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Glycobiology Advance Access originally published online on September 30, 2009
Glycobiology 2010 20(1):118-126; doi:10.1093/glycob/cwp154
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© The Author 2009. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oxfordjournals.org

Separation and identification of GM1b pathway Neu5Ac- and Neu5Gc gangliosides by on-line nanoHPLC-QToF MS and tandem MS: toward glycolipidomics screening of animal cell lines

Mostafa Zarei2,3, Johannes Müthing4, Jasna Peter-Katalinic3 and Laura Bindila1,3

3 Institute for Medical Physics and Biophysics, Robert-Koch-Str. 31
4 Institute for Hygiene, University of Münster, Robert-Koch-Str. 41, D-48149 Germany

1 To whom correspondence should be addressed: Tel: +49-2518355127; Fax: +49-2518355121; E-mail: bindila{at}uni-muenster.de

Received on April 20, 2009; revised on September 21, 2009; accepted on September 24, 2009

Monosialoganglioside fraction of YAC-1 lymphoma cells was comprehensively analyzed and structurally defined by nano-high-performance liquid chromatography (nanoHPLC) in on-line conjunction with electrospray ionization quadrupole time-of-flight mass spectrometry (ESI-QTOF MS). An efficient separation and sensitive detection of Neu5Gc-containing gangliosides from Neu5Ac-containing analogues was for the first time accomplished in a single nanoHPLC/ESI-QTOF MS run, as demonstrated for mouse hybridoma cell GM3 fraction containing GM3(Neu5Ac) and GM3(Neu5Gc) species and further applied for the analysis of YAC-1 lymphoma cell monosialoganglioside fraction. New insights into YAC-1 monosialoganglioside mixture heterogeneity were obtained: 31 distinct species, comprising 18 Neu5Gc-containing gangliosides and 13 Neu5Ac-containing species of GM1b and GalNAc-GM1b type were found to be expressed by YAC-1 cell line. On-line structural elucidation of individually separated Neu5Ac- and Neu5Gc-containing gangliosides provided strong evidence on the "GM1b-pathway" sourcing for monosialoganglioside synthesis. Such an analytical method is documented as superior to the classical approaches by increased speed of analysis, sensitivity and level of information, being thus a viable glycolipidomic tool.

Key words: YAC-1 lymphoma cells gangliosides / Neu5Ac- and Neu5Gc-containing gangliosides / separation of sialic acids / nanoHPLC / MS of gangliosides; glycosphingolipids

2 M.Z is presently affiliated to Freiburg Institute for Advanced Studies School of Soft Matter Research, Albertstr. 19 79104 Freiburg im Breisgau


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