Phospholipase A2 dependent release of fatty acids from peroxidized membranes

Unilamellar vesicles (liposomes) consisting of liver phosphatidylcholine and phosphatidylethanolamine were used as model membranes and subjected to lipid peroxidation. Following peroxidation samples were treated with phospholipase A2 from snake venom and subsequently analyzed for products of lipid peroxidation and of phospholipase A2 action. A significantly increased susceptibility to phospholipase A2 was noted for liposomes subjected to peroxidation, as compared to controls, which correlated with the extent of lipid peroxidation measured by the formation of thiobarbituric acid reacting products and conjugated dienes. Low levels of peroxides were detected in control liposomes and these peroxidases were rapidly cleaved by phospholipase A2, such that nearly 40% of the total peroxide content was associated with free fatty acids after l5 min incubation. Oxidized liposomes contained over seven fold the level of lipid peroxidation products, and the were also rapidly cleaved by phospholipase A2 where over 50% were recovered as free fatty acids following l5 min of exposure to phospholipase A2. Along with this high order of removal of oxidized fatty acids, a marked hydrolysis of intact fatty acids was also observed. The extent of fatty acid release was roughly correlated with the degree of fatty acid unsaturation. A substantial increase in the release of arachidonic acid was found when peroxidized membranes were analyzed.