Adsorption and Foaming Characteristics of Soy Globulins and Tween

2876

Ind. Eng. Chem. Res. 2008, 47, 2876-2885

Adsorption and Foaming Characteristics of Soy Globulins and Tween 20 Mixed Systems Victor Pizones Ruı´z-Henestrosa, Cecilio Carrera Sa´ nchez, and Juan M. Rodrı´guez Patino* Departamento de Ingenierı´a Quı´mica, Facultad de Quı´mica, UniVersidad de SeVilla, C/ Prof. Garcı´a Gonza´ lez, 1, E-41012-SeVilla (Spain)

The manufacture of food dispersions (emulsions and foams) with specific quality attributes depends on the selection of the most appropriate raw materials and processing conditions. In this contribution, we have analyzed the addition of Tween 20 (at 5 × 10-6 and 1 × 10-4 M, lower and higher than the critical micelle concentration, CMC, respectively) to improve the dynamic interfacial characteristics (adsorption and surface dilatational properties) and foam properties (foam capacity and stability) of soy globulin (7S and 11S at 0.1 wt %) acidic aqueous solutions. We have observed that (i) the adsorption (presence of a lag period, diffusion, and penetration at the air-water interface) of soy globulins depends on three factors: first the particular protein, second, the level of association/dissociation of these proteins by varying the pH (pH 7 was included as a control), and, finally, on the competitive adsorption between protein and Tween 20 in the aqueous phase. (ii) The surface dilatational properties reflect the fact that soy globulin and Tween 20 adsorbed mixed films exhibit viscoelastic behavior. The surface dilatational modulus decreases with the addition Tween 20 into the aqueous phase. (iii) The rate of adsorption and surface dilatational properties (surface dilatational modulus and phase angle) during adsorption at the air-water interface play an important role in the formation of foams generated from aqueous solutions of soy globulins. However, the stability of the foam correlates with the properties of the film at long-term adsorption. Introduction Foams are present in many foods, either in the finished product or incorporated during the production as a preliminary processing stage, which can be subjected to further processing before the product is completed. Among the functional effect of emulsifiers, foaming is of particular interest because it provides the texture for many traditional (ice cream, whipped topping, breads, cakes, meringues, beers, champagne, etc.) and new (cheese, butter, spreads, confectionary, sausages, etc.) aerated food products.1-5 The positive benefits of aerated food products are primarily to do with texture. However, foaming may also create important problems during processing.6,7 Thus, knowledge of the mechanism of foam formation and stabilization is essential if a foam of the required characteristics is to be produced. Foam formation and the stability of the resulting foam depend on the properties of the surface active components in the formulation.6-16 Foam formation is influenced by the adsorption of the foaming agent (emulsifier) at the air-water interface and its ability to reduce surface tension and develop a mechanical protection to new formed bubbles.8-11,17 Stability is an important property of food foams because consumer perception of quality is influenced by appearance. Foams are thermodynamically unstable, and their relative stability is affected by factors such as drainage of liquid previously present in the foam, disproportionation, and coalescence. Bubbles in a foam are stabilized by a bilayer of emulsifier molecules separated by the continuous aqueous phase. Thus, the characteristics of this thin film determine the formation and stability of the foam.9,17,15,17-20 An important topic is how the macroscopic characteristics of a foam can be improved by controlling the nanostructure formation at fluid interfaces (molecular or interfacial engineering). Two types

of emulsifiers are used in foods:2,21 low molecular weight emulsifiers (mainly lipids, phospholipids, surfactants, etc.) and macromolecules (proteins and some hydrocolloids). These surface-active substances are used as foaming agents because of their amphiphilic character. In this work, we have analyzed how formulation engineering can improve the interfacial and foaming characteristics of soy globulin acidic aqueous solutions by the addition of a surfactant (Tween 20). Vegetable proteins (i.e., soy globulins) are of equivalent quality to those of meat, milk, and eggs, and their production requires substantially fewer natural resources. The popularity of these protein and peptide derivatives is increasing, mainly because of their health benefits.1,22,23 They are grouped into two types according to their sedimentation coefficients, β-conglycinin (a 7S globulin) and glycinin (an 11S globulin). A notable feature of soy proteins is the strong pH dependence of the molecular conformation and the associated functional properties.24 Optimum functionality occurs at pH 95%) without further purification. Ionic strength was 0.05 M in all of the experiments. Tween 20 (polyoxy-ethylene sorbitan monolaurate (product no. 326896/1693) was acquired from Fluka. The isolation, solubility, molecular masses (determined by gel filtration chromatography, FPLC), the amino acid analysis (determined by high-performance liquid-chromatography, HPLC), and sodium dodecyl sulfate polyacrylamide gel electrophoresis of 7S and 11S soy globulins were determined as described elsewhere.25 Other structural characteristics (including scanning differential calorimetric analysis, surface hydrophobicity, and fluorescence spectroscopy) of 7S and 11S globulins have been also described elsewhere.35 The absence of active surface contaminants in the aqueous buffered solutions was checked by interfacial tension measurements before sample preparation. No aqueous solutions with a surface tension other than that accepted in the literature (7273 mN/m at 20 °C) were used. Sodium azide (Sigma) was added (0.05 wt %) as antimicrobial agent. Dynamic Surface Measurements. Measurements of timedependent surface pressure (π) and surface dilatational properties of adsorbed soy globulin and Tween 20 films at the air-water interface were performed simultaneously by an automatic drop tensiometer as described elsewhere.36,37 Protein solutions at 0.1 wt % as a function of pH (at pH 5 and 7) and in the absence and presence of Tween 20 (at 5 × 10-6 and 1 × 10-4 M) were prepared freshly to attain the desired concentration in solution and the composition of the aqueous phase, which was then stirred for 30 min. The protein-Tween 20 solution was placed in a 0.5 mL glass Hamilton syringe equipped with a stainless steel needle and then in a rectangular glass cuvette (5 mL) covered by a compartment, which was maintained at constant temperature (20 ( 0.2 °C) by circulating water from a thermostat. It was then allowed to stand for 30 min to achieve constant temperature and humidity in the compartment. Then a drop of emulsifier solution (5-8 µL) was delivered and allowed to stand at the tip of the needle for about 180 min to achieve adsorption at the air-water interface. An image of the drop was continuously recorded by a CCD camera and digitalized. The surface tension (σ) was calculated by analyzing the profile of the drop.36 The average standard accuracy of the surface tension for at least two measurements with different drops was roughly (0.5 mN/m. For surface dilatational property measurements of adsorbed emulsifier films at the air-water interface, the same automatic drop tensiometer was utilized as described elsewhere.38 Briefly, the method involved a periodic automatically controlled, sinusoidal interfacial compression and expansion performed by decreasing and increasing the drop volume at the desired amplitude (∆A/A) and angular frequency (ω)). The surface dilatational modulus (E ) Ed + iEv), its elastic (Ed ) E cos φ) and viscous (Ev ) E sin φ) components, and the phase angle (φ) were derived from the change in surface pressure, resulting from a small change in surface area (A) (eq 1). For a perfectly elastic material, the stress and strain are in phase, that is, the phase angle between stress and strain is zero (φ ) 0), and the imaginary term is zero. In the case of a perfectly viscous material, φ ) 90° and the real part is zero. The loss-angle tangent can be defined by the reason between imaginary and real components of the modulus (tan φ ) Ev/Ed). If the film is purely elastic, the loss-angle tangent is zero. The

Figure 1. Effect of Tween 20 concentration in the aqueous phase on the evolution of surface pressure with the square root of time for adsorption of 7S soy globulin at the air-water interface. Symbols: (O) 7S at pH 5, (b) 7S at pH 7, (4) Tween 20 at 5 × 10-6 M and at pH 5 (2) 7S + Tween 20 at 5 × 10-6 M and at pH 5, (]) Tween 20 at 1 × 10-4 M and at pH 5 ([) 7S + Tween 20 at 1 × 10-4 M and at pH 5; protein concentration in aqueous solution 0.1 wt %; ionic strength 0.05 M; temperature 20 °C.

surface dilatational properties were measured as a function of time, θ.

E)

dσ dπ )dA/A d ln A

(1)

The drop was subjected to repeated measurements of 5 sinusoidal oscillation cycles followed by a time corresponding to 50 cycles without any oscillation up to 180 min for protein adsorption. The deformation amplitude (∆A/A) and deformation frequency (ω) were maintained constants at 15% and 100 mHz, respectively. The percentage area change was determined in preliminary experiments to be in the linear region (data not shown). The average standard accuracy of the surface tension was roughly 0.1 mN/m. However, the reproducibility of the viscoelastic properties (for at least two measurements) was better than 5%. Adsorption Kinetics at the Air-Water Interface. The main features of the adsorption process include:39 (i) the diffusion of the emulsifier from the bulk onto the interface, (ii) adsorption (penetration) and interfacial unfolding, and (iii) aggregation (rearrangement) within the interfacial layer, multilayer formation, and even interfacial gelation. The third step is involved in biopolymer (protein or polysaccharide) adsorption but is normally absent during the adsorption of surfactants and lipids at fluid interfaces.13 However, adsorption of proteins is generally a complex process, often involving several types of conformational changes that may be either reversible or irreversible, and, in addition, time dependent.40,41 Because of its influence on foaming, the analysis of the adsorption kinetics of protein and Tween 20 will be centered on the diffusion of the emulsifier from the aqueous bulk phase onto the air-water interface. At low surface concentrations, the surface pressure is low and emulsifier molecules adsorb irreversibly by diffusion. In the case of diffusion-controlled adsorption, the first step of the adsorption process can be obtained from a modified form of the Ward and Tordai equation (eq 2).42

π ) 2C0kT(Dθ/3.14)1/2

(2)

where k is the Boltzmann constant, C0 is the concentration in the aqueous phase, D is the diffusion coefficient, and T is the absolute temperature. If the diffusion to the interface controls the adsorption process, a plot of π against θ1/2 will then be linear43,44 and the slope of this plot will be the diffusion rate constant (kdiff).

2878

Ind. Eng. Chem. Res., Vol. 47, No. 9, 2008

Figure 2. Effect of Tween 20 concentration in the aqueous phase on the evolution of surface pressure with square root of time for adsorption of 11S soy globulin at the air-water interface. Symbols: (O) 11S at pH 5, (b) 11S at pH 7, (4) Tween 20 at 5 × 10-6 M and at pH 5 (2) 11S + Tween 20 at 5 × 10-6 M and at pH 5, (]) Tween 20 at 1 × 10-4 M and at pH 5 ([) 11S + Tween 20 at 1 × 10-4 M and at pH 5; protein concentration in aqueous solution 0.1 wt %; ionic strength 0.05 M; temperature 20 °C.

Figure 4. Effect of Tween 20 concentration in the aqueous phase on the evolution of (A) surface dilatational modulus and (B) phase angle for 11S soy globulin adsorption at the air-water interface. Symbols: (O) 11S at pH 5, (b) 11S at pH 7, (4) Tween 20 at 5 × 10-6 M and at pH 5 (2) 11S + Tween 20 at 5 × 10-6 M and at pH 5, (]) Tween 20 at 1 × 10-4 M and at pH 5 ([) 11S + Tween 20 at 1 × 10-4 M and at pH 5. The deformation amplitude (∆A/A) and deformation frequency (ω) were 15% and 100 mHz, respectively. Protein concentration in aqueous solution 0.1 wt %; ionic strength 0.05 M; temperature 20 °C.

Figure 3. Effect of Tween 20 concentration in the aqueous phase on the evolution of (A) surface dilatational modulus and (B) phase angle for 7S soy globulin adsorption at the air-water interface. Symbols: (O) 7S at pH 5, (b) 7S at pH 7, (4) Tween 20 at 5 × 10-6 M and at pH 5 (2) 7S + Tween 20 at 5 × 10-6 M and at pH 5, (]) Tween 20 at 1 × 10-4 M and at pH 5 ([) 7S + Tween 20 at 1 × 10-4 M and at pH 5; protein concentration in aqueous solution 0.1 wt %; ionic strength 0.05 M; temperature 20 °C.

The rate of penetration and unfolding at the interface of adsorbed protein molecules was deduced from the application of a first-order phenomenological kinetic equation to the time evolution of π or E,45

ln(πf - πθ) /(πf - π0) ) -kiθ

(3)

where πf, π0, and πθ are the surface pressures at the final adsorption time of each step, at the initial time θ0, and at any time θ, respectively, and ki is the first-order rate constant. In practice, a plot of eq 3 usually yields two or more linear regions. The initial slope is taken to correspond to a first-order rate constant of penetration (kAds), whereas the second slope

corresponds to a first-order rate constant of protein rearrangement (kR).38 The fit of the experimental data to the mechanism was made at a time interval based on the best linear regression coefficient. However, because protein adsorption at fluid interfaces is very time-consuming, no attempt was made to discuss the experimental data for the second rearrangements step of previously adsorbed protein molecules. The application of eqs 2 and 3 to the adsorption kinetics of biopolymers (milk, soy proteins, sunflower protein hydrolysates, and polysaccharides) to evaluate the rates of diffusion and adsorption-penetration-rearrangement of the biopolymer at the air-water interface has been discussed elsewhere.13,37,38,46,47 After a rapid diffusion of biopolymer to the interface, the penetration, unfolding, and rearrangement of biopolymer at the interface control the rate of protein adsorption. Foaming Properties. The foaming properties of soy globulin and Tween 20 aqueous solutions were characterized according to their foam formation and stability measured in a commercial instrument, as described elsewhere.12 With this instrument, the foam formation and the foam stability can be determined by conductometric and optical measurements (through the foam volume). The foam is generated by blowing gas (nitrogen) at a flow of 45 mL/min through a porous glass filter (pore diameter 0.2 µm) at the bottom of a glass tube where 20 mL of the foaming agent solution under investigation is placed. The foam volume is determined using a CCD camera. The drainage of water from the foam was followed via conductivity measurements at different heights of the foam column. A pair of electrodes at the bottom of the column was used to measure the quantity of liquid that was not in the foam, whereas the volume of liquid in the foam was measured by conductimetry in three pairs of electrodes located along the glass column. In all experiments, the foam was allowed to reach a volume of

Ind. Eng. Chem. Res., Vol. 47, No. 9, 2008 2879 Table 1. Characteristics Parameters for Adsorption of 7S and 11S Soy Globulins at 0.1 wt %, Tween 20 at Two Representative Concentrations in Solution (as Some Examples) and Their Mixtures at the Air-Water Interface at 20 °C and at Ionic Strength 0.05 Ma θinduction (s) Tween 20 (10-4 M) Tween 20 (5 × 10-6 M) 7S (10-1%) 7S (10-1%) + Tween 20 (10-4 M) 7S (10-1%) + Tween 20 (5 × 10-6 M) 11S (10-1%) 11S (10-1%) + Tween 20 (10-4 M) 11S (10-1%) + Tween 20 (5 × 10-6 M)

0 0 72 0

kdiff (mN‚m-1‚s-0.5) (R) >55 0.65 (0.991) 0.4 (0.999) >60

θdiff (s)

kAds × 104 (s-1) (R)

π180 (mN/m)

E180 (mN/m)

0 500 1090 0

3.1 (0.993) 1.9 (0.990) 2.4 (0.998) 2.25 (0.974)

37.3 29.2 20.0 38.0

28.9 40.9 47.7 28.9

pH 5

0

1.0 (0.982)

130

2.85 (0.994)

26.1

32.4

180 0

0.4 (0.997) >60

900 0

2.3 (0.995) 3.1 (0.994)

18.0 38.1

34.9 29.7

120

2.55 (0.995)

20.6

28.4

20 21

8.8 (0.995) 3.1 (0.996)

27.1 26.3

47.9 57.0

0

0.78 (0.967)

0 7

2.5 (0.999) 3.9 (0.993)

pH 7

7S (10-1%) 11S (10-1%)

a Symbols: (θ induction) induction time, (kdiff) rate constant of diffusion, (kAds) rate constant of adsorption, (θdiff) period at which diffusion controls the kinetic of adsorption of emulsifiers at the air-water interface, (π180 and E180) surface pressure and surface dilatational modulus at 180 min of adsorption time, and (R) linear regression coefficient.

120 mL. The bubbling was then stopped and the evolution of the foam was analyzed. Foaming properties were measured at 20 °C. Three parameters were determined as a measure of foaming capacity. The overall foaming capacity (OFC, mL/s) was determined from the slope of the foam volume curve up to the end of the bubbling. The foam capacity (FC), a measure of gas retention in the foam, was determined by eq 4. The relative foam conductivity (Cf, %) is a measure of the foam density and was determined by eq 5.

FC ) Cf )

Vfoam(f) Vgas(f)

Cfoam(f) 100 Cliq(f)

(4)

(5)

where Vfoam(f) is the final foam volume, Vgas(f) is the final gas volume injected, and Cfoam(f) and Cliq(f) are the final foam and liquid conductivity values, respectively. The static foam stability was determined from the volume of liquid drained from the foam over time.48 The half-life time (θ1/2), referring to the time needed to drain a half of liquid in the foam, was used as a measure of the foam stability. The foam stability was also determined by the time evolution of the foam conductivity.49 The relative conductivity of the foam (Cθ/Ci, where Cθ and Ci are the foam conductivity values at time θ ) θ and at θ ) 0, respectively, of the foam rupture) as a function of time was fitted using a second-order exponential equation,

Cθ/Ci ) A1 exp(-θ/θd) + A2 exp(-θ/θdc)

(6)

where A1 and A2 are adjustable parameters and θd and θdc are the relaxation times, which can be related to the kinetics of liquid drainage (θd) from the foam (including gravitational drainage and marginal regeneration) and disproportionation and foam collapse (θdc), respectively.13,25-27 Results and Discussion Dynamics of Adsorption. The dynamics of the adsorption of 7S and 11S soy globulins at 0.1 wt %, Tween 20 at two representative concentrations in solutions (lower and higher than

the critical micelle concentration), as two examples, and their mixtures were followed by the time evolution of surface pressure (Figures 1 and 2) and surface dilatational properties (surface dilatational modulus and phase angle) (Figures 3 and 4). At the concentration studied, soy globulins are able to saturate the interface at equilibrium.50 For soy globulin adsorption at the air-water interface from protein solutions, we have observed that the rate of surface pressure (π) and surface dilatational modulus (E) change over time depends on the protein and, especially, on the pH and the addition of Tween 20. At Tween 20 concentrations higher than the critical micelar concentration (at CTween 20 ) 1 × 10-4 M), the interface is saturated by Tween 20 at equilibrium. The values of CMC for Tween 20 are between 1.7‚10-5 and 2.0‚10-5 M.51,52 The surface coverage is expected to increase with time, following the same time dependence of the surface pressure and surface dilatational modulus.53 Lag Period. A lag period (θinduction) was observed for 7S (Figure 1) and 11 S (Figure 2) soy globulin adsorption from aqueous solutions, especially at pH 5 (Table 1). For pure aqueous solutions, the lag period is higher for 11S compared to 7S globulin. The presence of an induction time could be related to the time required for the adsorption of sufficient protein molecules to make the interactions between adsorbed molecules appreciable. This lag period has been attributed to the molecular flexibility of the protein and its susceptibility to conformational changes.47,50,54,55 The differences in the lag period between 7S and 11 S globulins at pH 5 must be due to the higher molecular mass of 11S compared to 7S globulin, which can reduce the molecular flexibility of 11S globulin and its susceptibility to conformational changes. The presence of Tween 20 with the protein in aqueous solution also reduces the lag period to zero because the lag period is also absent in pure Tween 20 aqueous solutions, at Tween 20 concentrations lower and higher than the CMC. These results are of practical importance because the absence of a lag period for soy globulinTween 20 mixtures at pH 5 would be beneficial for the formation of the foam, as will be discussed later. Diffusion to the Interface. The kinetics of emulsifier diffusion to the air-water interface can be monitored by measuring changes in surface pressure with the square root of time, after the lag period (Figures 1 and 2). From the slope of the plot of π against θ1/2, we deduce the diffusion rate (kdiff) of emulsifier toward the interface according to eq 2.

2880

Ind. Eng. Chem. Res., Vol. 47, No. 9, 2008

Figure 5. Effect of Tween 20 concentration in the aqueous phase on the (A) overall foam capacity (OFC), (B) foam capacity (FC), and (C) relative foam conductivity (Cf %) of aqueous solutions of 7S soy globulin: (a) 7S at pH 5, (b) 7S at pH 7, (c) Tween 20 at 5 × 10-6 M and at pH 5, (d) 7S + Tween 20 at 5 × 10-6 M and at pH 5, (e) Tween 20 at 1 × 10-4 M and at pH 5, and (f) 7S + Tween 20 at 1 × 10-4 M and at pH 5; protein concentration in aqueous solution 0.1 wt %; bubbling gas, nitrogen; gas flow 45 mL/s; temperature 20 °C; ionic strength 0.05 M.

Figure 6. Effect of Tween 20 concentration in the aqueous phase on the (A) overall foam capacity (OFC), (B) foam capacity (FC), and (C) relative foam conductivity (Cf %) of aqueous solutions of 11S soy globulin: (a) 11S at pH 5, (b) 11S at pH 7, (c) Tween 20 at 5 × 10-6 M and at pH 5, (d) 11S + Tween 20 at 5 × 10-6 M and at pH 5, (e) Tween 20 at 1 × 10-4 M and at pH 5, and (f) 11S + Tween 20 at 1 × 10-4 M and at pH 5; protein concentration in aqueous solution 0.1 wt %; bubbling gas, nitrogen; gas flow 45 mL/s; temperature 20 °C; ionic strength 0.05 M.

It can be seen that (Table 1): (i) The constant rate of diffusion (kdiff) for soy globulins is higher at pH 7 compared to at pH 5, because at pH 7 7S globulin presents soluble aggregates of R, R′ and β forms and 11S presents aggregates and subunits of AB and polypeptides A and B.56 However, at pH 5, 7S and 11S globulins present aggregates without polypeptides (in the case of 7S) or with reduced amount of polypeptides A and B (in the case of 11S). In agreement with the penetration theory, the aggregation of soy globulins in acidic aqueous solution (at pH 5) may reduce its diffusion toward the air-water interface. However, the diffusion coefficient also depends on the molecular size and shape and chemical nature of the protein surface (such as the surface hydrophobicity), among other factors. At neutral pH, 11S exhibits greater surface hydrophobicity than 7S,47 a phenomenon that can explain the higher diffusion to the interface of 11S compared to 7S soy globulin (Table 1). (ii) The diffusion of Tween 20 at CTween 20 > CMC is too fast to be detected by the methods used because of the lower molecular masses of the soluble forms.13,51 (iii) The diffusion at pH 5 is also faster for mixed systems. Interestingly, in the presence of Tween 20 the values of kdiff are the same for 7S and 11S globulins at pH 5 and are a little higher than those for pure Tween 20. These results indicate that a competitive adsorption between soy globulins and Tween 20 takes place at the interface, with the higher rate of diffusion for Tween 20, because of its lower molecular mass.51

(iv) The period at which diffusion controls the kinetics of adsorption of soy globulins at the air-water interface (θdiffusion) is higher at pH 5 than at pH 7. That is, the protein requires more time to penetrate, adsorb, and unfold at the interface in the most aggregated forms at pH 5. At pH 5, θdiffusion is zero for Tween 20 at CTween 20 > CMC. For Tween 20 at CTween 20 < CMC, θdiffusion is lower than for 11S and 7S soy globulins at pH 5, in this order. Finally, the values of θdiffusion for soy globulin-Tween 20 mixtures are lower than those for pure components, which reflect a positive synergistic effect on θdiffusion for mixed systems. In summary, the results in Figures 1 and 2 and Table 1 reflect the fact that the diffusion of 7S and 11S globulins to a fluid interface depends on the modification in the 11S/7S ratio, the level of association/dissociation of these proteins by varying the pH and the competitive adsorption between protein and Tween 20 in the aqueous phase. Adsorption and Penetration at the Interface. At long-term adsorption, the rate of adsorption is lower than the rate of diffusion (Figures 1 and 2) because an energy barrier exists and the rate of protein penetration into the interfacial film starts to be rate-limiting.37,45 We find, for all experiments on protein adsorption, two linear regions in the plot of ln[(π180 - πθ)/ (π180 - π0)] versus θ according to eq 3 (data not shown). The values of the slope of the first linear region can be associated

Ind. Eng. Chem. Res., Vol. 47, No. 9, 2008 2881

Figure 7. Evolution of the overall foaming capacity (OFC) with the rate of diffusion to the air-water interface (kdiff) for aqueous solutions of (O) 7S and (4) 11S soy globulins in the absence and in the presence of Tween 20 at 5 × 10-6 and 1 × 10-4 M in the aqueous phase; protein concentration in aqueous solution 0.1 wt %; bubbling gas, nitrogen; gas flow: 45 mL/s, pH 5; temperature 20 °C; ionic strength 0.05 M. The line is dawn to guide the eye.

with the rate constant of adsorption, penetration, and unfolding at the air-water interface (kAds) for 7S and 11S globulins (Table 1). The rate of adsorption of soy globulins is higher at pH 7 compared to pH 5 because the proteins are adsorbed in a more aggregated state at pH 5. For Tween 20 the rate of adsorption is facilitated at the higher concentration, coinciding with a higher interfacial concentration after diffusion. For soy globulinTween 20 mixed films, the values of kAds are the same (for 11S) or a little lower (for 7S) than that for pure Tween 20 at CTween 20 > CMC. For CTween 20 < CMC, the values of kAds for mixed films are the same as those for pure soy globulins, which are higher than that for pure Tween 20 at CTween 20 < CMC. Because of their influence on the stability of foams,31 the values of surface pressure (π180) and surface dilatational modulus (E180) at long-term adsorption (at 180 min of adsorption time) are included in the Table 1. It can be seen that: (i) the values of π180 for soy globulins are lower at pH 5 compared to pH 7, but minor differences are observed between 7S and 11S proteins at pH 5 or 7. The values of E180 are lower for 11S at pH 7 compared to those at pH 5 but are the same for 7S at pH 5 and 7. (ii) For Tween 20, the values of π180 are higher at CTween 20 > CMC, as this surfactant saturated the interface, but the opposite is observed for the values of E180. (iii) For soy globulin-Tween 20 mixed films at CTween 20 > CMC, the values of π180 are the same as that for pure Tween 20, but at CTween 20 < CMC the values of π180 for 11S-Tween 20 mixed films are similar to that for pure 11S soy globulin or between those for pure components for 7S-Tween 20 mixed films. Thus, for pure components, the interfacial concentration, which is denoted by the values of π180, is higher for proteins at pH 7 compared to pH 5 and for Tween 20 and soy globulinTween 20 mixed systems at CTween 20 > CMC. The combined effects of interfacial concentration and emulsifier interactions at the interface, which are denoted by the values of E180, are higher for soy globulins compared to mixed systems because both emulsifiers saturated the air-water interface. The coadsorption of soy globulins and Tween 20 at the air-water interface weakens the mechanical properties of the mixed films in relation to those for pure components. Surface Dilatational Characteristics. From the transient surface viscoelastic properties (surface dilatational modulus, E, its elastic, Ed, and viscous, Ev, components and the loss angle, φ) for pure and soy globulins + Tween 20 mixed systems, it was observed (data not shown) that the values for the surface

Figure 8. Effect of Tween 20 concentration in the aqueous phase on the (A) half-life time (θ1/2, s), (B) relaxation time of drainage (θd, s), and (C) relaxation time of disproportionation/collapse (θdc, s) of foams generated from aqueous solutions of 7S soy globulin: (a) 7S at pH 5, (b) 7S at pH 7, (c) Tween 20 at 5 × 10-6 M and at pH 5, (d) 7S + Tween 20 at 5 × 10-6 M and at pH 5, (e) Tween 20 at 1 × 10-4 M and at pH 5, and (f) 7S + Tween 20 at 1 × 10-4 M and at pH 5; protein concentration in aqueous solution 0.1 wt %; bubbling gas, nitrogen; gas flow 45 mL/s; temperature 20 °C; ionic strength 0.05 M.

dilatational modulus were very similar to those for the dilatational elasticity and the dilatational viscosity values were low. Thus, over the adsorption period studied here the film behaved, from a rheological point of view, as viscoelastic (practically elastic) with a loss-angle tangent higher than zero. Timedependent surface dilatational modulus (E) and loss angle are plotted for adsorbed films of 7S (Figure 3) and 11S (Figure 4) soy globulins at 0.1 wt %, Tween 20 (at two representative concentrations in solution, as some examples), and their mixtures at the air-water interface. The increase in E with time (part A of Figure 3 and part A of Figure 4) may be associated with adsorption of emulsifiers at the interface. This behavior was similar to that observed for milk and vegetable proteins,37,53,57 Tween 20,51 lipid,57 and protein-lipid51,58 adsorption at the airwater interface. The results of time-dependent surface dilatational properties for soy globulins are consistent with the existence of proteinprotein interactions, which are thought to be due to protein adsorption at the interface via diffusion, penetration, and rearrangement (looping of the amino acid residues). The looping of the amino acid residues of soy globulin molecules is more closely packed, and the surface density is higher as the adsorption time increases.47 The closer packing of soy protein at higher adsorption time is a consequence of the existence of a molecular rearrangement of the previously adsorbed soy protein molecules, as is reflected by the significant increment

2882

Ind. Eng. Chem. Res., Vol. 47, No. 9, 2008

in E (part A of Figure 3 and part A of Figure 4). For 7S globulin, the values of E during the adsorption are higher at pH 7 compared to pH 5, but are the same at long-term adsorption. For 11S soy globulin, the maximum value for E over time was observed at pH 7, compared to pH 5. That is, as the molecular structure of these proteins was more denatured, less aggregated, and with higher possibilities of interactions between amino acid residues at interface.35 Under these conditions (at pH 7), the values of E over time were higher for 11S than for 7S globulin. However, the opposite was observed at pH 5 when the aggregation of these proteins (more for 11S than for 7S globulin) and the absence of significant amounts of free polypeptides does not favor the existence of interactions between amino acid residues. The mechanical properties of the adsorbed protein film (with high E values) must be emphasized for practical reasons because the bubbles can be protected against recoalescence during foaming at high E values. For Tween 20 at CTween 20 < CMC, E increases monotonically with time, but at CTween 20 > CMC the faster adsorption of Tween 20 is reflected by the presence of a plateau in E-time plots (part A of Figure 3 and part A of Figure 4). However, the faster adsorption of Tween 20 from micelar solutions (at CTween 20 > CMC) does not favor the existence of Tween 20-Tween 20 interactions at the air-water interface. Thus, the values of E are lower for Tween 20 at CTween 20 > CMC compared to those at CTween 20 < CMC. For soy globulin-Tween 20 mixed systems at CTween 20 < CMC, the values of E are lower than those for pure components. However, at CTween 20 > CMC the values of E for the mixed systems are the same as those for pure Tween 20, which denotes that Tween 20 dominates at the interface. The same phenomenon was observed for BSA + Tween 2051 and β-lactoglobulindiglycerol ester58 adsorbed films. These results can be explained by the competitive adsorption of protein and Tween 20 at the air-water interface. In fact, at CTween 20 < CMC the adsorption of Tween 20 weakens the protein-protein interactions giving an adsorbed mixed film with E values lower than those for pure components. At CTween 20 > CMC, Tween 20 predominates at the air-water interface. Thus, Tween 20-Tween 20 interactions are predominant but these interactions are weaker than proteinprotein interactions, which agree with the lower E values because of the presence of Tween 20. This Tween 20/protein ratio is a singular composition in mixed films with specific properties,29-31,48,59 which in turn affects the stability of food dispersions.60-63 As reflected by the time evolution of the phase angle (part B of Figure 3 and part B of Figure 4), the viscoelastic behavior of soy globulins, Tween 20, and its mixtures at short adsorption time leads to a more elastic film at long-term adsorption (7S soy globulin at pH 5 is an exception), as a consequence of the molecular rearrangement of previously adsorbed protein molecules and even to the formation of a gel-like protein film at higher adsorption time. The high values of the phase angle (part B of Figure 3 and part B of Figure 4) confirm that Tween 20 at CTween 20 > CMC hinders the formation of a soy globulin gel-like elastic film at pH 5. The viscoelastic behavior of soy globulins at pH 5, due to aggregation of the protein, was also observed at a microscopic level by the topography of spread films.64 However, Tween 20 at CTween 20 < CMC has the capacity to form elastic films by itself and in the presence of soy globulins because the angle values are very low, especially at long-term adsorption. The phase angle for soy globulin-Tween 20 mixed systems follows the same trends as that for pure Tween 20 at CTween 20

Figure 9. Effect of Tween 20 concentration in the aqueous phase on the (A) half-life time (θ1/2, s), (B) relaxation time of drainage (θd, s), and (C) relaxation time of disproportionation/collapse (θdc, s) of foams generated from aqueous solutions of 11S soy globulin: (a) 11S at pH 5, (b) 11S at pH 7, (c) Tween 20 at 5 × 10-6 M and at pH 5, (d) 11S + Tween 20 at 5 × 10-6 M and at pH 5, (e) Tween 20 at 1 × 10-4 M and at pH 5, and (f) 11S + Tween 20 at 1 × 10-4 M and at pH 5. Protein concentration in aqueous solution 0.1 wt %; bubbling gas, nitrogen; gas flow 45 mL/s; temperature 20 °C; ionic strength 0.05 M.

> CMC. However, at CTween 20 < CMC the phase angle of mixed systems is between those for pure 7S and Tween 20 or are even higher than for 11S and Tween 20 pure systems. Thus, the same conclusions in relation to compatibility and interactions, as those derived from E values, can be deduced here from the values of loss angle for soy globulins and Tween 20 adsorbed mixed films. Foaming Characteristics. Foaming Capacity. The overall foaming capacity (OFC, mL/s), the foam capacity (FC), and the relative foam conductivity (Cf, %) as a function of pH and the addition of Tween 20 in the aqueous phase at two representative concentrations in solution (as some examples) for 7S and 11S soy globulins at 0.1 wt % are shown in Figures 5 and 6, respectively. It can be deduced that (i) the overall foaming capacity (OFC), the gas and liquid retentions (FC and Cf, respectively) in the foam and the foam density (Cf) for 7S and 11S soy globulins are higher at pH 7 compared to pH 5, a pH at which these proteins do not produce foam because of the

Ind. Eng. Chem. Res., Vol. 47, No. 9, 2008 2883

Figure 10. Evolution with (A) surface pressure (π180) and (B) surface dilatational modulus (E180) at long-term adsorption (at 180 min of adsorption time) of half-life time (θ1/2, s) of foams generated from aqueous solutions of 7S (open symbols) and 11S (filled symbols) soy globulins as a function of pH (5 and 7), and Tween 20 concentration in solution (at 5 × 10-6 and 1 × 10-4 M); protein concentration in aqueous solution 0.1 wt %; ionic strength 0.05 M; bubbling gas, nitrogen; gas flow 45 mL/s; temperature 20 °C. The lines are drawn to guide the eye.

low protein solubility and aggregation.35 That is, under these conditions soy globulins do not foam enough to reach 120 mL of foam because the rate of formation and stabilization of new bubbles is lower than the rate of foam rupture. (ii) The foam capacity of Tween 20 at CTween 20 < CMC is also zero, but at CTween 20 > CMC the foam capacity of Tween 20 is similar to those for pure soy globulins at pH 7, which is included as reference (Figures 5 and 6). (iii) As for pure components, aqueous solutions of soy globulins and Tween 20 have the capacity to produce foam at pH 5 at CTween 20 > CMC. However, the foaming capacity of these solutions is the same as that for pure Tween 20 aqueous solution at CTween 20 > CMC. These results corroborate the idea that Tween 20 is adsorbed preferentially at the air-water interface and that the foaming capacity of aqueous solutions of soy globulins and Tween 20 is due to the presence of Tween 20 in the mixture. (iv) Soy globulin-Tween 20 mixed systems at CTween 20 < CMC and at pH 5 do not have the capacity to produce foam as observed for pure components at the same concentrations. These results agree qualitatively with observations deduced from dynamic characteristics of adsorbed films (Figures 1-4). The foaming capacity of soy globulin and Tween 20 aqueous solutions is determined by dynamic interfacial properties (presence of lag period and the rate of diffusion and dynamic dilatational modulus). These results confirm the hypothesis that there exists a relationship between the foaming capacity and presence of a lag period and the rate of diffusion of the protein toward the air-water interface (Figure 7).12,13,31 That is, in the presence of a lag period and as the rate of diffusion is lower (for 7S and 11S globulins at pH 5, Tween 20 at CTween 20 < CMC, and their mixtures), the foaming capacity is lower (it is practically zero) because the protein concentration at the

interface (Figures 1 and 2) and the surface dilatational modulus (part A of Figures 3 and part A of Figure 4) are also lower. In summary, the conditions (pH 7 for soy globulins, Tween 20 at CTween 20 > CMC, and their mixtures) that favor the absence of a lag period and a faster diffusion of the protein toward the interface (Table 1) coincide with the optimum foaming capacity, no matter what the protein, 7S, or 11S globulin (Figure 7). Foam Stability. The half-life time of volume of liquid drained from the foam (θ1/2) and the relaxation times related to the kinetics of liquid drainage from the foam (θd) and disproportionation and foam collapse (θdc) for 7S and 11S soy globulins, Tween 20, and their mixtures are shown in Figures 8 and 9. In most of the experiments performed in this study, the empirical equation (eq 6) fits the data of foam stability (Chi2 range within 0.00005-0.00022 and R2 range within 0.996-0.997), indicating that two microscopic processes (liquid drainage, disproportionation and/or collapse) are involved in foam decay. It can be seen that (i) the foam stability of soy globulins is lower (it is zero) at pH 5 compared to at pH 7, which may be related to the high aggregation of this protein at the interface at pH 5. At pH 7, the stability of the foam generated from 7S is higher than that for 11S globulin. (ii) The foam stability of Tween 20 at CTween 20 < CMC is also zero. At CTween 20 > CMC, the foam stability of Tween 20 is lower than those for aqueous solutions of soy globulins at pH 7. (iii) Although soy globulin at pH 5 in combination with Tween 20 at CTween 20 > CMC can produce foam, the foam has poor stability as observed for pure Tween 20. Again the stability of the foam generated from soy globulin + Tween 20 aqueous solutions is due mainly to the presence of preferential adsorption of Tween 20 at the air-water interface. The effect of surface properties at long-term adsorption on foam stability has been analyzed in the literature.12,13,25-27,63,65-67 The relationship between foam stability and the surface pressure at long-term adsorption may be due to increased interfacial adsorption. On the other hand, the combined effects of interfacial adsorption and interfacial interactions between adsorbed soy globulin molecules, which are reflected in the values of E also correlate with the foam stability. In this study we have observed than the effect of the surface pressure (π180) and surface dilatational modulus (E180) at long-term adsorption (Table 1) on foam stability is complex (Figure 10). The increased interfacial adsorption (at high π180 values) and the combined effects of interfacial adsorption and interfacial interactions between adsorbed soy globulin molecules (at high E180 values) can explain the higher stability of the foam (Figure 10). The main deviations were observed for soy globulin foams at pH 5 in the presence of Tween 20 (as the foam is preferentially stabilized by Tween 20, with a high surface density; but with low mechanical properties). Conclusions In many food dispersion formulations, proteins are used in acidic aqueous solutions. However, optimum functionality of soy globulins occurs at pH < 5, which limits their application as food ingredients. In this work, we have analyzed the effect of the addition of a surfactant (Tween 20) to improve the interfacial and foaming characteristics of soy globulins. The dynamics of adsorption and the surface dilatational characteristics and foam properties (foam capacity and stability) of aqueous solutions of β-conglycinin (a 7S globulin) and glycinin (an 11S globulin), at pH 5 and 7, were studied as a function of

2884

Ind. Eng. Chem. Res., Vol. 47, No. 9, 2008

the Tween 20 concentration in solution (CTween 20, higher and lower than the critical micelle concentration, CMC). We have observed that (i) the adsorption of soy globulins depends on the particular protein (7S or 11S soy globulin), the level of association/dissociation/aggregation of these proteins by varying the pH, and on the competitive adsorption between protein and Tween 20 in the aqueous phase. The adsorption of soy globulins is much improved by the addition of Tween 20 at CTween 20 > CMC. These results are of practical importance because of are necessary low Tween 20 concentrations in acidic aqueous solutions (at pH 5) to improve the rate of adsorption of these proteins at the air-water interface. (ii) The surface dilatational properties reflect the fact that soy globulin and Tween 20 adsorbed mixed films exhibit viscoelastic, practically elastic, behavior. The surface dilatational modulus decreases with the addition Tween 20 into the aqueous phase. (iii) The addition of Tween 20 at CTween 20 > CMC, which favor the adsorption of the protein toward the interface, coincide with the optimum foaming capacity, no matter what the protein, 7S or 11S globulin. There exists a relationship between the foaming capacity and the rate of diffusion of Tween 20 and soy globulins toward the air-water interface. (iv) The stability of the foam generated from soy globulin + Tween 20 acidic aqueous solutions (at pH 5) is due mainly to the preferential adsorption of Tween 20 at the air-water interface. The increased interfacial adsorption (at high π180 values) and the combined effects of interfacial adsorption and interfacial interactions between adsorbed soy globulin and Tween 20 molecules (at high E180 values) can explain the stability of the foam, with few exceptions. Acknowledgment The authors acknowledge the support of CICYT through Grant AGL2007-60045 and Junta de Andalucı´a through Grant PO6-AGR-01535. Literature Cited (1) Damodaran, S.; Paraf, A. Food Proteins and Their Application; Dekker: New York, 1997. (2) Dickinson E. An Introduction to Food Colloids; Oxford University Press: Oxford, U. K., 1992. (3) Dickinson, E. Interfacial, Emulsifying and Foaming Properties of Milk Proteins. In AdVanced Dairy Chemistry, Vol. 1 Proteins, parts A and B; Fox, P. F., McSweeney, P., Eds.; Kluwer Academic/Plenum Publishers: Dordrecht, The Netherlands, 2003; p 1229. (4) Campbell, G. M.; Mougeot, E. Creation and Characterization of Aerated Food Products. Trends Food Sci. Technol. 1999, 10, 283. (5) Bubbles in Food; Campbell, G. M., Webb, C., Padiela, S., Nirajan, K., Eds.; Eagan Press: St. Paul, MN, 1999. (6) Defoaming: Theory and Industrial Applications; Garret, P. R., Ed.; Marcel Dekker: New York, 1993. (7) Denkov, N. D. Mechanisms of Foam Destruction by Oil-Based Antifoams. Langmuir 2004, 20, 9463. (8) Dickinson, E. Protein Adsorption at Liquid Interfaces and the Relationship to Foam Stability. In Foams: Physics, Chemistry and Structure; Wilson, A. J., Ed.; Springer: London, 1989; p 39. (9) Prins, A. AdVances in Food Emulsions and Foams; Dickinson, E., Stainsby, G., Eds.; Elsevier Applied Science: London, 1989; p 91. (10) Prins, A.; van Kalsbeek, H. K. Foaming Behaviour and Dynamic Surface Properties of Liquids. Curr. Opin. Colloid Interface Sci. 1998, 3, 639. (11) Prins, A. Stagnant Surface Behaviour and its Effect on Foam and Film Stability. Colloids Surf., A 1999, 149, 467. (12) Carrera, C.; Rodrı´guez Patino, J. M. Interfacial, Foaming and Emulsifying Characteristics of Sodium Caseinate as Influenced by Protein Concentration in Solution. Food Hydrocolloids 2005, 19, 407. (13) A Ä lvarez, J. M.; Rodrı´guez, Patino, J. M. Formulation Engineering of Food Model Foams Containing Diglycerol Esters and β-Lactoglobulin. Ind. Eng. Chem. Res. 2006, 45, 7510.

(14) Rodrı´guez Patino, J. M.; Min˜ones, J., Jr.; Milla´n, H., Jr.; Pedroche, J. J.; Carrera, C.; Pizones, V.; Milla´n, F. Interfacial and Foaming Properties of Enzyme-Induced Hydrolysis of Sunflower Protein Isolate. Food Hydrocolloids 2007, 21, 782. (15) Murray, B. S. Stabilization of Bubbles and Foams. Curr. Opin. Colloid Interface Sci. 2007, 12, 232. (16) Damodaran, S. Protein Stabilization of Emulsions and Foams. J. Food Sci. 2005, 70, R54. (17) Wilde, P. J.; Clark, D. C. Methods in Testing Protein Functionality; Hall, G. M., Ed.; Blackie Academic and Professional: London, 1996; p 110. (18) Halling, P. J. Protein Stabilized Foams and Emulsions. Crit. ReV. Food Sci. Nutr. 1981, 13, 155. (19) Wilde, P. J. Interfaces: Their Role in Foam and Emulsion Behaviour. Curr. Opin. Colloid Interface Sci. 2000, 5, 176. (20) Murray, B. S. Interfacial Rheology of Food Emulsifiers and Proteins. Cur. Opin. Colloid Interface Sci. 2002, 7, 426. (21) McClements, D. J. Food Emulsions: Principles, Practice and Techniques, 2nd ed.; CRC Press: Boca Raton, Florida, 2005. (22) Vioque, J.; Clemente, A.; Pedroche, J.; Yust, M. M.; Milla´n, F. Obtencio´n y Aplicaciones de Hidrolizados Proteicos. Grasas Aceites 2001, 52, 132. (23) Moure, A.; Sineiro, J.; Domı´nguez, H.; Parajo, J. C. Functionality of Oilseed Protein Products: A Review. Food Res. Int. 2006, 39, 945. (24) Utsumi, S.; Matsumura, Y.; Mori, T. Structure-Function Relationships of Soy Proteins. In Food Proteins and Their Applications; Damodaran, S.; Paraf, A., Eds.; Dekker: New York, 1997; p 257. (25) Pizones, V.; Carrera, C.; Yust, M. M.; Pedroche, J.; Milla´n, F.; Rodrı´guez Patino, J. M. Limited Enzymatic Hydrolysis can Improve the Interfacial and Foaming Properties of Soy Globulins. J. Agric. Food Chem. 2007, 55, 1536. (26) Pizones, V.; Carrera, C.; Rodrı´guez Patino, J. M. Formulation Engineering can Improve the Interfacial and Foaming Properties of Soy Globulins. J. Agric. Food Chem. 2007, 55, 6339. (27) Pizones, V.; Carrera, C.; Yust, M. M.; Pedroche, J. J.; Milla´n, F.; Rodrı´guez Patino, J. M. Interfacial and Foaming Characteristics of Soy Globulins as a Function of pH and Ionic Strength. Colloids Surf., A 2007, 309, 202. (28) Larsson, K. Food Emulsions; Friberg. S., Larsson, K., Eds.; Marcel Dekker: New York, 1997; p. 39. (29) Rodrı´guez Patino, J. M.; Rodrı´guez Nin˜o, M. R.; Carrera, C. ProteinEmulsifier Interactions at the Air-Water Interface. Curr. Opin. Colloid Interface Sci. 2003, 8, 387. (30) Rodrı´guez Patino, J. M.; Rodrı´guez Nin˜o, M. R.; Carrera, C. Physico-Chemical Properties of Surfactant and Protein Films. Curr. Opin. Colloid Interface Sci. 2007, 12, 187. (31) Rodrı´guez Patino, J. M.; Rodrı´guez Nin˜o, M. R.; Carrera, C. Implications of Interfacial Characteristics of Food Emulsifiers in Foam Formulations. AdV. Colloid Interface Sci. 2007, doi: 10.1016/j.cis.2007.12.007. (32) Semenova, M. G.; Belyakova, L. E.; Polikarpov, Y. N.; Ii’in, M. M.; Istarova, T. A.; Anokhina, M. S.; Tsapkina, E. N. Thermodynamic Analysis of the Impact of the Surfactant-Protein Interactions on the Molecular Parameters and Surface Behaviour of Food Proteins. Biomacromolecules 2006, 7, 101. (33) Marze, S. P. L.; Saint-Jalmes, A.; Langenin, D. Protein and Surfactant Foams: Linear Rheology and Dilatancy Effect. Colloids Surf., A 2005, 263, 121. (34) Saint-Jalmes, A.; Peugeot, M.-L.; Ferraz, H.; Langevin, D. Differences Between Protein and Surfactant Foams: Microscopic Properties, Stability and Coarsening. Colloids Surf., A 2005, 263, 219. (35) Molina, S.; Carrera, C.; Rodrı´guez Nin˜o, M. R.; An˜o´n, C.; Rodrı´guez Patino, J. M. Structural Characterization and Surface Activity of Spread and Adsorbed Soy Globulin Films at Equilibrium. Colloids Surf., B 2003, 32, 57. (36) Labourdenne, S.; Gaudry-Rolland, N.; Letellier, S.; Lin, M.; Cagna, A.; Verger, R.; Rivie´re, C. The Oil-Drop Tensiometer: Potential Applications for Studying the Kinetics of (Phospho) Lipase Action. Chem. Phys. Lipids 1994, 71, 163. (37) Rodrı´guez Nin˜o, M. R.; Rodrı´guez Patino, J. M. Effect of the Aqueous Phase Composition on the Adsorption of Bovine Serum Albumin to the Air-Water Interface. Ind. Eng. Chem. Res. 2002, 41, 1489. (38) Min˜ones, J., Jr.; Rodrı´guez Patino, J. M. Rheological Properties of Hydrolysates of Proteins From Extracted Sunflower Flour Adsorbed at the Air-Water Interface. Ind. Eng. Chem. Res. 2005, 44, 7761. (39) MacRitchie F. Chemistry at Interfaces; Academic Press: San Diego, CA., 1990. (40) Makievski, A. V.; Fainerman, V. B.; Bree, M.; Wu¨stneck, R.; Kra¨gel, J.; Miller. R. Adsorption of Proteins at the Liquid/Air Interface. J. Phys. Chem. B 1998, 102, 417.

Ind. Eng. Chem. Res., Vol. 47, No. 9, 2008 2885 (41) Aksenenko, E. V.; Kovalchuk, V. I.; Fainerman, V. B.; Miller, R. Surface Dilational Rheology of Mixed Adsorption Layers at Liquid Interfaces. AdV. Colloid Interface Sci. 2006, 122, 57. (42) Ward, A. F. H.; Tordai, L. Time Dependence of Boundary Tensions of Solutions. J. Chem. Phys. 1946, 14, 453. (43) MacRitchie, F. Protein Adsorption/Desorption at Fluid Interfaces. Colloids Surf. 1989, 41, 25. (44) Xu, S.; Damodaran, S. Kinetics of Adsorption of Protein at the Air-Water Interface From a Binary Mixture. Langmuir 1994, 10, 472. (45) Graham, D. E.; Phillips, M. C. Proteins at Liquid Interfaces. I. Kinetics of Adsorption and Surface Denaturation, J. Colloid Interface Sci. 1979, 70, 403. (46) Horne, D. S.; Rodrı´guez Patino, J. M. Biopolymers at Interfaces; Malmsten, M, Ed.; Marcel Dekker: New York, 2003; p 857. (47) Rodrı´guez Patino, J. M.; Carrera, C.; Molina, S.; Rodrı´guez Nin˜o, M. R.; An˜o´n, C. Adsorption of Soy Globulin Films at the Air-Water Interface. Ind. Eng. Chem. Res. 2004, 43, 1681. (48) Rodrı´guez Patino, J. M.; Rodrı´guez Nin˜o, M. R.; Alvarez Go´mez, J. M. Interfacial and Foaming Characteristics of Protein-Lipid Systems. Food Hydrocolloids 1997, 11, 49. (49) Kato, A.; Takanashi, A.; Matsudomi, D.; Kobayashi, K. Determination of oaming Properties of Proteins by Conductivity Measurements. J. Food Sci. 1983, 48, 62. (50) Rodrı´guez Nin˜o, M. R.; Carrera, C.; Pizones, V.; Rodrı´guez Patino, J. M. Milk and Soy Protein Films at the Air-Water Interface. Food Hydrocolloids 2005, 19, 417. (51) Rodrı´guez Nin˜o, M. R.; Wilde, P. J.; Clark, D. C.; Rodrı´guez Patino, J. M. Rheokinetic Analysis of Bovine Serum Albumin and Tween 20 Mixed Films on Aqueous Solutions. J. Agric. Food Chem. 1997, 45, 3016. (52) Maldonado, M.; Martin Molina, A.; Martin Rodrı´guez, A.; Cabrerizo, M.; Ga´lvez, M. J.; Langevin, D. Surface Properties and Foam Stability of Protein/Surfactant Mixtures: Theory and Experiment. J. Phys. Chem. C 2007, 111, 2715. (53) Benjamins, J. Static and Dynamic Properties of Protein Adsorbed at Liquid Interfaces, Ph.D. Thesis, Wageningen University, 2000. (54) Miller, R.; Fainerman, V. B.; Makievski, A. V.; Kra¨gel, J.; Grigoriev, D. O.; Kazakov, V. N.; Sinyachenko, O. V. Dynamic of Protein and Mixed Protein/Surfactant Adsorption Layers at the Water/Fluid Interface. AdV. Colloid Interface Sci. 2000, 86, 39. (55) Razumosvky, L.; Damodaran, S. Surface Activity-Compressibility Relationship of Proteins at the Air-Water Interface. Langmuir 1999, 5, 1392. (56) Lakemond, C. M. M.; de Jongh, H. H. J.; Hessing, M.; Gruppen, H.; Voragen, A. G. J. Soy Glycinin: Influence of pH and Ionic Strength on Solubility and Molecular Structure at Ambient Temperatures. J. Agric. Food Chem. 2000, 48, 1985.

(57) A Ä lvarez, J. M.; Rodrı´guez Patino, J. M. Viscoelastic Properties of Diglycerol Ester and Protein Adsorbed Films at the Air-Water Interface. Ind. Eng. Chem. Res. 2007, 46, 2693. (58) A Ä lvarez, J. M.; Pizones, V.; Carrera; C.; Rodrı´guez Patino, J. M. The Role of Static and Dynamic Characteristics of Diglycerol Esters and β-Lactoglobulin Mixed Films on Foaming. 1. Dynamic phenomena at the air-water interface. Food Hydrocolloids 2008, 22, 1105–1116. (59) Wilde, P. J.; Rodrı´guez Nin˜o, M. R.; Clark, D. C.; Rodrı´guez Patino, J. M. Molecular Diffusion and Drainage of Thin Liquid Films Stabilized by Bovine Serum Albumin-Tween 20 Mixtures in Aqueous Solutions of Ethanol and Sucrose. Langmuir 1997, 13, 7151. (60) Chen, J.; Dickinson, E. Protein/Surfactant Interactions. Part 1. Flocculation of Emulsions Containing Mixed Protein+Surfactant. Colloids Surf., A 1995, 100, 255. (61) Chen, J.; Dickinson, E. Protein/Surfactant Interactions. Part 2. Electrophoretic Mobility of Mixed Protein+Surfactant. Colloids Surf., A 1995, 100, 267. (62) Chen, J.; Dickinson, E. Surface Shear Viscosity and Protein Surfactant Interactions in Mixed Protein Films Adsorbed at the Oil-Water Interface. Food Hydrocolloids 1995, 9, 35. (63) A Ä lvarez, J. M.; Pizones, V.; Carrera; C.; Rodrı´guez Patino, J. M. The Role of Static and Dynamic Characteristics of Diglycerol Esters and β-Lactoglobulin Mixed Films on Foaming. 2. Adsorption and Foaming. Food Hydrocolloids 2007, doi: 10.1016/j.foodhyd.2007.06.015. (64) Carrera, C.; Rodrı´guez Nin˜o, M. R.; Molina, S.; An˜o´n, C.; Rodrı´guez Patino, J. M. Soy Globulin Spread Films at the Air-Water Interface. Food Hydrocolloids 2004, 18, 335. (65) Martin, A. H.; Grolle, K.; Bos, M. A.; Cohen Stuart, M. A.; van Vliet, T. Network Forming Properties of Various Proteins Adsorbed at the Air/Water Interface in Relation to Foam Stability. J. Colloid Interface Sci. 2002, 254, 175. (66) Baeza, R.; Carrera, C.; Pilosof, A. M. R.; Rodrı´guez Patino. Interfacial and Foaming Properties of Propylenglycol Alginates. Effect of Degree of Esterification and Molecular Weight. Colloids Surf., B 2004, 36, 139. (67) Baeza, R.; Carrera, C.; Pilosof, A. M. R.; Rodrı´guez Patino, J. M. In Food Colloids: Interactions, Microstructure and Processing; E. Dikinson, Ed.; Royal Society of Chemistry: Cambridge, U. K., 2005; p 301.

ReceiVed for reView November 7, 2007 ReVised manuscript receiVed February 7, 2008 Accepted February 19, 2008 IE071518F