DEVELOPMENT OF ENTRAPMENT COLUMNS FOR THE STUDY OF AFFINITY BASED ANALYSIS OF DRUG-PROTEIN INTERACTIONS

High-performance affinity chromatography (HPAC) is a type of liquid chromatography in which solutes are separated based on their binding to a stationary phase that is a biologically-related agent. Because of the strong and selective nature of many biological interactions, this method has already become a powerful technique for the purification and analysis of solutes that are complementary to the immobilized binding agent. Human serum albumin (HSA), the most abundant protein in the blood with concentrations of 35-50 mg/mL in serum, has interactions with many drugs, which can affect the absorption, distribution, metabolism and excretion of such agents. The overall goal of this thesis is to examine the use of on-column entrapment methods based on hydrazide-activated silica and oxidized glycogen as a capping agent for the immobilization of proteins as affinity ligands in HPAC. Although this general type of entrapment method has been previously examined reported by our group, this method still needs further optimization for its use in an on-column format and in new applications based on HPAC. For example, it is necessary to conduct studies to further increase the amount of the entrapped affinity ligand that can be obtained by using alternative types of supports. Advisor: David Hag