Cytochrome P450 inactivation during reductive metabolism of 1,1-dichloro-2,2,2-trifluoroethane (HCFC-123) by phenobarbital- and pyridine-induced rat liver microsomes.
The reductive metabolic activation of 1,1-dichloro-2,2,2-trifluoroethane (HCFC-123), one of the potential substitutes for the ozone-depleting chlorofluorocarbons and a close structural analogue of the hepatotoxic anesthetic halothane, was investigated in vitro. During incubation of liver microsomes from phenobarbital-(PB) or pyridine-induced (PYR) rats with 0-20 mM HCFC-123 under anaerobic conditions, a dose- and time-dependent depletion of added exogenous glutathione was observed, indicating the formation of reactive metabolites. Under similar incubation conditions, except for the absence of glutathione, 1-chloro-2,2,2-trifluoroethane and 1-chloro-2,2-difluoroethene were detected as products of reductive metabolism of HCFC-123, as previously reported for halothane. As shown previously in our laboratory for halothane, under these conditions HCFC- 123 also caused a statistically significant loss of microsomal cytochrome P450 (P450) as indicated by a decrease of the classical absorption spectrum in the presence of CO. Both glutathione depletion and P450 loss were almost completely prevented by previous saturation of the incubation mixture with CO and were partially prevented by the presence of the free-radical scavenger N-t-butyl-alpha-phenylnitrone or the carbene trapping agent 2,3-dimethyl-2-butene, suggesting that both types of intermediates may be involved. The loss of P450 was associated with a quantitatively similar loss of microsomal heme, as measured by the pyridine hemochromogen reaction, with PB but not with PYR microsomes. Finally, both the P4502E1-specific p-nitrophenol hydroxylase activity in PYR microsomes and the P4502B1/2-specific pentoxyresorufin O-depentylase activity in PB microsomes were significantly inhibited (58 and 53%, respectively) by prior incubation with HCFC-123, suggesting that both isoforms are able to catalyze the activation of this halogenated compound. These results indicate that indeed HCFC-123, like its analogue halothane, is activated reductively to reactive metabolites by at least two P450 isoforms, namely P4502E1 and P4502B1/2. These metabolites, probably free radicals and/or carbene species, may attack the enzyme resulting in modification of the heme group and subsequent loss of catalytic activity.
Ferrara
R
R
Institute of Occupational Medicine, University of Padua Medical School, Italy.
Tolando
R
R
King
L J
LJ
Manno
M
M
D000818:Animals; D017402:Chlorofluorocarbons / Q000378:metabolism*; D064227:Chlorofluorocarbons, Ethane; D019392:Cytochrome P-450 CYP2E1 / Q000378:metabolism; D065691:Cytochrome P-450 CYP2E1 Inhibitors; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism; D003573:Cytochrome b Group / Q000037:antagonists & inhibitors / Q000378:metabolism; D005609:Free Radicals; D008401:Gas Chromatography-Mass Spectrometry; D005978:Glutathione / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism; D008297:Male; D008862:Microsomes, Liver / Q000187:drug effects / Q000378:metabolism*; D010084:Oxidation-Reduction; D010634:Phenobarbital / Q000494:pharmacology*; D011725:Pyridines / Q000494:pharmacology*; D051381:Rats; D017208:Rats, Wistar; D013997:Time Factors
D017402:Chlorofluorocarbons; D064227:Chlorofluorocarbons, Ethane; D065691:Cytochrome P-450 CYP2E1 Inhibitors; D065607:Cytochrome P-450 Enzyme Inhibitors; D003573:Cytochrome b Group; D005609:Free Radicals; D007527:Isoenzymes; D011725:Pyridines; C067411:2,2-dichloro-1,1,1-trifluoroethane; D003577:Cytochrome P-450 Enzyme System; D019392:Cytochrome P-450 CYP2E1; D005978:Glutathione; C023666:pyridine; D010634:Phenobarbital