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Updated: Mar 7, 2026

Cellular Lipid Extraction for Targeted Stable Isotope Dilution Liquid Chromatography-Mass Spectrometry Analysis
Published on: November 17, 2011
Comparison of derivatization/ionization techniques for liquid chromatography tandem mass spectrometry analysis of
Sven W Meckelmann1, Stefan Hellhake2, Maryvonne Steuck2
1Chair of Food Chemistry, Faculty of Mathematics and Natural Sciences, University of Wuppertal, Wuppertal, Germany; Institute of Infection and Immunity, School of Medicine, Cardiff University, Cardiff, UK.
Abstract:
The performance of two derivatization and ionization techniques for the quantitative reversed phase liquid chromatography (LC)- mass spectrometry (MS) analysis of hydroxy fatty acids (OH-PUFA) in plasma was evaluated: One used AMPP (N-(4-aminomethylphenyl)pyridinium chloride) leading to a positive charged amid-derivate which can be detected by electrospray ionization (ESI)-MS. Second yielded penta fluorobenzyl bromide (PFB) ester derivates allowing detection in electron capture atmospheric pressure chemical ionization (ecAPCI)-MS. The sensitivity of detection of a comprehensive set of hydroxy fatty acids of n6- and n3- poly unsaturated fatty acids was investigated. On the SCIEX3200 MS the applied PFB derivatization led to poor limits of detection (LOD) of 10-100nM (0.1-1pmol/0.03-0.3ng on column). By contrast, AMPP derivatization led to a similar sensitivity compared to the standard ESI(-) of non derivatized analytes (LOD about 1nM (10fmol/3pg on column)). For several analytes, including 9-HETE, 11-HETE and 17-HDHA the AMPP derivatization improved sensitivity enabling their detection in human plasma. However, precision was reduced by AMPP derivatization and variation in IS recovery indicated a strong matrix influence on the MS-signal. In sum, with the instrumentation used, neither of these derivatization methods improves in our hands the LC-MS based quantification of oxylipins.
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