Development of a new polysaccharide-based chiral stationary phase for high-performance liquid chromatography

In the summer of 1992 the Food and Drug Administration (FDA) released guidelines for the marketing of chiral drugs. While pharmaceutical manufacturers retain the authority to decide whether to develop an asymmetric or enantiomerically pure product, the approval of a racemic product now requires a manufacturer to provide rigorous details on the safety of the chiral mixture. These new requirements have given chiral resolution and the determination of enantiomeric content a legal importance in addition to their well-established biological importance.; The primary means of analyzing chiral drugs is by High Performance Liquid Chromatography (HPLC) utilizing chiral stationary phases (CSP). All known CSPs can be classified into five groups based on their mechanism of separation. The separation mechanism of one class, the helical polymers (Type III), has been extensively studied but not clearly established. The commercial Type III CSPs are coated phases subject to solvent and pressure limitations, and the selection of the proper stationary phase for the separation of a given chiral compound must be made by trial and error. The direct bonding of a suitable chiral polymer would alleviate the solvent and pressure restrictions and may clarify some of the questions regarding the mechanism of separation.; The objectives of this research project are to develop a method for directly bonding hyaluronic acid to a modified silica gel matrix and to determine the utility of the newly bonded phase for resolving chiral drug mixtures. Hyaluronic acid is a good candidate for direct bonding because it possesses native functional groups not available in the chiral polymers of the coated phases.; The bonded cyclodextrin stationary phases were developed by first demonstrating the chiral recognition ability of cyclodextrin as a mobile phase additive, and the same course was followed in this work. Initial studies focused on developing new methods for resolving the chiral test compounds using {dollar}beta{dollar}-cyclodextrin as a mobile phase additive. Each chiral compound was successfully separated, and subsequent experiments substituted D-glucuronic acid, N-acetylglucosamine, and hyaluronic acid oligosaccharides for the {dollar}beta{dollar}-cyclodextrin and the resolving power of each was determined.; The first step in bonding hyaluronic acid to modified silica gel was to determine the optimal conditions for the silanization of the silica. Experiments were carried out to establish the optimal silane concentration and the effects of using multi-ligand silylating agents. Following silanization, attempts were made at bonding glucuronic acid and hyaluronic acid oligosaccharides to the modified silica gels were made using a water soluble carbodiimide and N-hydroxysuccinimide as crosslinking agents.