Glycosphingolipid structural analysis and glycosphingolipidomics

Sphingosines, or sphingoids, are a family of naturally occurring long-chain hydrocarbon derivatives sharing a common 1,3-dihydroxy-2-amino-backbone motif. The majority of sphingolipids, as their derivatives are collectively known, can be found in cell membranes in the form of amphiphilic conjugates, each composed of a polar head group attached to an N-acylated sphingoid, or ceramide. Glycosphingolipids (GSLs), which are the glycosides of either ceramide or myo-inositol-(1-O)-phosphoryl-(O-1)-ceramide, are a structurally and functionally diverse sphingolipid subclass; GSLs are ubiquitously distributed among all eukaryotic species and are found in some bacteria. Since GSLs are secondary metabolites, direct and comprehensive analysis (metabolomics) must be considered an essential complement to genomic and proteomic approaches for establishing the structural repertoire within an organism and deducing its possible functional roles. The glycosphingolipidome clearly comprises an important and extensive subset of both the glycome and the lipidome, but the complexities of GSL structure, biosynthesis, and function form the outlines of a considerable analytical problem, especially since their structural diversity confers by extension an enormous variability with respect to physicochemical properties. This chapter covers selected developments and applications of techniques in mass spectrometric (MS) that have contributed to GSL structural analysis and glycosphingolipidomics since 1990. Sections are included on basic characteristics of ionization and fragmentation of permethylated GSLs and of lithium-adducted nonderivatized GSLs under positive-ion electrospray ionization mass spectrometry (ESI-MS) and collision-induced mass spectrometry (CID-MS) conditions; on the analysis of sulfatides, mainly using negative-ion techniques; and on selected applications of ESI-MS and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) to emerging GSL structural, functional, and analytical issues. The latter section includes a particular focus on evolving techniques for analysis of gangliosides, GSLs containing sialic acid, as well as on characterizations of GSLs from selected nonmammalian eukaryotes, such as dipterans, nematodes, cestodes, and fungi. Additional sections focus on the issue of whether it is better to leave GSLs intact or remove the ceramide; on development and uses of thin-layer chromatography (TLC) blotting and TLC-MS techniques; and on emerging issues of high-throughput analysis, including the use of flow injection, liquid chromatography mass spectrometry (LC-MS), and capillary electrophoresis mass spectrometry (CE-MS).