methylnitronitrosoguanidine and 2-nitrofluorene

The Escherichia coli mu operon was subcloned into a pKK233-2 vector containing rat glutathione S-transferase (GST) 5-5 cDNA and the plasmid thus obtained was introduced into Salmonella typhimurium TA1535. The newly developed strain S.typhimurium NM5004, was found to have 52-fold greater GST activity than the original umu strain S.typhimurium TA1535/pSK1002. We compared sensitivities of these two tester strains, NM5004 and TA1535/pSK1002, for induction of umuC gene expression with several dihaloalkanes which are activated or inactivated by GST 5-5 activity. The induction of umuC gene expression by these chemicals was monitored by measuring the cellular beta-galactosidase activity produced by umuC"lacZ fusion gene in these two tester strains. Ethylene dibromide, 1-bromo-2-chloroethane, 1,2-dichloroethane, and methylene dichloride induced umuC gene expression more strongly in the NM5004 strain than the original strain. 4-Nitroquinoline 1-oxide and N-methyl-N'-nitro-N-nitrosoguanidine were found to induce umuC gene expression to similar extents in both strains. In the case of 1-nitropyrene and 2-nitrofluorene, however, NM5004 strain showed weaker umuC gene expression responses than the original TA1535/pSK1002 strain. 1,2-Epoxy-3-(4'-nitrophenoxy)propane, a known substrate for GST 5-5, was found to inhibit umuC induction caused by 1-bromo-2-chloroethane. These results indicate that this new tester NM5004 strain expressing a mammalian GST theta class enzyme may be useful for studies of environmental chemicals proposed to be activated or inactivated by GST activity.

Topics: 4-Nitroquinoline-1-oxide; Animals; Biotransformation; Ethane; Ethylene Dibromide; Ethylene Dichlorides; Fluorenes; Gene Expression; Genetic Vectors; Glutathione Transferase; Hydrocarbons, Halogenated; Methylene Chloride; Methylnitronitrosoguanidine; Mutagenicity Tests; Operon; Pyrenes; Rats; Salmonella typhimurium; Transfection; Transformation, Bacterial

The mutational specificities of various chemical mutagens were compared in isogenic E. coli strains with different DNA repair capabilities (wild-type, uvrA, umuC, and uvrA umuC) in a reversion assay employing a set of mutant lacZ genes that can detect two types of transitions, four types of transversions, and five kinds of specific frameshift events. A uvrA derivative was more sensitive than the wild-type strain to 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone for +1G, -1G, -2(C-G), +1A and -1A frameshifts, G.C-->A.T transitions, and G.C-->T.A transversions. In a uvrA background, G.C-->T.A transversions and +1G, +1A, and -1A frameshifts appeared to be umuC-dependent, while G.C-->A.T transitions were not. N-Ethyl-N'-nitro-N-nitrosoguanidine was more mutagenic in a uvrA background for five kinds of frameshifts and G.C-->A.T transitions, but not for G.C-->T.A, A.T-->C.G, and A.T-->G.C base substitutions. A.T-->C.G transversions were totally dependent on umuC gene function. For the investigation of mutational specificities induced by frameshift mutagens, an rfa mutation was additionally introduced. The rfa strain responded to 2-nitrofluorene, which induced primarily -2(C-G) frameshift mutations. In an rfa uvrA background, benzo[a]pyrene induced +1G, -1G, +1A, and -1A frameshifts. 2-Aminoanthracene induced +1G, -1G, and +1A, but not -1A, frameshifts, with -1G frameshifts predominating. Ethidium bromide induced only two types of frameshifts, -1G and +1A. Frameshifts induced by ICR-170 were independent of umuC gene function, while those by induced 1-nitropyrene were partly umuC-dependent.

Topics: Adenosine Triphosphatases; Aminoacridines; Anthracenes; Bacterial Proteins; Benzo(a)pyrene; beta-Galactosidase; DNA Ligases; DNA Repair; DNA-Binding Proteins; DNA-Directed DNA Polymerase; Escherichia coli; Escherichia coli Proteins; Ethidium; Fluorenes; Frameshift Mutation; Furans; Genes, Bacterial; Glycosyltransferases; Lac Operon; Methotrexate; Methylnitronitrosoguanidine; Mutagenesis; Mutagenicity Tests; Mutagens; Nitrogen Mustard Compounds; Point Mutation; Pyrenes; Species Specificity; Suppression, Genetic

A heat-resistant factor in ethanol extracts of the fungus Craterellus cornucopioides completely inhibited the mutagenicity of aflatoxin B1, benzo[a]pyrene, the acridine half mustard ICR-191 and 2-nitrofluorene in a forward-mutation system using Salmonella typhimurium TM677 (screening for 8-azaguanine resistance). There was no inhibitory effect on the mutagenic activity of 4-nitroquinoline-N-oxide, methyl methanesulfonate or N-methyl-N'-nitro-N-nitrosoguanidine. Experiments performed to elucidate the mechanism of the antimutagenic effect showed that neither an alteration of cell viability nor an interference with the excision-repair and the inducible SOS-repair system was involved. The conceivable mechanisms for the antimutagenicity of the ethanol extract include direct chemical interaction with the mutagen and/or inhibition of the activation process in the case of the promutagens. The antimutagenic activity of Craterellus cornucopioides is not unique among mushroom species. The ethanol extracts of 6 other mushrooms showed a similar antimutagenic activity.

Topics: 4-Nitroquinoline-1-oxide; Aflatoxin B1; Aflatoxins; Aminacrine; Basidiomycota; Benzo(a)pyrene; DNA Repair; Ethanol; Fluorenes; Hot Temperature; Methyl Methanesulfonate; Methylnitronitrosoguanidine; Mutagenicity Tests; Mutation; Nitrogen Mustard Compounds

Since its development by Dr. Bruce Ames and his colleagues more than a decade ago, the Salmonella/mammalian microsome mutagenicity assay has become a widely accepted tool to assist in the identification of chemicals with mutagenic and carcinogenic potential. Several automated approaches to Salmonella testing have been proposed in recent years but have failed to gain acceptance in the scientific community due to poor performance or lack of demonstrated usefulness. In this paper we report on an automated system that successfully generates dose-response data and, moreover, reduces the labor, materials, and sample mass required to obtain such information. In the standard plate-incorporation assay, dose-response relationships are defined by testing discrete doses of the test agent on a series of agar plates. In contrast, the spiral Salmonella assay generates dose-response data from a continuous concentration gradient on a single agar plate. Upon analysis, each spiral plate yields a dose-response curve consisting of 13 data points that span a concentration range of about 15:1, which is equivalent to 5 two-fold serial dilutions. The performance of the spiral Salmonella assay was compared to that of the conventional plate-incorporation assay using 13 mutagens and 7 nonmutagens selected from a variety of chemical classes. Concordant qualitative responses were obtained for all compounds tested, and comparable dose-response relationships were generated by all mutagens with the exception of sodium azide and cyclophosphamide, which are highly water-soluble and, thus, are unable to maintain a well-defined concentration gradient on a spiral plate due to rapid diffusion. In general, toxicity was expressed at a lower dose in the spiral assay, and the mutagenic potencies (slopes of the dose-response curves) were greater in the spiral assay relative to the plate-incorporation assay. These differences will be discussed, as will the applicability of the spiral plating technique to routine screening and its relevancy to future mutagenesis testing.

Topics: 4-Nitroquinoline-1-oxide; Anthracenes; Biotransformation; Fluorenes; Image Processing, Computer-Assisted; In Vitro Techniques; Methylnitronitrosoguanidine; Mutagenicity Tests; Mutation; Salmonella typhimurium; Time Factors

The two secondary biliary acids (lithocholic and deoxycholic acids) were co-mutagenic when they were each co-incubated with dimethylhydrazine in the presence of Salmonella typhimurium TA 100. These observations were extended to other toxic chemicals, acting as direct (N-methyl-N'nitro-N-nitrosoguanidine and 2-nitrofluorene) or indirect (2-acetylaminofluorene) mutagens. Lithocholic and deoxycholic acids show a similar behavior towards genotoxic molecules. Nevertheless two differences must be noted. Lithocholic acid was the stronger co-mutagen. When lithocholic acid inhibited mutagenic activity of 2-nitrofluorene, deoxycholic acid did not modify it. Interactions between the two secondary bile acids occurred so that the co-mutagenic activity of the mixture of these two bile acids depended on the ratio of their concentrations. Besides their cancer-promoting effect, biliary acids also could have co-initiating effect that could depend upon the ratio of their concentration in the intestine. Calculating the ratio of fecal concentrations of deoxycholic/lithocholic acids thus could be a more sensitive index for cancer risk than simply measuring the fecal concentration of the two biliary acids taken separately.

Topics: 2-Acetylaminofluorene; Animals; Bile Acids and Salts; Colonic Neoplasms; Drug Synergism; Feces; Fluorenes; Male; Methylnitronitrosoguanidine; Mutagenicity Tests; Mutagens; Rats; Rats, Inbred Strains; Risk; Salmonella typhimurium

Topics: 2-Acetylaminofluorene; Acetylcysteine; Animals; Anthracenes; Azides; Benzo(a)pyrene; Biotransformation; Dose-Response Relationship, Drug; Fluorenes; Methylnitronitrosoguanidine; Microsomes, Liver; Mutagenicity Tests; Mutagens; Rats; Salmonella typhimurium; Sodium Azide