Skip to main content
Log in

A simple method to identify ether lipids in spermatozoa samples by MALDI-TOF mass spectrometry

  • Research Paper
  • Published:
Analytical and Bioanalytical Chemistry Aims and scope Submit manuscript

Abstract

Plasmalogens (alkenylacyl glycerophospholipids) are important lipid constituents of many tissues and cells (e.g., selected spermatozoa). Since the molecular weights of plasmalogens overlap with that of diacyl- or alkyl acyl lipids, sophisticated mass spectrometry (MS; including MS/MS) analysis is normally used for the unequivocal identification of plasmalogens. We will show here that a simple matrix-assisted laser desorption/ionization time-of-flight mass spectrometer (without MS/MS capability) in combination with acidic hydrolysis and subsequent derivatization with 2,4-dinitrophenylhydrazine (DNPH) and/or digestion with phospholipase A2 (PLA2) is sufficient to determine the contributions of ether lipids in spermatozoa extracts. As neither diacyl nor alkylacyl lipids are sensitive to acids and do not react with DNPH, the comparison of the mass spectra before and after treatment with acids and/or DNPH addition readily provides unequivocal information about the plasmalogen content. Additionally, the released aldehydes are readily converted into the 2,4-dinitrophenylhydrazones and can be easily identified in the corresponding negative ion mass spectra. Finally, PLA2 digestion is very useful in confirming the presence of plasmalogens. The suggested method was validated by analyzing roe deer, bovine, boar, and domestic cat spermatozoa extracts and comparing the results with isolated phospholipids.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

Abbreviations

DNPH:

2,4-Dinitrophenylhydrazine

DHB:

2,5-Dihydroxybenzoic acid

GPC:

Glycerophosphorylcholine

MS:

Mass spectrometry

MALDI-TOF:

Matrix-assisted laser desorption and ionization time-of-flight

PL:

Phospholipid

PLA2 :

Phospholipase A2

References

  1. Frisardi V, Panza F, Seripa D, Farooqui T, Farooqui AA (2011) Glycerophospholipids and glycerophospholipid-derived lipid mediators: a complex meshwork in Alzheimer’s disease pathology. Prog Lipid Res 50:313–330

    Article  CAS  Google Scholar 

  2. Murakami M (2011) Lipid mediators in life science. Exp Anim 60:7–20

    Article  CAS  Google Scholar 

  3. Schug ZT, Frezza C, Galbraith LC, Gottlieb E (2012) The music of lipids: how lipid composition orchestrates cellular behaviour. Acta Oncol 51:301–310

    Article  CAS  Google Scholar 

  4. Fuchs B, Schnapka-Hille L, Schiller J, Cross MA (2011) Oxygen and cytokine-dependent changes in choline phospholipid saturation in hematopoietic progenitor cells detected by MALDI-TOF mass spectrometry. Chem Phys Lipids 164:636–642

    Article  CAS  Google Scholar 

  5. da Silva TF, Sousa VF, Malheiro AR, Brites P (2012) The importance of ether-phospholipids: a view from the perspective of mouse models. Biochim Biophys Acta 1822:1501–1508

    Article  Google Scholar 

  6. Magnusson CD, Haraldsson GG (2011) Ether lipids. Chem Phys Lipids 164:315–340

    Article  CAS  Google Scholar 

  7. Wallner S, Schmitz G (2011) Plasmalogens: the neglected regulatory and scavenging lipid species. Chem Phys Lipids 164:573–589

    Article  CAS  Google Scholar 

  8. Lessig J, Fuchs B (2009) Plasmalogens in biological systems: their role in oxidative processes in biological membranes, their contribution to pathological processes and aging and plasmalogen analysis. Curr Med Chem 16:2021–2041

    Article  CAS  Google Scholar 

  9. Nagan N, Zoeller RA (2001) Plasmalogens: biosynthesis and functions. Prog Lipid Res 40:199–229

    Article  CAS  Google Scholar 

  10. Wiesner P, Leidl K, Boettcher A, Schmitz G, Liebisch G (2012) Lipid profiling of FPLC-separated lipoprotein fractions by electrospray ionization tandem mass spectrometry. J Lipid Res 50:574–585

    Article  Google Scholar 

  11. Schiller J, Müller K, Süss R, Arnhold J, Gey C, Herrmann A, Lessig J, Arnold K, Müller P (2003) Analysis of the lipid composition of bull spermatozoa by MALDI-TOF mass spectrometry—a cautionary note. Chem Phys Lipids 126:85–94

    Article  CAS  Google Scholar 

  12. Fuchs B, Süss R, Teuber K, Eibisch M, Schiller J (2011) Lipid analysis by thin-layer chromatography—a review of the current state. J Chromatogr A 1218:2754–2774

    Article  CAS  Google Scholar 

  13. Loizides-Mangold U (2013) On the future of mass-spectrometry-based lipidomics. FEBS J 280:2817–2829

    Article  CAS  Google Scholar 

  14. Berdyshev EV (2011) Mass spectrometry of fatty aldehydes. Biochim Biophys Acta 1811:680–693

    Article  CAS  Google Scholar 

  15. Kölliker S, Oehme M, Dye C (1998) Structure elucidation of 2,4-dinitrophenylhydrazone derivatives of carbonyl compounds in ambient air by HPLC/MS and multiple MS/MS using atmospheric chemical ionization in the negative ion mode. Anal Chem 70:1979–1985

    Article  Google Scholar 

  16. Fuchs B, Süss R, Schiller J (2010) An update of MALDI-TOF mass spectrometry in lipid research. Prog Lipid Res 49:450–475

    Article  CAS  Google Scholar 

  17. Brombacher S, Owen SJ, Volmer DA (2003) Automated coupling of capillary-HPLC to matrix-assisted laser desorption/ionization mass spectrometry for the analysis of small molecules utilizing a reactive matrix. Anal Bioanal Chem 376:773–779

    Article  CAS  Google Scholar 

  18. Teuber K, Federova M, Hoffmann R, Schiller J (2012) 2,4-dinitrophenylhydrazine as a new reactive matrix to analyze oxidized phospholipids by MALDI-TOF mass spectrometry. Anal Lett 45:968–976

    Article  CAS  Google Scholar 

  19. Hui SP, Chiba H, Kurosawa T (2011) Liquid chromatography-mass spectrometric determination of plasmalogens in human plasma. Anal Bioanal Chem 400:1923–1931

    Article  CAS  Google Scholar 

  20. Zemski Berry KA, Murphy RC (2004) Electrospray ionization tandem mass spectrometry of glycerophosphoethanolamine plasmalogen phospholipids. J Am Soc Mass Spectrom 15:1499–1508

    Article  CAS  Google Scholar 

  21. Fuchs B, Müller K, Göritz F, Blottner S, Schiller J (2007) Characteristic oxidation products of choline plasmalogens are detectable in cattle and roe deer spermatozoa by MALDI-TOF mass spectrometry. Lipids 42:991–998

    Article  CAS  Google Scholar 

  22. Fuchs B, Jakop U, Göritz F, Hermes R, Hildebrandt T, Schiller J, Müller K (2009) MALDI-TOF "fingerprint" phospholipid mass spectra allow the differentiation between ruminantia and feloideae spermatozoa. Theriogenology 71:568–575

    Article  CAS  Google Scholar 

  23. Lessig J, Gey C, Süss R, Schiller J, Glander HJ, Arnhold J (2004) Analysis of the lipid composition of human and boar spermatozoa by MALDI-TOF mass spectrometry, thin layer chromatography and 31P NMR spectroscopy. Comp Biochem Physiol B 137:265–277

    Article  Google Scholar 

  24. Bligh EG, Dyer WJ (1959) A rapid method of total lipid extraction and purification. Can J Biochem Physiol 3:911–917

    Article  Google Scholar 

  25. White T, Bursten S, Federighi D, Lewis RA, Nudelman E (1998) High-resolution separation and quantification of neutral lipid and phospholipid species in mammalian cells and sera by multi-one-dimensional thin-layer chromatography. Anal Biochem 258:109–117

    Article  CAS  Google Scholar 

  26. Schiller J, Arnhold J, Benard S, Müller M, Reichl S, Arnold K (1999) Lipid analysis by matrix-assisted laser desorption and ionization mass spectrometry: a methodological approach. Anal Biochem 267:46–56

    Article  CAS  Google Scholar 

  27. Schiller J, Süss R, Fuchs B, Müller M, Petković M, Zschörnig O, Waschipky H (2007) The suitability of different DHB isomers as matrices for the MALDI-TOF MS analysis of phospholipids: which isomer for what purpose? Eur Biophys J 36:517–527

    Article  CAS  Google Scholar 

  28. Arnhold J, Osipov AN, Spalteholz H, Panasenko OM, Schiller J (2002) Formation of lysophospholipids from unsaturated phosphatidylcholines under the influence of hypochlorous acid. Biochim Biophys Acta 1572:91–100

    Article  CAS  Google Scholar 

  29. Angelini R, Vitale R, Patil VA, Cocco T, Ludwig B, Greenberg ML, Corcelli A (2012) Lipidomics of intact mitochondria by MALDI-TOF/MS. J Lipid Res 53:1417–1425

    Article  CAS  Google Scholar 

  30. Teuber K, Schiller J, Jakop U, Lüpold S, Orledge JM, Blount JD, Royle NJ, Hoodless A, Müller K (2011) MALDI-TOF mass spectrometry as a simple tool to determine the phospholipid/glycolipid composition of sperm: pheasant spermatozoa as one selected example. Anim Reprod Sci 123:270–278

    Article  CAS  Google Scholar 

  31. Sun G, Yang K, Zhao Z, Guan S, Han X, Gross RW (2008) Matrix-assisted laser desorption/ionization time-of-flight mass spectrometric analysis of cellular glycerophospholipids enabled by multiplexed solvent dependent analyte-matrix interactions. Anal Chem 80:7576–7585

    Article  CAS  Google Scholar 

  32. Fuchs B, Bresler K, Schiller J (2011) Oxidative changes of lipids monitored by MALDI MS. Chem Phys Lipids 164:782–795

    Article  CAS  Google Scholar 

Download references

Acknowledgment

This work was supported by the German Research Council (DFG Schi 476/12-1, SFB 1052/B6, Mu 1520/4-1, Fu 771/1-2, and Ni 1396/3-1.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Jürgen Schiller.

Additional information

Ariane Nimptsch and Beate Fuchs contributed equally to this work.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Nimptsch, A., Fuchs, B., Süß, R. et al. A simple method to identify ether lipids in spermatozoa samples by MALDI-TOF mass spectrometry. Anal Bioanal Chem 405, 6675–6682 (2013). https://doi.org/10.1007/s00216-013-7147-z

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00216-013-7147-z

Keywords

Navigation