1992 年 21 巻 4 号 p. 249-258
We describe a new enzymatic assay method to measure sulfated bile acids in urine, in. which the essential components are a novel bile acid sulfate sulfatase (BSS) and β-hydroxysteroid dehydrogenase (β-HSD: EC 1.1.1.1.51). The BSS produced inducibly by Pseudomonas testosteroni was able to efficiently hydrolyze various kinds of bile acid 3α-sulfates to form 3β-hydroxyl bile acids and sulfuric acid. Total sulfated bile acid (TSBA) in urine was determined by the combined reaction of desulfation of bile acid 3α-sulfates by BSS, followed by dehydrogenation of desulfated substrates by β-HSD and conventional colorimetric assay of reduced nicotinamide adenine dinucleotide (NADH) as formazan dye in the presence of nitrotetrazolium blue and diaphorase. The calibration curve for glycolithocholic acid 3-sulfate as the standard was linear up to 250μmol/l. Analytical recovery of various sulfated bile acids in urine averaged 98% excepting the recovery of taurolithocholic acid 3-sulfate which was approximately 80%. The CV for intra-and inter-assay variations was ≤3.2% and ≤3.4%, respectively. This method is accurate and simple, and less time-consuming than those previously reported. We determined the concentration of TSBA in adult urine samples by this method, and observed that urinary TSBA of normal subjects was less than 7.6μmol/g creatinine (mean2.0±1.8μmol/g creatinine), while in patients with acute hepatitis, liver cirrhosis, and intra-and extra-hepatic biliary obstruction urinary TSBA was markedly increased.
