1. The urinary excretion of a metabolite of illudin S after oral administration to rat has been studied. 2. From an ethyl acetate extract of urine, metabolite 1, a cyclopropane ring-cleavage compound, was identified by liquid chromatographic-mass spectrometric analysis 3. A hplc method for determination of the metabolite in rat urine was developed with extraction using Sep-Pak C18 cartridge followed by liquid-liquid extraction. 4. The metabolite excreted during 3 days after administration of illudin S to rat amounted to approximately 10-19% of the dose as free form and 3-5% of the dose as glucuronide.