The very low mutagenic activity of sodium azide inArabidopsis thaliana

Sodium azide, reported to be a strong mutagen in barley, revealed a very weak mutagenic activity inArabidopsis.     

Genetic activity of allyl chloride

Allyl chloride (3-chloroprene) is mutagenic for Salmonella typhimurium and it induces gene conversions in Saccharomyces cereuisiae. It also displays DNA-modifying activity for E. coli. This is in contrast to a recent study which reported its lack of genetic activity for Salmonella typhimurium.     

Structural Requirements for Mutagenic Activities of N-Heterocyclic Bases in the Salmonella Test System

The mutagenic activities of quinoline, isoquinoline, phenanthridine, benzo(f)quinoline, benzo(h)quinoline and their α-amino derivatives were compared in relation to the effect of structural changes using the Salmonella typhimurium test system. All mutagenic compounds tested require the liver microsomal fraction for their mutagenic activity. Phenanthridine, two benzoquinolines and quinoline were mutagenic. α-Amination of two benzoquinolines and quinoline resulted to increase their mutagenic activity intensively. Addition of a benzene ring to the benzene moiety of 2-aminoquinoline, so that two carbon atoms are shared, affected distinctly the increase in the mutagenic activity. The co-existence of benzoquinoline series with 2-aminobenzo(f)quinoline showed the clear synergistic action.     

Mutagenicity of flavones,chromones and acetophenones in Salmonella typhimurium: New structure—activity relationships

28 flavones and 11 structurally-related flavonoids, chromones, and acetophenones, were tested for mutagenicity in the Salmonella typhimurium his reversion assay. 7 flavones, all of which were hydroxy- or methoxy-substituted at position 8, were moderate to strong mutagens in strain TA100 in the presence of rat liver S9 mix. In each case, the response of strain TA98 was either not significant or was very much weaker than that observed in strain TA100. The activation by S9 is not mediated by the microsomal cytochrome P₄₅₀ system, since activation was not diminished when microsomes were removed by centrifugation at 100000 × g. The observed strain specificity and structural requirements for activity indicate a mutagenic mechanism different from that associated with previously reported mutagenic flavonols (3-hydroxy-flavones) which are most active in strain TA98. The most mutagenic flavone investigated, 5,7,8-trihydroxy-flavone (norwogonin), had a potency of 17 revertants/nmole.Simplification of the chemical structures to hydroxy-substituted chromone and acetophenone systems revealed similar strain specificity, hydroxylation requirements, and S9 dependence within these structural classes, suggesting a similar activation pathway and mutagenic mechanism. The greatest mutagenic potency was observed within the flavone series, but significant potency was retained by similarly hydroxylated chromones and acetophenones. No mutagenic activity was observed in the absence of the aryl ketone moiety.     

5-Methyltryptophan resistance mutations in Escherichia coli K-12. Mutagenic activity of monofunctional alkylating agents including organophosphorus insecticides

The induction of 5-methyltryptophan (5-MT) resistance mutations was assayed as a test system for mutagenic chemicals in Escherichia coli. It is assumed that different premutational alterations in several genes of the Escherichia coli chromosome will lead to 5-MT-resistant mutants. The chemicals used were three monofunctional alkylating agents as reference compounds, namely β-propiolactone (β-PL), N-methyl-N′-nitro-N-nitrosoguanidine (MNNG), and methyl methanesulfonate (MMS), which are all mutagenic in the 5-MT system; of the eight organophosphorus insecticides tested, four have definite mutagenic activity (Dichlorvos, Oxydemetonmethyl, Dimethoate, and Bidrin), one is probably mutagenic (Methylparathion) and the remaining three (Parathion, Malathion and Diazinon) do not induce 5-MT resistance mutations in the conditions used here (< 30% survival). The relative mutagenic activity after a treatment time of 60 min is (in decreasing order) MNNG > MMS > Dichlorvos > Oxydemetonmethyl, Dimethoate and Bidrin. The concentration-dependent mutagenic activity of all mutagenic compounds is nearly linear when plotted on a log-log scale (with slopes varying from 1.0 to 1.5) and could be taken as an indication that one premutational reaction will be sufficient for the induction of one 5-MT-resistant mutant.     

Mutagenic activity of the methyl and phenyl derivatives of the food mutagen 2-amino-3-methylimidazo[4,5-f]quinoxaline (IQx) in the Ames test

The mutagenic activity of 15 different mono-, di-, tri-, and tetramethyl derivatives of the food mutagen IQx (2-amino-3-methylimidazo[4,5-f]quinoxaline), one diphenyl derivative of IQx and two phenyl derivatives of 5-MeIQx (2-amino-3,5-dimethylimidazo[4,5-f]quinoxaline) were studied in the Ames test with Salmonella typhimurium TA98 and enzymatic activation (S9). The number and positioning of the methyl groups strongly affected the mutagenic activity. The phenylated compounds showed weak mutagenic potency. It seems that both resonance stabilization of the nitrenium ion and steric effects are important in determining mutagenic potency.     

Metabolic activation of dimethylnitrosamine and diethyl-nitrosamine to mutagens

Dimethylnitrosamine (DMN) and diethylnitrosamine (DEN) are not mutagenic by themselves, but they can be converted by mammalian enzymes to highly mutagenic products. As indicators for mutagenic activity, Neurospora crassa and Salmonella typhimurium were used. The ad-3 forward-mutation system was used to detect specific locus mutations; mutants in this system can range from multi-locus deletions to leaky mutations. The induction of mutations in S. typhimurium is detected as induction of histidine revertants of the histidine-requiring strain G₄6. The activation of DMN is microsomal, inhibited by SKF 525-A, and requires co-factors. The activating enzyme is induced in mice by pretreatment with phenobarbital, 3-methylcholanthrene and butylated hydroxytoluene. The mutagenic activity of the reaction products is directly correlated with the metabolic formation of formaldehyde with and without induction by 3-methylcholanthrene and across strains of mine. Formaldehyde does not contribute to the mutagenic activity of the reaction products. It is clear from the data that the reversion sites in G₄6 are more sensitive than the ad-3 loci of Neurospora crassa to the mutagenic action of DMN metabolites formed by mammalian liver. The microsomal assay is a few orders of magnitude more sensitive than the intraperitoneal host-mediated assay, and the intrahepatic host-mediated assay is a few orders of magnitude more sensitive than the in vitro microsomal system.     

Mutagenicity of organophosphorus compounds in bacteria and drosophila

140 Organophosphorus compounds (OP's) have been tested for mutagenic activity in bacteria, principally by using two specially constructed sets of tester strains of the bacteria Salmonella typhimurium and Escherichia coli. It was found that 20% gave positive mutagenic responses and that this group of chemicals produce base substitutions rather than frame-shift mutations. In most cases the DNA repair genes exrA⁺ and recA⁺ were for mutagenic activity.Seven compounds were further tested in Drosophila melanogaster for the ability to induce recessive lethal mutations. In some of these cases the doses administered to the flies had to be very low due to the highly toxic nature of the compounds. To overcome this problem, the accumulation of recessive lethal mutations was measured in populations which were continually exposed to the compounds over a period of some 18 months. During this time the populations developed increased resistance to the compound and so the dose administered could gradually be increased. Six of the compounds were mutagenic.Of the compounds tested in both systems, those showing mutagenic activity in bacteria were also mutaganic in Drosophila, those mutagenic in bacteria were not mutagenic in Drosophila.     

Evaluation of mutagenic effect of the fungicide fenaminosulf in Drosophila melanogaster

Fenaminosulf (p-dimethylaminobenzenediazo sodium sulfonate, CAS registry No. 140-56-7) which is an active ingredient in several commercial fungicide was reported to be mutagenic in Salmonella typhimurium (McCann et al., 1975), Bacillus subtilis (Kada et al., 1974) and shown to cause chromosome aberrations in plants (Zutshi and Kaul, 1975). Since fenaminosulf has structural similarity to the potent carcinogen, butter yellow (p-dimethylaminoazobenzene, CAS registry No. 60-11-7), the present studies were undertaken to evaluate the mutagenic potential of this fungicide in Drosophila melanogaster. Fenaminosulf administered at 10 mg/100 ml food medium failed to induce sex-linked recessive mutations in Drosophila. Since Drosophila has drug-metabolizing enzymes similar to those of mammals (Vogel, 1975), it is suggested that the lack of mutagenic activity of fenaminosulf could be due to the conversion of fenaminosulf to non-mutagenic derivatives in Drosophila.     

Antifungal activity of gemini quaternary ammonium salts

A series of gemini quaternary ammonium chlorides and bromides with various alkyl chain and spacer lengths was synthesized. The most active compounds against fungi were chlorides with 10 carbon atoms within the hydrophobic chain. Among these compounds were few with no hemolytic activity at minimal inhibitory concentrations. None of the tested compounds were cytotoxic and mutagenic. Cationic gemini surfactants poorly reduced the adhesion of microorganisms to the polystyrene plate, but inhibited the filamentation of Candida albicans. One of the tested compounds eradicated C. albicans and Rodotorula mucilaginosa biofilm, what could be important in overcoming catheter-associated infections. It was also shown that gemini surfactants enhanced the sensitivity of C. albicans to azoles and polyenes, thus they might be potentially used in combined therapy against fungi.     

Isolation and studies of the mutagenic activity in the Ames test of flavonoids naturally occurring in medical herbs

Quercetin, rhamnetin, isorhamnetin, apigenin and luteolin were isolated from medicinal herbs: Erigeron canadensis L., Anthyllis vulneraria L. and Pyrola chloranta L. The mutagenicity of these naturally occurring flavonoids was tested by the Ames method with S. typhimurium strains TA1535, TA1538, TA97, TA98, TA100 and TA102 in the presence and absence of metabolic activation. Of the above flavonoids only quercetin and rhamnetin revealed mutagenic activity in the Ames test. Quercetin induced point mutations in strains TA97, TA98, TA100 and TA102 of S. typhimurium. The presence of S9 rat liver microsome fraction markedly enhanced the mutagenic activity of quercetin in these strains. Rhamnetin appeared to be a much weaker mutagen in the Ames test. The compound induced mutations in strains TA97, TA98 and TA100 of S. typhimurium but only in the presence of metabolic activation.Comparison of the structure of the studied flavonoids with their mutagenic activity indicates that the mutagenicity of flavonoids is dependent on the presence of hydroxyl groups in the 3′ and 4′ positions of the B ring, and that the presence of a free hydroxy or methoxy group in the 7 position of the A ring also probably contributes to the appearance of mutagenic activity of flavonoids in the Ames test. It also appeared that the presence of methoxy groups, particularly in the B ring of the flavonoid molecule, markedly decreases the mutagenic activity of the compound.     

Mutagenicity of alkylhydrazine oxalates in Salmonella typhimurium TA100 and TA102 demonstrated by modifying the growth conditions of the bacteria

Alkylhydrazines are important carcinogens. However, they show generally only weak mutagenicity and the activities reported from different laboratories are contradictory. We have developed a sensitive method to detect the mutagenicity of alkylhydrazines. The method is based on a modified preculturing procedures in the Ames test, the emphasis in the modification being a change in the growth period of tester strains. The optimal growth periods were found to be 11 h in Salmonella typhimurium TA100 and 5 h in Salmonella typhimurium TA102. We tested the mutagenic activity of 12 alkylhydrazines; 1,2-dimetehylhydrazine, 1,2-diethylhydrazine, 1,2-dipropylhydrazine. 1,2-dibutylhydrazine, 1,1-dimethylhydrazine, 1,1-diethylhydrazine, 1,1-dipropylhydrazine, 1,1-dibutylhydrazine, methylhydrazine, ethylhydrazine, propylhydrazine, and butylhdyrazine. All 12 alkylhydrazines were clearly mutagenic in Salmonella typhimurium TA102, and 10 hydrazines were mutagenic in Salmonella typhimurium TA100, both in the absence of S9 mix. The mutagenicity was inhibited by the addition of S9 mix or bovine serum albumin. This suggests deactivation of the mutagens by proteins.     

The mutagenicity and DNA-damaging activity of cyclic aliphatic sulfuric acid esters.

The cyclic aliphatic sulfuric acid esters 1,2-ethylene sulfate (ESF), 1,3-propylene sulfate (PSF) and 1,3-butylene sulfate (BSF) have been tested for their mutagenic and DNA-damaging activity. Mutagenicity of the compounds was established with his-auxotrophic indicator strains of Salmonella typhimurium using the in vitro plate test and the host-mediated assay technique with mice as host animals. The DNA-damaging activity was tested in a repair test with Proteus mirabilis mutants defective in DNA repair.In the repair test with a set of P. mirabilis strains (PG713 hcr^?rec^?: PG273 hcr⁺rec⁺) PSF and BSF showed a preferential growth inhibition of the repair-defective strain suggesting DNA-damaging activity of these chemicals. No such activity was found for ESF using the same concentrations of 5 and 15 μmol/plate.All cyclic sulfates revert the tester strain TA1535 of S. typhimurium in vitro indicating their ability to induce base substitutions. Compared with the reference compounds dimethyl sulfate (DMS), diethyl sulfate (DES), 1,3-propane sulfone (PPS) and 1,4-butane sulfone (BTS) the mutagenic activity in the plate test can be described as follows: PPS > PSF > BSF > BTS > ESF > DES > DMS.Dose-response studies in the host-mediated assay with tester strain TA1950 of S. typhimurium as genetic indicator system revealed a linear dosedependency of mutagenic activity. For PPS and PSF the lowest effective dose (LED) has been established as 10 μmol/kg. The LED for BSF and BTS was 50 μmol/kg, DMS and DES were mutagenic in doses of 2500 μmol/kg, while ESF was only weakly mutagenic with a LED of 5000 μmol/kg.The dose-response studies in the host-mediated assay and the results obtained in the in vitro spot test demonstrate similarities in the mutagenic action of the cyclic sulfates PSF and BSF and the respective sulfones, while the stronger alkylating compound ESF was a weak mutagen both in vitro and in vivo.     

Effects of N-nitrosopiperidine substitutions on mutagenicity in Drosophila melanogaster.

N-Nitrosopiperidine (NP) and various derivatives were fed to Drosophila melanogaster males over a wide concentration range in order to assess their mutagenic potency in the induction of X-linked recessive lethals and chromosome loss. NP was effective in inducing lethals, as were its halogen and methyl-substituted derivatives, with the exception of 2,6-dimethyl NP. (Methyl substitutions at the alpha carbon atoms reduce or eliminate mutagenic activity.) Substitution of halogen groups on the piperidine ring enhanced the mutagenic activity, with the 3-chloro compound being the most mutagenic. In contrast, substitutions with a hydroxyl, carboxyl, or keto group resulted in a loss of mutagenicity. None of the compounds tested increased the frequency of chromosome loss or breakage in mature sperm.     

Mutagenicity of potentially carcinogenic mycotoxins produced byFusarium moniliforme

The mutagenic behaviour of two potentially carcinogenic mycotoxins produced byFusarium moniliforme was investigated in theSalmonella mutagenicity test using tester strains TA97a, TA98, TA100, and TA102. The mutagenic response obtained with fusarin C (1, 5, and 10μg/plate) against tester strains TA98 and TA100 in the presence of microsomal activation confirmed previous observations on the mutagenic behaviour of this mutagen while that obtained against TA97a is reported for the first time. No dose-response relationship could be detected for the concentration levels (0.2, 0.5, 1, 5, 10 mg/plate) tested for FB₁, FB₂, and FB₃ against any of the tester strains used in either the plate incorporation and / or the pre-incubation tests. A cytotoxic effect was obtained at concentration levels of 5 and 10mg/plate in the absence of the microsomal activation mixture. From the studies it became evident thatF moniliforme produces two compounds, a mutagenic compound, fusarin C which has been shown to lack carcinogenic activity in rats and the non-mutagenic fumonisin B mycotoxins of which FB₁ is known to be responsible for the hepatocarcinogenicity of the fungus in rats.     

Structural specificity of aromatic compounds with special reference to mutagenic activity in Salmonella typhimurium — a series of chloro- or fluoro-nitrobenzene derivatives

The mutagenicity of 21 chloro- or fluoronitrobenzene compounds and 9 chloro- or fluorobenzene compounds in Salmonella typhimurium (strains TA98, TA1538, TA1537, TA100 and TA1535) was examined. The tests were carried out under the conditions of absence and presence of liver microsomal activation.15 nitro-group compounds had mutagenic activity; above all, compounds of fluoronitrobenzene were mutagenic for both types of strain. On the other hand, chloronitrobenzene compounds were mutagenic for base-pair substitution strains only. Mutagenic activity was exhibited by all compounds having a chloro or fluoro substituent at the para and ortho position in the nitrobenzene nucleus. All compounds without a nitro substituent showed no mutagenic activity.     

In vitro synthesis of a mutagenic azide metabolite by cell-free bacterial extracts

Cell-free extracts of Salmonella typhimurium synthesize a mutagenic azide metabolite from sodium azide and O-acetylserine. S. typhimurium mutant DW379 (O-acetylserine sulfhydrylase-deficient) extracts were neither able to carry out this reaction nor produce the mutagenic azide metabolite in vivo. The in vitro reaction was inhibited by sulfide but not by l-cysteine. The catalytic activity responsible for the mutagenic metabolite synthesis was stable to brief heating up to 55°C and had a pH optimum between 7–7.4. These results suggest that the enzyme O-acetylserine sulfhydrylase catalyzes the reaction of azide with O-acetylserine to form a mutagenic azide metabolite.     

Nitrosoguanidine-induced mutations inStreptomyces indicus

The mutagenic activities of five 1-alkyl derivatives of 3-nitro-l-nitrosoguanidine,viz. methyl, ethyl, propyl, isopentyl and octyl, were tested onStreptomyces indicus. All the compounds were found to be mutagenic. The methyl and ethyl derivatives were most effective regarding mutational frequency. The effectiveness decreased with increasing number of C-atoms in the alkyl group of the mutagens. The compounds tested had a much higher mutagenic efficiency than UV-rays. A marked variation existed in antibiotic activity among the biochemical mutants: increase, decrease or complete absence of such activity were seen.     

Mutagenic activity of methyl-substituted tri-and tetracyclic aromatic sulfur heterocycles

A number of polycyclic aromatic sulfur heterocycles have been identified in coal-derived products and in shale oils. The mutagenic activity of some of these compounds, including dibenzothiophene, benzo[b]naphtho[1,2-d]thiophene, benzo[b]naphtho[2,1-d]thiophene and benzo[b]naphtho[2,3-d]thiophene have been determined using the Salmonella/microsome mutagenicity test. These compounds demonstrated either very weak or no mutagenic activity. The methyl derivatives of each of these four compounds were assayed for mutagenic activity. Salmonella typhimurium TA98 was used as the tester strain. All assays required a rat-liver homogenate metabolic activator. Five of the methylated derivatives, 1-methylbenzo[b]naphtho[1,2-d]thiophene, 3-methylbenzo[b]naphtho[1,2,-d]thiophene, 1-methylbenzo[b]-naphtho[2,1-d]thiophene, 6-methylbenzo[b]naphtho[2,1-d]thiophene and 4-methylbenzo[b]naphtho[2,3-[d]thiophene demonstrated mutagenic activity. However, activity was observed only at high concentrations of the metabolic activator.     

In vitro cytotoxic and mutagenic evaluation of thirteen commercial herbal mixtures sold in KwaZulu-Natal,South Africa

Cytotoxic and mutagenic effects of thirteen commercial herbal mixtures sold in KwaZulu-Natal, South Africa were evaluated using the neutral red uptake (NRU) assay and the Ames test. The herbal mixtures tested included Umzimba omubi, Umuthi wekukhwehlela ne zilonda, Mvusa ukunzi, Umpatisa inkosi, Imbiza ephuzwato, Vusa umzimba, Ingwe® muthi mixture, Ibhubezi™, Supreme one hundred™, Sejeso herbal mixture Ingwe®, Lion izifozonke Ingwe®, Stameta™ BODicare® and Ingwe® special muti. The relative cytotoxicity of the herbal mixtures was established by determining their NI₅₀ values (50% inhibition of neutral red uptake). The test revealed that the most toxic herbal mixture was Umpatisa inkosi with an NI₅₀ value of 0.016 mg/mL and the least toxic mixture was Stameta™ BODicare® with an NI₅₀ value of 28.00 mg/mL. The herbal mixtures showed no mutagenic effects against Salmonella typhimurium tester strains TA98, TA100, TA102, TA1535 and TA1537 when the assay was done without S9 metabolic activation. However, four herbal mixtures, Umpatisa inkosi, Imbiza ephuzwato, Vusa umzimba and Stameta™ BODicare® showed mutagenic effects against TA98 but not the rest of the tester strains after using S9 metabolic activation. Umpatisa inkosi also exhibited weak mutagenic activity against TA1535 after metabolic activation. The remaining mixtures did not show mutagenic effects against all the tester strains after S9 metabolic activation. The cytotoxic and mutagenic results reported here offer a step toward determining the safety of commercial herbal mixtures in South Africa. Herbal mixtures showing higher cytotoxic and mutagenic effects need to be further investigated for their possible effects on humans.