Goto J, Suzaki K, Ebihara M, Nambara T, Masu A
J Chromatogr. 1985 Dec 13;345(2):241-50.
The separation and characterization of unconjugated and conjugated bile acid 3-glucuronides in biological fluids without prior deconjugation by high-performance liquid chromatography (HPLC) are described. A urine sample from a patient with obstructive jaundice was passed through a Sep-Pak C18 cartridge and was separated into groups by ion-exchange chromatography on a lipophilic gel, piperidinohydroxypropyl Sephadex LH-20, providing the glucuronide fraction. Subsequent resolution into individual 3-glucuronides was attained by HPLC on muBondapak C18 and Shodex ODS Pak F-411 columns. The 3-glucuronides of cholate, deoxycholate, chenodeoxycholate, glycocholate, glycochenodeoxycholate and taurochenodeoxycholate were identified on the basis of their behaviour in HPLC using mobile phases of different pH. The enzymatic hydrolysis of these glucuronides and derivatization of deconjugated bile acids with 1-anthroyl nitrile followed by chromatographic separation on a Cosmosil 5C18 column with fluorescence detection were carried out for unequivocal characterization. The ratio of unconjugated, glyco- and tauro-conjugated bile acid 3-glucuronides excreted in urine was found to be ca. 2:3:1.
本文描述了在不进行预先脱共轭的情况下，通过高效液相色谱法（HPLC）对生物流体中未共轭和共轭胆汁酸3-葡萄糖醛酸苷进行分离和表征的方法。将一名阻塞性黄疸患者的尿液样本通过Sep-Pak C18柱，并在亲脂性凝胶哌啶羟丙基葡聚糖凝胶LH-20上进行离子交换色谱分离，得到葡萄糖醛酸苷部分。随后，通过在muBondapak C18和Shodex ODS Pak F-411柱上进行HPLC，将其分离为各个3-葡萄糖醛酸苷。根据胆酸盐、脱氧胆酸盐、鹅去氧胆酸盐、甘氨胆酸盐、甘氨鹅去氧胆酸盐和牛磺鹅去氧胆酸盐的3-葡萄糖醛酸苷在不同pH值流动相的HPLC行为进行鉴定。对这些葡萄糖醛酸苷进行酶促水解，并将脱共轭胆汁酸与1-蒽腈衍生化，随后在Cosmosil 5C18柱上进行色谱分离并进行荧光检测，以进行明确的表征。发现尿液中排泄的未共轭、甘氨共轭和牛磺共轭胆汁酸3-葡萄糖醛酸苷的比例约为2:3:1。
