The in vitro biosynthesis and stability measurement with agyl-glycuronide isoformes of the main metabolite of ipriflavone

Katalin Jemnitz, Ferenc Lévai, Katalin Monostory, Istv́n Szatmári, Ĺszló Vereczkey

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The formation and stability of 1-β-glucuronide conjugate of the main metabolite of ipriflavone [7-(1-carboxy-ethoxy)-isoflavone] (CI) - were studied by using liver microsomes, hepatocytes, and isolated perfused liver of untreated and 3-methylcholanthrene (MC) treated dog and rat, and human liver microsomes. MC treatment enhanced the rate of conjugation twice as much as that of the control in the microsomes of both dogs and rats. Conjugation of CI by microsomes results in two metabolites, both sensitive to pH and temperature. Other two glucuronide forms appeared in experiments with hepatocytes and perfused liver. Mass spectrometry supported. The conclusion, assumption that both metabolites produced by microsomes are glucuronide conjugate isoforms of CI, and that they could be distinguished according to the intensity of peaks on FAB-MIKE spectra. The β-glucuronidase enzyme hydrolysed only the 1-β-glucuronide isomer, the other, migrated form remained unchanged. D-saccharic-acid-1,4-lactone, a specific inhibitor of β-glucuronidase enzyme, decreased the rate of enzymatic cleavage. Standard curves of CI were prepared by HPLC, and 1-β-CI-glucuronide was quantified according to the amount of CI formed by hydrolysis. The stability of conjugates greatty depends on pH and temperature, and the rate of degradation and isomerization is sensitive to the value of both. Lowering the pH from 7.4 to 5.0 and the temperature from 37°C to 18°C increased the stability of glucuronides. Increasing the pH to 12.0 results in very rapid acyl migration and hydrolysis.

Original languageEnglish
Pages (from-to)153-160
Number of pages8
JournalEuropean Journal of Drug Metabolism and Pharmacokinetics
Issue number3-4
Publication statusPublished - Jan 1 2000



  • Acyl-glucuronide
  • Ipriflavone

ASJC Scopus subject areas

  • Pharmacology
  • Pharmacology (medical)

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