Formation of PGF2-isoprostanes during the oxidative modification of low density lipoprotein

It has been recently reported that a series of prostaglandin F2-like compounds (PGF2-isoprostanes) are produced in vivo by non-enzymatic peroxidation of arachidonic acid. Of these, 8-epi-PGF2 alpha is the major component and has been shown to be a potent vasoconstrictor. Here we investigated the formation of PGF2-isoprostanes during copper-mediated oxidation of low density lipoprotein (LDL). Free and total (sum of free and esterified) levels of PGF2-isoprostanes were measured at times 0, 3, 6, 8 and 24 h, using a solid-phase extraction procedure and gas chromatography-mass spectrometry. In native LDL, free levels of PGF2-isoprostanes were between 0.06 and 0.10 ng/mg protein (n = 4), and the total levels ranged from 0.027 to 0.057 ng/mg protein (n = 4). Free levels of the isoprostanes were found to increase throughout the oxidation, whereas total levels reached a maximum after 3 h then gradually decreased. 8-epi-PGF2 alpha was the major isoprostane formed (free concentration after 24 h 1.8 +/- 0.1 ng/mg protein (n = 4); total concentrations of 8.8 +/- 1.8 and 6.1 +/- 3.6 ng/mg protein (n = 4) after 3 and 24 h, respectively). The levels of isoprostanes correlated well with other indices of lipid peroxidation (conjugated dienes, hydroperoxides, thiobarbituric reactive substances) measured at similar time points. The release of PGF2-isoprostanes from oxidised LDL in macrophages could be a contributory factor in the development of atherosclerosis.