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Journal of Liquid Chromatography and Related Technologies


A LC-MS/MS technique separated the bovine and mouse brain gangliosides monosialotetrahexosylgangliosides (GM1), disialotetrahexosylgangliosides (GD1a), trisialotetrahexosylgangliosides (GT1b) and tetrasialotetrahexosylgangliosides (GQ1b) using a phenyl-hexyl HPLC column and employing a linear methanol gradient in water, which is 0.028% in ammonium hydroxide. The gangliosides were separated according to sialic acid class, and within a particular class, gangliosides having different ceramide carbon chain lengths were also separated. All gangliosides of a particular sialic acid class eluted in characteristic retention time windows in the order of GQ1b, (earliest), GT1b, GD1a, and GM1 (latest). Within each specific retention time window for a particular ganglioside class, gangliosides were separated in the order of increasing ceramide carbon chain length. The phenyl-hexyl column separation of gangliosides is advantageous over established hydrophilic interaction and conventional reversed-phase chromatography techniques, in that the former separates gangliosides according to sialic acid class but not ceramide composition and the latter distributes all the sialic acid ganglioside classes throughout the entire chromatogram. The mechanism of separation of the ganglioside sialic acid classes is proposed to be a p-electron repulsion of negatively- charged gangliosides by the column phenyl moiety.


This work was conducted with supportfrom the Faculty and Research Development Program and theDissertation Research Award program at Cleveland State University.This work was also supported by the Neurodegenerative Diseases andAging Research Focus Area at Northeast Ohio Medical University. TheNational Science Foundation Major Research Instrumentation Grant(CHE-0923398) supported the requisition of the ABSciex QTrap 5500mass spectrometer instrument which was used in the optimizationstudies.




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