Hypochlorous acid-mediated modification of cholesterol and phospholipid: analysis of reaction products by gas chromatography-mass spectrometry.
Oxidative modification of membrane lipids by hypochlorous acid could be an important element in the mechanism of membrane disruption by activated neutrophils. We have previously shown that hypochlorous acid reacts with unsaturated fatty acids of membrane phospholipids to give fatty acid chlorohydrins (Winterbourn et al. 1992. Arch. Biochem. Biophys. 296: 547-555). In the present study, we examined the reaction of cholesterol in bilayers with an inert phospholipid carrier. Product separation and identification was performed using gas chromatography-mass spectrometry after trimethylsilyl-derivatization. Unlike the reaction of hypochlorous acid with unsaturated fatty acids, no chlorohydrin derivatives were found with cholesterol. Instead, the main oxidation products were identified as the epimeric cholesterol 5,6-epoxides and 4-hydroxycholesterol, while several other hydroxy- and keto-derivatives were also found in smaller amounts. Analysis of the products obtained after reaction of vesicles composed of a mixture of several unsaturated phospholipid species plus cholesterol revealed that the individual fatty acids and cholesterol all exhibit comparable susceptibilities toward hypochlorous acid. Using myeloperoxidase to generate hypochlorous acid, basically the same products and product distribution were obtained. These studies show that unsaturated phospholipids and cholesterol can be profoundly modified by reaction with hypochlorous acid. This warrants further investigation to define the role of lipid modifications in neutrophil-mediated membrane disruption.
van den Berg
J J
JJ
Children's Hospital Oakland Research Institute, CA 94609.
Winterbourn
C C
CC
Kuypers
F A
FA
D002784:Cholesterol / Q000031:analogs & derivatives / Q000737:chemistry*; D008401:Gas Chromatography-Mass Spectrometry; D006863:Hydrogen-Ion Concentration; D006888:Hydroxycholesterols / Q000737:chemistry; D006997:Hypochlorous Acid / Q000737:chemistry*; D008051:Lipid Bilayers / Q000737:chemistry; D008081:Liposomes / Q000737:chemistry; D010084:Oxidation-Reduction; D009195:Peroxidase / Q000378:metabolism; D010743:Phospholipids / Q000737:chemistry*
D006888:Hydroxycholesterols; D008051:Lipid Bilayers; D008081:Liposomes; D010743:Phospholipids; C002415:cholesterol alpha-oxide; D006997:Hypochlorous Acid; D002784:Cholesterol; D009195:Peroxidase