Aqueous Two-Phase Extraction: Theory and Experiment

当两种水溶性聚合物（如PEG和葡聚糖）的水溶液混合在一起，超过临界浓度时，就会发生液-液相分离。当聚乙二醇和盐混合在一起时，也会出现类似的情况。将蛋白质或酶添加到所得到的两相混合物中的任何一种中，将倾向于在两相之间不均匀地分配，从而允许提取特定的蛋白质。在过去的三年中，PI与位于西德Julich的酶技术研究所的Kula教授合作，并在NSF资助（CBT-872084）的支持下，开发了水相两相分配的统计力学理论，并对PEG/Dx系统的相行为进行了补充实验。实验涉及相平衡图的仔细组合，并确定两相之间的界面张力作为组成的敏感指标。理论上，已经建立了聚合物-聚合物、聚合物-蛋白质、离子-离子、蛋白质-蛋白质、蛋白质-离子和聚合物-离子相互作用的简单模型，可用于计算水中的平均力势。拟议的研究建议在相同的方向上进行额外的理论工作。利用已得到的平均力势，利用积分方程技术的闭包假设，求出径向分布函数。根据径向分布函数，可以得到热工性质，确定相分离行为。这些计算将由相平衡的Gibbs Monte-Carlo计算机模拟加以补充。这些模拟将允许验证通过积分方程途径获得的平衡，并将提供一种方法来获得积分方程方法证明不充分的相平衡。将进行模拟和积分相平衡计算，以发展能够预测聚合物/聚合物和聚合物/盐混合物相平衡的理论，以及蛋白质的分离，包括多重蛋白质分配和可能的亲和配体附着在聚合物分子上的影响。蛋白质分配的补充实验也将进行。

When two aqueous solutions of water soluble polymer such as PEG and dextran are mixed together, above critical concentrations, a liquid-liquid phase separation occurs. A similar situation occurs when PEG and salt are mixed together. Proteins or enzymes added to either of the resulting two phase mixtures will tend to partition unequally between the two phases, thus allowing for the extraction of a particular protein. Over the past three years, in collaboration with Professor Kula of the Institute for Enzymetechnology in Julich, West Germany, and under the auspices of an NSF grant (CBT-872084), the PI has been developing a statistical mechanical theory for aqueous two phase partitioning, and has been undertaking complementary experiments on the phase behavior of the PEG/Dx system. The experimentation involved the careful assemblage of phase equilibria diagrams, and the identification of the interfacial tension between the two phases as a sensitive indicator of composition. Theoretically, simple models of polymer-polymer, polymer-protein, ion-ion, protein-protein, protein-ion and polymer-ion interactions have been developed which can be used to calculate potentials of mean force in water. The proposed research suggests additional theoretical work along the same lines. The mean force potentials which have been developed are to be used to obtain the radial distribution function via the closure postulation of the integral equation technique. From the radial distribution function, thermodynamic properties can be obtained, and phase separations behavior identified. These calculations will be supplemented by Gibbs Monte-Carlo computer simulations of the phase equilibria. These simulations will allow verification of the equilibria obtained by the integral equation route, and will provide a method for obtaining phase equilibria for which the integral equation approach proves inadequate. Simulations and integral phase equilibria calculations will be carried out to develop theories capable of predicting phase equilibria in polymer/polymer and polymer/salt mixtures, and the segregation g of proteins including multiple protein partitioning and possibly the effect of affinity ligands attached to the polymer molecule.. Complementary experiments on protein partitioning will also be undertaken.