Kinetic deuterium isotope effects on deamination and N-hydroxylation of cyclohexylamine by rabbit liver microsomes.
Deuterium isotope effects on the kinetic parameters for deamination and N-hydroxylation of cyclohexylamine (CHA) catalyzed by rabbit liver microsomes with NADPH are investigated. Both reactions are inhibited by carbon monoxide and have the characteristics of typical cytochrome P450-dependent monooxygenase reactions. A small and significant deuterium isotope effect operates in the oxidative deamination of CHA. The apparent isotope effects, i.e., VH/VD and (V/K)H/(V/K)D ratios for deamination, are 1.75 and 1.8-2.3, respectively. On the basis of N-hydroxylation, the VH/VD and (V/K)H/(V/K)D ratios are 0.8-0.9. The N-hydroxylation rate of alpha-deuterated CHA (D-CHA) is somewhat higher than that of CHA. The increased increment of hydroxylamine formation seems to coincide with the decreased amount of deamination. Substitution of deuterium in the alpha-position of CHA results in metabolic switching of cytochrome P450 from deamination to N-hydroxylation with low deuterium isotope effects. The data are interpreted in terms of an initial one-electron abstraction from the nitrogen to form an aminium cation radical followed by recombination with iron-bound hydroxyl radical leading to N-hydroxylamine, or followed by alpha-carbon deprotonation to form a neutral carbon radical. The latter can lead to a carbinolamine intermediate for deamination by way of imine or recombination with nascent iron-bound hydroxyl radical. The relative rates of the reactions depend on the alpha-carbon deprotonation rates of amines.
Kurebayashi
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Division of Medical Chemistry, National Institute of Hygienic Sciences, Tokyo, Japan.
D000818:Animals; D002248:Carbon Monoxide / Q000494:pharmacology; D002849:Chromatography, Gas; D003514:Cyclohexylamines / Q000378:metabolism*; D003641:Deamination; D003903:Deuterium; D008401:Gas Chromatography-Mass Spectrometry; D006900:Hydroxylation; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology / Q000378:metabolism*; D009249:NADP / Q000378:metabolism; D010084:Oxidation-Reduction / Q000187:drug effects; D010100:Oxygen / Q000494:pharmacology; D011817:Rabbits
D003514:Cyclohexylamines; D009249:NADP; D002248:Carbon Monoxide; D003903:Deuterium; D010100:Oxygen