Mutagenicity profile of atmospheric particulate matter in a small urban center subjected to airborne emission from vehicle traffic and sugar cane burning

Atmospheric particulate matter (PM) is genotoxic and recently was classified as carcinogenic to humans by the International Agency for Research on Cancer. PM chemical composition varies depending on source and atmospheric conditions. The Salmonella/microsome assay is the most used mutagenicity test and can identify the major chemical classes responsible for observed mutagenicity. The objective of this work was to characterize the mutagenicity of PM samples from a countryside city, Limeira, Brazil, which is influenced by heavy traffic and sugar cane biomass burning. Six samples of total PM were collected. Air mass backward trajectories were calculated. Organic extracts were assayed using the Salmonella/microsome microsuspension mutagenicity assay using TA98, YG1041, and TA1538, with and without metabolic activation (S9). YG1041 was the most sensitive strain and mutagenicity reached 9,700 revertants per m³ without metabolic activation. Potency for TA1538 was higher than TA98, indicating that this strain should be considered in air mutagenicity studies. The increased response to YG1041 relative to TA98, and the decreased response with S9, suggests that nitroaromatics are the major contributors. Limeira is among the most mutagenic cities in the world. High mutagenicity in Limeira seems to occur when the air mass from the area of sugarcane production is mixed with air from the region impacted by anthropogenic activities such as traffic. An increase in the formation of nitro‐polycyclic aromatic hydrocarbons may result from longer contact time between the aromatic compounds and the atmosphere with high NOx and ozone concentration, although more studies are required to confirm this hypothesis. Environ. Mol. Mutagen. 57:41–50, 2016. © 2015 Wiley Periodicals, Inc.     

Increased mutagenicity of chromium compounds by nitrilotriacetic acid

Nitrilotriacetic acid trisodium salt (NTA), which is a substitute for polyphosphates in household laundry detergents, and N-nitrosoiminodiacetic acid (NIDA), a derivative of NTA produced by metabolism of soil microorganisms, were tested for in vitro mutagenicity in bacteria and yeasts. No gene reversions in five strains of Salmonella typhimurium (TA 1535, TA1537, TA1538, TA98, and TA100), no forward gene mutations in Schizosaccharomyces pombe P1, and no mitotic gene conversions at two loci in Saccharomyces cerevisiae D4 were induced by NTA (up to 870 micrograms/plate or 40 micrograms/ml) and NIDA (up to 2,000 micrograms/plate or 1,000 micrograms/ml), independently of the presence of rat liver metabolic activation. The influence of NTA on the mutagenic and clastogenic activity of several chromium compounds was examined in the Salmonella/microsome assay and in the sister chromatid exchange (SCE) assay in mammalian cell cultures (Chinese hamster ovary [CHO] line). NTA does not affect the genetic inactivity of water-soluble Cr(III) (Cr2[SO4]3) and the direct mutagenicity of soluble Cr(VI) (Na2CrO4,K2Cr2O7) compounds. The very insoluble Cr(VI) compounds PbCrO4 and PbCrO4 X PbO are instead clearly mutagenic in the Salmonella/microsome assay (TA100 strain) only in the presence of NTA or NaOH. The mutagenicity of lead chromates is correlated with the amounts of Cr(VI) solubilized by NTA or alkali, as detected by the colorimetric reaction with diphenylcarbazide and atomic absorption spectrophotometry. In the SCE assay, the insoluble lead chromates are directly clastogenic owing to prolonged treatment conditions and cellular endocytosis. The chromosome-damaging activity of PbCrO4 is significantly increased by NTA but not by NaOH.     

Effect of bacterial concentration on reversions induced in salmonella typhimurium TA1538 by N-hydroxy-2-acetylaminofluorene

Reversions induced by N-hydroxy-2-acetylaminofluorene (N-OH-AAF) were measured in the Salmonella/microsome quantitative plate assay using various concentrations of Salmonella typhimurium strain TA 1538. The number of induced revertants increased with an increasing number of bacteria/plate, but the variation in reversion frequency was not as great as the variation in bacterial concentration. The effects of bacterial concentration on reversion fixation, phenotypic expression, selection of his⁺ revertants, and the interaction of the mutagen with bacterial DNA were examined. Suspension cultures of TA 1538 were exposed to N-OHAAF and various dilutions were prepared and assayed for revertants/10⁸ bacteria. Reversion frequencies were very dependent on bacterial concentration between approximately 0.2 × 10⁸ and 2 × 10⁸ bacteria/plate. Revertants/10⁸ TA 1538 were reduced above about 2 × 10⁸ bacteria/plate, indicating that culture conditions limited reversion fixation, expression and/or selection at these concentrations. The number of spontaneous revertants/plate determined both from bacteria exposed to dimethyl sulfoxide in suspension culture and in the Salmonella/microsome assay were not greatly affected by bacterial concentration. To study the effect of bacterial concentration on the interaction of the mutagen with bacterial DNA, various concentrations of TA 1538 were exposed to the same dose of N-OH-AAF. Both revertants/10⁸ TA 1538 and DNA adducts varied inversely with bacterial concentration. The effect of bacterial concentration on both reversion fixation and/or expression and on mutagen binding to DNA may influence reversion frequencies in the Salmonella/microsome assay.     

SOS induction in Escherichia coli and Salmonella mutagenicity: a comparison using 330 compounds

To examine the concordance of two microbial genotoxicity short-termassays, 330 experimental results for the SOS chromotest usingtester strain Escherichia coli PQ37 were compared with the resultsof the Salmonella/mammalian microsome mutagenicity assay withSalmonella typhimurium TA97, TA98, TA100, TA102, TA104, TA1535,TA1537 and/or TA1538. With respect to qualitative features,the concordance between SOS chromotest and Salmonella mutagenicitytest results was 86.4% (sensitivity, 78.6%; specificity, 100%;     

Evaluation of the mutagenicity of an N-nitroso contaminant of the sunscreen Padimate O: N-nitroso-N-methyl-p-aminobenzoic acid, 2-ethylhexyl ester (NPABAO).

The nitrosamine contaminant, N-nitroso-N-methyl-p-aminobenzoic acid, 2-ethylhexyl ester (NPABAO), of the major sunscreen ingredient Padimate O (4-N,N'-dimethylamino-benzoic acid, 2-ethylhexyl ester) was synthesized and tested for mutagenicity in the Salmonella typhimurium and mouse lymphoma L5178Y TK +/- assays. In contrast to the previously reported positive responses in S. typhimurium tester strains TA100 and TA1535 [Loeppky et al., 1991], there were no increases in the number of revertants with strains TA98, TA100, TA1535, and TA1538 in either the Salmonella plate incorporation [Ames et al., 1975] or preincubation [Yahagi et al., 1977] assays. Additional testing with Salmonella, following the modified preincubation procedure [Rogan, 1990] that gave the initial positive response, was also negative. Data from the mouse lymphoma assays were also uniformly negative. During synthesis of NPABAO, small amounts of 4-N,N'-dimethylamino-3-nitrobenzoic acid, 2-ethylhexyl ester (DMANBAO) can be formed. To determine whether the reported positive mutagenicity response of NPABAO could be the result of trace amounts of DMANBAO in the NPABAO, that compound was also synthesized and tested for mutagenicity with Salmonella. Positive responses were obtained with tester strains TA98 and TA 1538 but not with TA100 and TA1535, indicating that DMANBAO was not responsible for the increase in revertants originally reported.     

A preliminary characterization of the mutagenicity of atmospheric particulate matter collected during sugar cane harvesting using the Salmonella/microsome microsuspension assay

During sugar cane harvesting season, which occurs from May to November of each year, the crops are burnt, cut, and transported to the mills. There are reports showing that mutagenic activity and PAH content increase during harvesting season in some areas of São Paulo State in comparison with nonharvesting periods. The objective of this work was to preliminarily characterize the mutagenic activity of the total organic extracts as well as corresponding organic fractions of airborne particulate matter (PM) collected twice from two cities, Araraquara (ARQ) and Piracicaba (PRB), during sugar cane harvesting season using the Salmonella/microsome microssuspension assay. One sample collected in São Paulo metropolitan area was also included. The mutagenicity of the total extracts ranged from 55 to 320 revertants per cubic meter without the addition of S9 and from not detected to 57 revertants per cubic meter in the presence of S9 in areas with sugar cane plantations. Of the three fractions analyzed, the most polar ones (nitro and oxy) were the most potent. A comparison of the response of TA98 with YG1041 and the increased potencies without S9 indicated that nitro compounds are causing the observed effect. More studies are necessary to verify the sources of the mutagenic activity such as burning of vegetal biomass and combustion of heavy duty vehicles used to transport the sugar cane to the mills. The Salmonella/microsome assay can be an important tool to monitor the atmosphere for mutagenicity during sugar cane harvesting season. Environ. Mol. Mutagen. 2008. © 2008 Wiley‐Liss, Inc.     

Use of genetically manipulated Salmonella typhimurium strains to evaluate the role of human sulfotransferases in the bioactivation of nitro‐ and aminotoluenes

Various nitro‐ and aminotoluenes demonstrated carcinogenic activity in rodent studies, but were inactive or weakly active in conventional in vitro mutagenicity assays. Standard in vitro tests do not take into account activation by certain classes of enzymes. This is true in particular for sulfotransferases (SULTs). These enzymes may convert aromatic hydroxylamines and benzylic alcohols, two major classes of phase‐I metabolites of nitro‐ and aminotoluenes, to reactive esters. Here it is shown that expression of certain human SULTs in Salmonella typhimurium TA1538 or TA100 strongly enhanced the mutagenicity of various nitrotoluenes and nitro‐ and amino‐substituted benzyl alcohols. Human SULT1A1, SULT1A2, and SULT1C2 showed the strongest activation. The observation that some nitrotoluenes as well as some aminobenzyl alcohols were activated by SULTs in the absence of cytochromes P450 implies that mutagenic sulfuric esters were formed at both the exocyclic nitrogen and the benzylic carbon, respectively. Nitroreductase deficiency (using strain YG7131 instead of TA1538 for SULT1A1 expression) did not affect the SULT‐dependent mutagenicity of 1‐hydroxymethylpyrene (containing no nitro group), moderately enhanced that of 2‐amino‐4‐nitrobenzyl alcohol, and drastically attenuated the effects of nitrobenzyl alcohols without other substituents. The last finding suggests that either activation occurred at the hydroxylamino group formed by nitroreductase or the nitro group (having a strong –M effect) had to be reduced to an electron‐donating substituent to enhance the reactivity of the benzylic sulfuric esters. The results pointed to an important role of SULTs in the genotoxicity of nitrotoluenes and alkylated anilines. Activation occurs at nitrogen functions as well as benzylic positions. Environ. Mol. Mutagen. 57:299–311, 2016. © 2016 Wiley Periodicals, Inc.     

Structure-mutagenicity relationships of four amino-imidazonaphthyridines and imidazoquinolines

We tested four isomeric imidazonaphthyridines and one imidazoquinoline compound for mutagenic activity in the Ames/Salmonella mutagenicity assay, using strain TA98 and strain YG1024, an analogue of strain TA98 with elevated O-acetyl-transferase levels. Their potency was related to calculated electronic parameters. Five compounds with a linear arrangement of 3 rings showed a positive response in strain YG1024. Compound 2 (1-methyl-imidazo[4,5-b][1,7]naphthyridin-2-amine) is the most mutagenic in both strains, giving specific activities of about 200 and 30 revertants per microgram in strains YG1024 and TA98, respectively. Three of the compounds were weak mutagens, giving a positive dose-response only in strain YG1024, with 3–5 revertants per microgram. A higher response of all five compounds in strain YG1024 as opposed to TA98 indicates that they require O-acetyltransferase activity for their metabolism. Mutagenic potencies in strain YG1024 were positively correlated to the energy of the LUMO (lowest unoccupied molecular orbital) of the nitrenium ion. © 1995 Wiley-Liss, Inc.     

1-nitropyrene as a marker for the mutagenicity of diesel exhaust-derived particulate matter in workplace atmospheres

The use of 1-nitropyrene (1-NP) as a marker for the occupational exposure to diesel exhaust (DE) mutagens was investigated in workplace atmospheres contaminated with DE from a variety of emission sources, such as power supplies, forklifts, trucks, caterpillar vehicles, trains, ships' engines, and vehicles in city traffic. Total suspended particulate matter was collected by area sampling. The 1-NP content of acetone extracts of these samples as determined by gas chromatography-high resolution mass spectrometry varied from 0.080 to 17 μg/g acetone extractable matter, corresponding to air concentrations of 0.012 to 1.2 ng/m³. A sample collected in a rural area contained 0.0017 ng/m³ 1-NP. The mutagenicity of the extracts was tested in the Salmonella typhimurium strains TA98 and TA1538, using the microsuspension assay with and without metabolic activation by an exogeneous metabolizing system (rat liver S9-fraction). In addition, the S. typhimurium strains YG1021 and YG1024 were used because of their high sensitivity towards the mutagenicity of nitro polycyclic aromatic hydrocarbons. When plotting the mutagenic potency of the air sample extracts as determined in the absence of liver S9 versus the particle-associated 1-NP level, a relatively high correlation (r = 0.80–0.91) was observed in all of the S. typhimurium strains. High correlations (r = 0.80–0.93) were also observed when plotting the results of mutagenicity testing after activation by S9 versus the outcome of chemical analysis. These results show that the 1-NP content of workplace air samples is associated with their mutagenic potency, suggesting that 1-NP may be used as a marker for occupational exposure to DE-de-rived particle-associated mutagens © 1995 Wiley-Liss, Inc.     

Mutagenic screening of marker grenade dyes by the Salmonella reversion assay, L5178Y/TK+/- mouse lymphoma assay, and in vivo sister chromatid exchange analysis in mice

Two dyes (C.I. Solvent Yellow No. 33 and a mixture of C.I. Solvent Yellow No. 33 and C.I. Solvent Green No. 3) were tested for mutagenicity in the Salmonella reversion assay and the L5178Y/TK+/- mouse lymphoma assay, and also for sister chromatid exchange (SCE) induction in vivo in C57B1/6J mice. In addition, a greater than 99.9% pure sample of the yellow dye [2-(2'-quinolyl)-1,3-indandione] was tested with and without exogenous activation in the Salmonella reversion assay and the L5178Y/TK+/- mouse lymphoma assay. Neither C.I. Solvent Yellow No. 33 nor the C.I. Solvent Yellow No. 33 and Solvent Green No. 3 mixture was positive for inducing SCEs in vivo. All three dyes were tested in the standard plate incorporation test in seven Salmonella strains TA98, TA100, TA102, TA104, TA1535, TA1537, and TA1538. The dyes were negative with and without exogenous activation in TA98, TA1535, and TA1538. One test with TA1537 was positive with the greater than 99.9% purified yellow dye. All three dyes gave weakly positive results (less than a twofold increase) with S-9 in TA100 and were clearly positive in TA102 and TA104 both with and without S-9. They also induced mutation at the thymidine kinase locus in mouse lymphoma cells, produced both large- and small-colony trifluorothymidine-resistant mutants, and were clastogenic. The purified yellow dye was further tested for SCE induction in mouse lymphoma cells and was determined to give a slightly positive response in the presence of S-9.     

Evaluation of the genotoxicity of river sediments from industrialized and unaffected areas using a battery of short-term bioassays

The present investigation evaluated the capacity of the Salmonella mutagenicity test, the comet assay, and the micronucleus assay to detect and characterize the genotoxic profile of river sediments. Three stations were selected on an urban river (Bouches du Rhône, France) exposed to various sources of industrial and urban pollution (StA, StB, and StC) and one station on its tributary (StD). One station in a nonurban river was included (REF). The concentrations of 16 polycyclic aromatic hydrocarbons (PAHs) were determined by HPLC, and the genotoxicity of the sediments was monitored by the Salmonella mutagenicity test (TA98 + S9, YG1041 ± S9), the comet assay, and the micronucleus assay on CHO cells. Chemical analysis showed that the total PAH concentrations ranged from 23 μg kg^?1 dw (REF) to 1285 μg kg^?1 dw (StD). All the sediments were mutagenic in the Salmonella mutagenicity test. The mutagenicity was probably induced by the presence of nitroarenes (StA, StB, StC, and StD) and aromatic amines (REF) as deduced from the mutagenicity profiles of strains YG1041 ± S9 and TA98 + S9. The comet assay revealed direct DNA lesions in REF, StA, and StB sediments and metabolization‐dependent DNA damage in StC and StD. The micronucleus assay showed an absence of clastogenicity for StA ± S9 and StC‐S9, and a significant clastogenicity ± S9 for the three other stations. The genotoxicity ranking determined by the comet assay + S9 matched the ranking of total and carcinogenic PAH concentrations, and this assay was found to be the most sensitive. Environ. Mol. Mutagen., 2008. © 2008 Wiley‐Liss, Inc.     

Light-dependent mutagenesis by benzo[a]pyrene is mediated via oxidative DNA damage

Benzo[a]pyrene (B[a]P) is an environmental carcinogenic polycyclic aromatic hydrocarbon (PAH). Mammalian enzymes such as cytochrome P-450s and epoxide hydrase convert B[a]P to reactive metabolites that can covalently bind to DNA. However, some carcinogenic compounds that normally require metabolic activation can also be directly photoactivated to mutagens. To examine whether B[a]P is directly mutagenic in the presence of light, we exposed Salmonella typhimurium strains with different DNA repair capacities to B[a]P and white fluorescent light at wavelengths of 370-750 nm. B[a]P plus light significantly enhanced the number of His+ revertants. Mutagenesis was completely light-dependent and required no exogenous metabolic activation. The order of mutability of strains with different DNA repair capacities was strain YG3001 (uvrB, mutMST) > strain TA1535 (uvrB) > strain YG3002 (mutMST) > strain TA1975. The uvrB gene product is involved in the excision repair of bulky DNA adducts, and the mutMST gene encodes 8-oxoguanine (8-oxoG) DNA glycosylase, which removes 8-oxoG from DNA. Introduction of a plasmid carrying the mOgg1 gene that is the mouse counterpart of mutMST substantially reduced the light-mediated mutagenicity of B[a]P in strain YG3001. B[a]P plus light induced predominantly G:C --> T:A and G:C --> C:G transversions. We propose that B[a]P can directly induce bulky DNA adducts if light is present, and that the DNA adducts induce oxidative DNA damage, such as 8-oxoG, when exposed to light. These findings have implications for the photocarcinogenicity of PAHs.     

Mutagenicity of blue rayon extracts of fish bile as a biomarker in a field study

Blue rayon (BR) in combination with the Salmonella/microsome assay was used to evaluate the mutagenicity of fish bile samples. Specimens of Mugil curema from two sites were collected over a 1‐year period. Piaçaguera channel contains high concentrations of total polycyclic aromatic hydrocarbons (PAHs) and other contaminants, while Bertioga channel was considered the reference sites in this study. Bile was extracted with BR and tested with TA98, TA100, and YG1041 strains with and without S9 in dose response experiments. PAH metabolite equivalents were analyzed using reverse‐phase high performance liquid chromatography /fluorescence. Higher mutagenic responses were observed for the contaminated site; YG1041 with S9 was the most sensitive strain/condition. Mutagenicity ranged from 3,900 to 14,000 rev./mg at the contaminated site and from 1,200 to 2,500 rev./mg of BR at the reference site. The responses of YG1041 were much higher in comparison with the TA98 indicating the presence of polycyclic compounds from the aromatic amine class that cause frameshift mutation. TA100 showed a positive mutagenic response that was enhanced following S9 treatment at both sites suggesting the presence of polycyclic compounds that require metabolic activation. benzo(a)pyrene, naphthalene, and phenanthrene metabolite equivalents were also higher in the bile of fish collected at the contaminated site. It was not possible to correlate the PAH metabolite quantities with the mutagenic potency. Thus, a combination of the Salmonella/microsome assay with YG1041 with S9 from BR bile extract seems to be an acceptable biomarker for monitoring the exposure of fish to mutagenic polycyclic compounds. Environ. Mol. Mutagen., 2010. © 2009 Wiley‐Liss, Inc.     

Mutagenicity of products from coal gasification and liquefaction in the salmonella/microsome assay

As a first step in the assessment of their possible bio-effects, coal-related materials were tested for mutagenicity in the Salmonella/microsome assay. Of three coal gasification by-products tested, only a tar was mutagenic for any of four Salmonella strains. The following liquefaction materials were mutagenic for strains TA1538, TA98, and/or TA100: A liquefaction vehicle oil and coal hydrogenation filtered liquid, separated bottoms, vacuum overhead, and vacuum bottoms. Neither powdered coal nor water produced as a by-product of the hydrogenation process was positive in the Salmonella test. No coal-related material was mutagenic for the missense mutant TA1535 or for any strain in the absence of metabolic activation provided by rat hepatic homogenates (S9). In all but one instance Aroclor 1254-induced S9 provided the maximum activation for mutagenesis. Fractionation of all samples was undertaken by serial extraction with organic solvents of increasing polarity (hexane, toluene, methylene chloride, acetonitrile). Highly mutagenic materials were found in fractions of the hydrogenation filtered liquid, vacuum overhead, and vacuum bottoms. Thus far non-mutagenic samples have not yielded mutagenic components upon fractionation.     

The SIMULTEST: a new approach to screening chemicals with the Salmonella reversion assay

A new Salmonella mutagenicity test method is under development to test a chemical with more than one strain simultaneously (the "SIMULTEST"), that is, different Salmonella typhimurium tester strains are used in combination on the same plate. Strains are combined in two sets: strains with plasmid pKM101 (TA97, TA98, TA100, and TA102) and strains without the plasmid (TA1535, TA1537, and TA1538). The SIMULTEST combinations successfully detect the mutagenic activity of five mutagens in different chemical classes. This approach may be useful in reducing the workload associated with mutagenicity testing with Salmonella.     

Mutagenicity of lichen constituents

Usnic acid (the most abundant lichen constituents), physodic, and physodalic acids isolated from Hypogymnia enteromorph (Ach.) Nyl. were tested for mutagenicity in the Ames Salmonella/microsome assay. Physodalic acid exhibited clear dose-related mutagenicity against Salmonella typhimurium strain TA 100 with or without S9 mix in both plate-incorporation and preincubation assays. The addition of S9 mix increased the number of revertants approximately threefold and fourfold in preincubation and plate-incorporation assays, respectively.     

Evaluation of di-(2-ethylhexyl)phthalate and its major metabolites in the ames test and L5178Y mouse lymphoma mutagenicity assay

Di-(2-ethylhexyl)phthalate (DEHP) and its two major metabolites, mono-(2-ethylhexyl)phthalate (MEHP) and 2-ethylhexanol (EH), were tested for genetic activity in both the Salmonella/mammalian microsome mutagenicity (Ames) assay and the L5178Y TK^+/— mouse lymphoma cell mutagenicity assay. All chemicals were tested in both the presence and absence of Aroclor-induced liver microsomes prepared from male Sprague-Dawley rats. Dose levels for both assays were selected from preliminary toxicity studies for each chemical. Neither DEHP, MEHP, nor EH exhibited any significant mutagenic activity in strains TA-98, TA-100, TA-1535, TA-1537, and TA-1538 in the Ames test or when tested in the L5178Y TK^+/ — mouse lymphoma cell mutagenicity assay.     

Role of nitroreduction in the synergistic mutational response induced by mixtures of 1- and 3-nitrobenzo[a]pyrene in Salmonella typhimurium

Previous studies showed that binary mixtures of the environmental pollutants 1- and 3-nitrobenzo[a]pyrene produced a synergistic mutational response in the Salmonella reversion assay. Since nitroreduction is believed to mediate the direct-acting mutagenicity of the individual compounds, we have examined the role of nitroreduction in the mutagenicity of mixtures of 1- and 3-nitrobenzo[a]pyrene in the Salmonella plate incorporation assay. While mixtures of 1- and 3-nitrobenzo[a]pyrene induced up to 183% more revertants in strain TA98 than produced by equivalent amounts of the individual compounds, in the nitroreductase-deficient strain TA98NR the same mixtures only induced up to 57% more revertants than the individual compounds. Analysis of mixtures of 1- and 3-nitrosobenzo[a]pyrene (the two-electron reduction products of 1- and 3-nitrobenzo[a]pyrene) for mutation induction in TA98 yielded no evidence of a synergistic effect between the compounds. The mutagenicity of the mixtures was dependent upon the amount of the more mutagenic component. Salmonella cultures were also incubated with mixtures of 1- and 3-nitrobenzo[a]pyrene, as well as with equivalent amounts of the individual compounds. In two experiments, nitroreductive ability, as measured by the amount of 1-nitropyrene metabolized to 1-aminopyrene in 1 hr, was increased 9 to 105% in cultures pretreated with the mixtures as compared with cultures pretreated with the individual compounds. The results of this study support the hypothesis that nitroreduction is a major factor in the synergistic mutational response induced by 1- and 3-nitrobenzo[a]pyrene in Salmonella typhimurium.     

Combining different assays and chemical analysis to characterize the genotoxicity of waters impacted by textile discharges

Waters receiving textile discharges can exhibit genotoxic and mutagenic activity, which has been related to the presence of dyes and aromatic amines as synthesis precursors or byproducts. The aim of this study was to identify dyes and aromatic amines in water samples impacted by textile discharges, and to evaluate the genotoxic responses of these samples using the Salmonella/microsome assay in strains TA98 and YG1041, and the Fpg‐modified comet assay in the RTL‐W1 fish cell line. The genotoxicity of river samples downstream of the discharge was greater than the upstream samples in both of the Ames tests. The Fpg‐modified comet assay detected similar levels of DNA damage in the upstream and downstream samples. Mutagenicity was not detected with TA98, except for the Quilombo River samples, but when YG1041 was used as the tester strain mutagenicity was detected for all sites with a very different profile in upstream sites relative to the other sites. The mutagenic response strongly indicated that aromatic amines or dyes were contributing to the mutagenic activity downstream. The impact of textile discharges was also confirmed by chemical analysis, because the highest concentrations of azo dyes and aromatic amines were detected in the river downstream. This study shows the value of combining assays measuring complementary endpoints to better characterize the mutagenicity of environmental samples, with the advantage that this approach provides an indication of what classes of compounds are responsible for the effect. Environ. Mol. Mutagen. 57:559–571, 2016. © 2016 Wiley Periodicals, Inc.     

A promising Ames battery for mutagenicity characterization of new dyes

When testing new products, potential new products, or their impurities for genotoxicity in the Ames test, the quantity available for testing can be a limiting factor. This is the case for a dye repository of around 98,000 substances the Max Weaver Dye Library (MWDL). Mutagenicity data on dyes in the literature, although vast, in several cases is not reliable, compromising the performance of the in silico models. In this report, we propose a strategy for the generation of high‐quality mutagenicity data for dyes using a minimum amount of sample. We evaluated 15 dyes from different chemical classes selected from 150 representative dyes of the MWDL. The purity and molecular confirmation of each dye were determined, and the microplate agar protocol (MPA) was used. Dyes were tested at the limit of solubility in single and concentration‐response experiments using seven strains without and with metabolic activation except for anthraquinone dyes which were tested with eight strains. Six dyes were mutagenic. The most sensitive was YG1041, followed by TA97a > TA98 > TA100 = TA1538 > TA102. YG7108 as well as TA1537 did not detect any mutagenic response. We concluded that the MPA was successful in identifying the mutagenicity of dyes using less than 12.5 mg of sample. We propose that dyes should be tested in a tiered approach using YG1041 followed by TA97a, TA98, and TA100 in concentration‐response experiments. This work provides additional information on the dye mutagenicity database available in the literature.