title: Determination of 3-oxo-delta4- and 3-oxo-delta4,6-bile acids and related compounds in biological fluids of infants with cholestasis by gas chromatography-mass spectrometry.
abstract: A method has been developed for the determination of 3-oxo-delta4- and 3-oxo-delta4,6-bile acids and related bile acids in biological fluids of infants by gas chromatography-mass spectrometry (GC-MS) of the methyl ester-dimethylethylsilyl ether-methoxime derivatives. The 7alpha-hydroxylated 3-oxo-delta4-bile acids were partially dehydrated to give the 3-oxo-delta4,6-bile acids by trimethylsilyl or dimethylethylsilyl derivatization and other pretreatments under acidic or alkaline conditions for GC-MS analysis. To prevent dehydration, the 3-oxo-delta4-bile acids were derivatized to the oximes by treatment with O-methylhydroxylamine prior to pretreatments such as solid-phase extraction, enzymatic solvolysis and hydrolysis of the conjugates, and silylation with dimethylethylsilylimidazole. Calibration curves for the bile acids were linear over a range of 5-250 ng and the detection limit was 100 pg for each 3-oxo-delta4-bile acid. Recoveries of the bile acids and their glycine and taurine conjugates from bile acid-free urine and serum ranged from 94.2 to 105.9% of their added amounts. The bile acids in urine and serum of four patients with severe cholestatic liver disease were measured by the analytical method, and the 3-oxo-delta4-bile acids were determined to be the major bile acids (59-68%) in the urines associated with 3-oxo-delta4-steroid 5beta-reductase deficiency or dysfunction.
authors.affiliation: Faculty of Pharmaceutical Sciences, Health Sciences University of Hokkaido, Ishikari-Tobetsu, Japan.
authors.forename: M
authors.initials: M
authors.lastname: Suzuki
authors.forename: T
authors.initials: T
authors.lastname: Murai
authors.forename: T
authors.initials: T
authors.lastname: Yoshimura
authors.forename: A
authors.initials: A
authors.lastname: Kimura
authors.forename: T
authors.initials: T
authors.lastname: Kurosawa
authors.forename: M
authors.initials: M
authors.lastname: Tohma
mesh_terms: D001647:Bile Acids and Salts / Q000097:blood* / Q000652:urine*; D002779:Cholestasis / Q000097:blood* / Q000652:urine*; D008401:Gas Chromatography-Mass Spectrometry; D006801:Humans; D007223:Infant
chemical_list: D001647:Bile Acids and Salts