Studies on steroids. CCXXXVIII. Determination of bile acids in liver tissue by gas chromatography-mass spectrometry with negative ion chemical ionization detection.

A method for the determination of bile acids in 2-10 mg of human liver tissue by gas chromatography (GC) in combination with negative ion chemical ionization (NICI) mass spectrometry is described. Unconjugated, glycine- and taurine-conjugated bile acids labelled with 18O and 2H were used as internal standards. The preparation of these compounds was attained by the exchange reaction of the carbonyl group with H218O, followed by metal hydride reduction. Bile acids in solubilized liver tissue were extracted with a Sep-Pak C18 cartridge, separated into the unconjugated, glycine- and taurine-conjugated fractions by ion-exchange chromatography on piperidinohydroxypropyl-Sephadex LH-20 and then derivatized to the pentafluorobenzyl ester-dimethylethylsilyl ethers. Subsequent resolution of each fraction into lithocholate, deoxycholate, chenodeoxycholate, ursodeoxycholate and cholate was attained by GC on a cross-linked 5% phenylmethyl silicone fused-silica capillary column where bile acids were monitored with a characteristic carboxylate anion [M - 181]-in the NICI mode using isobutane as a reagent gas. The newly developed method was applied to the quantitation of bile acids in liver tissue with satisfactory sensitivity and reliability.