Automated high-performance liquid chromatographic assay for the determination of 7-ethoxycoumarin and umbelliferone

J Chromatogr. 1992 Jul 1;578(1):141-5. doi: 10.1016/0378-4347(92)80237-k.


An improved high-performance liquid chromatographic assay is presented for the determination of 7-ethoxycoumarin O-deethylase activity. Following a 30-min microsomal incubation, 7-ethoxycoumarin, 4-methylumbelliferone (internal standard), and the metabolite umbelliferone were extracted with chloroform. Separation was achieved with an isocratic mobile phase using a microBondapak phenyl (300 mm x 3.9 mm I.D.) analytical column. The effluent was monitored by fluorescence detection with an excitation wavelength of 360 nm and an emission wavelength of 470 nm. The intra- and inter-assay coefficients of variation were 10 and 6%, respectively. A detection limit of 0.07 micrograms/ml was achieved, making this method suitable for characterizing P-450 activity of human livers.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • 7-Alkoxycoumarin O-Dealkylase / analysis
  • Chromatography, High Pressure Liquid / methods*
  • Coumarins / analysis*
  • Coumarins / metabolism
  • Humans
  • Microsomes, Liver / metabolism
  • Umbelliferones / analysis*
  • Umbelliferones / metabolism


  • Coumarins
  • Umbelliferones
  • 7-ethoxycoumarin
  • 7-hydroxycoumarin
  • 7-Alkoxycoumarin O-Dealkylase