Rapid and quantitative determination of metabolites from multiple cytochrome P450 probe substrates by gradient liquid chromatography-electrospray ionization-ion trap mass spectrometry

J Chromatogr B Analyt Technol Biomed Life Sci. 2002 Nov 25;780(2):371-9. doi: 10.1016/s1570-0232(02)00548-2.

Abstract

A rapid quantitative assay method, developed by combining fast gradient liquid chromatography and electrospray ionization-ion trap mass spectrometry, is described for the simultaneous determination of CYP450 probe substrate metabolites (4-aminophenol for CYP2E1, acetaminophen for CYP1A2, dextrorphan for CYP2D6, 4'-hydroxymephenytoin for CYP2C19, 4-hydroxytolbutamide for CYP2C9 and 6beta-hydroxytestosterone for CYP3A4) in microsomal incubations. Using this method Michaelis-Menten kinetic parameters K(m) and V(max) for the probe substrates in human liver microsomes were obtained. This LC-MS-MS method, developed with the use of LC-ESI-ion trap MS instrumentation, can efficiently be used to improve the throughput and cost-effectiveness in the preclinical drug metabolism studies.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Calibration
  • Chromatography, Liquid / methods*
  • Cytochrome P-450 Enzyme System / metabolism*
  • Humans
  • Isoenzymes / metabolism*
  • Microsomes, Liver / enzymology
  • Quality Control
  • Reproducibility of Results
  • Spectrometry, Mass, Electrospray Ionization / methods*
  • Substrate Specificity

Substances

  • Isoenzymes
  • Cytochrome P-450 Enzyme System