J. Lipid Res.
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Originally published In Press as doi:10.1194/jlr.D600007-JLR200 on April 25, 2006

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Journal of Lipid Research, Vol. 47, 1559-1571, July 2006
Copyright © 2006 by American Society for Biochemistry and Molecular Biology

Methods

Liquid chromatography-mass spectrometry for comprehensive profiling of ceramide molecules in human hair

Yoshinori Masukawa1, Hisashi Tsujimura and Hirofumi Narita

Tochigi Research Laboratories, Kao Corporation, Ichikai, Haga, Tochigi 321-0962, Japan

Published, JLR Papers in Press, April 25, 2006.

1 To whom correspondence should be addressed. e-mail: masukawa.yoshinori{at}kao.co.jp

Ceramides (CERs) play key roles in signal transduction and cell regulation, probably during the keratinization of human hair. Current methods using mass spectrometry (MS), however, are not sufficient to allow the comprehensive analysis of CER molecules, including isobaric and isomeric CERs. Therefore, a method for the comprehensive profiling of CERs was developed. The method developed is based on reversed-phase liquid chromatography (RPLC) coupled to atmospheric pressure chemical ionization (APCI)-MS. Comprehensive identification and profiling of CERs is achieved using two sets of multimass chromatograms obtained from two channel detections that monitor both molecular-related and sphingoid-related ions under two different in-source collision-induced dissociation conditions and using retention times obtained from RPLC. The application of this method revealed that human hair contains 73 species of CER molecules, which were all corroborated by structural analysis using tandem mass spectrometry. The results further revealed that the composition is characterized by predominant molecules consisting of even carbon atom-containing saturated/unsaturated nonhydroxy or {alpha}-hydroxy fatty acids and C18 dihydrosphingosine, a minor but distinct content of isobaric/isomeric and odd chain-containing CERs. This successfully developed RPLC-APCI-MS technique allows the comprehensive profiling of CER molecules in hair for the investigation of their physicochemical and physiological roles.

Abbreviations: ADS, ceramides consisting of {alpha}-hydroxy fatty acid and dihydrosphingosine moieties; APCI, atmospheric pressure chemical ionization; AS, ceramides consisting of {alpha}-hydroxy fatty acid and sphingosine moieties; CER, ceramide; CID, collision-induced dissociation; ESI, electrospray ionization; FAM, fatty acid moiety; LC, liquid chromatography; MS, mass spectrometry; MS/MS, tandem mass spectrometry; NDS, ceramides consisting of nonhydroxy fatty acid and dihydrosphingosine moieties; NS, ceramides consisting of nonhydroxy fatty acid and sphingosine moieties; RPLC, reversed-phase liquid chromatography; RT, retention time; SPM, sphingoid moiety; TIC, total ion chromatogram

Supplementary key words fatty acid moiety • identification • isobaric • isomeric • molecular-related ion • multimass chromatogram • retention time • sphingoid moiety • sphingoid-related ion


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