A high performance liquid chromatographic (HPLC) assay for unchanged paracetamol and its glucuronide, sulphate, cysteine and mercapturic acid conjugates in the urine of man, mouse and rat is described. The method is simple, rapid and reproducible. The metabolite assay has been used to characterize the male C3H mouse, which shows sensitivity to paracetamol toxicity similar to man, as a model for paracetamol metabolism studies. In male C3H mice there was no evidence to suggest saturability of the glucuronidation pathway on increasing the paracetamol dose from 50 to 300 mg/kg. By contrast, the metabolic ratio and fractional excretion of both the sulphate and glutathione-derived conjugates decreased with increasing paracetamol dose. For animals administered a 200 mg/kg dose of paracetamol, pretreatment with phenobarbitone or 3-methylcholanthrene increased the fractional excretion and metabolic ratio of the glutathione-derived and glucuronic acid conjugates. Piperonyl butoxide pretreatment of animals administered the same dose of paracetamol inhibited glutathione and glucuronic acid conjugation.