The Effect of Superoxide Anion in the Production of Seven Major Cholesterol Oxidation Products in Aptoric and Protic Conditions

Abstract

The reaction of cholesterol with superoxide anion was investigated in aprotic (dry) and in protic (aqueous) conditions. Superoxide anion was produced by electrochemical reduction of molecular oxygen in a solution of 0.1 M tetrabutylammonium bromide in acetonitrile. The cholesterol and cholesterol oxidation products, 7-ketocholesterol, 7α-hydroxycholesterol, 7β-hydroxycholesterol and 25-hydroxycholesterol were measured by high performance liquid chromatography using a μ-Porasil normal phase column and a UV detector. Cholesterol triol, 5α and 5β cholesterol epoxides were measured by capillary gas chromatography. None of these cholesterol oxidation products was detected from the reaction of cholesterol with superoxide anion in aprotic conditions. This indicates that cholesterol was not oxidized by direct attack of superoxide anion. When 4% water (v/v) was added to the aprotic reaction mixture, to induce the dismutation of superoxide anions and the production of hydroxy free radicals, it was found that cholesterol was not oxidized by the active oxygen species thus produced. The effects of hydrogen peroxide, on the reaction of cholesterol with superoxide anion in the protic condition was also investigated. When hydrogen peroxide was added to superoxide anion in aqueous solution, only 7-ketocholesterol, 7α-hydroxycholesterol, 7β-hydroxycholesterol were formed from cholesterol.