LC-MS/MS analysis of permethylated N-glycans facilitating isomeric characterization

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• 作者：Shiyue Zhou ; Xue Dong ; Lucas Veillon ; Yifan Huang…
• 关键词：N ; Glycan isomers ; Tandem mass spectrometry ; Permethylation ; Porous graphitic carbon column ; LC ; MS/MS
• 刊名：Analytical and Bioanalytical Chemistry
• 出版年：2017
• 出版时间：January 2017
• 年：2017
• 卷：409
• 期：2
• 页码：453-466
• 全文大小：
• 刊物类别：Chemistry and Materials Science
• 刊物主题：Analytical Chemistry; Biochemistry, general; Laboratory Medicine; Characterization and Evaluation of Materials; Food Science; Monitoring/Environmental Analysis;
• 出版者：Springer Berlin Heidelberg
• ISSN：1618-2650
• 卷排序：409

文摘

The biosynthesis of glycans is a template-free process; hence compositionally identical glycans may contain highly heterogeneous structures. Meanwhile, the functions of glycans in biological processes are significantly influenced by the glycan structure. Structural elucidation of glycans is an essential component of glycobiology. Although NMR is considered the most powerful approach for structural glycan studies, it suffers from low sensitivity and requires highly purified glycans. Although mass spectrometry (MS)-based methods have been applied in numerous glycan structure studies, there are challenges in preserving glycan structure during ionization. Permethylation is an efficient derivatization method that improves glycan structural stability. In this report, permethylated glycans are isomerically separated; thus facilitating structural analysis of a mixture of glycans by LC-MS/MS. Separation by porous graphitic carbon liquid chromatography at high temperatures in conjunction with tandem mass spectrometry (PGC-LC-MS/MS) was utilized for unequivocal characterization of glycan isomers. Glycan fucosylation sites were confidently determined by eliminating fucose rearrangement and assignment of diagnostic ions, achieved by permethylation and PGC-LC at high temperatures, respectively. Assigning monosaccharide residues to specific glycan antennae was also achieved. Galactose linkages were also distinguished from each other by CID/HCD tandem MS. This was attainable because of the different bond energies associated with monosaccharide linkages.