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  • Rapid nanoscale quantitative analysis of plant sphingolipid long-chain bases by GC-MS.

Rapid nanoscale quantitative analysis of plant sphingolipid long-chain bases by GC-MS.

Analytical and bioanalytical chemistry (2012-05-12)
Jean-Luc Cacas, Su Melser, Frédéric Domergue, Jérôme Joubès, Brice Bourdenx, Jean-Marie Schmitter, Sébastien Mongrand
ABSTRACT

In eukaryotic organisms, sphingolipids are major structural lipids of biological membranes and perform additional essential functions as signalling molecules. While long-chain bases (LCB), the common precursor to all sphingolipid classes, is represented by only one major molecular species in animals and fungi, up to nine LCB have been found in plants. In the absence of genuine plant sphingolipid references required for proper quantification, we have reinvestigated and optimized a protocol destined to the quantification of total plant LCB that relies on the use of gas chromatography-mass spectrometry (GC-MS). This rapid three-step protocol sequentially involves (1) the release of LCB from biological samples using barium hydroxide solution, (2) their oxidation into aldehydes by metaperiodate, and (3) the subsequent identification/quantification of these aldehydes by GC-MS. It is simple and reliable and enables separation of aldehydes upon their stero-specificity. It further enables the quantification of total LCB from a wide variety of samples including yeast and animal cell cultures.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Barium hydroxide, technical grade, ~95%
Avanti
D-ribo-phytosphingosine (C17 base), Avanti Research - A Croda Brand 860602P, powder
Avanti
Sphinganine (d20:0), Avanti Research - A Croda Brand 860674P, powder
Sigma-Aldrich
Barium hydroxide solution, 0.3 N
Sigma-Aldrich
D-erythro-Dihydrosphingosine, ≥98%
Avanti
Glucosylceramide (Soy), Avanti Research - A Croda Brand
Sigma-Aldrich
Barium hydroxide octahydrate, ACS reagent, ≥98%
Sigma-Aldrich
Barium hydroxide monohydrate, 98%
Sigma-Aldrich
Barium hydroxide octahydrate, ≥98%