Quantification of dinor,dihydro metabolites of F 2-isoprostanes in urine by liquid chromatography/tandem mass spectrometry

Abstract

The F 2-isoprostanes (F 2-IsoP) are a series of prostaglandin (PG)-F 2-like compounds that are produced by free-radical-mediated oxidation of arachidonic acid. One F 2-IsoP with potent biological activity is 15-F 2t-IsoP and increased levels of 15-F 2t-IsoP have been measured in several diseases. The major urinary metabolite of 15-F 2t-IsoP (8-iso-PGF 2α) is 2,3-dinor-5,6-dihydro-15-F 2t-IsoP (15-F 2t-IsoP-M). Previously, we developed a stable isotope dilution gas chromatography/negative chemical ionization/mass spectrometry (MS) assay for 15-F 2t-IsoP-M, which, while highly sensitive, required time-consuming derivatization and thin-layer chromatography purification. We now report the development of a more rapid high-performance liquid chromatography method coupled to electrospray ionization–tandem mass spectrometry (LC/MS/MS) to analyze all of the dinor,dihydro metabolites of the F 2-IsoP isomers (F 2-IsoP-M). The precision of this assay was ±5.0% and the accuracy 80%. The assay remained linear over a range of 1–100 ng injected onto the LC column. Levels of F 2-IsoP-M determined by the LC/MS/MS assay method significantly correlated with levels of 15-F 2t-IsoP-M determined by the GC/MS assay ( R = 0.77 y = 67.2 x − 0.5). The levels of F 2-IsoP-M detected in spot urines from 40 normal subjects were 38.1 ± 19.1 ng/mg creatinine (mean ± SD). This method provides an accurate and rapid assay to assess oxidative status in vivo.