Identification of 5 beta-cholestane-3 alpha,7 alpha,12 alpha,24,25,26-hexol in human urine

Abstract

A new bile alcohol, 5 beta-cholestanehexol, was identified in the urine of healthy humans as the glucuronide. The bile alcohol glucuronide fraction was isolated by an ion exchange chromatography on piperidinohydroxypropyl Sephadex LH-20. After enzymatic hydrolysis, the bile alcohols were converted into trimethylsilyl ether derivatives and analyzed by a combination of gas-liquid chromatography and mass spectrometry. The major bile alcohol was 27-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24,25-pentol. As minor constituents the following C26 and C27 bile alcohols were identified: 27-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24,25,26-hexol, 5 beta-cholestane-3 alpha,7 alpha,12 alpha,24,25-pentol, 5 beta-cholestane-3 alpha,7 alpha,12 alpha,24,26-pentol, 5 beta-cholestane-3 alpha,7 alpha,12 alpha,25,26-pentol. In addition to these bile alcohols, a new bile alcohol was identified as a sixth component of the urinary bile alcohols. The structure was assigned as (24S)-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24,25,26-hexol by the direct comparison of mass spectral data and chromatographic properties with synthetic standard. The average daily excretion of the new bile alcohol was 28.6 micrograms and 3.0% of the total bile alcohols. The presence of 27-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24,25-pentol and 27-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24,25,26-hexol suggests that 26-hydroxylation of 5 beta-cholestane-3 alpha,7 alpha,12 alpha,24,25-pentol is most likely for the biosynthesis of this new bile alcohol.