Biotransformation of [14C]midazolam in the rat in vitro and in vivo.

1. Aerobic incubation of the maleate salt of midazolam, 8-chloro-6-(2-fluorophenyl)-1-methyl-4H-imidazo[1,5-a][1,4]benzodiazepine, with rat liver 9000 g supernatant fluid fortified with an NADPH-generating system yielded three metabolites. These were identified with an 1-hydroxymethyl derivative formed by oxidation of the methyl substituent on the imidazo moiety, the derivative hydroxylated at C-4 of the diazepine ring, and the corresponding dihydroxy derivative. 2. Pretreatment of rats with phenobarbital stimulated the oxidative metabolism in vitro of midazolam by 9000 g liver supernatant. 3. Rats given 2.5 mg/kg of [14C]midazolam maleate i.v. excreted 81% of the administered radioactivity in the faeces and 10% in the urine in 24 h. The highest levels of 14C were in the liver and the lowest in the brain during the first day. 4. Four phenolic derivatives of midazolam were identified in rat bile. These were the 4'-hydroxy and the 4'-hydroxy-1-hydroxymethyl derivatives of midazolam and two benzophenones. Evidence that the benzophenones were artifacts derived from the 4-hydroxy derivatives of the two biliary benzodiazepine metabolites is presented. These biliary metabolites were excreted as glucuronide and/or sulphate conjugates. 5. The Propensity of the rat to excrete phenolic metabolites of midazolam in bile is in agreement with the metabolism of several other benzodiazepines in this species.