Interfaces mixed protein films

Mixed-Protein Films Adsorbed at the Oil-Water Interface... 

Dickinson, E., Rolfe, S.E., Dalgleish, D.G. (1990). Surface shear viscometry as a probe of protein-protein interactions in mixed milk protein films adsorbed at the oil-water interface. International Journal of Biological Macromolecules, 12, 189-194. 

Though most studies on protein adsorption at interfaces have been conducted in solutions having a single well characterized protein, evidence has emerged in recent years that film properties in mixed protein systems are much more complex than in single protein systems. 

There are two principal problems with penetration experiments the adsorption characteristics of the protein have to be understood, and the amount of protein that adsorbs to the interface when lipid is present has to be determined. Previously, most researchers used the change in film pressure (Atr) as a measure of the amount of protein that interacted with the lipid monolayer. However, this approach implicitly assumes that the adsorption of protein can be described by Gibbs adsorption equation, but as pointed out by Colacicco (6), this is invalid for proteins which adsorb irreversibly. Because the surface concentration of protein is unknown, radiolabeled proteins have been used (8, 9, 10). This work has been concerned exclusively with highly water-soluble proteins whose prime mode of interaction with monolayers (and bilayers) is electrostatic. In these cases a simple description of the packing in the mixed lipid-protein films was impossible (6). 

The now classic experiments of Graham and Phillips showed (5) inter alia that the surface rheology of pure protein films is very dependent on the type and amount of protein adsorbed, as well as on the conditions of adsorption. In this paper, with films formed from mixed protein solutions, we shall show that the surface viscosity is an extremely sensitive probe of the time-dependent structural and compositional changes taking place during competitive adsorption at the oil—water interface. While steady-state tensions can invariably... 

Figure 1.4 Mixed protein and surfactant interfaces Weak protein interactions and restricted diffusion of surfactants result in reduced stability and probable film rupture.

Protein Adsorption. Films were kept submerged in bulfer in individual bottles. Concentrated protein solution in the same buflFer (and at the same temperature and pH) was added with a pipette to avoid exposing the films to the air/water interface. The solutions were mixed by swirling the films. In studies at 37°C, the solutions were kept in a water bath regulated to ztI°C. 

The existence of protein-LMWE interactions depends on the interfacial composition and on the protein/LMWE ratio. In general, the surface activity of the mixed films is determined by the LMWE as the surface pressure of the mixed film is the same as the LMWE equilibrium spreading pressure, and the monolayer is not saturated by the protein. However, the protein determines the surface activity of mixed films as the protein saturates the monolayer. In the intermediate region there exists coexistence of protein and LMWE at the interface. 

When a soluble LMWE (like Tween 20) as well as a protein is present in water both components will form adsorbed films at the air-water interface. At low LMWE concentrations, protein reduces the surface tension to a greater extent than protein-LMWE mixed systems. However, the opposite was observed at high LMWE concentrations, above the critical micelle concentration (CMC), because the protein molecules are displaced from the interface by the LMWE. Over the intermediate region, close to the CMC, both protein and LMWE coexist at interface. However, tensiometric studies indicate that the compatibility of proteins and nonionic emulsifier at fluid interfaces is very poor, in contrast to mixtures of ionic-surface-active homologues. 

Using a unique device that incorporates different interfacial techniques, such as surface film balance and Brewsfer angle microscopy (BAM), we have analyzed fhe sfrucfural characferisfics of profein-LMWE mixed films spread on fhe air-water interface (Pafino ef al., 2003 Lucero, in press). At surface pressures lower than that for profein collapse a mixed monolayer of LMWE and protein may exist. At surface pressures higher than that for protein collapse, collapsed protein residues may be displaced from the interface by LMWE molecules fhaf is, fhe mixed film is practically dominated by LMWE molecules the tt-A isotherms of fhe mixed film are parallel to that of the lipid. 

Surface dilatational rheology is a very sensitive technique to analyze the competitive adsorption/displacement of protein and LMWE emulsifier at the air-water interface (Patino et al., 2003). A common trend is that the surface dilatational modulus increases as the monolayer is compressed and is a maximum at the highest surface pressures, at the collapse point of the mixed film, and as the content of LMWE in the mixture increases. At higher TT, the collapsed protein residues displaced from the interface by LMWE molecules have important influence on the dilatational characteristics of the mixed films. The mechanical properties of the mixed films also demonstrate that, even at the highest tt, the LMWE is unable to displace completely protein molecules from the air-water interface. 

Recent works by the Norwich group (Gunning et al., 1999 Mackie et ak, 1999, 2000 Wilde, 2000) have demonstrated how surfactants disrupt and displace proteins from an interface by a three-stage "orogenic" mechanism. We have adopted this mechanism to describe spread protein-LMWE mixed films (Figure 14.8) (Patino et al., 2003). The results suggest that for spread... 

In the mixed systems, the behavior was similar to that observed for surface pressure. In the presence of surface-active PGA (Figure 25.3a and b) at low concentrations in the bulk phase (0.1 wt%), competition between the biopolymers at the interface results in a lower Ed than that expected from the observation of the single components. However, at higher concentrations of PS and long adsorption times, a cooperative adsorption can be deduced. This result could be explained by a concentration of (3-lg at the interface caused by the incompatibility with different biopolymers (that is more evident at higher concentrations). These phenomena would lead to an increase in the protein association in the film with the resultant increase in viscoelasticity. 

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