Tissue, species, and substrate concentration differences in the position-selective hydroxylation of N-nitrosodibutylamine. Relationship to the distribution of cytochrome P-450 isozymes 2 (IIB) and 5 (IVB).
The relative rates of 3- and 4-hydroxylation of N-nitrosodibutylamine in microsomal preparations depend upon species, tissue, and substrate concentration. With rabbits, at a substrate concentration of 200 microM, the two reactions differ by less than 30% in preparations from liver, lung, or intestine, whereas the ratio of 4- to 3-hydroxylation is 2.0 with bladder and only 0.4 with kidney. As the substrate concentration is lowered, this ratio increases to a maximum of about 2.0 in hepatic and pulmonary preparations. The sum of the rates of 3- and 4-hydroxylation of N-nitrosodibutylamine in microsomal preparations from untreated rabbits is highest in those from lung (about 2-fold greater than liver). Following treatment of rabbits with phenobarbital, however, the highest rate is with hepatic microsomes (about 2-fold greater than pulmonary microsomes). Antibodies to cytochrome P-450 isozymes 2 (IIB) and 5 (IVB) together inhibit greater than 90% of the microsomal 3- and 4-hydroxylation of N-nitrosodibutylamine. The ratio of 4- to 3-hydroxylation with antibodies to isozyme 2 present is the same (2.0) as that obtained with purified isozyme 5, and the ratio with antibodies to isozyme 5 present is the same (0.2) as that with purified isozyme 2. The Km with isozyme 5 is less than 10 microM, whereas the Km with isozyme 2 is 55 microM, a difference that explains why position selectivity in microsomal incubations is dependent upon substrate concentration. Also, differences in the relative and absolute rates of hydroxylation among various tissues reflect differences in the contents of isozymes 2 and 5.(ABSTRACT TRUNCATED AT 250 WORDS)
Schulze
J
J
Laboratory of Cellular and Molecular Pharmacology, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, NC 27709.
Richter
E
E
Philpot
R M
RM
D000818:Animals; D006224:Cricetinae; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D006168:Guinea Pigs; D006900:Hydroxylation; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000378:metabolism*; D007700:Kinetics; D008647:Mesocricetus; D051379:Mice; D008810:Mice, Inbred C57BL; D008861:Microsomes / Q000201:enzymology; D008862:Microsomes, Liver / Q000201:enzymology; D009602:Nitrosamines / Q000378:metabolism* / Q000493:pharmacokinetics; D009928:Organ Specificity; D011817:Rabbits; D051381:Rats; D013045:Species Specificity; D013379:Substrate Specificity
D007527:Isoenzymes; D009602:Nitrosamines; C012279:dibutylnitrosamine; D003577:Cytochrome P-450 Enzyme System