Formation of hydroxy derivatives, aldehydes, and nitrite from N-nitrosomethyl-n-amylamine by rat liver microsomes and by purified cytochrome P-450 IIB1.
The metabolism was examined of the esophageal carcinogen N-nitrosomethylamylamine (NMAA) by liver microsomes and slices from adult male Sprague-Dawley rats. Hydroxylation at C-2 to C-5 of the amyl group to give stable hydroxy-NMAAs was studied by gas chromatography-thermal energy analysis to determine the products. Microsomal metabolism produced mainly 4-hydroxy-NMAA, proceeded optimally in 100 mM phosphate at pH 7.4, and showed no sex differences. Induction by phenobarbital (PB) and 3-methylcholanthrene produced effects which were similar in slices and microsomes, with PB inducing hydroxylation at all positions and 3-methylcholanthrene specifically inducing 3-hydroxylation by factors of 2- and 6-fold. Clofibrate and isoniazid treatments did not affect NMAA metabolism by liver slices. Aroclor-1254 strongly induced microsomal 2- and 3-hydroxylation. For 2- to 5-hydroxylation, Km values for uninduced microsomes were, respectively, 1.6, 1.2, 0.3, and 1.1 mM, with Vmax of 0.08, 0.26, 1.06, and 0.15 nmol/min/mg protein. With PB-induced microsomes, all 4 Km values were 0.4-0.7 mM. Liver microsomal production of nitrite and aldehydes from NMAA was determined colormetrically or (for pentaldehyde) by high-pressure liquid chromatography of the 2,4-dinitrophenylhydrazone. Uninduced microsomes produced nitrite, formaldehyde and pentaldehyde from 0.6 mM NMAA at rates that were, respectively, 0.15, 0.72, and 1.15 times that for 4-hydroxylation. PB especially induced depentylation, whereas 3-methylcholanthrene induced depentylation and denitrosation, but suppressed demethylation. A reconstituted system containing cytochrome P-450 IIB1 gave metabolite ratios similar to those in PB-induced microsomes. The results account for most of the possible primary metabolites of NMAA and demonstrate the selectivity for metabolism at each position.
Eppley Institute for Research in Cancer, University of Nebraska Medical Center, Omaha 68105.
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D000447:Aldehydes / Q000378:metabolism; D000818:Animals; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D006900:Hydroxylation; D008862:Microsomes, Liver / Q000378:metabolism*; D009573:Nitrites / Q000378:metabolism; D009602:Nitrosamines / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains
D000447:Aldehydes; D009573:Nitrites; D009602:Nitrosamines; C025033:N-amyl-N-methylnitrosamine; D003577:Cytochrome P-450 Enzyme System