Mutagenicity of the mycotoxin emodin in the salmonella/microsome system.

The mycotoxin emodin was found to be a frameshift mutagen for Salmonella typhimurium strain TA 1537 after metabolic activation in a mammalian microsome system.     

Mutagenicity of the mycotoxin emodin in the salmonella/microsome system.

The mycotoxin emodin was found to be a frameshift mutagen for Salmonella typhimurium strain TA 1537 after metabolic activation in a mammalian microsome system.     

Mutagenicity of anthraquinone and hydroxylated anthraquinones in the Ames/Salmonella microsome system.

The mutagenicity of anthracene, anthraquinone, and four structurally similar compounds of each was evaluated in the Ames/Salmonella microsome assay. Anthraquinone was shown to be mutagenic for strains TA1537, TA1538, and TA98 in the absence of rat liver homogenate. The four anthraquinone derivatives tested were mutagenic for TA1537 exclusively. None of the anthracenes exhibited mutagenic activity.     

Evaluation of 1,3-pentadiene for mutagenicity by the Salmonella/mammalian microsome assay

1,3-Pentadiene, a food contaminant produced by some molds when they metabolize sorbic acid, was tested for mutagenicity, using variations of the Salmonella/mammalian microsome assay. The chemical was incorporated into the test system (with and without S9 mix) by 3 methods: (a) the standard plate incorporation assay, (b) a liquid preincubation procedure and (c) exposure of test bacteria in the soft agar overlay to gaseous 1,3-pentadiene. The chemical was extremely toxic to the test bacteria with amounts as low as 2.0 microgram/plate causing cell death. However, none of the nonlethal concentrations tested by any of the methods was mutagenic to Salmonella typhimurium strains TA97, TA98, TA100, TA1535, TA1537 or TA1538.     

Lack of mutagenicity and putative carcinogenicity of several novel benzopyrene derivatives.

Several novel benzopyrene derivatives with the same gross structure and the same electronic periphery as benzo(a)pyrene, but with some alteration in the complete electronic structure, when tested in the Ame's Salmonella/microsome test (TA 1537, TA 100 and TA 98], were found to lack mutagenicity and, therefore, putative carcinogenicity.     

Mutagenicity of plant flavonols in the Salmonella/mammalian microsome test: activation of flavonol glycosides by mixed glycosidases from rat cecal bacteria and other sources.

Over 70 naturally occurring and synthetic flavonoids were screened for mutagenicity with 5 tester strains in the Salmonella/mammalian microsome assay: TA1535, TA100, TA1537, TA1538 and TA98. Frameshift mutagenicity was confined to the flavonols (flavon-3-ols) in strain TA98, TA1537 and TA100. The two most mutagenic falvonols, namely, quercetin (3,3',4',5,7-pentahydroxyflavone) and kaempferol (3,4',5,7-tetrahydroxyflavone), exhibiting 12 and 7 revertants/nmol in TA98 respectively, are also the most common flavonols occurring in plants. Other flavonols exhibited less activity (revertants/nmol): galangin (2.0), rhamnetin (0.45), kaempferide (0.24), fisetin (0.14), myricetin (0.12), robinetin (0.06) and morin (0.05). All of these flavonols apparently exhibited significant activation by Aroclor 1254 induced rat-liver microsome preparations (S9). However, subsequent study revealed that only those flavonols either lacking or possessing one B ring hydroxyl group had an absolute requirement for microsomal activation. Alternatively, quercetin with two B-ring OH groups is not activated by microsomal enzymes, but by soluble (S100) enzymes from liver which are apparently constitutive and not subject to the usual chemical induction. 3 flavonol glycosides, namely, quercetrin (quercetin-3-O-rhamnoside), rutin (quercetin-3-O-rutinoside) and robinin (kaempferol-3-O-galactosido-rhamnoside-7-O-rhamnoside), were found to be nonmutagenic. They could, however, be activated by a variety of mixed glycosidases incorporated in the usual pour plate procedure. The most effective enzyme mixtures were obtained from rat cecal bacteria and from the snail Helix pomatia.     

Absence of mutagenic activity of ticlopidine in the Ames Salmonella/microsome test

Ticlopidine hydrochloride, 5-(o-chlorobenzyl)-4,5,6,7-tetrahydrothieno[3,2-c]pyridine hydrochloride, a platelet aggregation inhibitor, was tested for mutagenic activity in the Ames Salmonella/mammalian microsome test. Salmonella typhimurium strains TA98, TA100, TA1535, TA1537 and TA1538 were employed. Two of these strains (TA1535 and TA100) are sensitive to base-pair substitution mutagens, and the remaining 3 are sensitive to frame-shift mutagens. There was no evidence that ticlopidine hydrochloride had any mutagenic activity either in the presence or absence of a liver microsomal supplement.     

Absence of mutagenic activity of benorylate, paracetamol and aspirin in the Salmonella/mammalian microsome test

Benorylate and its two major hydrolysis products, paracetamol and aspirin were examined for mutagenicity in the Salmonella/mammalian microsome screening test. The compounds were tested in 6 strains of Salmonella typhimurium (TA1535, TA1537, TA1538, TA100, TA97 and TA98) in the presence and absence of a rat-liver microsome activation system. Benorylate did not show evidence of mutagenic activity in the 6 strains tested with or without metabolic activation at concentrations ranging from 0.006 to 3 mg per plate. Paracetamol and aspirin likewise did not show any evidence of mutagenic activity at concentrations ranging from 0.1 to 50 mg per plate for the former and 0.01 to 50 mg per plate for the latter.     

A new Salmonella tester strain,TA97, for the detection of frameshift mutagens: A run of cytosines as a mutational hot-spot

We have developed a new Salmonella tester strain, TA97, for use in the Salmonella/microsome mutagenicity test. DNA sequencing has shown that this strain contains and added cytosine, resulting in a run of six cytosines at the mutated site in the histidine D gene. Its mutagenic specificity is similar to that of the frameshift mutagen tester strain, TA1537, which also contains an added cytosine in a run of cytosines and is currently among the five standard tester strains used for general mutagen screening. We assessed the mutagenic potency of 21 frameshift mutagens for TA1537 and TA97. TA97 was considerably more sensitive than TA1537 to reversion by these frameshift mutagens. In addition, one agent, PR toxin (from Penicillium roqueforti), which was not detected by any of the previously existing standard tester strains, did revert TA97; and two substituted aryl-alkyl triazenes, which had not been reported previously to be frameshift mutagens, were mutagenic in this new tester strain. We suggest that TA1537 be replaced by TA97 for general screening of mutagenicity.     

Mutagenicity of islandicin and chrysophanol in the Salmonella/microsome system.

Chrysophanol and islandicin, two anthraquinones which are structurally related to emodin, were found to be frame-shift mutagens for Salmonella typhimurium strain TA 1537 after metabolic activation.     

Genotoxicity of naturally occurring hydroxyanthraquinones

A variety of structurally related hydroxyanthraquinones (HA) were investigated in a test battery for the evaluation of mutagenicity and cell-transforming activity. The tests were: (1) the Salmonella typhimurium mutagenicity assay, (2) the V79-HGPRT mutagenicity assay, (3) the DNA-repair induction assay in primary rat hepatocytes and (4) the in vitro transformation of C3H/M2 mouse fibroblasts. In Salmonella, most of the tested compounds were mutagenic in strain TA1537, but only a few were active in other strains. Among these were HA with a hydroxymethyl group, such as lucidin and aloe-emodin. In V79 cells, only HA with 2 hydroxy groups in the 1,3 positions (1,3-DHA, purpurin, emodin) or with a hydroxymethyl sidechain (lucidin and aloe-emodin) were mutagenic. The compounds found to be active in V79 cells were also active in the DNA-repair assay and in the C3H/M2 transformation assay. Thus, it appears that the genotoxicity of HA is dependent on certain structural requirements.     

Absence of mutagenic activity of benorylate,paracetamol and aspirin in the Salmonella//mammalian microsome test

Benorylate and its two major hydroyssis products, paracetamol and aspirin were examined for mutagenicity in the Salmonella/mammalian microsome screening test. The compounds were tested in 6 strains of Salmonella typhimurium (TA1535, TA1537, TA1538, TA100, TA97 and TA98) in the presence and absence of a rat-liver microsome activation system. Benorylate did not show evidence of mutagenic activity in the 6 strains tested with or without metabolic activation at concentrations ranging from 0.006 to 3 mg per plate. Paracetamol and aspirin likewise did not show any evidence of mutagenic activity at concentrations ranging from 0.1 to 50 mg per plate for the former and 0.01 to 50 mg per plate for the latter.     

Investigations on DNA binding in rat liver and in Salmonella and on mutagenicity in the Ames test by emodin, a natural anthraquinone

Emodin (1,6,8-trihydroxy-3-methylanthraquinone), an important aglycone found in natural anthraquinone glycosides frequently used in laxative drugs, was mutagenic in the Salmonella/mammalian microsome assay (Ames test) with a specificity for strain TA1537. The mutagenic activity was activation-dependent with an optimal amount of S9 from Aroclor 1254-treated male Sprague-Dawley rats of 20% in the S9 mix (v/v) for 10 micrograms emodin per plate. Heat inactivation of the S9 for 30 min at 60 degrees C prevented mutagenicity. The addition of the cytochrome P-448 inhibitor 7,8-benzoflavone (18.5 nmoles per plate) reduced the mutagenic activity of 5.0 micrograms emodin per plate to about one third, whereas the P-450 inhibitor metyrapone (up to 1850 nmoles per plate) was without effect. To test whether a metabolite binds covalently to Salmonella DNA, [10-(14)C]emodin was radiosynthesized, large batches of bacteria were incubated with [10-(14)C]emodin and DNA was isolated. [G-3H]Aflatoxin B1 (AFB1) was used as a positive control mutagen known to act via DNA binding. DNA obtained after aflatoxin treatment could be purified to constant specific activity. With emodin, the specific activity of DNA did not remain constant after repeated precipitations so that it is unlikely that the mutagenicity of emodin is due to covalent interaction of a metabolite with DNA. The antioxidants vitamin C and E or glutathione did not reduce the mutagenicity. Emodin was also negative with strain TA102. Thus, oxygen radicals are probably not involved. When emodin was incubated with S9 alone for up to 50 h before heat-inactivation of the enzymes and addition of bacteria, the mutagenic activity did not decrease. It is concluded that the mutagenicity of emodin is due to a chemically stable, oxidized metabolite forming physico-chemical associations with DNA, possibly of the intercalative type. In order to check whether an intact mammalian organism might be able to activate emodin to a DNA-binding metabolite, radiolabelled emodin was administered by oral gavage to male SD rats and liver DNA was isolated after 72 h. Very little radioactivity was associated with the DNA. Considering that DNA radioactivity could also be due to sources other than covalent interactions, an upper limit for the covalent binding index, CBI = (mumoles chemical bound per moles DNA nucleotides)/(mmoles chemical administered per kg body weight) of 0.5 is deduced. This is 10(4) times below the CBI of AFB1.(ABSTRACT TRUNCATED AT 400 WORDS)     

The effect of alkoxy substituents on the mutagenicity of some aminoazobenzene dyes and their reductive-cleavage products

15 aminoazobenzene dyes and 7 of their reductive-cleavage products were examined in the Salmonella/microsome assay with strains TA98, TA100, TA1535, TA1537 and TA1538. Dyes tested included 5 derivatives of 4-aminoazobenzene with different alkoxy substituents (-OCH3, -OCH2CH3, -OCH2CH2 CH3, -OCH2CH2CH2CH3 or -OCH2CH2OH) in the 8-position as well as the corresponding derivatives of 4-[(4-aminophenyl)azo]-N,N-diethylaniline and 4-[(4-aminophenyl)azo]-N,N-bis(2-hydroxyethyl)aniline. In general, as the size of the substituent ortho to the primary amino group of the dyes was increased, the mutagenicity decreased. A similar trend was observed for the reductive-cleavage products. The results from the latter aspect of this study suggest that the mutagenicity of aminoazobenzene dyes can not be accounted for solely from the properties of their reductive-cleavage products.     

In vitro mutagenicity and cell-transformation screening of N-(2,3-epoxy-propyl)-phthalimide.

N-(2,3-Epoxy-propyl)-phthalimide (EPP) was tested for genetic activity in the Salmonella/microsome mutagenicity test. Concentration-dependent mutagenicity was demonstrated in S. typhimurium strains TA1535, TA1537 and TA100 with and without rat S9. It was inactive in strain TA1538, and active without rat S9 in TA98 at the high dose. EPP induced 6-thioguanine-resistant mutants of Chinese hamster ovary cells in the absence of an exogenous activating system. EPP produced dose-dependent enhancement of SA7 virus transformation of primary hamster-embryo cells, and transformed secondary hamster-embryo cells in a non-dose-related fashion. At a dose of 5 g/kg p.o. or i.m., EPP was inactive in the host-mediated assay using C57Bl/6XC3H mice and S. typhimurium strain TA1535. Murine testicular DNA synthesis was not inhibited by oral administration of EPP at 1000 mg/kg.     

Mutagenicity of dipyridyls and their methylated derivatives in Salmonella typhimurium/rat liver microsome system

The Salmonella/microsome assay with strains TA97, TA98, TA100, TA1535, TA1537 and TA1538 was used to examine the potential mutagenicity of 5 dipyridyls, 1 tripyridyl, 3 dipyridinium diiodides and 2 pyridinium monoiodides. The widely used herbicide paraquat (1,1-dimethyl-4,4'-dipyridinium diiodide) and its precursor 4,4'-dipyridyl gave weak and marginal mutagenic activity to Salmonella typhimurium TA1535 and TA1538 in the presence of S9-mix. Significantly high mutagenicity was obtained with 2,2'-, 3,3'-, 2,3'-, and 2,4'-dipyridyls, 2,2',2"-tripyridyl, and 5 pyridinium salts under the same conditions. The positive mutagenic response of 2,2',2"-tripyridyl suggests that higher polymers of pyridine contaminating paraquat preparations might be mutagenic. The dose-response curves of 1,1-dimethyl-3,3'-dipyridinium diiodide and 1,1'-dimethyl-2,2'-dipyridinium diiodide revealed an exponential relationship between the number of induced revertants and the compound concentrations. The results suggested that the mechanism of mutation induced by these two compounds might be attributed to the chain reactions of their free-radicals with molecular oxygen.     

Ames Salmonella/mammalian-microsome testing of peptides and peptide synthesis reagents

The Ames Salmonella/mammalian-microsome assay was used to evaluate the bacterial mutagenicity of 6 bioactive peptides and of 11 chemical reagents used in peptide synthesis. Samples of 2 reagents, bis(2-oxo-3-oxazolidinyl)phosphinic chloride and fluoren-9-ylmethyl chloroformate, showed mutagenic activity with strains TA100 and TA1535, and with TA1537, respectively. No mutagenic activity was found with the bioactive peptides or with the other 9 peptide synthesis reagents.     

Mutagenic activity of nine N,N-disubstituted hydrazines in the Salmonella/mammalian microsome assay.

The mutagenic activity of N,N-dimethyl-, N,N-diethyl-, N,N-dibutyl-, N,N-diisobutyl-, N,N-di(p-tolyl)-, N-ethyl-N-phenyl-, N,N-dibenzyl-, N,N-diphenyl- and N,N-diisopropylhydrazine was examined in the Salmonella/mammalian microsome assay using the strains TA1535, TA1537, TA97, TA98, TA100, TA102 and TA1530. All nine hydrazines were mutagenic in at least one tester strain, although of borderline significance for some of the compounds. The mutagenic potencies of the hydrazines varied 2-3 orders of magnitude, from very weak to moderate mutagenic activity. In general, the addition of S9 resulted in a lowering of the mutagenic activity and a lowering of the toxic properties of the hydrazines. The test results were relatively difficult to evaluate due to toxic effects of many of the test compounds on the test bacteria which may have resulted in an underestimation of the mutagenic potencies of some of the compounds. The pattern of mutagenic activity of the hydrazines in the different tester strains indicates that more than one mechanism of action may be involved in the mutagenicity.     

Mutagenicity of the coccidiostat diaveridine in the Salmonella/mammalian microsome assay

The coccidiostat diaveridine was tested for mutagenicity in the Salmonella/microsome assay with tester strains TA100 and TA98. This compound was not mutagenic in either tester strain in the presence and absence of rat S9 mix, but was found to be mutagenic in strain TA100 after metabolic activation with hamster S9 mix.     

Mutagenicity testing of condensates of smoke from titanium dioxide/hexachloroethane and zinc/hexachloroethane pyrotechnic mixtures

Condensates of smoke from titanium dioxide/hexachloroethane and zinc/hexachloroethane pyrotechnic mixtures were investigated for their potential to produce genetic damage in the tester strains TA98, TA100, TA1535 and TA1537 of Salmonella typhimurium and in the mouse bone marrow micronucleus assay. Both smoke condensates contained several chlorinated hydrocarbons among which tetrachloroethylene, hexachloroethane, hexachlorobutadiene and hexachlorobenzene were identified by GC/MS. Condensate of smoke from titanium dioxide/hexachloroethane showed a dose-related positive response in the Salmonella assay with strains TA98 and TA100 in the absence of metabolic activation from rat liver S9 fraction. Both smoke condensates were negative in the micronucleus assay but produced a small but significant depression of erythropoietic activity. The results indicate that smoke condensate from titanium dioxide/hexachloroethane mixtures contains unidentified compound(s) that may be considered mutagenic in the Salmonella assay.