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The genotoxicity of organotin compounds in SOS chromotest and rec-assay.

In these days pollution by organotin compounds in the environment extends widely and effects on human health are feared. We studied the genotoxicity of various organotin compounds (butyltins, phenyltins, methyltins) and inorganic tin (SnCl4), which are present in the environment, with the SOS chromotest and the rec-assay. Mono-n-butyltin oxide, n-butyltin trichloride and di-n-butyltin dichloride showed high SOS-inducing potency in the SOS chromotest with Escherichia coli PQ37. Di-n-butyltin dichloride, tri-n-butyltin chloride, bis(tri-n-butyltin)oxide, dimethyltin dichloride and trimethyltin chloride were recognized as genotoxic chemicals by the rec-assay.

4-Nitroquinoline-1-oxide↗

Genotoxicity of polycyclic aromatic hydrocarbons in Escherichia coli PQ37.

In the present investigation, 32 polycyclic aromatic hydrocarbons (PAHs) were tested for genotoxicity in E. coli PQ37 using the standard tube assay of the SOS chromotest. PAHs such as benzo[ghi]fluoranthene, benzo[j]fluoranthene, benzo[a]pyrene, chrysene, dibenzo[a,l]pyrene, fluoranthene and triphenylene exhibited high genotoxicity when incubated in the presence of an exogenous metabolic activation mixture. The results were compared to those obtained with the Salmonella/microsome test.

Alkaline Phosphatase↗

Genotoxicity assessment of waste products of aluminum plasma etching with the SOS chromotest.

We evaluated 36 characteristic waste products from the plasma etching of aluminum for genotoxicity with the SOS chromotest. The majority of the samples showed genotoxic activity in tester strain Escherichia coli PQ37 without metabolic activation using S9 mix. In the presence of S9, a deactivation of the samples was regularly observed. Comparable studies with the Salmonella/microsome (Ames) test using tester strains Salmonella typhimurium TA98 and TA100 indicated actual mutagenicity of waste products. Gas chromatograms of the organic constituents of all waste products were performed in parallel with the genotoxicity assays. In contrast to the similarity of the peak patterns of all chromatograms, the biological effects of individual waste samples showed large differences. Information on chemical composition and the SOS chromotest results of a representative sample recovered over a period of 2 years is given. For this sample, the influences of sample preparation and cytotoxic matrix effects on the test parameters are also shown.

Aluminum↗

Study of the genotoxic activity of five chlorinated propanones using the SOS chromotest, the Ames-fluctuation test and the newt micronucleus test.

Three short-term assays (the SOS chromotest, the Ames-fluctuation test and the newt micronucleus test) were carried out to evaluate the genotoxicity of five chlorinated propanones identified in several chlorinated waters (monochloropropanone, 1,1-dichloropropanone, 1,3-dichloropropanone, 1,1,1-trichloropropanone and 1,1,3-trichloropropanone). In the SOS chromotest, all the compounds except monochloropropanone were found to induce primary DNA damage in Escherichia coli. With the fluctuation test, all five chloropropanones showed mutagenic activity on Salmonella typhimurium strain TA100. The newt micronucleus assay detected a clastogenic effect on the peripheral blood erythrocytes of Pleurodeles waltl larvae only for 1,3-dichloropropanone and 1,1,3-trichloropropanone. Moreover, two structure-activity relationships are noticeable: (1) chloropropanones with chlorine substituents on both carbon positions (1,3-DCP and 1,1,3-TCP) are by far more genotoxic than chloropropanones substituted only on one carbon position (1,1-DCP and 1,1,1-TCP); (2) the increase of the number of chlorine substituents decreases the mutagenic activity (fluctuation test) of the chlorinated propanones studied.

Acetone↗

Genotoxicity of kraft pulp spent liquors from different types of chlorination procedures.

The genotoxicity of spent liquors from kraft softwood and hardwood pulp bleaching processes was studied using the Ames Salmonella test and the SOS chromotest. The induction of micronuclei, in vivo, was assayed in bone marrow erythrocytes of B6 mice treated with softwood first chlorination stage spent liquor. The softwood bleaching process used a combination of Cl2 and ClO2 at the first chlorination stage. During the study the amount of free chlorine at the first chlorination stage in the softwood bleachery was gradually decreased, although the amount of active chlorine remained the same. Enzymatic bleaching was also used in a softwood process together with chlorine (Cl2 + ClO2). The hardwood bleaching plant used only ClO2 at the first chlorination stage. A decrease in genotoxicity, corresponding to the decrease in Cl2, was observed in the Ames Salmonella assays of the softwood bleaching plant effluents. A similar decrease was observed in the SOS chromotest. The highest decrease in mutagenic activity was observed when enzymatic bleaching was used together with chlorine.

Animals↗

Genotoxicity of 2-halosubstituted enals and 2-chloroacrylonitrile in the Ames test and the SOS-chromotest.

2-Chloroacrolein and 2-bromoacrolein are very potent direct mutagens not requiring metabolic activation in Salmonella typhimurium strains TA 100 and TA 1535. Mutagenic activities decrease with increasing degree of methyl substitution at carbon atom C-3 of the acrolein moiety from 2-chloroacrolein via 2-chlorocrotonaldehyde to 2-chloro-3,3-dimethylacrolein. With 2-chloroacrylonitrile equivocal results are obtained in strain TA 100 without S9-mix and unequivocal with S9-mix. In the SOS-chromotest the 2-chloroenals are also very strong genotoxins and the structure-activity relationships found in the Ames test are clearly confirmed. 2-Chloroacrylonitrile is not positive in the SOS-chromotest. The mutagenic mechanisms are discussed, and indications are provided that genotoxicity/mutagenicity depends on formation of DNA adducts, e.g., 1,N2-cyclic deoxyguanosine adducts.

Acrolein↗

Genotoxicity and DNA adduct formation of incense smoke condensates: comparison with environmental tobacco smoke condensates.

Indoor air pollution has now been recognized as a potentially important problem for public health, since people spend most of their day in closed environments. Incense burning is possibly associated with elevated risks of leukemia and brain tumor in children from the epidemiological studies. Thus, evaluation of the genotoxicity of smoke condensates from incense burning is needed. We examined the genotoxicity of incense smoke condensates (ISC) using the Ames test in S. typhimurium strains with different mutagenic specificity and level of metabolic enzyme, the SOS chromotest in E. coli PQ37, and sister chromatid exchange assay in Chinese hamster ovary cells (SCE/CHO). The genotoxicity of environmental tobacco smoke condensates (TSC) was also evaluated by the three assays to compare with the genotoxicity of ISC. ISC showed a positive response in TA98, but not in TA100. It suggested that ISC only contained frame shift mutagens. The mutagenicity of ISC in both strains of TA98NR with deficient nitroreductase and TA98/1,8-DNP6 with deficient O-acetyltransferase was markedly decreased compared to that in TA98 strain. However, the mutagenicity was enhanced in YG1024 with overexpression of O-acetyltransferase activity. Thus, nitroarenes seemed to be responsible in part for the mutagenicity of ISC. Interestingly, all of the four ISC and two TSC samples showed a dose-dependent genotoxic response in the SOS chromotest with E. coli PQ37 but a low SCE induction of those samples were observed in CHO cells. When the genotoxicity was analyzed based on the condensates per one gram of original samples, the genotoxicity of two TSC condensates in prokaryotic cells was higher than that of four ISC samples except for the genotoxicity of TSC-2 in TA98 strain. However, the genotoxicity of certain ISC in eukaryotic cells based on the SCE/CHO assay was higher than that of TSC. To compare the covalent binding of DNA reactive intermediates of ISC and TSC to S. typhimurium TA98, the DNA adducts were evaluated by the 32P-postlabeling method with butanol extraction version. Similar diagonal radioactive zone (DRZ) was observed between ISC and CSC. However, DNA adduct levels induced by TSC were much greater than that of ISC.

Acetyltransferases↗

Study of the genotoxic activity of six halogenated acetonitriles, using the SOS chromotest, the Ames-fluctuation test and the newt micronucleus test.

Three short-term assays (the SOS chromotest, the Ames-fluctuation test and the newt micronucleus test) were carried out to evaluate the genotoxicity of six halogenated acetonitriles identified in chlorinated waters (monochloro-, dichloro-, trichloro-, monobromo-, dibromo- and bromochloroacetonitrile). With the SOS chromotest, three of the chemicals studied (dichloro-, dibromo- and bromochloroacetonitrile) were found to induce primary DNA damage in Escherichia coli PQ37. In the Ames-fluctuation test, all the compounds except dibromoacetonitrile showed mutagenic activity on Salmonella typhimurium strain TA100. The newt micronucleus assay detected a clastogenic effect on the peripheral blood erythrocytes of Pleurodeles waltl larvae for all the six haloacetonitriles studied. Moreover, two structure-activity relationships were noted: (1) the genotoxic activity of haloacetonitriles containing bromine substituents appeared higher than the corresponding chlorinated acetonitriles and (2) the clastogenic activity of the chlorinated acetonitriles increased with the number of chlorine substituents.

Acetonitriles↗

SCP1-dependent Weigle activation of the VP5 phage in Streptomyces coelicolor A3(2).

The kinetics of induction of the UV-irradiated bacteriophage VP5 (Weigle reactivation) in Streptomyces coelicolor A3(2) strains with and without plasmid was investigated. Chloramphenicol (CAF) inhibits Weigle reactivation (WR) in UF strains (SCP1 absent) but not in SCP1+ strains of IF fertility (free plasmid). CAF, moreover, inhibits protein synthesis in non-irradiated UF and IF strains. In UV-irradiated IF strains, on the other hand, protein synthesis takes place irrespective of CAF. Weigle reactivation appears to require protein synthesis: the SCP1 plasmid, by protecting protein synthesis from CAF inhibition in UV-irradiated strains, allows WR. The proteins synthesized after UV induction during the pre-incubation period were investigated and the results suggest that a new UV-induced protein, coded by a gene localized on the plasmid, interacts with the cellular SOS system.

Bacterial Proteins↗

Validation of the SOS/Umu test with mutagenic complex mixtures.

The SOS/Umu test, a rapid system for detecting genotoxic agents by monitoring SOS responses, was evaluated with the extracts of 8 mutagenic complex mixtures (airborne particles, coal dust, tobacco snuff, fried beef, fried shredded pork, airborne particles from polyurethane plants). In this system, the SOS function induced by genotoxicants is detected by a colorimetric measurement of beta-galactosidase in tester cells carrying a umuC-lacZ-fused gene on the plasmid. Results from the study show that a higher beta-galactosidase activity was found when the enzyme substrate and treated cells were added simultaneously into the enzyme reaction mixture and post-treatment dilution (10 X dilution with fresh medium) and incubation (for 2 h) were incorporated. The post-treatment dilution is necessary to reduce a possible false positive due to the color of test substances. The extracts of all mutagenic complex mixtures tested were found to induce dose-related SOS responses, indicating that the SOS/Umu test is potentially useful for the detection of mutagenic complex environmental mixtures.

Air Pollutants↗

Suppressive effects of coffee on the SOS responses induced by UV and chemical mutagens.

SOS-inducing activity of UV or chemical mutagens (AF-2, 4NQO and MNNG) was strongly suppressed by instant coffee in Salmonella typhimurium TA1535/pSK1002. As decaffeinated instant coffee showed a similarly strong suppressive effect, it would seem that caffeine, a known inhibitor of SOS responses, is not responsible for the effect observed. The suppression was also shown by freshly brewed coffee extracts. However, the suppression was absent in green coffee-bean extracts. These results suggest that coffee contains some substance(s) which, apart from caffeine, suppresses SOS-inducing activity of UV or chemical mutagens and that the suppressive substance(s) are produced by roasting coffee beans.

4-Nitroquinoline-1-oxide↗

Evidence that N-methyl-N'-nitro-N-nitrosoguanidine induces adaptive response in Bacillus thuringiensis.

Bacillus thuringiensis is shown to have an inducible error-free repair system for alkylation damage as found in Escherichia coli and Bacillus subtilis. Growth of cells in the presence of low concentrations of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) induces an adaptive response which is characterized by an increase in resistance to killing and mutagenesis by challenge with higher concentrations of MNNG. In addition, we have noted with interest that adaptive low doses seem to produce lesions at a rate sufficient to induce an increase of mutation frequency, and inhibition of cell division. The possibility of an interaction between SOS and adaptive responses with these low doses of MNNG is discussed.

Adaptation, Physiological↗

SOS-inducing activity of chemical carcinogens and mutagens in Salmonella typhimurium TA1535/pSK1002: examination with 151 chemicals.

The umu test system is a newly developed method to evaluate genotoxic activities of a wide variety of environmental carcinogens and mutagens (Oda et al., 1985). In the present study, we further examined the abilities of 151 chemicals to induce umu gene expression in Salmonella typhimurium TA1535/pSK1002. Among the chemicals examined, 72 compounds induced umu gene expression, which could be defined on a basis of increased beta-galactosidase activity by 2-fold over the background level. The potent genotoxic compounds without metabolic activation were adriamycin, bleomycin, daunorubicin, 1,3-dinitropyrene, 1,6-dinitropyrene, 1,8-dinitropyrene, N-ethyl-N'-nitro-N-nitrosoguanidine, furylfuramide, methyl methanesulfonate, N-methyl-N'-nitro-N-nitrosoguanidine, mitomycin C, 1-nitropyrene and 4-nitroquino-line-1-oxide. In the presence of S9, aflatoxin B1, 2-aminoanthracene, Glu-P-1, IQ, MeIQ, MeIQx, Trp-P-1 and Trp-P-2 also induced umu gene expression markedly. Several chemicals such as 2-acetylaminofluorene, 9-aminoacridine, azobenzene, benzanthracene, benzidine, diethyl nitrosamine, 1-nitronaphthalene, paraquat, potassium dichromate and sodium nitrite were weakly genotoxic and the induction by these compounds could be detected only when the incubation time was prolonged from 2 h to 5 h. Data are also presented that some of the chemicals such as dimethyl sulfoxide, m-dioxan, 5-fluorouracil and paraquat, which have been reported to be non-mutagenic in Ames/Salmonella assay, were found to be active in inducing umu gene expression, while the known mutagenic compounds including acrylonitrile, 4,4'-dinitrobiphenyl, furfural, methylene chloride, 1-naphthylamine, sodium azide, o-tolidine and o-toluidine were non-genotoxic in the present assay system.

Biotransformation↗

Role of rec genes in SOS-induced inhibition of cell division in Escherichia coli.

The inhibition of cell division induced by bleomycin (BM) and UV irradiation in the set of rec mutants of E. coli K12 was studied. Data presented in this work indicate that BM treatment requires mainly the RecBC pathway for the induction of cell filamentation. In the recB21 mutant cell filamentation is delayed and reduced compared to the wild type. Cell filamentation is BM-induced with similar kinetics in strains with a proficient RecBC recombination pathway (rec+, recF143 and recN262), as well as in the strain with a fully expressed RecF pathway (recB21recC22sbcB15). Induction is completely abolished in the recB21recF143 double mutant. On the other hand cell filamentation was induced similarly by UV irradiation in all strains with a functional recF gene and in the strain with a fully operative RecF pathway, but it was delayed in the recF143 and recB21recF143 mutants.

Bleomycin↗

Nitrous acid induced damage in T7 DNA and phage.

T7 phage was exposed to 56 mM nitrous acid at pH 4.6 causing a 90% decrease in survival for each 10 min duration of exposure. The survival of phage made by encapsulating nitrous acid treated DNA into empty phage heads was nearly the same as the survival of phage exposed to nitrous acid in vivo. In contrast to previous reports, growth of SOS-induced wild-type E.coli showed no increase in survival. The survival of nitrous acid treated phage was not lowered when grown on E.coli strains deficient in DNA polymerase I, exonuclease III, and the uvrA component of the nucleotide excision-repair endonuclease. Therefore, these enzymes are not vital for repair of nitrous acid induced damage in bacteriophage T7.

DNA Damage↗

Induction of the SOS response by hydroxyurea in Escherichia coli K12.

Hydroxyurea at concentrations higher than 10(-2) M induced the recA and sfiA genes of E. coli as well as the lambda prophage by a pathway independent of the recBC genes. In addition, the hydroxyurea-mediated induction of the SOS response is accompanied by a recA-dependent decrease on the cellular ATP pool. The presence of the multicopy plasmid pPS2, harboring the nrdAB genes (encoding the ribonucleoside reductase enzyme), abolished the hydroxyurea-induced expression of the recA gene. These data lead us to suggest that induction of the SOS response by hydroxyurea is due to the blocking of DNA replication by the inhibition of the ribonucleoside reductase complex activity.

Bacterial Proteins↗

Influence of S9 mix in the induction of SOS system by quercetin.

The induction of recA, umuC and sfiA genes by quercetin was studied in the presence and in the absence of S9 mix. The inducing activity of quercetin is higher for sfiA than for recA and umuC genes in the absence of S9 mix. The putative genotoxic metabolites of quercetin produced by S9 mix display different inducing activities of the three SOS genes as compared to quercetin. The induction of sfiA gene is decreased by the presence of S9 mix, whereas an opposite effect was observed concerning umuC and recA. These data suggest that the error-prone repair pathway participates in mutagenesis by quercetin and its metabolites. Moreover, the type of DNA damage exerted by quercetin seems to be determined by its metabolic fate. The importance of testing for the induction of other SOS genes, together with sfiA, in the study of SOS functions as a genotoxic index is emphasized.

Animals↗