Glucuronides of drugs often accumulate during long term therapy. The hydrolysis of glucuronides can be catalysed by beta-glucuronidase, an enzyme expressed in many tissues and body fluids in humans. The possible contribution of beta-glucuronidase to drug disposition in humans has not been assessed in a systematic manner, but this enzyme may be able to release, locally or systemically, the active or inactive parent compound from drug glucuronides, thereby modifying the disposition and action of these drugs. Based on the information available on the localisation, expression and variability of beta-glucuronidase, the concept of beta-glucuronidase-mediated drug metabolism is outlined in this article using examples from the literature. Since some issues surrounding the beta-glucuronidase-mediated deconjugation of drug glucuronides still need to be clarified in humans, additional data from animal models supporting this concept have been included. Moreover, as beta-glucuronidase has already been proven to be useful in tumour specific bioactivation of glucuronide prodrugs of anticancer agents, we also focus on anticancer prodrug approaches utilising beta-glucuronidase. This review summarises the role of beta-glucuronidase in drug disposition and drug targeting in humans.