Gas Chromatograhy-Tandem Mass Spectrometry Determination of 8-Iso-PGF2α,a Biomarker of in vivo Lipid Peroxidation,in Human Plasma and Urine

Abstract

The measurement of isoprostanes is a promising assay that is specific and sensitive enough to detect in vivo lipid peroxidation. We present here a gas chromatography-tandem mass spectrometry (GC/MS/MS)method that enables determination of 8-iso-prostaglandin F2α (8-iso-PGF2α)in human plasma and urine. After the addition of [²H₄]-PGF2α as the internal standard to acidified plasma or urine, the samples are purified on C₁₈ and silica cartridges, derivatised as pentafluorobenzyl esters, extracted with diethyl ether, purified on silica gel TLC plates and finally silylated. Then, 8-iso-PGF2α and its internal standard are measured by GC/MS/MS in selective-reaction monitoring mode using the transition [M −181]⁻ to [M −181 – (3 × 90)]⁻. The detection limit of this method is 5 pg mL⁻¹. Its application is presented in two situations of oxidative stress: in vitro low-density lipoprotein oxidation and in smokers. Measurement of urinary 8-iso-PGF2α levels provides a non-invasive in vivo index of free radical generation that appears not to be confounded by changes in diet.