Identification of metabolites from type III F2-isoprostane diastereoisomers by mass spectrometry

F₂-isoprostanes (F₂-iPs) are prostaglandin (PG)-like products of non-enzymatic free radical-catalyzed peroxidation of arachidonic acid that are now widely used as indices of lipid peroxidation in vivo. Knowledge of the metabolic fate of F₂-iPs in vivo is still scant, despite its importance for defining their overall formation and biological effects in vivo. Type III F₂-iPs, which are diastereoisomers of cyclooxygenase-derived PGF_2α, may be metabolized through the pathways of PG metabolism. We therefore studied the in vitro metabolism of eight synthetic Type III F₂-iP diastereoisomers in comparison with PGF_2α. We used gas chromatography-mass spectrometry and high performance liquid chromatography-electrospray-tandem mass spectrometry for structural identification of metabolites formed after incubation of the various compounds with isolated rat hepatocytes. PGF_2α was metabolized to several known products, resulting from a combination of β-oxidation, reduction of Δ⁵ and/or Δ¹³ double bonds, and 15-OH oxidation, plus other novel products deriving from conjugation with taurine of PGF_2α and its metabolites. Of the eight F₂-iP diastereoisomers, some were processed similarly to PGF_2α, whereas others showed peculiar metabolic profiles according to specific stereochemical configurations. These data represent the first evidence of biodegradation of selected Type III F₂-iP isomers other than 8-epi-PGF_2α, through known and novel pathways of PGF_2α metabolism. The analytical characterization of these products may serve as a basis for identifying the most significant products formed in vivo.