| Recovery and purification of proteins are of vital importance in biotechnology. Design and optimization of separation processes require an understanding of the behavior of proteins in complex aqueous solutions, where salts, polymers or other solutes may be present at high concentrations. The objective of this research is to develop a molecular-thermodynamic description of protein-protein interactions in such systems, to facilitate design and operation of protein-purification processes.;Specific attention is given to salt-induced precipitation of proteins, a frequently-used separation technique in the pharmaceutical and biotechnology industries. Protein-solution behavior is described on a molecular level, in terms of a two-body potential of mean force (PMF). The protein-protein PMF is a measure of the overall interaction between two protein molecules; it is generally determined by the physicochemical properties of the protein, the precipitating agent(s) and the solvent, by the concentrations of all solutes present, and by the temperature. Working PMF models for protein interactions in aqueous solutions are formulated over a broad range of pH and ionic strength. The DLVO potential for colloid interactions in low-ionic-strength solutions, which combines Debye-Huckel-type Coulomb repulsion with van-der-Waals dispersion attraction, is supplemented with various other contributions that are important in protein-precipitation solutions. Requisite PMF parameters are regressed from experimental data (diffusivity measurements and hydrogen-ion titration curves) gathered in dilute and semi-dilute protein solutions.;A statistical-mechanical approach has been applied to describe precipitation phase equilibrium in concentrated aqueous protein solutions. Using the Random Phase Approximation to describe the effect of protein interactions (expressed in terms of the potential of mean force) on solution structure and stability, simple analytical expressions are derived for the chemical potential of protein and for the equation of state of the protein solution. With the hypothesis that protein precipitation is a liquid-liquid equilibrium, these thermodynamic relations are used to calculate phase-equilibrium data (e.g., protein concentrations in both liquid phases) for precipitating systems. Phase equilibria are predicted for two globular proteins, hen-egg-white lysozyme and bovine $alpha$-chymotrypsin. Semi-quantitative agreement with experimental phase-equilibrium data is achieved, demonstrating the usefulness of molecular theories for describing macromolecular phase-equilibrium phenomena in protein solutions. |
