Cholesterol oxides II. Measurement of the 5,6-epoxides during cholesterol oxidation in aqueous dispersions

Cholesterol in aqueous dispersion with sodium stearate or Triton surfactants was aerated at various pH values at 50 and 80 C. Analysis of the reaction mixtures by TLC during the oxidation produced qualitatively similar patterns regardless of pH or temperature. Major oxidation products observed were 7-ketocholesterol, the isomeric 7-hydroperoxy- and 7-hydroxycholesterols, the isomeric 5,6-epoxycholestanols and 3β,5α,6β-trihydroxycholestanol. The concentrations of 3β,5α,6β-trihydroxycholestanol and an unknown compound increased greatly at the lower pH values. Recovery of the 5,6-epoxide isomers by preparative TLC followed by capillary GC allowed theα- andβ-epoxide isomers to be quantitated. Oxidations at pH 8 and 12 produced increasing amounts of the epoxides with time, without significant changes in theα/β-epoxide ratio. However, oxidations at the acidic pH values of 5.5 and 3 showed large changes in theα/β-epoxide ratio during the oxidation. Measurement of the hydrolysis rates of the 5,6-epoxides at pH 5.5 showed that theβ-epoxide isomer is more labile than theα-isomer by a factor of 2.5. The rate constant for the hydrolysis of theα-epoxide isomer was 5.3 × 10⁻⁷ sec⁻¹ and that of theβ-isomer 13 × 10⁻⁷ sec⁻¹.