Women and smokers have elevated urinary F2-isoprostane metabolites: A novel extraction and LC-MS methodology

Abstract

F2-Isoprostanes (F2-IsoPs), regio- and stereoisomers of prostaglandin F2α (PGF2α), and urinary F2-IsoP metabolites including 2,3-dinor-5,6-dihydro-8-iso-PGF 2α [2,3-dinor-8-iso-PGF1α (2,3-dinor-F1)] and 2,3 dinor-8-iso-PGF2α (2,3-dinor-F2), have all been used as biomarkers of oxidative stress. A novel method was developed to measure these biomarkers using a single solid phase extraction (SPE) cartridge, separation by HPLC, and detection by negative mode selected reaction monitoring (SRM) mass spectrometry (MS), using authentic standards of PGF2α; 8-iso-PGF2α; 2,3-dinor-F1 and 2,3-dinor-F2 to identify specific chromatographic peaks. The method was validated in a population of healthy, college-aged nonsmokers (n = 6 M/8F) and smokers (n = 6 M/5F). Urinary F 2-IsoP concentrations were ∼0.2-1.5 μg/g creatinine, 2,3-dinor-F1 was ∼1-3 μg/g and 2,3-dinor-F2 was ∼3-5 μg/g. Additional F2-IsoPs metabolites were identified using SRM. The sum of all urinary F2-IsoP metabolites was 50-100 μg/g creatinine indicating their greater abundance than F2-IsoPs. Women had higher F2-IsoP metabolite concentrations than did men (MANOVA, main effect P = 0.003); cigarette smokers had higher concentrations than did nonsmokers (main effect P = 0.036). For men or women, respectively, smokers had higher metabolite concentrations than did nonsmokers (P < 0.05). Thus, our method simultaneously allows measurement of urinary F2-IsoPs and their metabolites for the determination of oxidative stress. © 2008 AOCS.