Sphingolipids comprise a highly diverse and complex class of molecules that serve
as both structural components of cellular membranes and signaling molecules capable
of eliciting apoptosis, differentiation, chemotaxis, and other responses in mammalian
cells. Comprehensive or "sphingolipidomic" analyses (structure specific, quantitative
analyses of all sphingolipids, or at least all members of a critical subset) are required
in order to elucidate the role(s) of sphingolipids in a given biological context because
so many of the sphingolipids in a biological system are inter-converted structurally
and metabolically. Despite the experimental challenges posed by the diversity of sphingolipid-regulated
cellular responses, the detection and quantitation of multiple sphingolipids in a
single sample has been made possible by combining classical analytical separation
techniques such as high-performance liquid chromatography (HPLC) with state-of-the-art
tandem mass spectrometry (MS/MS) techniques. As part of the Lipid MAPS consortium
an internal standard cocktail was developed that comprises the signaling metabolites
(i.e. sphingoid bases, sphingoid base-1-phosphates, ceramides, and ceramide-1-phosphates)
as well as more complex species such as mono- and di-hexosylceramides and sphingomyelin.
Additionally, the number of species that can be analyzed is growing rapidly with the
addition of fatty acyl Co-As, sulfatides, and other complex sphingolipids as more
internal standards are becoming available. The resulting LC-MS/MS analyses are one
of the most analytically rigorous technologies that can provide the necessary sensitivity,
structural specificity, and quantitative precision with high-throughput for "sphingolipidomic"
analyses in small sample quantities. This review summarizes historical and state-of-the-art
analytical techniques used for the identification, structure determination, and quantitation
of sphingolipids from free sphingoid bases through more complex sphingolipids such
as sphingomyelins, lactosylceramides, and sulfatides including those intermediates
currently considered sphingolipid "second messengers". Also discussed are some emerging
techniques and other issues remaining to be resolved for the analysis of the full
sphingolipidome.