This double-blind, placebo-controlled, randomized, crossover study was designed to evaluate the effects of catechol-O-methyltransferase (COMT) inhibition by tolcapone on the pharmacokinetics of levodopa given as four different formulations of levodopa/benserazide: 50/12.5 mg, 100/25 mg, 200/50 mg (all standard release), or 100/25 mg (controlled release). Sixteen healthy volunteers, in two groups of 8, were given two different levodopa/benserazide formulations with and without tolcapone in random order on 4 days, each separated by a 7-day washout period. On each treatment day, 200 mg tolcapone or placebo (blinded) was taken orally 1 h before and 3 and 7 h after a single (unblinded) dose of levodopa/benserazide. All treatment combinations were well tolerated. Continuous inhibition of erythrocyte COMT activity by approximately 75% was observed over 13 h with tolcapone; this was unaffected by levodopa/benserazide formulation. Tolcapone had similar effects on plasma levodopa concentrations with the standard-release formulations: half-life and bioavailability increased approximately 2-fold compared with placebo, and maximum plasma concentration (Cmax) and time to Cmax (tmax) were unaffected, except for a slight increase in Cmax with the levodopa/benserazide 200/ 50 mg formulation. With the controlled-release formulation, tolcapone increased the levodopa area under the plasma concentration/time curve approximately 2-fold. Although levodopa tmax appeared delayed, the absorption phase was unaffected. Onset of levodopa effect is therefore not likely to be delayed when tolcapone is coadministered with this formulation. Regardless of levodopa/benserazide formulation, 3-O-methyldopa formation was reduced by 90% with tolcapone compared with placebo. These results show that tolcapone could potentiate the antiparkinsonian effects of levodopa independently of levodopa/benserazide formulation.