The biotransformation of minoxidil (2,4-diamino-6-piperidinopyrimidine 3-oxide) was studied in the rat, dog, and monkey and compared to reported results in the human. Chromatographic profiles of urinary metabolites show that each species excreted substantially the same metabolites but in quite different relative amounts. The monkey and the human exhibited similar metabolite profiles, whereas the dog and rat were quantitatively different from each other and from the monkey and human. The major excretory product for the monkey and human was a glucuronide conjugate of minoxidil. Substantially smaller amounts of unchanged minoxidil, 2,4-diamino-6-(4'-hydroxypiperidino)pyrimidine 3-oxide, and more polar metabolites also were excreted by these two species. The major excretory product in the rat was unchanged minoxidil. Almost as much (combined) of the two acidic metabolites, 2,4-diamino-6-(4'-carboxy-n-butylamino)pyrimidine and its 3-oxide, also were produced. Smaller amounts of the glucuronide of minoxidil, 2,4-diamino-6-(4'-hydroxypiperidino)pyrimidine 3-oxide, its 3'-hydroxy isomer, and 2,4-diamino-6-piperidinopyrimidine also were excreted by the rat. The major metabolite of minoxidil excreted by the dog was the 4'-hydroxy metabolite. Smaller amounts of unchanged minoxidil and polar metabolites and much smaller amounts of the glucuronide of minoxidil, the 3'-hydroxy metabolite, and 2,4-diamino-6-piperidinopyrimidine also were excreted by the dog. Evidence was obtained for a glucuronide conjugate of the 4'-hydroxy metabolite in this species. The major circulatory material in dog plasma was the 4'-hydroxy metabolite, whereas it was the glucuronide of minoxidil in monkey plasma.