Objectives: To investigate the effects of grapefruit juice on absolute bioavailability of scopolamine in healthy subjects and to evaluate differences in pharmacokinetics and pharmacodynamics between genders. Subjects, material and methods: 14 healthy subjects (7 men and 7 women) received scopolamine 0.5 mg as intravenous infusion, and as oral ingestion with and without grapefruit juice on separate occasions. Serum and urine samples were analyzed using gaschromatography-ion trap tandem mass spectrometry. Changes in subjective state were determined up to 24 hours after drug administration. Results: After oral administration, pretreatment with grapefruit juice led to a 30% increase in systemic bioavailability of scopolamine (p = 0.005) and a significant increase in time to reach peak serum concentration (t(max)) of scopolamine (p < 0.001). The C(max) value (6.61 ± 0.63 ng/ml) of scopolamine after i.v. administration in male subjects was Significant higher compared with the value in female subjects (3.93 ± 0.04 ng/ml; geometric mean ± SEM; p = 0.007). No differences were found in urinary excretion rate of scopolamine and scopolamine glucuronide across genders and between the three different routes of scopolamine administration. Scopolamine produced timedependent decrements in subjective alertness while contentment and Calmness were not influenced. Conclusions: Pretreatment with grapefruit juice delayed the absorption and increased the bioavailability of scopolamine, whereas elimination was not significantly affected. This study identified an influence of gender on C(max) of scopolamine after i.v. infusion.