Regioisomeric distribution of cholesteryl linoleate hydroperoxides and hydroxides in plasma from healthy humans provides evidence for free radical-mediated lipid peroxidation in vivo

We have previously reported the detection of cholesteryl ester hydroperoxid es, consisting mainly of cholesteryl linoleate hydroperoxides (Ch18:2-OOH), at nM levels in plasma from healthy humans (Y. Yamamoto and E. Niki, 1989, Biochem. Biophys. Res, Commun. 165: 988-993), To elucidate their productio n mechanism in vivo, we examined the distribution of Ch18:2-O(O)H regioisom ers in blood plasma from nine healthy young subjects using a sequential met hod consisting of methanol/hexane extraction in the presence of antioxidant , reductant, and internal standard, solid phase extraction to remove unoxid ized cholesteryl Linoleate, purification by reversed-phase high-performance Liquid chromatography (HPLC), and detection by normal phase HPLC, Furtherm ore, we confirm that little artifactual oxidation of cholesteryl linoleate occurred during analytical procedures indicated by the absence of oxidation products of cholesteryl 11Z,14Z-eicosadienoate (Ch20:2) when provided as a n exogenous substrate to the experimental procedure. We detected nhl levels of all free radical-mediated oxidation products, 13ZE-, 13EE-, 9-EZ, and 9 -EE-forms of Ch18:2-O(O)H, hi blood plasma, whereas the 13ZE-isomer resulti ng from enzymatic 15-lipoxygenase oxidation was not evident as a major prod uct.jlr These results indicate that free radical chain oxidation of lipids occurs even in healthy young individuals.