F4-Isoprostanes: A Novel Class of Prostanoids Formed during Peroxidation of Docosahexaenoic Acid (DHA)

Isoprostanes are prostaglandin (PG)-like compounds derived from free radical-catalyzed peroxidation of polyunsaturated fatty acids. F2-isoprostanes are produced in vivo by a non-cyclooxygenase mechanism involving free radical peroxidation of arachidonic acid. Peroxidation of eicosapentaenoic acid produces F3-isoprostanes. In this study, we explore the possibility of formation of F4-isoprostanes during peroxidation of docosahexaenoic acid (DHA) in vitro. DHA-liposomes were exposed at 37°C to either 2, 2′-azobis-(2-amidinopropane) hydrochloride (AAPH) or copper ions at final concentrations of 10 mM and 50 μM, respectively. Sample processing involved solid-phase extraction on a C18and an NH2- cartridge. After conversion to pentafluorobenzyl ester and trimethylsilyl derivatives, F4-isoprostanes were analysed by negative ion-chemical ionisation mass spectrometry using tetradeuterated PGF2α(PGF2-d4) as the internal standard. Quantitative analysis was carried out by selected ion monitoring (SIM) of the carboxylated anion [M-180]−at m/z 593 and 573 for the F4-isoprostanes and PGF2-d4, respectively. DHA oxidised by AAPH or by copper ions gave rise to a similar family of F4-isoprostanes. Formation of F4-isoprostanes increased throughout the oxidation period and was correlated with other indices of lipid peroxidation (hydroperoxides and thiobarbituric acid reactive substances). The possibility of analyzing F4-isoprostanes should provide new opportunities for studying the role of lipid peroxidation in nutritional studies and in the pathogenesis of neurodegenerative diseases.