Separation and Detection of Bile Acid 3-Glucuronides in Human Urine by Liquid Chromatography/Electrospray Ionization–Mass Spectrometry

✍ Scribed by Shigeo Ikegawa; Naoaki Murao; Takashi Motoyama; Takamitsu Yanagihara; Toshifumi Niwa; Junichi Goto

• Publisher: John Wiley and Sons
• Year: 1996
• Tongue: English
• Weight: 366 KB
• Volume: 10
• Category: Article
• ISSN: 0269-3879
• DOI: 10.1002/(sici)1099-0801(199611)10:6<313::aid-bmc603>3.0.co;2-f

✦ Synopsis

A method for the separation and detection of bile acid 3-glucuronides by liquid chromatography (LC)/ electrospray ionization (ESI)-mass spectrometry (MS) has been developed. On the ESI mode, glucuronides were characterized by an intense pseudo-molecular ion [M-HI-with a doubly charged ion [M-2HI2-, and the ratio of these negative ions were markedly influenced by an acidic component of salt added to a mobile phase, according to a pKa value of an acidic moiety at C-24. Bile acid 3-glucuronides in human urine were extracted with a Sep-Pak Clo cartridge, followed by purification on lipophylic ion exchange gel, piperidinohydroxypropyl Sephadex LH-20. Subsequent resolution into unconjugated, glycine and taurine conjugated bile acid 3-glucuronides was attained by LC on a Develosil ODs-HG-5 column using 20 mM ammonium acetate (pH 7.0)-methanol(lO12, v/v) as a mobile phase, where 3-glucuronides were monitored with a characteristic negative ion [M-HI-. The newly developed method was applied to the quantitatiation of bile acid 3-glucuronides in human urine with satisfactory sensitivity and reliability.