Effect of 2,4-dinitrophenol on the oxidative metabolism of hexobarbital by cytochrome P-450 in perfused rat liver.

Abstract

The effect of 2,4-dinitrophenol (2,4-DNP) on the oxidative metabolism of hexobarbital by cytochrome P-450 was investigated in perfused rat liver. In the livers from fed, phenobarbital (PB)-treated rats, 2,4-DNP (50 microM) had no effect on the redox state of cytochrome P-450 or on oxygen uptake during mixed-function oxidation of hexobarbital. In the livers from fasted, PB-treated rats, 2,4-DNP (50 microM) significantly decreased the amount of reduced (oxygenated) cytochrome P-450 and the drug-induced oxygen uptake by about 50%. 2,4-DNP caused a decrease of metabolites of hexobarbital in perfusate, in the fasted but not in the fed state. These results suggest that in fed, PB-treated rats NADPH for mixed-function oxidation of hexobarbital can be predominantly supplied from an extramitochondrial source (most probably via the cytosolic pentose phosphate shunt), but in fasted, PB-treated rats, about 50% of the NADPH required for the mixed-function oxidation is supplied from an intramitochondrial source. In the livers from PB-treated rats, infusion of sorbitol (4 mM), a glycogenic substrate in fasted rats, stimulated the rate of drug-induced oxygen uptake and the steady-state level of reduced (oxygenated) cytochrome P-450 increased during mixed-function oxidation of hexobarbital. These effects of sorbitol were almost completely abolished in the presence of 2,4-DNP. Complete inhibition of gluconeogenesis was also observed in the livers from fasted, PB-treated rats in the presence of 2,4-DNP (50 microM). The amount of metabolites of hexobarbital in the perfusate was increased by the addition of sorbitol in the fasted but not in the fed state. The effect of sorbitol on drug metabolism was inhibited by 2,4-DNP. These data may be explained by assuming that ATP is required for the conversion of sorbitol to metabolites (e.g. glucose-6-phosphate) which can produce NADPH in the cytosol.