Quantitation of lipid peroxidation products by gas chromatography-mass spectrometry

A method for the quantitation of lipid peroxidation products in total hepatic lipid has been developed. Lipid extracts are reduced and subsequently transmethylated with sodium methoxide. The hydroxy fatty acid methyl esters are isolated by silicic acid chromatography and derivatized to their trimethylsilyl ethers prior to analysis by gas chromatography-mass spectrometry. Three isomers, 11-, 12-, and 15-hydroxyeicosatetraenoic acid (HETE), are quantitated using selected ion monitoring techniques relative to the internal standard, methyl 15-hydroxyarachidate. In mice treated with carbon tetrachloride (2 ml/kg), the HETE levels in total hepatic lipid were 20-fold greater than those found in control animals. HETE levels were also elevated (5- to 10-fold) in hepatic lipid from rats treated with the same dose of carbon tetrachloride. Studies on subcellular fractions with this methodology show that these lipid peroxidation products are 5- to 6-fold higher in hepatic plasma membrane vesicles isolated from rats treated with carbon tetrachloride when compared with those isolated from control animals.