| dc.description | Capillary zone electrophoresis-electrospray mass spectrometry (CZE-ESMS) combines high efficiency separation with mass spectral detection. The principal objective of this work was to develop CZE-ESMS techniques for the characterization of glycoconjugates such as glycoproteins, glycopeptides and lipooligosaccharides (LOS). The characteristics of three different interface designs were evaluated. The co-axial sheath flow interface proved to be the most flexible due to its ease of use and universal applicability. However, a sheathless interface design was developed, and provided a significant gain in sensitivity compared to the co-axial interface. The O-deacylated LOS from Hemophilus influenzae type b mutant strains were characterized by anionic CZE-ESMS though the performance of the technique was compromised by adsorption of the analytes on the capillary walls. Oligosaccharides derived from these strains were amenable to both anionic and cationic CZE-ESMS and separation efficiencies in excess of 180,000 theoretical plates were typically obtained. CZE-ESMS resolved the majority of components in these samples including thbse differing only by a single hexose residue. CZE-ESMS using dynamically coated capillaries and concentrated organic acid electrolytes was effective at resolving and characterizing the glycoforms of N-linked glycoproteins and the glycopeptides arising from their chemical and proteolytic digests. The selective identification of the glycopeptides in these digests was facilitated by the formation of diagnostic saccharide oxonium ions using collisional activation in the orifice/skimmer region of the mass spectrometer. The same technique was used successfully to separate oligosaccharide mixtures derivatized with 2-aminopyridine (PA). Enhanced sample loadings were obtained using on-line capillary isotachophoresis (CITP) prior to CZE-ESMS analysis. Resolution of the PA-derivatized oligosaccharide mixtures was also possible using this technique although the analysis times were long. In addition, problems associated with unstable electrophoretic and electrospray conditions were encountered when samples containing high concentrations of contaminants from the derivatization procedure were analyzed by CITP-CZE-ESMS. | en_US |
