An On-Line Platform for the Analysis of Permethylated Oligosaccharides Using Capillary Liquid Chromatography-Electrospray Ionization-Tandem Mass Spectrometry.

Permethylation is one of the most practical modifications for mass spectrometric analysis of carbohydrates. In this study, we showed a facile platform for the on-line permethylation and analysis of acidic N-glycans using capillary liquid chromatography-electrospray ionization-tandem mass spectrometry to elucidate the N-glycan structures derived from glycoprotein samples. Two capillaries, individually packed with NaOH and C18 powder, were coupled to electrospray ion trap mass spectrometer. Prior to mass spectrometry, oligosaccharides were permethylated in NaOH-packed capillary by methyl iodide, and the permethylated glycans were consequently extracted from residual reagents, such as NaOH and methyl iodide, in C18-capillary, instead of using conventional chloroform extraction. After permethylation and purification, the permethylated glycans were released from C18 capillary by acetonitrile, and sprayed into mass spectrometer. Using this on-line strategy, a picomolar range of sialylated N-glycans from bovine fetuin were analyzed by tandem mass spectrometry.