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Titolo:
CHARACTERIZATION OF THE NADPH-DEPENDENT COVALENT BINDING OF [C-14] HALOTHANE TO HUMAN LIVER-MICROSOMES - A ROLE FOR CYTOCHROME P4502E1 AT LOW SUBSTRATE CONCENTRATIONS
Autore:
MADAN A; PARKINSON A;
Indirizzi:
UNIV KANSAS,MED CTR,DEPT PHARMACOL TOXICOL & THERAPEUT,CTR ENVIRONM &OCCUPAT HLTH KANSAS CITY KS 66160 UNIV KANSAS,MED CTR,DEPT PHARMACOL TOXICOL & THERAPEUT,CTR ENVIRONM &OCCUPAT HLTH KANSAS CITY KS 66160
Titolo Testata:
Drug metabolism and disposition
fascicolo: 12, volume: 24, anno: 1996,
pagine: 1307 - 1313
SICI:
0090-9556(1996)24:12<1307:COTNCB>2.0.ZU;2-M
Fonte:
ISI
Lingua:
ENG
Soggetto:
HALOTHANE-ASSOCIATED HEPATITIS; METABOLISM; 2E1; HEPATOTOXICITY; NEOANTIGENS; ANTIBODIES; RAT; IDENTIFICATION; DEFLUORINATION; EXPRESSION;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Citazioni:
28
Recensione:
Indirizzi per estratti:
Citazione:
A. Madan e A. Parkinson, "CHARACTERIZATION OF THE NADPH-DEPENDENT COVALENT BINDING OF [C-14] HALOTHANE TO HUMAN LIVER-MICROSOMES - A ROLE FOR CYTOCHROME P4502E1 AT LOW SUBSTRATE CONCENTRATIONS", Drug metabolism and disposition, 24(12), 1996, pp. 1307-1313

Abstract

Activation of halothane to trifluoroacetyl halide, followed by covalent binding to proteins (neoantigen formation) has been proposed to be the mechanism by which halothane causes immune hepatitis. The aim of this study was to identify the cytochrome P450 (CYP) enzyme primarily responsible for the NADPH-dependent covalent binding of [C-14]halothaneto human liver microsomes. Human liver microsomes were incubated in the absence and presence of NADPH with various concentrations of halothane (from 4.6 to 3,300 micrometers) to examine the effects of substrate concentration on the nonspecific and specific (NADPH-dependent) binding of [C-14]halothane to microsomal protein. As a function of substrate concentration, the specific binding of [C-14]halothane to human liver microsomes was biphasic, suggesting that the activation of halothane is catalyzed by a high-affinity enzyme(s) at low substrate concentrations (<150 micrometers) and by a low-affinity enzyme(s) at high substrate concentrations (>150 micrometers). For the high-affinity enzyme, the apparent K-M for the covalent binding of [C-14]halothane was approximately equal to 10 micrometers, and V-max was approximately equal to32 pmol equivalents of halothane bound/mg protein/min under conditions where covalent binding was directly proportional to incubation time and protein concentration. Ten individual samples of human liver microsomes were incubated with a low concentration of halothane (35 micrometers) to determine the sample-to-sample variation in the specific binding of [C-14]halothane to microsomal protein. Covalent binding ranged from 10 to 40 pmol equivalents of halothane bound/mg protein/min and was highly correlated (r(2)=0.93) with the sample-to-sample variation in chlorzoxazone 6-hydroxylase activity, which reflects the levels of CYP2E1. These results suggest that CYP2E1 is the high-affinity enzyme in human liver microsomes responsible for activating halothane to a reactive metabolite. This is supported by the observation that 4-methylpyrazole, a CYP2E1 inhibitor, inhibited the NADPH-dependent binding of [C-14]halothane to microsomal protein. The sample-to-sample variation in the covalent binding of [C-14]halothane at high substrate concentrations did not correlate with any known CYP enzyme activity. This suggests that several enzymes catalyze the oxidation of halothane at higher substrate concentrations. In conclusion, at pharmacologically relevantconcentrations, the covalent binding of halothane to human liver microsomes is primarily catalyzed by CYP2E1.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 28/09/20 alle ore 20:08:19