Binding of water and solute to protein-mixture and protein-coated alumina.

Extent of water vapour adsorption (n1) of gelatin and bovine serum albumin and their mixtures in different proportion respectively has been measured by isopiestic vapour pressure methods at various values of water activity (a1) ranging between zero and unity. Similar measurements have also been carried out with gelatin and BSA coated alumina powder. At a given value of a1, n1 for the protein mixture is found to be significantly less than their ideal value obtained from the additivity rule. Such decrease is probably due to the protein-protein interaction as a result of which some of the water binding sites become unavailable for water vapour adsorption. On the other hand when a protein is mixed with alumina powder, the water vapour adsorption of the protein coated alumina surface at a given water activity is found to be 2 to 3 times larger than its ideal value obtained from the additivity rule. The standard free energy changes for hydration of protein mixtures and protein-coated alumina have been evaluated using Bull equation. The extent of excess hydration of these proteins and their mixtures as well as protein-coated alumina in the presence of excess neutral salts and urea respectively have been evaluated using the isopiestic method. In all cases, the moles of water and solute respectively bound in absolute amount to biopolymers, biopolymer mixtures and protein-coated alumina have been evaluated in the limited range of solute concentrations in the medium. Based on the Gibbs-Duhem equations, a rigorous expression for the standard free energy change for binding of excess solute and solvent to biopolymer have been evaluated with reference to unit solute mole fraction as standard state. Free energies of excess hydration of different biopolymer systems have been evaluated using this equation.

Protein adsorption at solid-liquid interfaces: Part IV--Effects of different solid-liquid systems and various neutral salts.

Adsorption isotherms of BSA at the solid-water interfaces have been studied as a function of protein concentration, ionic strength of the medium, pH and temperature using silica, barium sulphate, carbon, alumina, chromium, ion-exchange resins and sephadex as solid interfaces. In most cases, isotherms for adsorption of BSA attained the state of adsorption saturation. In the presence of barium sulphate, carbon and alumina, two types in the isotherms are observed. Adsorption of BSA is affected by change in pH, ionic strength and temperature of the medium. In the presence of metallic chromium, adsorbed BSA molecules are either denatured or negatively adsorbed at the metallic interface. Due to the presence of pores in ion-exchange resins, adsorption of BSA is followed by preferential hydration on resin surfaces in some cases. Sometimes two steps of isotherms are also observed during adsorption of BSA on the solid resins in chloride form. Adsorption of BSA, beta-lactoglobulin, gelatin, myosin and lysozyme is negative on Sephadex surface due to the excess adsorption of water by Sephadex. The negative adsorption is significantly affected in the presence of CaCl2, KSCN, LiCl, Na2SO4, NaI, KCl and urea. The values of absolute amounts of water and protein, simultaneously adsorbed on the surface of different solids, have been evaluated in some cases on critical thermodynamic analysis. The standard free energies (delta G0) of excess positive and negative adsorption of the protein per square meter at the state of monolayer saturation have been calculated using proposed universal scale of thermodynamics. The free energy of adsorption with reference to this state is shown to be strictly comparable to each other. The magnitude of standard free energy of transfer (delta G0B) of one mole of protein or a protein mixture at any type of physiochemical condition and at any type of surface is observed to be 38.5 kJ/mole.

Protein adsorption at solid-liquid interfaces: Part III--Adsorption from ternary protein mixture.

Simultaneous adsorption of bovine serum albumin (BSA), beta-lactoglobulin and gelatin from aqueous solutions of their ternary mixture to the alumina-water interface has been studied as a function of protein concentration at different values of pH, ionic strength, temperature and weight fraction ratios of proteins. At a fixed weight fraction of beta-lactoglobulin, preferential adsorption (gamma w(lac)) of this protein significantly depends on the amounts of BSA and gelatin present in the solution before adsorption. At higher ranges of protein concentrations, extent of adsorption (gamma w(ser)) of BSA decreases sharply with increase of gamma w(lac) until gamma w(ser) becomes significantly negative, thereby indicating that beta-lactoglobulin and water preferentially adsorbed at the interface are responsible for complete displacement of BSA from the surface. On the other hand, adsorption (gamma w(gel)) of gelatin under similar situation increases mutually with increase in the values of gamma w(lac) in many systems. In few systems, gamma w(gel) also decreases with increase of gamma w(lac) depending upon solution parameters. At pH 5.2, increase of ionic strength and temperature, respectively, increases the extent of adsorption of each protein in the mixture considerably. Extents of adsorption of all proteins are observed to increase when pH is changed from 5.2 to 6.4. The affinities of different proteins in the mixture are expressed in unified scales either in terms of maximum extents of total adsorption or in terms of standard free energies of adsorption of protein mixtures with respect to surface saturation.

Protein adsorption at solid-liquid interfaces: Part II--Adsorption from binary protein mixture.

Extent of adsorption of proteins at alumina-water interface from solutions containing binary mixture of beta-lactoglobulin and bovine serum albumin (BSA), beta-lactoglobulin and gelatin, and gelatin and bovine serum albumin has been estimated as functions of protein concentrations at varying pH, ionic strength, temperature and weight fraction ratios of protein mixture. The extent of adsorption (gamma lacw) of lactoglobulin in the presence of BSA increases with increase of protein concentration (Clac) until it reaches a maximum but a fixed value gamma lacw(m). Extent of adsorption gamma serw also initially increases with increase of protein concentrations until it reaches maximum value gamma serw(m). Beyond these protein concentrations, adsorbed BSA is gradually desorbed due to the preferential adsorption of lactoglobulin from the protein mixture. In many systems, gamma serw at high protein concentrations even becomes negative due to the strong competition of BSA and water for binding to the surface sites in the presence of lactoglobulin. For lactoglobulin-gelatin mixtures, adsorption of both proteins is enhanced as protein concentration is increased until limiting values for adsorption are reached. Beyond the limiting value, lactoglobulin is further accumulated at the interface without limit when protein concentration is high. For gelatin-albumin mixtures, extent of gelatin adsorption increases with increase in the adsorption of BSA. The limit for saturation of adsorption for gelatin is not reached for many systems. At acid pH, adsorbed BSA appears to be desorbed from the surface in the presence of gelatin. From the results thus obtained the role of electrostatic and hydrophobic effects in controlling the adsorption process has been analysed.

Protein adsorption at solid-liquid interfaces: Part I--Affinities of proteins for alumina surface.

Extent of adsorption (gamma pw) of bovine serum albumin, beta-lactoglobulin, gelatin and myosin at the alumina-water interface has been measured as function of protein concentration (Cp) at several temperatures, pH, and ionic strengths of the medium. gamma pw for proteins in most cases increases with increase of protein concentration but it attains maximum value gamma pw(m) when Cp is high. Values of maximum adsorption have been examined in terms of molecular orientation, molecular size and shape and unfolding of the packed proteins at the interface. In few cases, gamma pw increases with increase of Cp without reaching a real state of saturation as a result of aggregation of molecules or extensive unfolding of the protein at the interface. In the case of beta-lactoglobulin at pH 5.2 and ionic strength 0.05, gamma pw in high concentration region decreases to zero value when Cp increases. For myosin at 45 degrees C and pH 6.4, and also at 27 degrees and pH 7.8, the values of gamma pw are all negative and these negative values increase with increase of Cp. All these results have been explained in terms of significant competitions of water and protein for binding to the surface sites of the powdered alumina. Adsorption of myosin has also been found to be affected in the presence of NaCl, KCl, CaCl2, KI, Na2SO4, LiCl and urea. The relative affinities of the adsorption of various proteins for the surface of alumina at different physical conditions of the system have been compared in terms of maximum values of adsorption attained when gamma pw is varied with Cp. The affinities are shown to be compared more precisely in terms of the standard free energy decrease for the saturation of the surface by protein as a result of the change in its concentration from zero to unity in the mole fraction scale.