Journal
BIOMEDICAL CHROMATOGRAPHY
Volume 23, Issue 5, Pages 516-523Publisher
WILEY
DOI: 10.1002/bmc.1147
Keywords
N-linked oligosaccharide; 1-pyrenesulfonyl chloride; ultra-performance liquid chromatography; fluorescence detection; time-of-flight mass spectrometry; ovalbumin; ribonuclease B; fetuin
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Funding
- Ministry of Education, Science, Sports and Culture of Japan
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Rapid, selective and sensitive determination of N-linked oligosaccharides in glycoproteins (ovalbumin, ribonuclease B and fetuin) was performed by ultra-performance liquid chromatography (UPLC) with fluorescence (FL) and electrospray ionization time-of-flight mass spectrometry (ESI-TOF-MS). The asparaginyl-oligosaccharide moiety was first liberated from each glycoprotein by pronase E (a proteolitic enzyme). The oligosaccharide fractions separated by gel-permeation chromatography were labeled with 1-pyrenesulfonyl chloride (PSC, a fluorescence reagent), separated by UPLC in a short run time, and then detected by FL and TOF-MS. The PSC-labeled oligosaccharides were selectively identified from the FL detection and then sensitively determined by ESI-TOF-MS. As the results, 15, eight and four kinds of N-linked oligosaccharides were detected from ovalbumin, ribonuclease B and fetuin, respectively. Because the present method is rapid (within 9 min), selective and sensitive (approximate 60 fmol, S/N = 5), the determination of N-linked oligosaccharides in various glycoproteins seems to be possible. Copyright (C) 2008 John Wiley & Sons, Ltd.
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