Characterization of ceramides by low energy collisional-activated dissociation tandem mass spectrometry with negative-ion electrospray ionization



Negative-ion electrospray ionization tandem quadrupole mass spectrometry provides a useful method for the structural characterization of ceramides. Fragment ions referring to the identities of the fatty acid substituent and of the long chain base of the molecules are readily available and the structure of ceramides can be easily determined. A unique fragmentation pathway which leads to formation of the fatty acid carboxylate anions (RCO2) was observed. This fragmentation is initiated by cleavage of the C2-C3 bond of the LCB to yield a N-acylaminoethanol anion ([RCONHCH2CH2O]), followed by rearrangement to a carboxyethylamine ([RCO2CH2CH2NH]) intermediate, which further dissociates to a RCO2+ ion. This pathway is confirmed by the CAD tandem mass spectrum of the synthetic N-acylaminoethanol standard and of the deuterated analogs of ceramides obtained by H-D exchange. The observation of RCO2 ion species permits an unambiguous identification of the fatty acyl moiety of ceramides. Tandem mass spectrometry methods for characterization of structural isomers of ceramides using product-ion scanning and for identification of specific ceramide subclasses in biological mixtures using neutral loss scanning are also demonstrated.


  1. 1.
    Hsu F. F.; Turk J.; Stewart M. E.; Downing D. T. Structural Studies on Ceramides as Lithiated Adducts by Low Energy Collisional-Activated Dissociation Tandem Mass Spectrometry with Electrospray Ionization. J. Am. Soc. Mass Spectrom., in press.Google Scholar
  2. 2.
    Gu, M.; Kerwin, J. L.; Watts, J. D.; Aebersold, R. Ceramide Profiling of Complex Lipid Mixtures by Electrospray Ionization Mass Spectrometry. Anal. Biochem. 1997, 244, 347–356.CrossRefGoogle Scholar
  3. 3.
    Raith, K.; Neubert, R. H. H. Liquid Chromatography-Electrospray Mass Spectrometry and Tandem Mass Spectrometry of Ceramides. Anal. Chim. Acta 2000, 403, 295–303.CrossRefGoogle Scholar
  4. 4.
    Raith, K.; Neubert, R. H. H. Structural Studies on Ceramides by Electrospray Tandem Mass Spectrometry. Rapid Commun. Mass Spectrom. 1998, 12, 935–938.CrossRefGoogle Scholar
  5. 5.
    Vietzke, J. P.; Brandt, O.; Abeck, D.; Rapp, C.; Strassner, M.; Schreiner, V.; Hintze, U. Comparative Investigation of Human Stratum Corneum Ceramides. Lipids 2001, 36, 299–304.CrossRefGoogle Scholar
  6. 6.
    Costello, C. E.; Vath, J. E. Methods in Enzymology; In: McCloskey, J. A., Ed.; Academic Press: San Diego, CA, 1990; p. 738–768 Vol. CXCIII.Google Scholar
  7. 7.
    Ann, Q.; Adams, J. Structural Determination of Ceramides and Neutral Glycosphingolipids by Collisional Activation of [M + Li]+ ions. J. Am. Soc. Mass Spectrom. 1992, 3, 260–263.CrossRefGoogle Scholar
  8. 8.
    Ann, Q.; Adams, J. Structural-Specific Collision-Induced Fragmentations of Ceramides Cationized with Alkali-Metal Ions. Anal. Chem. 1993, 65, 7–13.CrossRefGoogle Scholar
  9. 9.
    Hsu, F. F.; Bohrer, A.; Turk, J. Electrospray Ionization Tandem Mass Spectrometric Analysis of Sulfatide. Determination of Fragmentation Patterns and Characterization of Molecular Species Expressed in Brain and in Pancreatic Islets. Biochim. Biophys. Acta 1998, 1392(23), 202–216.Google Scholar
  10. 10.
    Hsu, F. F.; Turk, J. Studies on Phosphatidylglycerol with Triple Quadrupole Tandem Mass Spectrometry with Electrospray Ionization: Fragmentation Processes and Structural Characterization. J. Am. Soc. Mass Spectrom. 2001, 12, 1036–1043.CrossRefGoogle Scholar
  11. 11.
    Zhu, J.; Cole, R. B. Formation and Decompositions of Chloride Adduct Ions, [M + Cl], in Negative Ion Electrospray Ionization Mass Spectrometry. J. Am. Soc. Mass Spectrom. 2000, 11, 932–941.CrossRefGoogle Scholar
  12. 12.
    Zhu, J.; Cole, R. B. Ranking of Gas-Phase Acidities and Chloride Affinities of Monosaccharides and Linkage Specificity in Collision-Induced Decompositions of Negative Ion Electrospray-Generated Chloride Adducts of Oligosaccharides. J. Am. Soc. Mass Spectrom. 2001, 12, 1193–1204.CrossRefGoogle Scholar
  13. 13.
    Harrison, A. G. Chemical Ionization Mass Spectrometry. CRC Press: Florida, 1983 [chapter II].Google Scholar

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© American Society for Mass Spectrometry 2002

Authors and Affiliations

  1. 1.Mass Spectrometry Resource, Division of Endocrinology, Diabetes, and Metabolism, Department of MedicineWashington University School of MedicineSt. LouisUSA

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