RAT HEPATIC-MICROSOMAL METABOLISM OF ETHYLENETHIOUREA - CONTRIBUTIONS OF THE FLAVIN-CONTAINING MONOOXYGENASE AND CYTOCHROME-P-450 ISOZYMES

被引:22
作者
DECKER, CJ [1 ]
DOERGE, DR [1 ]
机构
[1] UNIV HAWAII,DEPT ENVIRONM BIOCHEM,HONOLULU,HI 96822
关键词
D O I
10.1021/tx00022a013
中图分类号
R914 [药物化学];
学科分类号
100701 ;
摘要
The contributions of the rat hepatic flavin-containing monooxygenase (FMO) and cytochrome P-450 isozymes (P-450) in the ethylenethiourea (ETU) mediated inactivation of P-450 isozymes and covalent binding of the compound to microsomal proteins were investigated. In vitro, ETU was found to inhibit P-450 marker activities in microsomes obtained from untreated (UT) and phenobarbital (PB), beta-naphthoflavone (BNF), and dexamethasone (DEX) pretreated rats. This inhibition was dependent on the presence of NADPH and was completely abolished by coincubation with glutathione (GSH). Heat treatment of microsomes prior to ETU-mediated P-450 inactivation led to diminished loss of P-450 marker activities in microsomes obtained from UT and PB-pretreated, but not BNF- or DEX-pretreated rats, suggesting FMO involvement in the inactivation of some P-450 isozymes. Covalent binding of [C-14]ETU to microsomal proteins was found to be NADPH-dependent and enhanced with BNF or DEX pretreatment of rats. This binding was completely inhibited by coincubation with GSH. Heat treatment of microsomes and P-450 inactivation studies indicated a predominant role of FMO in the observed covalent binding. Addition of the sulfhydryl reagents dithiothreitol (DTT) or GSH after the incubation of microsomes, [C-14]ETU, and NADPH resulted in the complete release of bound ETU, suggesting the reduction of disulfide bonds between oxidized ETU and protein sulfhydryls. Microsomal heme content was not decreased following incubation of microsomes with ETU and NADPH, and P-450 appeared to be converted to P-420. Metabolism of ETU in the presence of GSH resulted in the formation of oxidized glutathione (GSSG), suggesting the initial formation of a GSH-ETU adduct and subsequent disulfide exchange reaction. These results suggest that, under normal physiological conditions, reactive metabolites from ETU generated by either FMO or P-450 are preferentially trapped by endogenous GSH and do not interact with cellular targets.
引用
收藏
页码:482 / 489
页数:8
相关论文
共 46 条
[1]   REACTIVE INTERMEDIATES .I. SYNTHESIS AND OXIDATION OF 1- AND 2-AMINOBENZOTRIAZOLE [J].
CAMPBELL, CD ;
REES, CW .
JOURNAL OF THE CHEMICAL SOCIETY C-ORGANIC, 1969, (05) :742-&
[2]   A CONVENIENT RADIOMETRIC ASSAY FOR FLAVIN-CONTAINING MONOOXYGENASE ACTIVITY [J].
CASHMAN, JR .
ANALYTICAL BIOCHEMISTRY, 1987, 160 (02) :294-300
[3]   ROLE OF THE MICROSOMAL FAD-CONTAINING MONOOXYGENASE IN THE LIVER TOXICITY OF THIOACETAMIDE S-OXIDE [J].
CHIELI, E ;
MALVALDI, G .
TOXICOLOGY, 1984, 31 (01) :41-52
[4]   COVALENT BINDING OF N-HYDROXY-N-ACETYL-2-AMINOFLUORENE AND N-HYDROXY-N-GLYCOLYL-2-AMINOFLUORENE TO RAT HEPATOCYTE DNA - INVITRO AND CELL-SUSPENSION STUDIES [J].
CORBETT, MD ;
LIM, LO ;
CORBETT, BR ;
JOHNSTON, JJ ;
WIEBKIN, P .
CHEMICAL RESEARCH IN TOXICOLOGY, 1988, 1 (01) :41-46
[5]   STUDIES OF METABOLISM OF CARBON-DISULFIDE BY RAT-LIVER MICROSOMES [J].
DALVI, RR ;
POORE, RE ;
NEAL, RA .
LIFE SCIENCES, 1974, 14 (09) :1785-1796
[6]  
DALVI RR, 1975, CHEM-BIOL INTERACT, V10, P347, DOI 10.1016/0009-2797(75)90057-5
[7]   OXIDATIVE-METABOLISM OF SPIRONOLACTONE - EVIDENCE FOR THE INVOLVEMENT OF ELECTROPHILIC THIOSTEROID SPECIES IN DRUG-MEDIATED DESTRUCTION OF RAT HEPATIC CYTOCHROME-P450 [J].
DECKER, CJ ;
RASHED, MS ;
BAILLIE, TA ;
MALTBY, D ;
CORREIA, MA .
BIOCHEMISTRY, 1989, 28 (12) :5128-5136
[8]  
DECKER CJ, UNPUB FORMATION GLUT
[9]   AUTOCATALYTIC ALKYLATION OF THE CYTOCHROME-P-450 PROSTHETIC HEME GROUP BY 1-AMINOBENZOTRIAZOLE - ISOLATION OF AN NN-BRIDGED BENZENE-PROTOPORPHYRIN IX ADDUCT [J].
DEMONTELLANO, PRO ;
MATHEWS, JM .
BIOCHEMICAL JOURNAL, 1981, 195 (03) :761-764
[10]  
DEMONTELLANO PRO, 1988, PROG DRUG METAB, V2, P99