Analytical and Bioanalytical Chemistry

, Volume 407, Issue 5, pp 1403–1414 | Cite as

Comparison of sample preparation methods for the quantitative analysis of eicosanoids and other oxylipins in plasma by means of LC-MS/MS

  • Annika I. Ostermann
  • Ina Willenberg
  • Nils Helge Schebb
Research Paper


Oxylipins are potent lipid mediators. For the evaluation of their biological roles, several LC-MS based methods have been developed. While these methods are similar, the described sample preparation procedures for the extraction of oxylipins differ considerably. In order to deduce the most appropriate method for the analysis of non-esterified oxylipins in human plasma, we evaluated the performance of seven established sample preparation procedures. Six commonly used solid phase extraction (SPE) and one liquid-liquid extraction (LLE) protocol were compared based on the recovery of 13 added internal standards, extraction efficacy of oxylipins from plasma and reduction of ion-suppressing matrix. Dramatic differences in the performance in all three parameters were found. LLE with ethyl acetate was overall not a sufficient sample preparation strategy. The protocols using Oasis- and StrataX-material insufficiently removed interfering matrix compounds. Extraction efficacy of oxylipins on anion-exchanging BondElut cartridges was low, while removal of matrix was nearly perfect. None of the protocols led to a high extraction efficacy of analytes while removing all interfering matrix components. However, SPE on a C18-material with removal of matrix by water and n-hexane prior elution with methyl formate showed the best performance for the analysis of a broad spectrum of oxylipins in plasma.

Graphical Abstract

TOC art: Scheme of oxylipin extraction from plasma and ion suppression analysis of PGE2


Eicosanoids Lipidomics Liquid-liquid extraction Solid phase extraction Oxylipins LC-MS 



Arachidonic acid




Collision-induced dissociation




Docosahexaenoic acid


Dihydroxy docosapentaenoic acid


Dihydroxy eicosatetraenoic acid


Dihydroxy eicosatrienoic acid


Dihydroxy octadecadienoic acid


Dihydroxy octadecenoic acid


Ethyl acetate


Ethylenediaminetetraacetic acid


Eicosapentaenoic acid


Epoxy docosapentaenoic acid


Epoxy eicosatetraenoic acid


Epoxy eicosatrienoic acid


Epoxy octadecadienoic acid


Epoxy octadecenoic acid


Electrospray ionization


Acetic acid


Hydroxy docosahexaenoic acid


Hydroxy eicosapentaenoic acid


Hydroxy eicosatetraenoic acid


Hydroxy octadecadienoic acid


Hydroxy octadecatrienoic acid


Internal standard


Liquid chromatography mass spectrometry


Liquid-liquid extraction


Limit of detection










Oxo eicosatetraenoic acid




Polyunsaturated fatty acid


Reversed phase


Relative standard deviation


Standard deviation


Solid phase extraction


Selected reaction monitoring


Trihydroxy-ocadecenoic acid





This study was supported the Fonds der Chemischen Industrie, a Marie Curie Career Integration Grant (CIG 293536) of the European Union and a Grant of the German Research Foundation (SCHE 1801).

Supplementary material

216_2014_8377_MOESM1_ESM.pdf (1.7 mb)
ESM 1 (PDF 1.70 MB)
216_2014_8377_MOESM2_ESM.xlsx (115 kb)
ESM 2 (XLSX 115 kb)


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Copyright information

© Springer-Verlag Berlin Heidelberg 2014

Authors and Affiliations

  • Annika I. Ostermann
    • 1
  • Ina Willenberg
    • 1
  • Nils Helge Schebb
    • 1
    • 2
  1. 1.Institute for Food Toxicology and Analytical ChemistryUniversity of Veterinary Medicine HannoverHannoverGermany
  2. 2.Institute of Food ChemistryUniversity of WuppertalWuppertalGermany

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