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Mutagenesis vol. 12 no. 4 pp. 245-254, 1997
© 1997 UK Environmental Mutagen Society/Oxford University Press

research-article

Characterization of enzyme activities and cofactors involved in bioactivation and bioinactivation of chemical carcinogens in the tester strains Escherichia coli K12 MX100 and Salmonella typhimurium LT2 TA100

M. Kranendonk1,2, J.N.M. Commandeur1, A. Laires3, J. Rueff2 and N.P.E. Vermeulen1,4

1Leiden/Amsterdam Center for Drug Research (LACDR), Department of Pharmacochemistry, Division of Molecular Toxicology, Vrije Universiteit De Boelelaan 1083, 1081 HV Amsterdam, The Netherlands 2department of Genetics, Faculty of Medical Sciences, New University of Lisbon Rua de Junqueira 96, 1300 Lisboa 3Faculty of Sciences and Technology, New University of Lisbon P-2825 Monte de Caparica, Portugal

MX100 is an Escherichia coli K12 genotoxicity tester strain, especially developed for mechanistic studies of chemical mutagens and carcinogens. For the study of the role of specific enzymes in the bioactivation and bioinactivation of carcinogens, it is necessary to characterize MX100 as far as its metabolic bio(in)activation capacities are concerned. In this study such a characterization is performed in two types of cell-free lysates, one derived from stationary phase cells, grown in rich medium (SR-lysates) and one from exponentially growing cells (log phase), cultured in minimal medium (LM-lysates). Six Phase I enzyme activities of aromatic NADPH hydroxylase, NADH hydroxylase, flavin-containing monooxygenase (FMO), nitroreductase, DT-diaphorase and NADPH ferredoximoxidoreductase were determined. Activities of six Phase II enzymes glutathione S-transferases (GSTs), N-aryl acetyltransferase (NAT), arylamine sulphotransferase, UDPglucuronyltransferase and epoxide hydratase and of the Phase III enzyme cysteine conjugate (J-lyase were subsequently assessed. In addition, five antioxidant enzymes: superoxide dismutase (SOD), catalase, glutathione (GSH)-reductase, GSH-peroxidase and alkyl hydroperoxide reductase; as well as concentrations of glutathione (GSH) and its disulphide (GSSG), were measured. The activity parameters of all enzymes were compared with those obtained in similar lysates of the Salmonella strain TA100 and in rat liver preparations. The results indicate that MX100 as well as TA100 contain relatively low oxidative but high reductase Phase I activities. Both strains demonstrated low activities for the Phase II conjugation enzymes except for GSTs. In MX100, relatively high activities were detected for all antioxidative enzymes, activities which were lower in TA100. Significant differences in activities were observed between the SR-lysates derived from stationary phase/rich medium and LM-lysates from log phase/minimal medium cells for nitroreductase, GST, SOD, catalase, NADPH ferredoxin: oxidoreductase as well as in GSH content. In general, we described for the first time a metabolic characterization of the E.coli tester strain MX100 and the Salmonella typhimurium strain TA100 and discussed the results in terms of its significance for carcinogen bioactivation and bioinactivation capacities.

4To whom correspondence should be addressed


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