Determination of UDP-glucuronosyltransferase UGT2B7 activity in human liver microsomes by ultra-performance liquid chromatography with MS detection

Hui Xin Liu, Yu Qi He, Ying Hu, Yong Liu, Jiang Wei Zhang, Wei Li, Zheng Tao Wang, Ling Yang

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

A rapid and specific ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS) method was developed for the qualitative and quantitative determination of UGT2B7 activity using 3′-azido-3′-deoxythymidine (AZT) as probe substrate in human liver microsomes (HLMs). The method was validated for the determination of AZT glucuronidation (AZTG) with respect to specificity, linearity, detection limit, recovery, stability, precision and accuracy. The chromatographic separation was achieved on a UPLC BEH C18 column (50 mm × 2.1 mm i.d., 1.7 μm), with phase of acetonitrile-water (ratio 6:94). Selective ion reaction (SIR) monitor was specific for AZT, AZTG and I.S. The method was linear over the concentration range 0.5-500 μM for AZTG in spiked HLMs. Good precision and accuracy were obtained for concentrations over the standard curve range. AZTG was stable at 4 °C for at least 72 h in spiked liver microsomes samples. The method was successfully used to determine the kinetics of UGT activities toward AZT in HLMs. In addition, the method could determine the effects of fluconazole, a known UGT2B7 selective inhibitor, on AZTG in HLMs. Therefore, this method is suitable for in vitro studies using AZTG formation as an index reaction for UGT2B7 activity.

Original languageEnglish (US)
Pages (from-to)84-90
Number of pages7
JournalJournal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
Volume870
Issue number1
DOIs
StatePublished - Jul 1 2008

Fingerprint

Liquid chromatography
Liver Microsomes
Human Activities
Liquid Chromatography
Liver
Zidovudine
Fluconazole
Mass spectrometry
Tandem Mass Spectrometry
Limit of Detection
human UGT2B7 protein
Ions
Recovery
Kinetics
Water
Substrates

All Science Journal Classification (ASJC) codes

  • Analytical Chemistry
  • Biochemistry
  • Clinical Biochemistry
  • Cell Biology

Cite this

Determination of UDP-glucuronosyltransferase UGT2B7 activity in human liver microsomes by ultra-performance liquid chromatography with MS detection. / Liu, Hui Xin; He, Yu Qi; Hu, Ying; Liu, Yong; Zhang, Jiang Wei; Li, Wei; Wang, Zheng Tao; Yang, Ling.

In: Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences, Vol. 870, No. 1, 01.07.2008, p. 84-90.

Research output: Contribution to journalArticle

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