Mutagenic and cytotoxic effectiveness of diisopropyl xanthogen polysulphide in human lymphocyte cultures

The mutagenic and cytotoxic effectiveness of the new rubber vulcanisation accelerator diisopropyl xanthogen polysulphide (Robac AS 100) was tested in human lymphocyte cultures of four healthy probands. The concentrations of Robac AS 100 were 0.57, 5.7 and 57.0 microg/ml. Higher concentrations showed too high cytotoxicity to be evaluable.Without external activation, incubation time with Robac AS 100 was 21 h. In the presence of rat liver microsomes from aroclor-induced rats (2mg microsomal protein/ml), incubation of the test compound was 2h.Mutagenicity testing was performed by analysis of micronuclei (MN), structural chromosome aberrations (CAs) and sister chromatid exchanges (SCEs). The MN-rate was determined using the cytochalasin B (cyt B) block method. For evaluation of cytotoxicity, mitotic index (MI) and nuclear division index (NDI) were determined. The validity of the test methods was ascertained by positive controls: mitomycin C (MMC) and bleomycin (BLM) were used in experiments without exogenous activation and cyclophosphamide (CP) in experiments with exogenous activation.The presence of rat liver microsomes increased the mutagenic effect of Robac AS 100 in the SCE- and MN-test. But only the highest Robac AS 100-concentration (57.0 microg/ml) showed significantly increased mutagenic activity in all tests. However, cytotoxicity at this concentration was already substantial. Therefore, we consider the evidence for mutagenicity of Robac AS 100 as limited.     

Mutagenic effectiveness and efficiency of EMS,DES and gamma-rays in rice

Summary Data on chlorophyll mutation frequency after treatment with EMS, DES and gamma-rays and sequential administration of gamma-rays and the two alkylating agents in three varieties of rice have been used to work out quantitatively the effectiveness and efficiency of each mutagen and combination treatment. For effectiveness, the order is EMS > DES and for efficiency it is EMS > DES > gamma-rays. In some sequential treatments (Gamma-rays + DES in IR8 and Basmati; DES + gamma-rays in IR8 and Jhona; Gamma-rays + EMS in IR8 and Basmati; and EMS + gamma-rays in IR8, Jhona and Basmati) mutation frequency is more than additive (synergistic) but these treatments are decisively less efficient because of their relatively high injurious effects in the M₁. generation. EMS induces more albinas than gamma-rays do. The mutational spectrum patterns induced by gamma-rays and DES are alike. In general, combination treatments tend to increase the frequency of albinas over other types of chlorophyll mutants.     

Mutagenic and cytotoxic effectiveness of zinc dimethyl and zinc diisononyldithiocarbamate in human lymphocyte cultures

The mutagenic and cytotoxic effectiveness of the vulcanisation accelerators zinc dimethyldithiocarbamate (ZDMC; ziram) and zinc diisononyldithiocarbamate (ZDINDC; arbestab Z) was tested in lymphocyte cultures of five healthy probands. ZDMC and ZDINDC (c=0.1, 1.0 and 10.0microg/ml) were studied in lymphocyte cultures without external metabolic activation. Additionally, incubation of the compounds (c=10.0microg/ml) was performed in the presence of liver microsomes from aroclor-induced rats (1 and 2h, 1 and 2mg microsomal protein). Genotoxicity testing was performed by analysis of chromosomal aberrations (CA), sister chromatid exchanges (SCEs) and micronuclei (MN). For evaluation of antiproliferative effects, mitotic index (MI) and cell cycle kinetics (CCK) were determined. In contrast to earlier investigations we found no significantly increased mutagenic or cytotoxic activity of ZDMC; ZDINDC also was inactive under these conditions.     

p-benzoquinone and p-hydroquinone as histochemical reagents

Summary Studies on the rate controlling factors of the MTT-Hydroquinone lipoprotein reaction show that the formazan production is quantitatively linear on time of incubation, and dependent on temperature and hydroquinone concentration. Experiments indicate that the reproducibility is sufficient for the reaction to be used as a reference parameter in the numerical expression of dehydrogenase activities in tissue sections. A ratio system is described in which the formazan produced by the dehydrogenase is expressed in terms of the formazan from the non-enzymatic lipoprotein reaction, and a nomenclature for a unit measure is proposed. The method has been tested on guinea pig liver and heart, and the range of variation investigated.     

Modifications of ribonuclease A induced by p-benzoquinone

The nature of ribonuclease A (RNase) modifications induced by p-benzoquinone (pBQ) was investigated using several analysis methods. SDS-PAGE experiments revealed that pBQ was efficient in producing oligomers and polymeric aggregates when RNase was incubated with pBQ. The fluorescence behavior and anisotropy changes of the modified RNase were monitored for a series of incubation reactions where RNase (0.050 mM) was incubated with pBQ (0.050, 0.25, 0.50, 1.50 mM) at 37 °C in phosphate buffer (pH 7.0, 50 mM). The modified RNase exhibited less intense fluorescence and slightly higher anisotropy than the unmodified RNase. UV-Vis spectroscopy indicated that pBQ formed covalent bonds to the modified RNase. Confocal imaging analysis confirmed the formation of the polymeric RNase aggregates with different sizes upon exposure of RNase to high concentrations of pBQ. The interaction between the modified RNase and salts affecting biomineralization of salts was also investigated by scanning electron microscopy. Overall, our results show that pBQ can induce formation of both RNase adducts and aggregates thus providing a better understanding of its biological activity.     

Mutagenic sterol hydroperoxides

Sterol hydroperoxides 3 beta-hydroxy-5 alpha-cholest-6-ene-5-hydroperoxide and 3 beta-hydroxycholest-5-ene-7 alpha-hydroperoxide show weak dose-response direct mutagenicity towards Salmonella typhimurium strain TA 1537 in a liquid medium incubation bioassay. Responses were compromised by metabolism of the sterol hydroperoxides and by phase separation during the incubation period. Mutagenicity responses were increased by added superoxide dismutase but diminished by added rat liver S9 enzymes and abolished by added catalase. Catalase also abolished the stimulatory effect of superoxide dismutase. These results indicate that superoxide and peroxide be implicated in the mutagenicity responses.     

Mutagenic activity of radiosensitizers

Seven radiosensitizers, six derivatives of nitroimidazole (coded P1 to P5 and one imidazole derivative--P6 were investigated for mutagenicity using 3 short-term tests: Ames test, prophage lambda induction and tryptophan reversion test. Out of seven investigated compounds five were not mutagenic. Only P1 derivative induces base pair substitutions. Another derivative of nitroimidazole: metronidazole induces base pair substitution and frameshift mutations. Its positive response in the prophage lambda induction test suggests that metronidazole provokes also epigenetic changes.     

Mutagenic effectiveness of 14.1 MeV neutrons and 200 kV X-rays at the dumpy complex locus of Drosophila melanogaster.

The effectiveness of 14.1 MeV neutrons relative to 200 kV X-rays for the induction of the various kinds of dumpy mutation in mature sperm of Drosophila melanogaster was investigated. The estimated RBE values are: 0.52 for all complete mutations; 0.64 for the (olv, ov) types; 0.33 for the (ol, lv, o, v, c) types; 0.33 for all fractional mutations. These data lend support to the thesis that (1) complete dumpy mutations of the olv and ov types are more frequently associated with chromosomal aberrations than those of the ol, lv, o, v and c types, and (2) fractional mutations and complete mutations of the (ol, lv, o, v, c) types are most probably point mutational events.     

Glucosamine-mediated detoxification of p-benzoquinone and its removal by chitosan

p-Benzoquinone non-enzymatically reacted with d-glucosamine at physiological pH and moderate temperature. The reaction of p-benzoquinone with glucosamine was signaled by changes in the UV and visible spectra. The reactivity proceeded fastest at pH values above 7, with a sharp drop from pH 6.5 to 7.0, and the reaction was negligible in acidic conditions. The order of reactivity of amino sugars was d-mannosamine > d-glucosamine > d-galactosamine. From the reaction mixture, four conversion products were isolated and none was toxic to Escherichia coli even at 500–700 g ml^–1, while p-benzoquinone was cytotoxic to E. coli at 20 g ml^–1. Chitosan could react with p-benzoquinone efficiently and remove this toxicant in aqueous solution.     

Mutagenic activity of chloramines

Mutagenesis by chloramines and hypochlorous acid (HOCl) was studied to determine whether these agents could contribute to the mutagenic and potentially carcinogenic activity of stimulated leukocytes and whether environmental exposure to these agents is a cause for concern. Mutagenic activity was measured using the S. typhimurium TA97a, TA100 and TA102 tester strains. Because chloramines and HOCl are bactericidal, react rapidly with cell components, and can destroy the histidine and biotin required for the mutagenesis assay, activity can't be compared directly with that of less toxic or reactive agents. Nevertheless, chloramines were mutagenic when tested under appropriate conditions. TA100 was the most sensitive strain, and the most active mutagens were lipophilic dichloramines (RNCl2) including derivatives of histamine, ethanolamine and putrescine. Lipophilic monochloramines (RNHCl) such as histamine-monochloramine and NH2Cl were less active. Hydrophilic chloramines such as taurine-chloramines had low activity, and HOCl was inactive. The metabolic state of the bacteria was critical. Chloramines were mutagenic when added to bacteria with glucose at 37 degrees C, but killing predominated when chloramines were added at 4 degrees C or 25 degrees C, or at 37 degrees C without glucose. Production of chloramines and HOCl by leukocytes in vivo could contribute to the association of chronic inflammation and cancer as a result of: (1) the entry of membrane-permeable chloramines into normal cells followed by attack on intracellular components including DNA, and (2) the production of secondary mutagens such as compounds with carbonyl groups or carbon-chlorine bonds. On the other hand, chlorination of water supplies is perhaps more likely to destroy than create mutagens, and chloramines from the environment are unlikely to penetrate the skin and mucous membranes.     

Mutagenic evaluation of ronidazole

Ronidazole was evaluated for mutagenic potential using in vitro microbial tests and in vivo studies in mice. The microbial test used the histidine requiring mutants of Salmonella typhimurium with and without a rat liver microsomal activation system (Ames test). The studies in mice included the dominant lethal test, micronucleus test and cytogenetic assays.Ronidazole was given orally in doses of 50, 100 and 200 mg/kg/day in the in vivo studies. In the dominant lethal test, groups of male mice were treated for five consecutive days before being mated with untreated females. In the micronucleus test, the mice were administered the compound for 2 or 5 consecutive days; they were killed 6 h after the last dose and bone marrow was examined for the presence of micronuclei in developing erythrocytes. In the cytogenetic assays, bone marrow cells in metaphase were examined for chromosome aberrations, 6, 24 and 48 h after mice were treated acutely with the test compound. In addition, similar examinations of chromosomes were made on mice given five consecutive dosages of ronidazole and killed 6 h after the last dose. The results of the various in vivo studies did not suggest that ronidazole would be mutagenic for the mammal.Ronidazole at concentrations of 10 and 50 μg/plate was found to increase the number of back mutations of missense mutants in the in vitro bacterial test. This finding confirms the results of Voogd et al. [19]. Incorporation of the microsomal activation system had no effect on the mutagenic capability of the test compound.In conclusion, although ronidazole was shown to be mutagenic in in vitro bacterial systems, the in vivo systems did not suggest that the compound would be mutagenic for the mammal.     

Mutagenic cholesterol preparations.

Naturally air-aged commercial samples of USP or reagent-grade cholesterol contain components which are mutagenic towards Salmonella typhimurium strains TA1537, TA1538 and TA98. These mutagenic components are associated with the polar cholesterol autoxidation products, but identity of the mutagenic components has not been achieved. Pure crystalline nonmutagenic cholestrol free from autoxidation products becomes mutagenic towards these strains upon heating at 70 degrees in air or following exposure to 60 Co gamma-radiation.     

Mutagenic alterations of antibody biosynthesis

The investigations of changes of antibody affinities were carried out with 182 immuned CBA mice under the influence of the mutagens. The mice were injected with ethylene imine and its derivatives, 6-azauridine, cyanurchloride, theobromine, glyoxal, at a dose of 100 mg/kg 2 days before, simultaneously and in 2 and 5 days after the immunization. The indices of the functional affinity of antibodies (the mean constant of true association, the standard free energy, the concentration of the hapten binding sites, the heterogeneity index, the level of IgG and its Fab- and Fc-fragments) were determined within 5, 10 and 20 days. Three groups of changes of antibody affinities in immuned mice were observed under the influence of mutagens: a) disassotiative changes with acute fall of affinities; b) erase changes; c) variable changes with discrete and corrective displacement of some affinities indices. Analysis of the data obtained suggests the existence of two forms of the mutagen immunodepression: 1) real immunodepression with a parallel fall of primary and secondary indices of antigen-intibody interaction; and 2) functional immunodepression with a fall of secondary indices. Polyfunctionality of the inhibition effect of alkyl mutagens is demonstrated.     

Mutagenic potential of anti-herpes agents

A number of anti-herpes agents which are either licensed for clinical use (acyclovir) or subject of clinical studies (bromovinyldeoxyuridine, fluoroiodoaracytidine, dihydroxypropoxymethylguanine) or under preclinical investigation (i.e., fluoroiodoarauridine), fluoromethylarauridine, dihydroxybutylguanine, bromovinyldeoxycytidine, bromovinylarauridine and carbocyclic bromovinyldeoxyuridine) were evaluated for their ability to induce sister chromatid exchange (SCE), an indicator of mutagenesis. SCE was scored on metaphase chromosomes of human lymphocytes which had been exposed to 5-bromo-2'-deoxyuridine and varying concentrations of the test compounds. The antiviral assays were based on the inhibition of the cytopathogenicity of herpes simplex virus for human diploid fibroblasts. Most compounds, i.e. acyclovir, bromovinyldeoxyuridine or carbocyclic bromovinyldeoxyuridine, did either not induce SCE or only so at concentrations far above their minimum antiviral concentrations. However, fluoroiodoaracytidine and dihydroxypropoxymethylguanine were found to affect the SCE rate at a concentration (greater than or equal to 4.5 micrograms/ml) that is readily achievable in blood following intravenous injection.     

Mutagenic activity of swimming-pool water

Swimming pool water, being chlorinated and exposed to trace organics from use was investigated as a possible source of mutagens using the Salmonella/mammalian-microsome test. Procedures previously described for the extraction of trace organics from water using XAD-2 macroreticular resin were modified to allow quantitative extraction of mutagens. These procedures were superior to freeze-drying and solvent-extraction. Using a base-pair histidine mutant, strain TA100, of Salmonella typhimurium significantly mutagenic responses were observed using concentrates from 3 variations of the extraction procedure. Acidified pool-water extracts eluted with ether or acetone were mutagenic, the former enhanced in the presence of the induced microsomal fraction from rat livers. Non-acidified pool-water extracts eluted with acetone were mutagenic without microsomal activation. These results indicate the presence of more than one mutagen in what is likely a complex mixture of organic molecules in swimming-pool water.