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

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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.

Rationale for rapid LCB analysis by GC-MS

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Acknowledgments

We would like to warmly thank Dr. Christelle Guillier (INRA, Dijon, France) for providing alfalfa DIM samples. We are grateful to Dr. Charles Rudin and Dr. Jenni Brace (University of Chicago, Chicago, IL, USA) for sharing results about the LCB composition of yeast sur2Δ mutants. We also wish to warmly thank Dr. Eric Brenner (Université de Strasbourg, Strasbourg, France) and Karen Gaudin (Université Bordeaux 2, Bordeaux, France) for enthusiastic and stimulating discussions. We thank Dr. François-Xavier Felpin (Université Bordeaux 1, Bordeaux, France) for helping with the synthesis of long-chain fatty aldehydes. We are grateful to Dr. Patrick Moreau, Dr. René Lessire, and Dr. Jean-Jacques Besoule (CNRS, Bordeaux, France) for critical reading and stimulating discussions. We acknowledge the platforms Métabolome-Lipidome-Fluxome of Bordeaux (https://www.bordeaux.inra.fr/umr619/RMN_index.htm) for contribution to mass spectrometry equipment. This work has been funded by the French National Agency for Research, ANR program (NT09_517917, ANR PANACEA to SM and JMS). We declare no conflict of interest with any work cited in this study.

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Author notes

  1. Jean-Luc Cacas

    Present address: Centre INRA de Dijon, UMR 1347 Agroécologie, ERL6300 CNRS/INRA/Université de Bourgogne, Pole Interaction Plante-Microorganisme (IPM), 17 Rue Sully, 21000, Dijon, France

  2. Su Melser

    Present address: Laboratoire MRGM, Rare Diseases: Genetics and Metabolism, Université Bordeaux 1–2, Deuxième Etage, Ecole des Sages-Femmes, CHU Bordeaux, France

  3. Brice Bourdenx

    Present address: Fermentalg SA, 4 bis rue Rivière, 33500, Libourne, France

Authors and Affiliations

  1. Laboratoire de Biogenèse Membranaire, UMR 5200 CNRS-Université Victor Segalen Bordeaux 2, 146 rue Léo Saignat, 33076, Bordeaux, France

    Jean-Luc Cacas, Su Melser, Frédéric Domergue, Jérôme Joubès, Brice Bourdenx & Sébastien Mongrand

  2. Chimie Biologie des Membranes et Nanoobjets (CBMN)-UMR 5248, Centre de Génomique Fonctionnelle, Université de Bordeaux, BP 68, Université Bordeaux 2-Victor Segalen, 146 rue Léo Saignat, 33076, Bordeaux Cedex, France

    Jean-Marie Schmitter

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  1. Jean-Luc Cacas
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Correspondence to Jean-Luc Cacas.

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Cacas, JL., Melser, S., Domergue, F. et al. Rapid nanoscale quantitative analysis of plant sphingolipid long-chain bases by GC-MS. Anal Bioanal Chem 403, 2745–2755 (2012). https://doi.org/10.1007/s00216-012-6060-1

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  • DOI: https://doi.org/10.1007/s00216-012-6060-1

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