A randomized, single-center, open-label study of posaconazole (POS) was performed to determine the concentration of POS in the skin of 30 healthy adult human subjects receiving 400 mg POS oral suspension twice daily for 8 days with a high-fat meal. Blood samples for plasma POS level determination were collected at prespecified times on day 1 and day 8. From each subject, two 4-mm skin punch biopsy samples were obtained, one immediately before or after both the first and last doses of POS. A MIC(90) value of 250 ng/ml, which encompasses the majority of common dermatophytes, was used to calculate the time above the MIC(90) in plasma and skin. On days 1 and 8, POS attained peak plasma concentrations at median times of 8 and 5 h, respectively. On days 1 and 8, POS peak skin concentrations were attained at 12 and 3 h, respectively; peak skin concentrations were produced from a single composite profile. On day 8, POS concentrations in skin and plasma for the entire dosing interval were severalfold higher than the MIC(90). POS dosed at 400 mg twice daily per os was well tolerated in healthy subjects. Two subjects reported increased alanine aminotransferase (ALT) levels. The findings of this study demonstrate adequate skin penetration and have certain implications for the treatment of dermatophytic skin and nail infections.