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Liquid Chromatography/Mass Spectrometry for the Identification and Quantification of Rhamnolipids

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Pseudomonas Methods and Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1149))

Abstract

Rhamnolipids (RL) are surface-active glycolipids produced by Pseudomonas aeruginosa. They are always produced by this bacterium as a complex mixture of congeners, each composed of one or two rhamnose molecules linked to a dimer of 3-hydroxyfatty acids with a chain length of 8–12 carbons. Increasing interest for RL drives the need for efficient analytical methods to characterize these mixtures of molecules.

High-performance liquid chromatography (HPLC) coupled with tandem mass spectrometry (MS/MS) is a very precise and relatively high-throughput method for the identification of each congener and their quantification in bacterial cultures. Using 13C-labeled RL as internal standards can further enhance the precision of the quantification. Collision-induced dissociation (CID) experiments by MS/MS is a powerful tool for the detection and identification of structural variations in RL produced by various Pseudomonas strains or by a specific strain under different culture conditions. CID even allows the discrimination between isomers with subtle structural variations, like Rha-C8-C10 and Rha-C10-C8, which are almost inseparable chromatographically. We are presenting here the detailed protocols for HPLC/MS and HPLC/MS/MS analysis of RL and their lipid precursors, the 3-(3-hydroxyalkanoyloxy)alkanoic acids (HAA), directly in bacterial culture supernatants.

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Authors and Affiliations

  1. INRS-Institut Armand Frappier, Laval, QC, Canada, H7V 1B7

    Ahmad Mohammad Abdel-Mawgoud, François Lépine & Eric Déziel

Authors
  1. Ahmad Mohammad Abdel-Mawgoud
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  2. François Lépine
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  3. Eric Déziel
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Correspondence to Eric Déziel .

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Editors and Affiliations

  1. MRC Centre for Molecular Bacteriology and Infection (CMBI) Department of Life Sciences, Imperial College, London, UK

    Alain Filloux

  2. Estación Experimental del Zaidín, Consejo Superior de Investigaciones Científicas, Granada, Spain

    Juan-Luis Ramos

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Abdel-Mawgoud, A.M., Lépine, F., Déziel, E. (2014). Liquid Chromatography/Mass Spectrometry for the Identification and Quantification of Rhamnolipids. In: Filloux, A., Ramos, JL. (eds) Pseudomonas Methods and Protocols. Methods in Molecular Biology, vol 1149. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-0473-0_30

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  • DOI: https://doi.org/10.1007/978-1-4939-0473-0_30

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-0472-3

  • Online ISBN: 978-1-4939-0473-0

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