Quantitation of hemoproteins in rat small intestinal mucosa with identification of mitochondrial cytochrome P-450.

Hemoprotein content of isolated rat small intestinal mucosa cells was studied by optical spectroscopy.In- doleamine 2,3-&oxygenase, and cytochromes u + as, c + el, bael + ba, and P-450 accounted for essentially aIl of the heme after correction for contamination by hemoglobin.Catalase, methemoglobin, myoglobin, and cytochrome P-420 were not detectable.Mitochondrial cytochromes accounted for about 60 to 65% of the cellular heme, i n d o l e ~i n e 2,3-&oxygenase for about 308, and cytochromes P-450 and bs for most of the remainder.Addition of tryptophan to the cells produced a spectral change that was characterized by an absorbance maximum at 418 nm that was similar qualitatively and quantitatively to the previously reported oxygenated indoleamine dioxygenasetryptophan complex spectrum.Cytochrome P-450 in control cells was found to be largely in the mitochondrial fraction.Pretreatment with 3-methylcholanthrene increased the microsomal cytochrome P-450 content without effect on the mitochondrial content.Assay for benzo(u)pyrene metabolism in the two fractions from control cells indicated that it was not a substrate for the mitochondrial cytochrome P-450 and was a poor substrate for the microsomal enzyme.Pretreatment with 3-methylcholanthrene greatly stimulated the microsomal activity with no effect on the mitochondrial activity.A comparison of estimated turnover numbers of the 3-methylcholanthrene-induced forms of cytochrome P-450 in intestine and liver indicates that the forms are comparable.