Abstract
A novel high-performance liquid chromatographic (HPLC) method with post-column detection was developed to simultaneously analyze the lipid hydroperoxides of polar and nonpolar lipids. The HPLC method uses a reversed-phase column (octyl-bonded silica) and a mobile phase of water/acetonitrile/methanol. Hydroperoxides were detected at 592 nm by using a ferrous (II)/xylenol orange (FeXO) reagent. This method enabled the separation of fatty acid hydroperoxides, phosphatidylcholine hydroperoxides, and hydroperoxides of neutral lipids, including triacylglycerol hydroperoxides and cholesterol ester hydroperoxides, by chromatography. These hydroperoxides could be quantified in a range between 40 pmol and 2 nmol. This new method was applied to estimate the lipid hydroperoxides formed during the photosensitized oxidation of rat plasma.
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