| A general introduction and background information about mass spectrometry, structures of glycolipids from high plants, on target cleanup and detection of biological samples with solid phase extraction coupled with MALDI-MS analysis, and glycosylation of glycoproteins, are discussed in chapter 1.;There are three phases of research work in the thesis. In the first phase, chapter 2, new method development, extraction processes and separation procedures were undertaken to isolate glycophosposphingolipids from both tobacco plant tissues and suspension tobacco cultured cells. Structural characterization of glycophosposphingolipids from both tobacco plants and suspension-cultured cells were performed by gas chromatography-mass spectrometry (GC-MS), fast atom bombardment-mass spectrometry (FAB-MS), FAB-MS/MS, and nuclear magnetic resonance (NMR).;In the second phase of work, a new class of MALDI targets was developed for on-probe cleanup and detection of biological samples in the present of high concentration of impurities. Targets with surface coating through covalent linkage of poly-L-lysine with a variety of molecular weights was prepared for solid-phase extraction/matrix-assisted laser desorption ionization mass spectrometry (SPE/MALDI-MS). Targets modified in this manner have higher binding capacity towards biological molecules than targets modified by SAMs. In chapter 3, poly-L-lysine modified targets have been demonstrated to bind peptides/proteins through ionic interactions in the presence of high concentration of contaminants such as inorganic salts, buffers, detergents, chaotropic reagents by SPE/MALDI-MS. In chapter 4, the targets were successfully applied for the on-probe cleanup and detection of oligonucleotides by SPE/MALDI-MS in the presence of high concentration of inorganic salts.;In the third phase of my research, I concentrated on characterization of glycosylation of glycoproteins from a variety of biological systems.;N-linked glycosylation sites and carbohydrate structures of a recombinant human matrix metalloproteinase-9 (MAV-9), a major endogenous laccase from the lignin degrading white-rot fungus Pycnoporus cinnabarinus , and a major endogenous manganese dependent peroxidase (MnP) from monokaryotic strain 52J of the lignin degrading white-rot fungus Trametes Versicolor were described in chapter 5, 6, and 7 respectively. N-linked glycosylation sites and their oligosaccharide structures of all three glycoproteins were determined by a combination of by specific endoproteinases (trypsin, chymotrypsin, and Asp-N) digestions, microbore RP-HPLC, endo-, exoglycosidase digestions, and MALID-TOF-MS analysis. |
