Rabbit liver microsomal system: study of interaction with two model N-nitrosamines and their metabolism.
Rabbit liver microsomes of control (non-treated) or animals induced either by ethanol (EtOH) or phenobarbital (PB) were incubated with N-nitrosodimethylamine (NDMA) or N-nitrosomethylaniline (NMA). Difference spectroscopy showed that NMA is bound to the substrate-binding site of cytochrome P-450 (CYP) isoforms as heme ligand in control and EtOH pre-treated microsomes. On the other hand, PB-induced microsomes exhibit with NMA substrate type of spectra. NDMA does not provide any type of binding spectra with used microsomal systems. Oxidative bio-activation of N-nitrosamines by the microsomal CYP isoforms was measured as formaldehyde formation. Analysis of reaction kinetics in control microsomes revealed, for both substrates, two values of Michaelis-Menten constant (K(m)) for, K(m) values of 0.03 and 0.13 mmol/l for NDMA, and 0.30 and 0.82 mmol/l for NMA. Induction of animals with EtOH resulted in a decrease in the K(m) value for both substrates. In contrast, PB treatment caused an elevation of K(m) value for NDMA. Based on these data, we conclude that EtOH-inducible microsomal CYP isoforms (mainly CYP2E1) are responsible for binding and N-demethylation metabolism of both studied N-nitrosamines in rabbit liver microsomal system. The role of the other CYP isoforms involved in the metabolism of mentioned N-nitrosamines is discussed.
Sulc
M
M
Department of Biochemistry, Faculty of Science, Charles University, Hlavova 2030, 128 43 Prague 2, Czech Republic. mis@natur.cuni.cz
Kubícková
B
B
Máslová
V
V
Hodek
P
P
D000818:Animals; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004128:Dimethylnitrosamine; D000431:Ethanol / Q000008:administration & dosage*; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D008862:Microsomes, Liver / Q000187:drug effects / Q000378:metabolism*; D009602:Nitrosamines / Q000378:metabolism*; D010634:Phenobarbital / Q000008:administration & dosage*; D011817:Rabbits
D009602:Nitrosamines; D000431:Ethanol; D003577:Cytochrome P-450 Enzyme System; C014102:N-methyl-N-nitrosoaniline; D004128:Dimethylnitrosamine; D010634:Phenobarbital