Effect of protein thermo aggregation on the binding of BSA to gelatin type A.

We study the effect of limited heat-induced aggregation of BSA on structure development in the water-gelatin-thermally aggregated BSA (BSA(TA)) system. The pH is set at 5.4 and the temperature is higher than the conformation transition temperature of gelatin, but lower than the denaturation temperature of BSA. Dynamic light scattering, circular dichroism, and fluorescence measurements are used to monitor structure changes. Interaction of gelatin with BSA(TA) leads to formation of large complex particles with an average radius ∼1500 nm. BSA-gelatin complex formation accompanies partial destabilization of the secondary and tertiary structures of BSA and an additional exposure of hydrophobic tryptophan residues on the surface of the globule. It is shown that electrostatic interaction of the oppositely charged groups of BSA(TA) and gelatin is responsible for formation of such complex particles, whereas the secondary forces (hydrophobic interaction and hydrogen bonds) play an important role in stabilization of the complex particles. The zeta potentials of the native and the thermally aggregated BSA samples were determined, and the solvent quality has been quantified by determining the activity of the protein samples in their saturated solutions. It was shown that steric reasons (large size of the thermally aggregated BSA(TA) particles), and uncomplete charge compensation of the positively charged gelatin molecules by the negatively charged BSA(TA) particles are the main factors in determining structure formation, while the levels of the activity of the native BSA and BSA(TA) have a smaller effect on the structure of complex.

Inducing demixing of semidilute and highly compatible biopolymer mixtures in the presence of a strong polyelectrolyte.

The weak intermacromolecular interactions caused by the presence of a complexing agent in a two-phase biopolymer mixture can affect its phase equilibrium and morphology. In this communication, the attempt was made to induce demixing in semidilute and highly compatible sodium caseinate/sodium alginate system (SC-SA) mixtures in the presence of a sodium salt of dextran sulfate (DSS) at pH 7.0 (above the isoelectrical point of caseins), and to characterize phase equilibrium, intermacromolecular interactions, and structure of such systems by rheo-small angle light scattering (SALS), optical microscopy (OM), phase analysis, dynamic light scattering (DLS), fast protein liquid chromatography (FPLC), ESEM, and rheology. Addition of dextran sulfate sodium salt (DSS) to the semidilute single-phase SC-SA system, even in trace concentrations (10(-3) wt %), leads to segregative liquid-liquid phase separation and a substantial increase in storage and loss moduli of the system. The degree of the protein conversion in the complex grows when the concentration of SC in the system increases from 1 to 2 wt %. It is also established here that demixing of semidilute biopolymer mixtures, induced by the minor presence of DSS, is a rather common phenomenon, because it also was observed here for other biopolymer pairs. At high shear rates SC becomes even less compatible with SA in the presence of DSS than at rest. Experimental observations suggest that the approach for inducing demixing of semidilute and highly compatible biopolymer mixtures by physical interactions of the constituents is a promising tool for regulation of biopolymer compatibility and achieving better predictions of phase behavior of aqueous protein-charged polysaccharide systems.

Interfacial tension of aqueous biopolymer mixtures close to the critical point.

Proteins and polysaccharides, being the main constructional materials in many biological structures, have a limited compatibility in aqueous media. At sufficiently high concentrations, they form water-in-water emulsions. Interfacial tension is an important parameter in such systems since it is a controlling factor in the morphology development during processing. In this work a rheo-optical methodology, based on the analysis of small angle light scattering (SALS) patterns during fibril break-up, is used to study the interfacial tension of water-sodium caseinate-sodium alginate systems located close to and relatively far from the binodal. The interfacial tension close to the critical point was approximately 10(-8) N/m, and it increased considerably, to a value of up to 5.2 x 10(-6) N/m farther from the critical point. For the scaling of the interfacial tension with the density difference between the phases, a scaling exponent of 3.1 +/- 0.3 was found, in agreement with the critical mean-field scaling exponent of 3.

Effect of shear flow on the phase behavior of an aqueous gelatin-dextran emulsion.

A rheo-optical methodology, based on small angle light scattering and transmitted light intensity measurements, has been used to study in situ and on a time resolved basis the shear induced morphology in ternary two-phase water-gelatin-dextran mixtures. Emulsions close to the binodal line as well as far from it have been investigated. It is shown that above a critical shear rate, shear-induced mixing occurs at the length scales probed by the laser light. It is hypothesized that the shear-induced homogenization is due to the shear forces that exceed the intermolecular forces of the self-association process of the gelatin. The isothermal phase diagram at a fixed shear rate has been determined. In addition, the structure evolution after cessation of flow has been studied. When flow is stopped after homogenization, phase separation occurs almost instantaneously. When subsequently applying a low shear rate, the structure coarsens due to coalescence of the dispersed droplets. The kinetics of this coalescence process is strain controlled.

Phase state of aqueous gelatin-polysaccharide (1)-polysaccharide (2) systems.

Optical microscopy, ultracentrifugation, phase analysis and turbidimetric titration methods were used to study phase state and phase equilibria of quaternary water-gelatin-pectin-dextran system in the absence of salts and at pH higher than the isoionic point. It was found that these systems are two-phase ones, contrary to the single-phase behaviour of the ternary water-gelatin-pectin and water-gelatin-dextran systems under the same conditions. The observed phase separation is the result of incompatibility of gelatin with pectin, dextran molecules being distributed practically uniformly between coexisting phases. This phenomenon is rather typical for many water-gelatin-polysaccharide-1-polysaccharide-2 systems under the conditions when all the pairs of biopolymers are compatible. The high compatibility of gelatin with pectin or dextran in the ternary systems under given conditions is due to the formation of weakly bonded interpolymer complexes. The incompatibility of gelatin with pectin in the presence of dextran can be explained by the blockage of the reactive gelatin groups due to their competitive interactions with dextran.

Phase equilibria and mechanical properties of gel-like water-gelatin-locust bean gum systems.

The establishment of phase equilibrium in aqueous gelatin-locust bean gum (LBG) systems in the process of cooling from 313 to 291 K in specific conditions, passes ahead of the gelation process(.) This allows the suggestion that macrostructure and mechanical properties of the system can be predicted on the basis of knowledge of its phase diagram, obtained for the liquid gelatin-LBG systems comprising gelatin molecular aggregates. Textural and rheological analysis of gel-like gelatin-LBG systems demonstrate the effect of two factors determining their mechanical properties and acting opposite each other when the concentration of LBG in the system increases: concentration of gelatin by LBG in the process of phase separation, and decrease in the density of the gel network.