Bio-Molecular Thermodynamics of Protein Precipitation in Aqueous Solutions

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Experimental and theoretical bio-processing studies are directed toward the development of a molecular-thermodyamic model for separationg a target protein from a mixture of proteins in aqueuous solution. Protein separation is achieved by precipitation induced by salts, nonionic polymers, or both. The resulting system consists of two liquid phases in equilbrium, with one enriched in the target protein. A molecular-thermosynamic model is outlined for describing protein-precipitation phase equilibria. This model is derived from an understanding of the intermolecular forces in the precipitating solutions; it uses the mean-spherical- approximation to define the reference system as a mixture of charged hard spheres. Pertubations to this reference system result from the dispersion forces, dipole-dipole forces and perhaps, specific association forces. Osmotic pressures calculated from the model will be compared to osmotic-pressure data obtained from membrane osmometry and to osmotic second virial coefficients measured by low-angle laser-light scattering. Experimental phase-equilibrium (precipitation) measurements will provide data against which the molecular- thermodynamic model can be compared. Protein aggregate size distribution data from light scattering will be used to augment the theory when significant aggregation occurs. The goal of this reech is to establish an engineering-oriented correlation for the rational design of protein-precipitation processes. The long-range benefit of this research is that it is likely to provide useful knowledge for a better understanding of protein crystallization bioprocesses.