explanation of heat induced aggregation of protein molecules is the following two-step process (Ferry, 1948):

Native protein -> denatured protein (long chains) -> aggregated protein (associated network)

The first step is considered a denaturation process and the second step an aggregation process. Comparison of the rate of the denaturation step vs that of the aggregation step helps determine gel characteristics. For example, Ferry (1948) suggested that for a given rate of denaturation the rate of aggregation will be slow if the attractive forces between the denatured protein chain are small. The resulting gel will be a finer network of protein chains, will be less opaque, and will exhibit less syneresis than one with a faster rate of aggregation. A coarser network of protein chains yields an opaque gel with large interstices capable of holding solvent which is easily expressed from the matrix. Hermansson (1979) suggested that conditions favoring denaturation, such as high or low pH, have the opposite effect on aggregation of globular proteins, possibly due to the fact that at high net charge, protein-solvent interactions such as denaturation are favored, rather than protein-protein interactions such as aggregation. A gel network with a certain degree of order can be attained if the aggregation step occurs more slowly than the denaturation step. thus giving the denatured protein molecule, time to orient themselves before aggregation; this is lower in opacity and higher in elasticity then one where aggregation is not suppressed. Schmidt (1981) suggested that if aggregation occurs simultaneously with denaturation, an opaque. less elastic gel results.

Since the kinetics of the denaturation step relative to the aggregation step appear to he important in determining the type of gel produced, it is useful to review some kinetic terms that aid in describing the gelation process:

l. Reaction Rate Constant. The first is the reaction rate constant k (min -1), which is obtained from the first-order relationship (Land, 1975):


    -   --  = kc


where c=concentration and -dc/dt=the rate at which concentration decreases. Integrating between limits et at time l=U and c et time t gives:


A plot of ln c vs t gives a line of slope -k.

The rate constant may be temperature dependent, end this dependence of reaction rate constant on temperature can best be described by the Arrhenius equation:

    k=S exp (-Ea/RT)

where s=frequency factor (min -1), e.=activation energy (cal/mole). r=gas constant (1.987 cal/mole.°K), and t=absolute temperature (°K). A plot of In k vs 1/T gives e straight line of slope -Ea/R.

2. Z Value. Another useful kinetic quantity in describing the temperature dependency of the reaction rate is the Z value, which a defined as the necessary rise in heating temperature (°C) needed for a 10-fold increase in the reaction rate (Dagerskod, 1977). If some parameter of the gelation process is measured, say heating time to reach a certain gel strength, then the Zc value is the temperature rise needed to increase the heating time or reaction rate 10-fold, This value is useful in calculating the cook value, which is the time required at temperature T for a sample with a certain Zc value to receive a heat treatment equivalent to 1 min at 100°C (Dagerskod, 1977). To quantitate gelation, it in useful to coagulate samples to equal cook values to ensure equivalent heat treatment

Factors Affecting Gelation

Many researchers have tried to identify the factors which effect aggregation of proteins, since altering the rate of aggregation relative to the rate of denaturation appears to affect gel characteristics. The following are some of the factors which have been studied:

Table 2-APPARENT VISCOSITY AND YIELD FORCE (20°C) for albumen at pH 9.0 heated at various temperatures and for albumen at different pH levels heated at 80°C
Set pH Heating temperature (°C) Apparent viscosity (Poise) Yield force (dynes x 106
I 9.0 75 80 ± 12b 3.6 ± 0.9b
80 156 ± 59bc 7.5 ± 0.1c
85 230 ± 13cd 7.7 ± 1.3c
90 235 ± 15d 11.6 ± 1.4d
95 333 ± 29e 10.2 ± 1.6d
II 7.0 80 117 x 39b 5.1 ± 1.1b
8.0 80 128 ± 39bc 6.3 ± 1.5b
9.0 80 156 ± 59bc 7.5 ± 0.1b
10.0 80 278 ± 29cd 12.2 ± 2.4bc
10.5 80 330 ± 122de 19.7 ± 9 3c
11.0 80 476 ± 153e 18.5 ± 8.4c


Measurement of Heat-Induced Gelation

Many methods for measuring gelation involve testing individual samples after discrete time intervals of heating until coagulation or gelation is completed. Some of the methods which have been used are discussed below:

New Method Developed

The above methods for measuring heat-induced coagulation do have certain limitations, one of which is the need to make single-point measurements on many samples at different time intervals throughout the process to obtain a continuous view of the phenomenon. These samples must be [cooled?] instantly; otherwise the coagulation process con[tinues?] another limitation of some gel-strength or gravimetric measurements is that the sample is destroyed during the measurement. Yet another limitation is that for some turbidity measurements, once the sample becomes opaque, no further change is registered by the spectrophotometer despite continuing changes in gel characteristics. Furthermore, a gel can be clear end still exhibit increasing gel strength with time.

With these limitations in mind, we devised a new method to quantitate, the gelation process (Gossett et al., (1983b). This nondestructive technique requires only one sample to observe the total process. It involves continuous monitoring of the force exerted by the gel as coagulation takes place and can be used to obtain kinetic data on the heat-induced gelation of egg albumen.

The technique uses a recording electrobalance from which a wire probe is suspended into a jacketed cylinder containing the albumen sample. The cylinder is connected to a water bath maintained at the appropriate temperature, and, as coagulation occurs with time, the force exerted by the gel on the probe as the matrix forms is recorded on e linear recorder.

Typical electrobalance force-vs-time curves (Fig. 1) for albumen heated at temperatures from 60 to 95°C show that the force increases until a maximum force is observed. For some treatments, however, there are decreases in force after the maximum force is attained; this is thought to be due to expansion of the gel once the gel coagulates—the expansion may have raised the probe and caused a decrease in observed force.

Table 3-FIRST-ORDER RATE CONSTANTS for heat coagulation or albumen under various conditionsa
Set Albumen pH Water bath temperature (°C) k1 (min -1 ) k2 (min -1 ) Timeb (min)
I 9.0 60 0.329 t±0.100d 0.037 -± 0.008d 4.83 ± 1.36
65 0.377 ± 0.150d 0.036 ± 0.006d 5.44 ± 1.88
70 0.421 ± 0.143de 0.046 ± 0.012d 4.42 ± 1.11
75 0477 ±.0179'def 0.051 ± 0.011d 4.54 ± 1.51
80 0.617 ± 0.083ef 0.064 ± 0.026 3.01 ± 0.49
85 0.662 ± 0.04f 0.067 ± 0.021 3.09 v 0.14
90 0.690 ±0.277f 0.157 ± 0.010e 2.04 ± 0.33
95 1.223 v 0.12g 0.231 ± 0.089f 1.59 ± 0.45
Ave. r2 for regression 0.92 0 94
II 7.0 80 0.347 ± 0.145d 0.102 ± 0.021 d 4.42 ±1.13
8.0 80 0.434 ± 0.056d 0.071 ± 0.020e 3.69 ± 0.69
9.0 80 0.617 ± 0.083de 0.064 ± 0.026 e 3.01 ±0.49
10.0 80 0.923 ± 0 419ef 0.049 ± 0.020 e 1.80 ± 0.23
10.5 80 0.983 ± 0.328ef 0.044 ± 0 0.015 e 1.76 ± 0.41
11 0 80 0.820 ± 0.175'f 0.036 ± 0.018 e 2.02 ± 0.20
Ave. r2 for regression 0.95 0 93


a Quadruplicate determinations. From Gossett )1983), Gossett et al. (1983b)
b Time at which first step "ends" and second step "starts"
defg Means in the same column within a set that are followed by different superscripts are significantly different (P < 0.05)


Fig. 2-PLOT OF ln FORCE VS TIME showing rate constants k1 and k2 and the time t where the first step "stops" and the second step "starts." From Gossett at al. (1983b)

Rate constants for albumen (at pH 9) heated at 60-9.5°C and for albumen (pH 7.0-11.0) heated at 80°C are shown in Table 3. First-order kinetics and the existence of two reactions during the heat-coagulation of the albumen, as su[illegible] by the broken-line curve in Figure 2, yielded rate [constants] k1 and k2 and were obtained by best r2 fit If k1 is interpreted to be the rate constant for the denaturation process and k2 the rate constant for the aggregation process, then the data suggest that the denaturation end aggregation rates both increase with increasing heating temperatures. The time at which the first step "ends" and the second step "starts" is found to decrease with increasing heating temperature; this suggests that at higher temperatures, denaturation occurs faster and the onset of aggregation is expedited. With increase in pH, rate constants for denaturation increase but those for aggregation decrease; the results agree with those of Nakamura et al. (1978) and Hermansson (1979), who suggested that aggregation is decelerated at higher pH levels.


Fig 3-ARRHENIUS PLOTS for rate constants k1 (r=0.94) and k2 r=0.931) From Gossett et al (1983b)

Activation energies for the rate constants were calculated using the Arrhenius relationship; Figure 3 shows Ea to be 8.7 kcal/mole for the denaturation step and 14.4 kcal/mole for the aggregation step (Dwek and Navon, 1972, estimated Ea for the denaturation of egg albumen to be approximately 24 kcal/mole). In addition, a Zc value of 41.7°C was calculated by plotting the ln of the heating time required to reach a certain force value vs the heating temperatures.

Although this method offers the advantage of using just one a ample to continuously observe the gelation process, there are presently some limitations to the technique. The true source and physical meaning of the measured force are not yet understood and should be a topic of future research. Change. in the geometry of the gelling apparatus need to be explored to differentiate between the contribution of gelation and that of thermal expansion to the force. Lastly, optimization of probe design for various gel types would be desirable.

The gelation of food proteins is important to textural and rheological characteristics of a food product. Future, research should be directed toward finding quick, reliable methods that quantitate the gelation phenomenon so that food scientists can better manipulate the process to obtain desirable, high-quality foods.


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