The pharmacokinetics, tissue distribution and metabolism of caffeine were studied in male New Zealand White rabbits after an i.v. dose of 4 mg/kg. The mean (n = 4) distribution half-life was 0.2 hr and the mean elimination half-life was 3.8 hr. The mean clearance was 0.20 liters/kg/hr and the mean volume of distribution was 0.82 liters/kg. Concurrent samples of blood and semen from three rabbits, trained to ejaculate into an artificial vagina, were analyzed. The mean semen/blood concentration ratio of caffeine was 1.0. The concentrations of caffeine in the tissues of three rabbits were examined at 1 hr after an i.v. dose of 4 mg/kg. Most tissues exhibited a tissue/blood concentration ratio of approximately 1.0. Exceptions to this included fat, adrenals, liver and bile in which the ratios were 0.2, 0.6, 1.5 and 2.7, respectively. Urinary metabolites were investigated after an i.v. dose of 4 mg/kg of [14C]caffeine. The metabolites of caffeine were separated by high-pressure liquid chromatography and quantified by liquid scintillation counting. The major urinary metabolites of caffeine in the rabbit were 1-methylxanthine (22%), 1-methyluric acid (19%), 7-methylxanthine (16%) and 1,7-dimethylxanthine (14%).