Excretion of pentachlorophenol-14C in the urine of rats and mice after oral and intraperitoneal administration (10 to 25 mg/kg) was studied. 41 to 43% of the excretion activity was found to be present as unchanged pentachlorophenol. By means of gas chromatography-mass spectrometry, one metabolite was identified as tetrachlorohydroquinone, representing 5 and 24% of the excreted activity in rats and mice, respectively. Conjugation with glucuronic acid could not be demonstrated by enzymatic hydrolysis since tetrachlorohydroquinone was found to be a potent inhibitor of β-glucuronidase, the I50 being 1.6 to 2.0×10−6 M. Boiling the urine with hydrochloric acid was shown to convert all of the excreted activity to pentachlorophenol and tetrachlorohydroquinone, the latter representing 43 and 46% in rats and mice, respectively. Tetrachlorohydroquinone was found in the urine of workers occupationally exposed to pentachlorophenol.