Abstract
Amberlite XAD-2 was used to extract bile acids from urine or diluted serum of patients with hepatobiliary diseases. Columns containing Sephadex LH-20 were then used to separate the sulfated and nonsulfated bile acids. Thin-layer chromatography of the sulfated bile acid fraction obtained from urine revealed several spots with R(F) values different from those of the taurine or glycine conjugates. According to thin-layer chromatographic mobilities, gas-liquid chromatographic analyses, infrared spectra, and elementary analysis of the sulfated material, one of these sulfated bile acids was identified as glycochenodeoxycholic acid monosulfate, and the others were presumed to be taurochenodeoxycholic acid sulfate and glycocholic acid sulfate. A large amount of bile acid sulfate was found in urine of patients with hepatobiliary diseases. They accounted for 35.5-93.3% of total urinary bile acids and consisted of both di- and trihydroxycholanoic acids, with chenodeoxycholic acid as the major acid. Sulfated bile acids were also found in serum, and accounted for 1.8-21.2% of the total bile acids. Only dihydroxycholanoic acids (mainly chenodeoxycholic) were identified.
Highlights
E revealed several spots with RF values different from those of the taurine or glycine conjugates
The butanol was washed with water and evaporated.The residue was dissolved in a smallamount of chloroform-methanol 1:l (v/v) containing0.01 M NaCl and applied to a Sephadex LH-20 column
The methanol eluate was evaporated, and the residue was dissolved in 20 ml of 0.1 N NaOH. This sample was applied to an Amberlite XAD-2 column, which was washed with water
Summary
Allsolvents wereanalyticalgrade.AmberliteXAD-2 resin(RohmandHaas,Philadelphia,Pa.)was washed with 5-10 vol of water, methanol, acetone, methanol, and water, in that order. (25-100 pm)was purchased from Pharmacia, Uppsala, Sweden, and alumina (activity 1)fromMerckA.G.,Darmstadt,Germany. Gas-liquid chromatography A gas chromatograph, model GC-2C (Shimazu Manufacturing Co., Kyoto, Japan), with a hydrogenflame detector was used. T h e glass U-shaped column (150 cm X 4 mm ID) was packed with a commercial preparation (Shimazu Manufacturing Co.) of 1.5% QF-1 onChromosorb W. T h e column temperature was21 5°C
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