Determination of 1β-Hydroxylated Bile Acids and Related Compounds in Human Biological Fluids by Gas Chromatography-Mass Spectrometry

Abstract

A highly sensitive and specific quantitative assay of the 1β-hydroxylated bile acids and related compounds has been developed by gas chromatography-selected ion monitoring of their methyl ester-trimethylsilyl ether derivatives using the deuterium labeled internal standards: [²H₇] DCA, [²H₇] CA, [²H₅] 3β-hydroxy- and [²H₆]3β,12α-dihydroxy-5-cholen-24-oic acids. Calibration curves for these bile acids are useful over the range of 0.01–100 μg in serum (0.2 ml), urine (5 ml), or lyophilized meconium (1 mg). Recoveries of the bile acids and their conjugates ranged from 92 to 101% of the added amounts of their standard samples. In the analysis of bile acids in human meconium by the developed method, novel 1β,3α,7α-trihydroxy- (II) and 1β,3α,7α,12α-tetrahydroxy- (IV) 5β-cholan-24-oic acids were determined in significant amounts of 0.24 μg/mg (5.3% of the total bile acids) and 0.27 μg/mg (6.3%), respectively.