Optimization of preparative LC-MS method for fractionation of oligosaccharides of hyaluronan

Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biophysic and Physical Chemistry Candidate: Bc. Martina Dvořáková Supervisor: Doc. Ing. Alice Lázníčková, CSc. Consultant: Mgr. Martina Hermannová, Ph.D. Title of diploma thesis: Optimization of preparative LC-MS method for fractionation of oligosaccharides of hyaluronanu This diploma thesis deals with optimization of LC-MS method for analysis of hyaluronan oligosaccharides in preparative mode. The theoretical part summarizes available information about biological and chemical properties of hyaluronic acid. Hyaluronic acid is easily enzymatically degradable by mammalian hyaluronidases that produce hyaluronan oligosaccharides. The biological function of these degradation products depend on their molecular weight. High-performance liquid chromatography is mainly used for separation and purification of hyaluronan oligosaccharides. A new method for the determination of hyaluronan oligosaccharides is based on a combination of separation techniques and mass spectrometry. The experimental part deals with optimization of ionisation conditions for electrospray ionization mass spectrometry in positive and negative ion mode. In the first step, we focused on setting of capillary voltage, cone voltage, desolvation temperature, flow..