Genotoxic activity of potassium permanganate in acidic solutions

Potassium permanganate (KMnO4) combined with sulfuric acid is a strongly oxidizing mixture which has been recommended for the destruction and the decontamination of various mutagens/carcinogens in the publication series of the International Agency for Research on Cancer. Evaluation of the genotoxicity of 4 potassium permanganate solutions was performed using a microtechnique of the Ames test with the tester strains TA97, TA98, TA100 and TA102 with and without metabolic activation. Presence of direct-acting mutagens was detected in all the samples with the tester strain TA102 without S9 mix (163-357 revertants/microliters of the solutions). Three samples containing either acetone or ethanol as an organic solvent also induced a mutagenic response on tester strain TA100 without S9 mix (167-337 revertants/microliters). In addition, DNA damage in human peripheral blood lymphocytes was also measured for one of the mixtures by a new technique: the single-cell gel assay (SCGA). A sample with no organic solvent induced DNA damage in human lymphocytes with a dose-response relationship as determined by SCGA. The major mutagenic agent generated by the permanganate solutions was found to be manganese ion (Mn2+). Both manganese sulfate (MnSO4) and manganese chloride (MnCl2) gave mutagenic dose-response relationships on tester strain TA102 without S9 mix. The mutagenic potencies were 2.8 and 2.4 revertant/nmole for MnSO4 and MnCl2 respectively. MnCl2 also induced DNA damage in human lymphocytes as determined by the SCGA. The genotoxic effects of KMnO4 in acidic conditions were probably mediated by the conversion of MnO4- to Mn2+. KMnO4 in alkaline solutions did not produce mutagenic species and may offer an alternative for the degradation of genotoxic compounds.     

Rec effect of certain textile dyes in Bacillus subtilis

A large number of compounds are toxic, genotoxic, mutagenic, teratogenic and/or carcinogenic. The genotoxicity of four textile dyes commonly used in India namely Sulphur Red Brown 360 (SRB), Jade Green 2G (JG), Reactofix Turquoise Blue 5GFL (RTB) and Direct Scarlet 4BS (DS) was determined by Bacillus subtilis spore Rec assay, both in the presence and absence of metabolizing activation mixture (S9 mix). Each dye was toxic at higher dose levels. A dose-dependent increase in the depth of growth inhibition zones was observed for all dyes. Zones of inhibition were usually clearer at higher doses of the dyes and with Rec- bacteria, but were translucent with Rec+ bacteria. SRB and DS were toxic to Rec+ and Rec- bacteria. JG was less genotoxic in the absence of S9 mix, however, its genotoxic potential increased in the presence of S9 mix. Reactofix T blue was more genotoxic in the absence of S9 mixture.     

Comparison of two extraction procedures for the assessment of sediment genotoxicity: implication of polar organic compounds

Four sediment samples (Va?ne Airport VA, Va?ne Center VC, Va?ne North VN and Reference North RN) were collected in the Berre lagoon (France). Sediments were analyzed for polycyclic aromatic hydrocarbons (PAHs) by use of pressurized fluid extraction with a mixture of hexane/dichloromethane followed by HPLC with fluorescence detection analysis. Organic pollutants were also extracted with two solvents for subsequent evaluation of their genotoxicity: a hexane/dichloromethane mixture intended to select non-polar compounds such as PAHs, and 2-propanol intended to select polar contaminants. Sediment extracts were assessed by the Salmonella/microsome mutagenicity test with Salmonella typhimurium TA98+S9 mix and YG1041±S9 mix. Extracts were also assessed for their DNA-damaging activity and their clastogenic/aneugenic properties by the comet assay and the micronucleus test with Chinese Hamster ovary (CHO) cells. The PAH concentrations were 611ngg(-1)dw, 1341ngg(-1) dw, 613ngg(-1)dw and 482ngg(-1)dw for VA, VC, VN and RN, respectively. Two genotoxic profiles were observed, depending on the extraction procedure. All the non-polar extracts were mutagenic for TA98+S9 mix, and VA, VC, VN sediment samples exerted a significant DNA-damaging and clastogenic activity in the presence of S9 mix. All the polar extracts appeared mutagenic for TA98+S9 mix and YG104±S9 mix, and VA, VC, VN were genotoxic and clastogenic both with and without S9 mix. These results indicate that the genotoxic and mutagenic activities mainly originated from PAHs in the non-polar extracts, while these activities came from other genotoxic contaminants, such as aromatic amines and nitroarenes, in the polar extracts. This study focused on the important role of uncharacterized polar contaminants such as nitro-PAHs or aromatic amines in the global mutagenicity of sediments. The necessity to use appropriate extraction solvents to accurately evaluate the genotoxic hazard of aquatic sediments is also highlighted.     

Differential mutagenic activity of IQ (2-amino-3-methylimidazo[4,5-f]quinoline) in Salmonella typhimurium strains in vitro and in vivo, in Drosophila, and in mice

IQ, a heterocyclic aromatic amine which is formed during the frying of meat, was prepared by chemical synthesis. Its genotoxic potential was studied in bacteria, Drosophila and in mice. A mutagenic effect of IQ (frameshift induction) was detected in Salmonella typhimurium in experiments without metabolic activation; this effect was several orders of magnitude lower than that observed in the presence of an activation system. Ames tests with liver-homogenate S9 fraction from PCB-induced mice and rats confirmed the high mutagenic potency of IQ metabolites (Kasai et al., 1980a). Comparative studies on diagnostic Salmonella strains revealed that the high frameshift-inducing activity is independent of the plasmid pkM101; it is, however, greatly reduced by an intact excision-repair system for DNA lesions. The mutagenic activity of the metabolite(s) formed in vitro by S9 mix has a half-life of ca. 14 min. In the fruit fly, Drosophila melanogaster, IQ induced when used at sublethal concentrations, X-chromosomal recessive lethal mutations in male germ cells in a dose-dependent manner. In mice, tests were performed to detect somatic mutations: chromosomal anomalies (micronuclei) in bone marrow, and gene mutations (affecting coat pigmentation) in mice exposed to IQ in utero. No genotoxic effects were observed in these assays. However, the formation of mutagenic metabolites in the liver of IQ-treated mice was unequivocally demonstrated in host-mediated assays using Salmonella as mutagen probes in mice. The data demonstrate genotoxic activity of IQ in prokaryotic and eukaryotic organisms. The possible reasons for the different response of mammalian systems in vivo and the Salmonella system are discussed.     

Desmutagenic effect of humic acid

In the present study, humic acid was not found to be mutagenic and did not inhibit spontaneous mutation. It did, however, inhibit the mutagenicities of benzo[a]pyrene and 3-aminoanthracene (+S9 mix), but not the mutagenicities of 4NQO, AF-2 and MNNG (-S9 mix). 2-Nitrofluorene and 1-nitropyrene do not need S9 mix for activation of their mutagenicities, but an inhibitory effect was observed. Humic acid exerts a desmutagenic effect on mutagens directly before they act on cells. It does not act as an antimutagen which blocks the processes changing normal cells to mutants. The desmutagenic effect was not decreased by heat treatment (120 degrees C, 15 min). Humic acid was fractionated according to molecular weight and the desmutagenic effect increased with an increase in molecular weight. This effect in the fraction with molecular weight exceeding 300 000 was decreased by centrifugation. The desmutagenic ability of humic acid may result from soluble components and adsorption to small particles.     

Mutagenicity of chloro-olefins in the Salmonella/mammalian microsome test. I. Allyl chloride mutagenicity re-examined

Contrary to findings published up to now, allyl chloride, a well known directly acting mutagen for Salmonella typhimurium, is efficiently activated by rat-liver homogenate (S9 mix) under non-standard mutagenicity testing conditions. Its indirect, S9-mediated mutagenic activity is greatly enhanced when longer than standard preincubation times are applied. The indirect mutagenicity of allyl chloride, thus revealed, greatly exceeds its direct mutagenic activity. Obviously, standard mutagenicity testing conditions cannot be regarded as reliable tools for the evaluation of the full genotoxic potential of allyl chloride and, possibly, of other related compounds.     

The genotoxic and anti-genotoxic effects of <Emphasis Type="Italic">Stachys petrokosmos</Emphasis> leaf extract in human lymphocytes using microsomal fractions

The genotoxic and anti-genotoxic effects of Stachys petrokosmos leaf extracts (Sp) were investigated in human lymphocytes. The cells were treated with 1.5, 3.0 and 6.0 μL/mL concentrations of Sp leaf extracts for 24 and 48 h treatment periods in the absence and presence of metabolic activator (S9mix). In the absence of S9mix, Sp alone did not induce chromosome aberrations and formation of micronucleus while inducing the mean sister chromatid exchange at the highest concentration. In addition, Sp decreased the mutagenic effect of mitomycin-c. Sp alone showed a cytotoxic effect determined by a decrease in the proliferation index, mitotic index and nuclear division index. On the other hand a mixture of Sp and mitomycin-c resulted in a higher cytotoxic effect especially for 48 h treatment period. In the presence of S9mix, Sp was not genotoxic and cytotoxic however, it showed an anti-genotoxic effect by decreasing the effects of cyclophosphamide.     

Mutagenicity and cytotoxicity of 2-methoxyethanol and its metabolites in Chinese hamster cells (the CHO/HPRT and AS52/GPT assays)

2-Methoxyethanol (ethylene glycol monomethyl ether) (EGME), is one of the most commonly used solvents for industrial and consumer products. Although the solvent has been shown to be a reproductive toxin the genotoxic activities of EGME especially its metabolites, have not been adequately investigated. The mutagenicity and cytotoxicity of EGME and its major metabolites, methoxyacetaldehyde (MALD) and methoxyacetic acid (MAA) in Chinese hamster ovary (CHO) cells were therefore examined by us. We have determined the mutagenicity of these compounds at the hypoxanthine-guanine phosphoribosyltransferase (hprt) locus in CHO-K1-BH4 cells (CHO/HPRT assay) and the xanthine-guanine phosphoribosyl transferase (gpt) locus in CHO AS52 cells (AS52/GPT assay). The results show that these chemicals are not mutagenic to the hprt locus in CHO-K1-BH4 cells either with or without rat liver S9 mix as the metabolic activating system. With AS52 cells, only MALD is mutagenic in the absence of S9. It induced a dose-dependent mutagenic response. A dose-dependent cytotoxicity was induced by all compounds in both cell lines. MALD is the most and EGME is the least cytotoxic compounds. Our study shows that a metabolite of EGME, MALD, is highly cytotoxic and likely induces deletion-type mutations in AS52 cells. The genotoxic effect of EGME is, therefore, dependent upon its metabolism and its detection is dependent upon the assays used.     

Mutagen formation on nitrite treatment of indole compounds derived from indole-glucosinolate

The mutagenicities of 8 indole compounds (indole-3-acetonitrile, indole-3-carbinol, indole-3-acetamide, indole-3-acetic acid, 3-methylindole, indole-3-aldehyde, indole-3-carboxylic acid and indole) derived from indole glucosinolate were studied by mutation tests on Salmonella typhimurium TA98 and TA100 and Escherichia coli WP2 uvrA/pKM101 with and without S9 mix. None of the 8 indole compounds were mutagenic, but they became mutagenic on these 3 tester strains when treated with nitrite at pH 3. The nitrite-treated indole compounds were mutagenic without metabolic activation system (S9 mix), and their mutagenicities were decreased by the addition of S9 mix.     

Induction of sister chromatid exchanges and chromosome aberrations in cultured mammalian cells treated with aminophenylnorharman formed by norharman with aniline

Aminophenylnorharman (APNH) is a newly identified mutagenic heterocyclic amine formed by coupling of norharman with aniline in the presence of S9 mix. Furthermore, mutagenic amino-3'-methylphenylnorharman (AMPNH) and aminophenylharman (APH) have been identified from a reaction mixture of norharman and o-toluidine and that of harman and aniline, respectively, with S9 mix. Among these three heterocyclic amines, APNH shows most potent mutagenic activity towards Salmonella typhimurium TA98 and YG1024 with S9 mix. In the present study, the induction of sister chromatid exchanges (SCEs) by APNH was examined in Chinese hamster lung (CHL) cells in vitro, comparing it to those of AMPNH and APH. On incubation with rat S9 for 6h, followed by a recovery culture period of 18h, a dose-dependent effect was found at concentrations between 0.00125 and 0.01 microg/ml for APNH and between 0.3125 and 5 microg/ml for AMPNH and APH. The approximate chemical concentrations leading to a three-fold of control SCE levels calculated from slopes of the linear regressions of induced SCEs were 0.005 for APNH, 0.51 for AMPNH and 1.7 microg/ml for APH. Because of the very strong SCE-causing ability of APNH, we further explored its genotoxicity by examining the induction of chromosome aberrations in CHL cells. A dose-dependent effect was found for chromosome aberrations at concentrations between 0.00125 and 0.04 microg/ml of APNH. The aberrations observed were primarily chromatid exchanges (cte) and breaks (ctb). In conclusion, the potency of SCE induction and clastogenic activity induced by APNH is stronger than Actinomycin D, Mitomycin C (MMC) or 1,8-dinitropyrene which are considered to be the potent clastogens in the literature. Further studies are needed for elucidating mechanisms of the genotoxic actions of these compounds and for evaluating their potential hazards to human health.     

The genotoxicity of lucidin,a natural component of Rubia tinctorum L., and lucidinethylether,a component of ethanolic Rubia extracts

The genotoxic activity of lucidin (1,3-dihydroxy-2-hydroxymethyl-9,10-anthraquinone), a natural component of Rubia tinctorum L., was tested in a battery of short-term tests. The compound was mutagenic in five Salmonella typhimurium strains without metabolic activation, but the mutagenicity was increased after addition of rat liver S9 mix. In V79 cells, lucidin was mutagenic at the hypoxanthine-guanine phosphoribosyl transferase gene locus and active at inducing DNA single-strand breaks and DNA protein cross-links as assayed by the alkaline elution method. Lucidin also induced DNA repair synthesis in primary rat hepatocytes and transformed C3HI M2-mouse fibroblasts in culture. We also investigated lucidinethylether, which is formed from lucidin by extraction of madder roots with boiling ethanol. This compound was also mutagenic in Salmonella, but only after addition of rat liver S9 mix. Lucidinethylether was weakly mutagenic to V79 cells which were cocultivated with rat hepatocytes. The compound did not induce DNA repair synthesis in hepatocytes from untreated rats, but positive results were obtained when hepatocytes from rats pretreated with phenobarbital were used. We conclude that lucidin and its derivatives are genotoxic.Abbreviations DMBA 7,12-dimethylbenz(a)anthracene - HA hydroxyanthraquinones - LUE lucidinethylether - PRH primary rat hepatocytes - UDS unscheduled DNA synthesis     

Mutagenicity of products formed by ozonation of naphthoresorcinol in aqueous solutions

The mutagenicity of products formed by ozonation of naphthoresorcinol in aqueous solution was assayed with Salmonella typhimurium strains TA97, TA98, TA100, TA102 and TA104 in the presence and absence of S9 mix from phenobarbital- and 5,6-benzoflavone-induced rat liver. Ozonated naphthoresorcinol was mutagenic in TA97, TA98, TA100 and TA104 without S9 mix. By the addition of S9 mix, the mutagenic activity of ozonated naphthoresorcinol was markedly suppressed in TA98 and TA100, but became positive in TA102. High-performance liquid chromatography (HPLC) after derivatization to 2,4-dinitrophenylhydrazones demonstrated the formation of glyoxal as an ozonation product of naphthoresorcinol. Ion chromatographic technique also demonstrated the formation of o-phthalic acid, muconic acid, maleic acid, mesoxalic acid, glyoxylic acid and oxalic acid as ozonation products. The mutagenicity assays of these identified products with five Salmonella showed that glyoxal and glyoxylic acid were directly mutagenic; the former in TA100, TA102 and TA104, the latter in TA97, TA100 and TA104. In the presence of S9 mix, glyoxylic acid gave a positive response of mutagenicity for TA102. The experimental evidence supported that glyoxal and glyoxylic acid may contribute to the mutagenicity of ozonated naphthoresorcinol.     

Genotoxicity of apomorphine and various catecholamines in the Salmonella mutagenicity test (Ames test) and in tests for primary DNA damage using DNA repair-deficient B. subtilis strains (rec assay)

Apomorphine, N-nor-N-propyl-apomorphine, dopamine, L-DOPA, 6-hydroxydopamine and adrenaline were evaluated for genotoxicity using the Ames test and DNA repair-deficient and DNA repair-proficient Bacillus subtilis strains (rec assay, H17/M45; HLL3g/HJ-15). In the absence of an S9 liver homogenate, apomorphine induced frame-shift mutations in Salmonella typhimurium, mainly in strain TA1537; no indication of DNA-damaging effects in B. subtilis was observed. N-Nor-N-propyl-apomorphine was tested using strain TA1537 only and found to be mutagenic. Dopamine, L-DOPA, 6-hydroxydopamine and adrenaline were non-mutagenic when tested without S9, whereas they were all more toxic for DNA repair-deficient than for DNA repair-proficient B. subtilis strains, indicating a DNA-damaging potential. In a second set of experiments the mode of action of apomorphine and the relevance of the positive Ames test data were investigated. Glutathione in physiological concentrations reduced the mutagenic effect of apomorphine in a dose-dependent way, both in the presence and the absence of S9. S9 also reduced the mutagenicity of apomorphine. By comparing the effects of a complete S9 mix with those of a preparation without glucose-6-phosphate and NADP, it became clear that S9 also had an activating effect, overshadowed under standard conditions by its deactivating activity. Apomorphine was not mutagenic under anaerobic conditions. Superoxide dismutase and catalase reduced the mutagenic effect of apomorphine. All test conditions which reduced the mutagenic effect also inhibited the dark discoloration of the tester plates, indicating a retardation of apomorphine oxidation. It can, therefore, be concluded that oxidation of apomorphine leads to mutagenic products which induce frame-shift mutations in Salmonella typhimurium. This oxidation was prevented both by glutathione in concentrations well below physiological levels and/or by catalase and superoxide dismutase. Under these conditions, apomorphine was non-mutagenic in therapeutic concentrations as well as at higher dose levels. The possibility of genotoxic side effects occurring in patients treated with apomorphine as an emetic drug is therefore considered to be very unlikely.     

Evaluation of genotoxity and mutagenicity of DL-p-chlorophenylalanine, its methyl ester and some N-acyl derivatives

DL-p-chlorophenylalanine (PCPA) and its derivatives were evaluated for genotoxic effects using Escherichia coli and Bacillus subtilis strains lacking various DNA-repair mechanisms in spottest and in suspension test. The mutagenic activity of studied compounds was determined by the Ames test. Reverse mutation test was performed with Salmonella typhimurium strains TA98, TA100, TA1535 and TA1537 without S9 mix. 0.02 M nitrosomethylurea (NMU) standard mutagen was used as a positive control. The results showed that the parent nonessential amino acid PCPA had no detectable genotoxic and mutagenic activities in bacteria. The methyl ester of this amino acid and its N-phenylacetyl derivative possessed weak genotoxicity. Meanwhile N-sec-butyloxycarbonyl, N-benzyloxycarbonyl, N-(p-nitrophenylacetyl) and N-(p-nitrophenoxyacetyl) derivatives of DL-p-chlorophenylalanine exhibited appreciable genotoxicity. Among the seven tested compounds only N-benzyloxycarbonyl and N-(p-nitrophenoxyacetyl) derivatives of DL-p-chlorophenylalanine have been found to be mutagenic. Only parent PCPA possessed antimutagenic properties in respect of nitrosomethylurea. The structural modification, which strongly affects genotoxicity and mutagenicity perhaps may be due to steric hydrance of the substituents, causing interference with enzyme and DNA interactions.     

Salmonella/mammalian microsome assay with tetranitromethane and 3-nitro-L-tyrosine

The nitrosating agent tetranitromethane (TNM) and the nitrosation product 3-nitro-L-tyrosine (NT) were tested for mutagenic activity in the Salmonella/mammalian microsome assay. TNM showed strong genotoxic activity: it was mutagenic in all tester strains used (TA97, TA98, TA100, and TA102). The maximum mutagenic activity was reached between 16 and 32 micrograms/plate using the standard plate test; higher amounts led to distinct bactericidal effects. The mutagenicity was independent of an in vitro activation system. In the preincubation assay an increased bactericidal effect was observed. In contrast to TNM, NT, the nitrosation product, was non-mutagenic and non-toxic in the standard plate test and with the preincubation method up to 5000 micrograms/plate with and without S9 mix and with all tester strains used. Although TNM is a strong direct-acting mutagen, its nitrosating effect on proteins does lead to nongenotoxic nitro products of tyrosine in proteins.     

Mutagenic activities of cyclophosphamide (NSC-26271) and its main metabolites in Salmonella typhimurium, human peripheral lymphocytes and Chinese hamster ovary cells

Cyclophosphamide (CPA) and its main metabolites were analyzed with respect to their mutagenic activities in Salmonella, human peripheral lymphocytes (PL), and Chinese hamster ovary (CHO) cells. In Salmonella, the compounds were activated with S9 mix from rat livers, which were unstimulated or stimulated with Aroclor 1254 or phenobarbital. For the enzyme inducers the following order of efficiency was found for all test compounds except carboxyphosphamide: phenobarbital greater than Aroclor 1254 greater than non-induced. The most potent mutagens in all 3 test systems were 4-OH-CPA, PAM and nor-HN2. S9 mix transforms 4-OH-CPA to strong mutagenic compounds in the Salmonella assay. All metabolites tested in the Salmonella assay were activated by S9 mix to higher mutagenic potential.     

The mutagenic and clastogenic activity of tobacco smoke

Employing the Salmonella/microsome mutagenicity assay it was established that the mutagenic effect of tobacco smoke (TS) (240 cm3 in a 16-l glass chamber, at 1 min or 5 min exposure time) in S. typhimurium TA98 depended on the type of S9 mix used. Addition of S9 mix obtained from the liver of 3-methylcholanthrene- or Aroclor-1254-pretreated rats but not from the liver of phenobarbital-pretreated or untreated rats was required to demonstrate the mutagenic activity of TS. One might suggest that polycyclic aromatic hydrocarbons were involved in TS-induced mutagenesis in S. typhimurium TA98. In addition, treatment of BDF1 mice with TS (600 cm3 TS in a 14-l glass chamber, 2-6 exposures of 30 min each with a 1-min interval between them during which a total change of the air was made) caused an up to 3.5-fold increase of the number of micronucleated polychromatic erythrocytes (PCE) in mouse bone marrow detected 24 h after the TS exposure. Furthermore, a stable 2-5-fold elevation of the number of micronucleated normochromatic erythrocytes (NCE) was detected in the peripheral blood of mice treated daily (2 x 30 min) with TS, starting 48 h after the first TS exposure. The application of the micronucleus test in mouse peripheral blood, a more convenient and useful approach for detecting the chronic clastogenic activity of TS, allowed us to establish the cumulative genotoxic effect of TS in mice.     

Evaluation of the mutagenic potential of endosulfan using the Salmonella/mammalian microsome assay

The mutagenic potential of endosulfan, a polychlorinated insecticide, was assessed using the highly sensitive Salmonella tester strains TA97(a), TA98, TA100 and TA102. It exhibited a toxic effect at dose levels of 50 micrograms/plate and higher. Plate incorporation studies did not show mutagenic response with any of the tester strains used. A modification of the assay using a preincubation procedure showed mutagenic activity with and without metabolic activation with TA97(a) only. Increased toxicity was observed after addition of phenobarbital-induced S9 mix.     

Studies by means of the SCE assay in V79-E Chinese hamster cells on the mode of action of tri-substituted nitrosoureas

The genotoxic activity of 3,3-diethyl-1-methyl-1-nitrosourea ( DEMNU ), 1,3-dimethyl-3-phenyl-1-nitrosourea ( DMPNU ) and 1-chloroethyl-3-methyl-3-phenyl-1-nitrosourea ( CEMPNU ) was studied in the SCE assay in V79-E cells in vitro. These compounds are very stable in aqueous solutions, but are directly acting genotoxins . The SCE rates increase linearly with the length of the incubation period. This direct activity is presumably due to an intracellular catalytic decomposition. Whereas the SCE-inducing effect of DMPNU and CEMPNU is not influenced by addition of S9 mix, that of DEMNU is strongly potentiated by rat and Syrian hamster S9 mix. This DEMNU activation is an NADPH-dependent enzymatic reaction and is inducible by phenobarbital. The absence of a direct mutagenic effect of DEMNU in the Ames test, as reported by other authors, is probably caused by a striking insensitivity to tri-substituted nitrosoureas of the Salmonella assay. This assumption was substantiated by long-term application of very low DMPNU doses to V79-E. Long-term simultaneous treatment with DMPNU and bromodeoxyuridine (BUdR) significantly diminished the rate of SCE induction.     

Factors affecting the mutagenic activity of quercetin for Salmonella typhimurium TA98: metal ions, antioxidants and pH

The mutagenic activity of quercetin for Salmonella typhimurium TA98 was inhibited by addition of metal salts. MnCl2 was a potent inhibitor, followed by CuCl2, FeSO4, and FeCl3, the probable mechanism being facilitated catalytic oxidation of quercetin. With quercetin incorporated at a level of 100 nmoles/plate, approximate doses (nmoles/plate) to give 50% inhibition of mutagenic activity were: MnCl2 less than 10 (-S9), 18 (+S9); CuCl2 65 (-S9), greater than 100 (+S9); FeSO4 190 (-S9), greater than 300 (+S9); or FeCl3 275 (-S9), greater than 300 (+S9). Ascorbate, superoxide dismutase, and, to a lesser extent, NADH and NADPH, all enhanced the mutagenic activity of quercetin in the absence of the mammalian-microsome (S9) system, but had no significant effect in the presence of the S9 mix. The maximum enhancement of activity by ascorbate or superoxide dismutase was approximately 87% of the increase achieved by addition of the S9 mix. Tyrosinase (catechol oxidase) substantially reduced the mutagenic activity of quercetin in the absence of the S9 mix. At lower levels of tyrosinase, activity was restored by incorporation of the S9 mix. It is proposed that the S9 mix enhances the mutagenic activity of quercetin by scavenging superoxide radicals, thus inhibiting the autoxidation of quercetin, and possibly by reducing quinone oxidation products of quercetin. The mutagenic activity of quercetin increased substantially when the pH of the media was decreased. This may be due in part to a decrease in ionization of quercetin at lower pH, thereby increasing its absorption by the tester strain, to a decrease in the rate of autoxidation of quercetin at lower pH, or to a combination of these.