Identification and Characterization of Novel F2-Isoprostane Metabolites
Résumé
F2-isoprostanes (F2-IsoPs) are formed from the free radical
oxidation of arachidonic acid and are widely used as biomarkers of
endogenous oxidative damage. Sixty-four F2-IsoP isomers are
generated during lipid peroxidation; however, most studies have
focused only on one isomer, 15-F2t-IsoP (commonly referred to as
8-iso-PGF2α). Immunoassay and mass spectrometric (MS)
techniques have been developed to quantify 15-F2t-IsoP in
biological fluids. Even though this molecule has a short half-life of
only a few minutes, most studies quantify 15-F2t-IsoP directly
rather than its’ urinary metabolite (2,3-dinor-5,6-dihydro-15-F2tIsoP, 15-F2t-IsoP-M). An increasing body of evidence
demonstrates the efficacy of 15-F2t-IsoP-M quantification. For
example, 15-F2t-IsoP-M, but not 15-F2t-IsoP, was positively
associated with age and postmenopausal status in a large
population study of middle-aged and older women. Urinary 15-F2tIsoP-M also shows associations with plasma antioxidant levels,
carbohydrate intake, glycemic index, and environmental
exposures while urinary 15-F2t-IsoP does not. The utility of 15-F2tIsoP measurement is further limited by the fact that this molecule
can also be generated enzymatically via the cyclooxygenases. On
the other hand, 5-series F2-IsoPs are only generated via
nonenzymatic mechanisms. And, it has been demonstrated that
these molecules are present in greater abundance in plasma, tissue,
and urine than 15-series F2-IsoPs. Yet, 5-series F2-IsoP metabolism
remains unexplored. We have thus undertaken experiments to
identify and characterize metabolites of 5-epi-5-F2t-IsoP and 5-epi-
5-F2c-IsoP, two abundantly produced 5-series F2-IsoPs, in human
liver microsomes. Using targeted and untargeted metabolomic
approaches, glucuronide and sulfate metabolites of F2-IsoPs have
been identified by MS. Metabolism of these molecules by specific
enzymes has also been tested. Collectively, these findings indicate
that F2-IsoP metabolism is more complex than previously
appreciated. The presence of these metabolites in human urine is
under investigation. We anticipate that the completion of this work
will redefine F2-IsoP metabolism and outline a strategy to
comprehensively evaluate endogenous formation of these
important molecules