Urinary 8-epi-PGF{sub 2{alpha}} and its endogenous {beta}-oxidation products (2,3-dinor and 2,3-dinor-5,6-dihydro) as biomarkers of total body oxidative stress

Although measurements of plasma F{sub 2}-isoprostanes are established markers of oxidative stress, their quantification only reflects acute non-enzymatic lipid peroxidation. In this study, a new approach is described for the rapid isolation and measurement of urinary 8-epi-PGF{sub 2{alpha}} and its endogenous {beta}-oxidation metabolites (2,3-dinor-8-epi-PGF{sub 2{alpha}} and 2,3-dinor-5,6-dihydro-PGF{sub 2{alpha}}) for use as index of total body oxidative stress. Isoprostanes were partitioned with ethyl acetate and subsequently purified by chromatography on an aminopropyl (NH{sub 2}) and silica (Si) cartridge. Final analysis of F{sub 2}-isoprostanes as trimethylsilyl-ester/pentafluorobenzyl ester derivatives was carried out by stable isotope dilution mass spectrometry. Overall recovery of F{sub 2}-isoprostanes was 80 {+-} 4%. Inter- and intra-assay coefficients of variation were 5% and 7%, respectively. In a group of healthy humans, the mean excretion rates expressed as nmol/mmol creatinine for 2,3-dinor-8-epi-PGF{sub 2{alpha}}, 2,3-dinor-5,6-dihydro-8-epi-PGF{sub 2{alpha}}, and 8-epi-PGF{sub 2{alpha}} were 5.43 {+-} 1.93, 2.16 {+-} 0.71, and 0.36 {+-} 0.16, respectively. Correlations were obtained between 8-epi-PGF{sub 2{alpha}} and 2,3-dinor-8-epi-PGF{sub 2{alpha}} or 2,3-dinor-5,6-dihydro-8-epi-PGF{sub 2{alpha}} (r = 0.998 and r = 0.937, respectively). A strong relationship was also seen between 2,3-dinor-8-epi-PGF{sub 2} and 2,3-dinor-5,6-dihydro-8-epi-PGF{sub 2{alpha}} (r = 0.949). The new technique allows for high sample throughput and avoids the need for HPLC and/or other expensive equipment required for the initial sample preparation. Simultaneous analysis of urinary 8-epi-PGF{sub 2{alpha}} and its metabolites should provide unique tool in clinical trials exploring the role of oxidant injury in human disease.