Characterization of N-linked glycans by supercritical fluid chromatography-mass spectrometry.
N-Linked glycans have been characterized by supercritical fluid chromatography (SFC) and SFC-MS using positive- and negative-ion chemical ionization. Four common oligosaccharide derivatives have been prepared and their chromatographic properties assessed on three SFC columns of varying polarity. Carbon dioxide has been used as the SFC mobile phase, with ammonia or CO2 added to the ion source for positive- and negative-ion chemical ionization, respectively. Direct SFC-MS interfacing allows the analytical manipulations of single-ion monitoring, total-ion plots, background subtraction, library searches, and spectral reconstruction algorithms. Positive ammonia chemical ionization yields abundant molecular-weight information, (MH)+, and (MNH4)+ with little or no fragmentation. To capitalize on sensitivity, samples were prepared with the pentafluorobenzyl aminobenzoate reagent, acetylated, and analyzed by SFC-NICI-MS. This modification improves column efficiency and resolution and greatly enhances detecting sensitivity. These "soft" ionization conditions provide abundant molecular-weight-related anions for collision-induced dissociation and subpicogram detection.
Sheeley
D M
DM
Department of Nutrition, Harvard School of Public Health, Boston, Massachusetts 02115.
Reinhold
V N
VN
D000818:Animals; D002240:Carbohydrate Sequence; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods; D037102:Lectins / Q000737:chemistry; D008358:Mannose / Q000737:chemistry; D008361:Mannosidases / Q000037:antagonists & inhibitors; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry*; D037121:Plant Lectins; D011134:Polysaccharides / Q000737:chemistry* / Q000302:isolation & purification; D012756:Sheep; D030262:Soybean Proteins; D010835:Phytohemagglutinins
D037102:Lectins; D008358:Mannose; D008361:Mannosidases; D009844:Oligosaccharides; D037121:Plant Lectins; D011134:Polysaccharides; D030262:Soybean Proteins; C028824:soybean lectin; D010835:Phytohemagglutinins