Advertisement

Analytical and Bioanalytical Chemistry

, Volume 408, Issue 3, pp 733–745 | Cite as

Development and validation of an HPLC-MS method for the simultaneous quantification of key oxysterols, endocannabinoids, and ceramides: variations in metabolic syndrome

  • Valentin Mutemberezi
  • Julien Masquelier
  • Owein Guillemot-Legris
  • Giulio G. Muccioli
Research Paper

Abstract

Oxysterols, ceramides, and endocannabinoids are three families of bioactive lipids suggested to be involved in obesity and metabolic syndrome. To facilitate the quantification of these potentially interconnected lipids, we have developed and validated a liquid chromatography coupled to mass spectrometry method allowing for their simultaneous quantification from tissues. Sample purification is of great importance when quantifying oxysterols due to the potential artifactual conversion of cholesterol into oxysterols. Therefore, we developed a novel solid-phase extraction procedure and demonstrated that it allowed for good recoveries of the three families of analytes without artifactual oxidation of cholesterol. The oxysterols, ceramides, and endocannabinoids and their respective internal standards were chromatographically separated by HPLC and ionized using the atmospheric pressure chemical ionization (APCI) source of an LTQ-orbitrap mass spectrometer. The repeatability and bias were within the acceptance limits for all 23 lipids of interest. The sensitivity (limit of detection (LOD) and limit of quantification (LOQ)) and specificity of the method allowed us to quantify all the analytes in the liver and adipose tissue of control and high-fat diet-fed C57BL/6 mice. We found that 16 weeks of high-fat diet strongly impacted the hepatic levels of several oxysterols, ceramides, and endocannabinoids. A partial least-squares discriminant analysis (PLS-DA) based on the variations of the hepatic levels of these 23 bioactive lipids allowed differentiating the lean mice from the obese mice.

Keywords

Ceramides Endocannabinoids Oxysterols HPLC-MS Validation Metabolic syndrome 

Abbreviations

cer

Ceramide

dhcer

Dihydroceramide

di-OHC

Dihydroxycholesterol

eCBs

Endocannabinoids

EIC

Extracted ion chromatogram

Epoxychol

Epoxycholesterol

Ketochol

Ketocholesterol

NAEs

N-Acylethanolamines

OHC

Hydroxycholesterol

OHCnone

Hydroxycholestenone

ROS

Reactive oxygen species

SPE

Solid-phase extraction

Notes

Acknowledgments

The authors thank Dr. Alhouayek for her helpful suggestions. V.M. and O.G.-L. are research fellows of the “Fonds pour la recherche dans l’industrie et l’agriculture” (FRIA, Belgium). G.G.M. is the recipient of subsidies from the Fonds Spéciaux de Recherches (FSR, Université catholique de Louvain) and from the FRS-FNRS, Belgium (grants CC 1.5.034.10, FRFC 2.4555.08, J.0160.13). The MASSMET platform is acknowledged for the use of the mass spectrometer.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no competing interests.

Supplementary material

216_2015_9150_MOESM1_ESM.pdf (670 kb)
ESM 1 (PDF 670 kb)

References

  1. 1.
    Nagao K, Yanagita T (2008) Prog Lipid Res 47:127–146CrossRefGoogle Scholar
  2. 2.
    Muccioli GG, Naslain D, Backhed F, Reigstad CS, Lambert DM, Delzenne NM, Cani PD (2010) Mol Syst Biol 6:392CrossRefGoogle Scholar
  3. 3.
    Perry RJ, Samuel VT, Petersen KF, Shulman GI (2014) Nature 510:84–91CrossRefGoogle Scholar
  4. 4.
    Asrih M, Jornayvaz FR (2015) Mol Cell EndocrinolGoogle Scholar
  5. 5.
    Gouveia-Figueira S, Nording ML (2014) Anal Chem 86:1186–1195CrossRefGoogle Scholar
  6. 6.
    Basit A, Piomelli D, Armirotti A (2015) Anal Bioanal ChemGoogle Scholar
  7. 7.
    Griffiths WJ, Crick PJ, Wang Y (2013) Biochem Pharmacol 86:3–14CrossRefGoogle Scholar
  8. 8.
    Griffiths WJ, Crick PJ, Wang Y, Ogundare M, Tuschl K, Morris AA, Bigger BW, Clayton PT, Wang Y (2013) Free Radic Biol Med 59:69–84CrossRefGoogle Scholar
  9. 9.
    Poli G, Biasi F, Leonarduzzi G (2013) Redox Biol 1:125–130CrossRefGoogle Scholar
  10. 10.
    Murdolo G, Bartolini D, Tortoioli C, Piroddi M, Iuliano L, Galli F (2013) Free Radic Biol Med 65:811–820CrossRefGoogle Scholar
  11. 11.
    Larsen PJ, Tennagels N (2014) Mol Metab 3:252–260CrossRefGoogle Scholar
  12. 12.
    Bjorkhem I, Diczfalusy U (2002) Arterioscler Thromb Vasc Biol 22:734–742CrossRefGoogle Scholar
  13. 13.
    Iuliano L (2011) Chem Phys Lipids 164:457–468CrossRefGoogle Scholar
  14. 14.
    Bodin K, Andersson U, Rystedt E, Ellis E, Norlin M, Pikuleva I, Eggertsen G, Bjorkhem I, Diczfalusy U (2002) J Biol Chem 277:31534–31540CrossRefGoogle Scholar
  15. 15.
    Olsen BN, Schlesinger PH, Ory DS, Baker NA (2012) Biochim Biophys Acta 1818:330–336CrossRefGoogle Scholar
  16. 16.
    Olkkonen VM, Beaslas O, Nissila E (2012) Biomolecules 2:76–103CrossRefGoogle Scholar
  17. 17.
    Olkkonen VM, Li S (2013) Prog Lipid Res 52:529–538CrossRefGoogle Scholar
  18. 18.
    Tremblay-Franco M, Zerbinati C, Pacelli A, Palmaccio G, Lubrano C, Ducheix S, Guillou H, and Iuliano L (2015) SteroidsGoogle Scholar
  19. 19.
    Wooten JS, Wu H, Raya J, Perrard XD, Gaubatz J, Hoogeveen RC (2014) Cholesterol 2014:843468CrossRefGoogle Scholar
  20. 20.
    Castro BM, Prieto M, Silva LC (2014) Prog Lipid Res 54:53–67CrossRefGoogle Scholar
  21. 21.
    Maceyka M, Spiegel S (2014) Nature 510:58–67CrossRefGoogle Scholar
  22. 22.
    Cinar R, Godlewski G, Liu J, Tam J, Jourdan T, Mukhopadhyay B, Harvey-White J, Kunos G (2014) Hepatology 59:143–153CrossRefGoogle Scholar
  23. 23.
    Hanamatsu H, Ohnishi S, Sakai S, Yuyama K, Mitsutake S, Takeda H, Hashino S, Igarashi Y (2014) Nutr Diabetes 4, e141CrossRefGoogle Scholar
  24. 24.
    Turpin SM, Nicholls HT, Willmes DM, Mourier A, Brodesser S, Wunderlich CM, Mauer J, Xu E, Hammerschmidt P, Bronneke HS, Trifunovic A, LoSasso G, Wunderlich FT, Kornfeld JW, Bluher M, Kronke M, Bruning JC (2014) Cell Metab 20:678–686CrossRefGoogle Scholar
  25. 25.
    Raichur S, Wang ST, Chan PW, Li Y, Ching J, Chaurasia B, Dogra S, Ohman MK, Takeda K, Sugii S, Pewzner-Jung Y, Futerman AH, Summers SA (2014) Cell Metab 20:687–695CrossRefGoogle Scholar
  26. 26.
    Alhouayek M, Muccioli GG (2014) Drug Discov Today 19:1632–1639CrossRefGoogle Scholar
  27. 27.
    Silvestri C, Di Marzo V (2013) Cell Metab 17:475–490CrossRefGoogle Scholar
  28. 28.
    Osei-Hyiaman D, DePetrillo M, Pacher P, Liu J, Radaeva S, Batkai S, Harvey-White J, Mackie K, Offertaler L, Wang L, Kunos G (2005) J Clin Invest 115:1298–1305CrossRefGoogle Scholar
  29. 29.
    Matias I, Gonthier MP, Orlando P, Martiadis V, De Petrocellis L, Cervino C, Petrosino S, Hoareau L, Festy F, Pasquali R, Roche R, Maj M, Pagotto U, Monteleone P, Di Marzo V (2006) J Clin Endocrinol Metab 91:3171–3180CrossRefGoogle Scholar
  30. 30.
    Cristino L, Becker T, Di Marzo V (2014) Biofactors 40:389–397CrossRefGoogle Scholar
  31. 31.
    Kajiwara K, Ikeda A, Aguilera-Romero A, Castillon GA, Kagiwada S, Hanada K, Riezman H, Muniz M, Funato K (2014) J Cell Sci 127:376–387CrossRefGoogle Scholar
  32. 32.
    Schiffmann S, Birod K, Mannich J, Eberle M, Wegner MS, Wanger R, Hartmann D, Ferreiros N, Geisslinger G, Grosch S (2013) Int J Biochem Cell Biol 45:1886–1894CrossRefGoogle Scholar
  33. 33.
    Meljon A, Theofilopoulos S, Shackleton CH, Watson GL, Javitt NB, Knolker HJ, Saini R, Arenas E, Wang Y, Griffiths WJ (2012) J Lipid Res 53:2469–2483CrossRefGoogle Scholar
  34. 34.
    Meljon A, Wang Y, Griffiths WJ (2014) Biochem Biophys Res Commun 446:768–774CrossRefGoogle Scholar
  35. 35.
    Maroof N, Ravipati S, Pardon MC, Barrett DA, Kendall DA (2014) J Alzheimers Dis 42:227–245Google Scholar
  36. 36.
    Muccioli GG, Stella N (2008) Anal Biochem 373:220–228CrossRefGoogle Scholar
  37. 37.
    AOAC OFFICIAL METHODS OF ANALYSIS. Guidelines for standard method performance requirements, appendix F. 1–17. 2012. AOAC INTERNATIONAL. Ref Type: Online SourceGoogle Scholar
  38. 38.
    Sonawane LV, Poul BN, Usnale SV, Waghmare PV, Surwase LH (2014) Pharm Anal Acta 5:1–7CrossRefGoogle Scholar
  39. 39.
    Cummings J, Raynaud F, Jones L, Sugar R, Dive C (2010) Br J Cancer 103:1313–1317CrossRefGoogle Scholar
  40. 40.
    Lee JW, Devanarayan V, Barrett YC, Weiner R, Allinson J, Fountain S, Keller S, Weinryb I, Green M, Duan L, Rogers JA, Millham R, O’Brien PJ, Sailstad J, Khan M, Ray C, Wagner JA (2006) Pharm Res 23:312–328CrossRefGoogle Scholar
  41. 41.
    Hoffman D, Kringle R (2007) Pharm Res 24:1157–1164CrossRefGoogle Scholar
  42. 42.
    Shabir GA (2003) J Chromatogr A 987:57–66CrossRefGoogle Scholar
  43. 43.
    Triba MN, Le ML, Amathieu R, Goossens C, Bouchemal N, Nahon P, Rutledge DN, Savarin P (2015) Mol Biosyst 11:13–19CrossRefGoogle Scholar
  44. 44.
    Karuna R, Christen I, Sailer AW, Bitsch F, Zhang J (2015) Steroids 99:131–138CrossRefGoogle Scholar
  45. 45.
    Narayanaswamy R, Iyer V, Khare P, Bodziak ML, Badgett D, Zivadinov R, Weinstock-Guttman B, Rideout TC, Ramanathan M, Browne RW (2015) PLoS One 10, e0123771CrossRefGoogle Scholar
  46. 46.
    Razzazi-Fazeli E, Kleineisen S, Luf W (2000) J Chromatogr A 896:321–334CrossRefGoogle Scholar
  47. 47.
    Georgiou CA, Constantinou MS, Kapnissi-Christodoulou CP (2014) Food Chem 145:918–926CrossRefGoogle Scholar
  48. 48.
    Helmschrodt C, Becker S, Thiery J, Ceglarek U (2014) Biochem Biophys Res Commun 446:726–730CrossRefGoogle Scholar
  49. 49.
    Miyoshi N, Iuliano L, Tomono S, Ohshima H (2014) Biochem Biophys Res Commun 446:702–708CrossRefGoogle Scholar
  50. 50.
    Cyster JG, Dang EV, Reboldi A, Yi T (2014) Nat Rev Immunol 14:731–743CrossRefGoogle Scholar
  51. 51.
    Balvers MG, Wortelboer HM, Witkamp RF, Verhoeckx KC, Basit A, Piomelli D, Armirotti A (2013) Anal Biochem 434:275–283CrossRefGoogle Scholar
  52. 52.
    Amano SU, Cohen JL, Vangala P, Tencerova M, Nicoloro SM, Yawe JC, Shen Y, Czech MP, Aouadi M (2014) Cell Metab 19:162–171CrossRefGoogle Scholar
  53. 53.
    Teodoro JS, Varela AT, Rolo AP, Palmeira CM (2014) Genes Nutr 9:406CrossRefGoogle Scholar
  54. 54.
    de Aguiar Vallim TQ, Tarling EJ, Edwards PA (2013) Cell Metab 17:657–669CrossRefGoogle Scholar
  55. 55.
    Engeli S, Lehmann AC, Kaminski J, Haas V, Janke J, Zoerner AA, Luft FC, Tsikas D, Jordan J (2014) Obesity (Silver Spring) 22:E70–E76CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  • Valentin Mutemberezi
    • 1
  • Julien Masquelier
    • 1
  • Owein Guillemot-Legris
    • 1
  • Giulio G. Muccioli
    • 1
  1. 1.Bioanalysis and Pharmacology of Bioactive Lipids Research Group, Louvain Drug Research InstituteUniversité catholique de LouvainBrusselsBelgium

Personalised recommendations