Phenobarbital species differences

There may be species differences in the induction of drug metabolism in extra-hepatic tissues. For example, treatment of rabbits with phenobarbital significantly increases the hydroxylation and N-demethylation of N-methylaniline and the microsomal protein and cytochrome P-4S0 content of liver and kidney microsomes, but not in lung and small intestine. However, treatment of rats with phenobarbital, 3,4-benzpyrene, or DDT does not significantly affect the levels of cytochrome P-4S0 or drug metabolism in kidney microsomes. 

Recently Hansson and Wikvall studied 12 -hydroxylation in a reconstituted system consisting of highly purified cytochrome P-450 LM4 from rabbits [101]. The cytochrome P-450 fractions used were electrophoretically homogenous, but the cytochrome P-450 from starved rabbits had up to 4 times higher capacity to catalyse 12a-hydroxylation than had cytochrome P-450 from untreated, phenobarbital-treated, or -naphthoflavone-treated rabbits. It should be mentioned that treatment with /8-naphthoflavone increases the amount of cytochrome P-450 LM4 in the liver. Amino acid analyses, peptide-mapping experiments as well as absorption spectral and circular dichroism spectral analyses revealed physical differences between cytochrome P-450 LM4 preparations from starved and phenobarbital-treated animals. It was concluded that the cytochrome P-450 LM4 fraction was heterogenous and contained a species of cytochrome P-450 specific for 12a-hydroxylation. 

Conclusive evidence that a species of cytochrome P-450 was involved in the hydroxylation was presented by Okuda et al., who showed that the photochemical action spectrum for reversal of the carbon monoxide inhibition of 26-hydroxylation of 5)8-cholestane-3a,7a,12a-triol in rat liver exhibited a maximum at 450 nm [134]. Pedersen et al. [135] and Sato et al. [136] reported simultaneously that small amounts of cytochrome P-450 could be solubilized from the inner membranes of rat liver mitochondria that was active towards cholesterol as well as 5)8-cholestane-3a,7a,12a-triol in the presence of ferredoxin, ferredoxin reductase and NADPH. The mechanism of hydroxylation is thus the same as that operative in the biosynthesis of steroid hormones in the adrenals and in the la-hydroxylation of 25-hydroxyvitamin D in the kidney (Fig. 8). The liver mitochondrial cytochrome P-450 was not active in the presence of microsomal NADPH-cytochrome P-450 reductase [135,136]. Ferredoxin reductase as well as ferredoxin were active regardless of whether they were isolated from rat liver mitochondria or bovine adrenal mitochondria [133]. The partially purified cytochrome P-450 had a carbon monoxide difference spectrum similar to that of microsomal cytochrome P-450 from liver microsomes and adrenal mitochondria. In the work by Pedersen et al. [133], the concentration of mitochondrial cytochrome P-450 in rat liver mitochondria from untreated rats was calculated to be only about 0.1 nmole/mg protein. Treatment of rats with phenobarbital increased the specific content of cytochrome P-450 in the mitochondria more than 2-fold, without significant increase in the 26-hydroxylase activity. The carbon monoxide spectrum of the reduced cytochrome P-450 solubilized from liver mitochondria of phenobarbital-treated rats exhibited a spectral shift of about 2 nm as compared to the corresponding spectrum obtained in analysis of preparations from untreated rats. This was taken as evidence that more than one species of cytochrome P-450 was present in the preparation. It was later shown by Pedersen et al. [137] and Bjbrkhem et al. [138] that the preparation was also able to catalyse 25-hydroxylation of vitamin D3 and that different enzymes are involved in... 

---