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Nitro-fatty Acids Occur in Human Plasma in the Picomolar Range: a Targeted Nitro-lipidomics GC–MS/MS Study

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Lipids

Abstract

First studies on the occurrence of nitrated fatty acids in plasma of healthy subjects revealed basal concentrations of 600 nM for free/nonesterified nitro-oleic acid (NO2-OA) as measured by liquid chromatography tandem mass spectrometry (LC–MS/MS). We recently showed by a gas chromatography tandem mass spectrometry (GC–MS/MS) method the physiological occurrence of two isomers, i.e., 9-NO2-OA and 10-NO2-OA, at mean basal plasma concentrations of 880 and 940 pM, respectively. In consideration of this large discrepancy we modified our originally reported method by replacing solid-phase extraction (SPE) by solvent extraction with ethyl acetate and by omitting the high-performance liquid chromatography (HPLC) step for a more direct detection and with the potential for lipidomics studies. Intra-assay imprecision and accuracy of the modified method in human plasma were 1–34% and 91–221%, respectively, for added NO2-OA concentrations in the range 0–3,000 pM. This method provided basal plasma concentrations of 306 ± 44 pM for 9-NO2-OA and 316 ± 33 pM for 10-NO2-OA in 15 healthy subjects. Nitro-arachidonic acid and nitro-linolenic acid were not detectable in the plasma samples. In summary, our studies show 9-NO2-OA and 10-NO2-OA as endogenous nitrated fatty acids in human plasma in the pM range; HPLC is recommendable as a sample clean-up step for reliable quantification of nitro-oleic acids by GC–MS/MS.

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Abbreviations

CID:

Collision-induced dissociation

GC–MS/MS:

Gas chromatography tandem mass spectrometry

HAc:

Acetic acid

LC–MS/MS:

Liquid chromatography tandem mass spectrometry

MeCN:

Acetonitrile

MeOH:

Methanol

m/z :

Mass-to-charge

NO2-OA:

Nitro-oleic acid

PFB:

Pentafluorobenzyl

PFB-Br:

Pentafluorobenzyl bromide

QC:

Quality control

S/N:

Signal-to-noise

SPE:

Solid-phase extraction

SRM:

Selected reaction monitoring

TSQ:

Triple-stage quadrupole

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Authors and Affiliations

  1. Institute of Clinical Pharmacology, Hannover Medical School, Carl-Neuberg-Strasse 1, 30625, Hannover, Germany

    Dimitrios Tsikas, Alexander A. Zoerner, Anja Mitschke & Frank-Mathias Gutzki

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  1. Dimitrios Tsikas
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  2. Alexander A. Zoerner
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  3. Anja Mitschke
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  4. Frank-Mathias Gutzki
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Correspondence to Dimitrios Tsikas.

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Tsikas, D., Zoerner, A.A., Mitschke, A. et al. Nitro-fatty Acids Occur in Human Plasma in the Picomolar Range: a Targeted Nitro-lipidomics GC–MS/MS Study. Lipids 44, 855–865 (2009). https://doi.org/10.1007/s11745-009-3332-4

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