The glycine conjugate of 3 beta-hydroxy-5-cholen-24-oic acid and its sulfate labeled with deuterium at the C-2, -4, and -23 positions were synthesized. A highly sensitive and specific quantitative assay of the bile acid has been developed by selected ion monitoring in gas chromatography-mass spectrometry of the methyl ester trimethylsilyl ether derivatives using the deuterium labeled conjugates as internal standards. Calibration curves for the bile acid and its sulfate exhibited a linear relationship over the range of 0.01-100 micrograms/ml in human serum.