Toxicological impact of strobilurin fungicides on human and environmental health: a literature review.

Fungicides are specifically used for controlling fungal infections. Strobilurins, a class of fungicides originating from the mushroom Strobilurus tenacellus, act on the fungal mitochondrial respiratory chain, interrupting the ATP cycle and causing oxidative stress. Although strobilurins are little soluble in water, they have been detected in water samples (such as rainwater and drinking water), indoor dust, and sediments, and they can bioaccumulate in aquatic organisms. Strobilurins are usually absorbed orally and are mainly eliminated via the bile/fecal route and urine, but information about their metabolites is lacking. Strobilurins have low mammalian toxicity; however, they exert severe toxic effects on aquatic organisms. Mitochondrial dysfunction and oxidative stress are the main mechanisms related to the genotoxic damage elicited by toxic compounds, such as strobilurins. These mechanisms alter genes and cause other dysfunctions, including hormonal, cardiac, neurological, and immunological impairment. Despite limitations, we have been able to compile literature information about strobilurins. Many studies have dealt with their toxic effects, but further investigations are needed to clarify their cellular and underlying mechanisms, which will help to find ways to minimize the harmful effects of these compounds.

Elucidating the effects of pure glyphosate and a commercial formulation on early life stages of zebrafish using a complete biomarker approach: All-or-nothing!

The search for new methods in the toxicology field has increased the use of early life stages of zebrafish (Danio rerio) as a versatile organism model. Here, we use early stages of zebrafish to evaluate glyphosate as pure active ingredient and within a commercial formulation in terms of oxidative stress. Biomarkers involved in the oxidative status were evaluated along with other markers of neurotoxicity, genotoxicity, cytotoxicity, energy balance and motor performance, and the selected tools were evaluated by its sensitivity in determining early-warning events. Zebrafish embryos exposed to glyphosate active ingredient and glyphosate-based formulation were under oxidative stress, but only the commercial formulation delayed the embryogenesis, affected the cholinergic neurotransmission and induced DNA damage. Both altered the motor performance of larvae at very low concentrations, becoming larvae hypoactive. The energy balance was also impaired, as embryos under oxidative stress had lower lipids reserves. Although data suggest that glyphosate-based formulation has higher toxicity than the active ingredient itself, the most sensitive biomarkers detected early-warning effects at very low concentrations of the active ingredient. Biochemical biomarkers of defense system and oxidative damage were the most sensitive tools, detecting pro-oxidant responses at very low concentrations, along with markers of motor performance that showed high sensitivity and high throughput, suitable for detecting early effects linked to neurotoxicity. Alterations on morphology during embryogenesis showed the lowest sensitivity, thus morphological alterations appeared after several alterations at biochemical levels. Tools evaluating DNA damage and cell proliferation showed mid-sensitivity, but low throughput, thus they could be used as complementary markers.

Deleterious effects of benzotriazoles on zebrafish development and neurotransmission: 5-Chloro-benzotriazole versus 1H-benzotriazole.

Benzotriazoles are heterocyclic compounds typically presenting a benzene ring fused with a triazole molecule. The industry uses these compounds as anti-corrosion agents and recently, they have been employed in the pharmaceutical industry and in detergent formulations. Benzotriazoles persist in the environment, and water treatment plants cannot degrade them completely. Consequently, these compounds have been detected in rivers, lakes, and drinking water, which makes assessing their safety for the human and aquatic animal populations crucial. Here, we have evaluated and compared how exposure to 1H-benzotriazole or 5-chloro-benzotriazole affect the zebrafish embryo-larval stages. We have determined the acute toxicity, morphometric alterations, and acetylcholinesterase activity on zebrafish embryos, as well as behavioral endpoints using the tail coiling assay. The estimated LC50 of 5-chloro-benzotriazole was 19 mg/L, whereas 1H-benzotriazole caused no mortality. The zebrafish embryos exposed to 20 and 25 mg/L 5-chloro-benzotriazole had decreased hatching rate and exhibited pericardial and yolk sac edemas. Furthermore, the embryo length and eye area were decreased, in contrast with an increased yolk sac after exposure to 20 mg/L 5-chloro-benzotriazole. In turn, 1H-benzotriazole also decreased the eye area of zebrafish embryos, but no other significant morphological alterations were observed. The tail coiling assay showed that the zebrafish embryos increased the percentage of time moving and the number of embryonic movements per minute after exposure to 1H-benzotriazole (15 mg/L) or 5-chloro-benzotriazole (20 and 25 mg/L), indicating that these compounds were potentially neurotoxic. However, acetylcholinesterase activity was not significantly altered in embryos exposed to 1H-benzotriazole, but significantly decreased when exposed to 0.05 mg/L 5-chloro benzotriazole confirming its neurotoxicity at a much lower concentration. Our findings showed that 5-chloro-benzotriazole seems to induce more harmful alterations to zebrafish embryos than 1H-benzotriazole. Nevertheless, 1H-benzotriazole seems to induce a direct effect on eye development for concentrations lower than the ones of 5-chloro-benzotriazole affecting zebrafish embryos.

Setting limits for N-nitrosamines in drugs: A defined approach based on read-across and structure-activity relationship for N-nitrosopiperazine impurities.

This paper describes DARAN (Defined Approach for Risk Assessment of New Nitrosamines), an new defined approach that uses lines of reasoning based on structure-activity relationship (SAR) patterns and Read-Across (RAx) to set transparent and acceptable limits for new N-nitrosamines for which no toxicological data exist. We selected the compound 1-methyl-4-nitrosopiperazine (MeNP) as a target to calculate a new acceptable limit on the basis of a more transparent and scientifically reasoned RAx. We used publicly available databases and datasets to retrieve experimental in vitro mutagenicity and in vivo carcinogenicity data for N-nitrosopiperazine compounds and to form the chemical category for an RAx. We carried out SAR analyses to try to understand patterns and to obtain interpretable inferences of variation in carcinogenic potency among the N-nitrosopiperazines compounds and their differences with the potent nitrosamines NDMA (N-nitrosodimethylamine) and NDEA (N-nitrosodiethylamine). To estimate an acceptable limit for the target MeNP, we used the scientifically based hypotheses and the evidence lines of about the influence of structural attributes for a robust RAx. On the basis of the criteria proposed in the Assessment Report EMA/369136/20202 and by using the SAR hypotheses obtained by the analysis, we obtained a robust RAx, scientifically supported assumptions, which resulted in TD50 values predicted from the closest structurally related compounds and a worst-case approach.

Phenothiazinium dyes for photodynamic treatment present lower environmental risk compared to a formulation of trifloxystrobin and tebuconazole.

The widespread use of conventional chemical antifungal agents has led to worldwide concern regarding the selection of resistant isolates. In this scenario, antimicrobial photodynamic treatment (APDT) has emerged as a promising alternative to overcome this issue. The technique is based on the use of a photosensitizer (PS) and light in the presence of molecular oxygen. Under these conditions, the PS generates reactive oxygen species which damage the biomolecules of the target organism leading to cell death. The great potential of APDT against plant-pathogenic fungi has already been reported both in vitro and in planta, indicating this control measure has the potential to be widely used in crop plants. However, there is a lack of studies on environmental risk with ecotoxicological assessment of PSs used in APDT. Therefore, this study aimed to evaluate the environmental toxicity of four phenothiazinium PSs: i) methylene blue (MB), ii) new methylene blue N (NMBN), iii) toluidine blue O (TBO), and iv) dimethylmethylene blue (DMMB) and also of the commercial antifungal NATIVO®, a mixture of trifloxystrobin and tebuconazole. The experiments were performed with Daphnia similis neonates and zebrafish embryos. Our results showed that the PSs tested had different levels of toxicity, with MB being the less toxic and DMMB being the most. Nonetheless, the environmental toxicity of these PSs were lower when compared to that of NATIVO®. Furthermore, estimates of bioconcentration and of biotransformation half-life indicated that the PSs are environmentally safer than NATIVO®. Taken together, our results show that the toxicity associated with phenothiazinium PSs would not constitute an impediment to their use in APDT. Therefore, APDT is a promising approach to control plant-pathogenic fungi with reduced risk for selecting resistant isolates and lower environmental impacts when compared to commonly used antifungal agents.

A review of the success and challenges in characterizing human dermal exposure to flame retardants.

Since organic flame retardants (FRs) have several industrial applications, they have been largely detected in environmental and biological samples, and humans have been highly exposed to them. Although the effects of oral and inhaled FRs have been well studied, dermal exposure to them has only recently been pointed out as a potential route of human exposure. Consequently, the effects of FRs on the skin and secondary target organs have been poorly investigated. This review article summarizes the main findings regarding dermal exposure to FRs, points the limitation of the published studies, and suggests future perspectives for better understanding of how dermal exposure to FRs impacts the human health. This review lists some gaps that must be filled in future studies, including characterization of the bioavailable fraction and assessment of exposure for new FRs, to establish their physiological significance and to improve the development of 3D dermal tissue for more reliable results to be obtained.

Early Life Stage Assays in Zebrafish.

Fish embryo toxicity (FET) test using zebrafish (Danio rerio) has been established as an alternative assay to animal experimentation. The FET assay enables the assessment of multiple morphological endpoints during the development of zebrafish early life stages, showing high impact to the field of ecotoxicology on risk assessment of chemicals and pollutants. Moreover, it is also applied to screening drug-induced toxicity and human diseases, due to the high genetic and physiological orthology between zebrafish and humans. Here, we describe FET test, with all steps and several adaptations involved in the methodological procedures. To demonstrate the efficiency of this method, results using the reference substance 3,4-dichloroaniline (DCA) were included to demonstrate sublethal and teratogenic malformations on zebrafish embryos. Thus, there is a strong tendency for using FET tests as a replacement strategy of traditional tests in toxicology and ecotoxicology.

Acetylcholinesterase (AChE) Activity in Embryos of Zebrafish.

Acetylcholinesterase (AChE) is a useful biomarker for organophosphate and carbamate pesticides exposure. The inhibition of this enzyme has been associated with neurotoxicity and alterations at higher levels of biological organization, such as behavior and development impairments. In this chapter, we describe the methodologies for analyses of AChE activity in pools of 96 h of embryos of zebrafish (Danio rerio) using a spectrophotometric method adapted to 96-well microtiter plates.

In vitro characterization of cutaneous immunotoxicity of immortalized human keratinocytes (HaCaT) exposed to reactive and disperse textile dyes.

Several synthetic dyes are used by textile industry for supplying the market of colored clothes. However, these chemicals have been associated with a variety of adverse human health effects, including textile dermatitis. Thus, there is a growing concern to identify textile dyes potentially as skin immunotoxicants. The aim of this in vitro study was to characterize the immunotoxic potential of reactive (Reactive Green 19 [RG19], Reactive Blue 2 [RB2], Reactive Black 5 [RB5]) and disperse (Disperse Red 1 [DR1]) textile dyes using a dermal cell line. For this purpose, a cell-based approach was conducted with immortalized human keratinocytes (KC) (HaCaT) using selected biomarkers of cutaneous inflammation including modulation of matrix metalloproteinases (MMP), oxidative stress such as reactive oxygen species (ROS) generation, and inflammatory cytokine profile. DR1 was the only dye able to trigger an immune response such as release of IL-12 cytokine, a potent co-stimulator of T helper 1 cell, which may be considered as a skin immunotoxicant. The reactive dyes including RB5 that were previously reported as skin sensitizers failed to induce inflammatory reactions under the conditions tested. The reactive dyes studied may pose a risk to human KC by induction of effects related to modulation of MMP-2 (RB5) and -9 (RB5 and RB2) and generation of ROS (RG19 and RB2). Thus, all these dyes need to be used with caution to avoid undesirable effects to consumers who may be exposed dermally.

Identification of biotransformation products of disperse dyes with rat liver microsomes by LC-MS/MS and theoretical studies with DNA: Structure-mutagenicity relationship using Salmonella/microsome assay.

Azo dyes are known as a group of substances with DNA damage potential that depend on the nature and number of azo groups connected to aromatic rings (benzene and naphthalene), chemical properties, e.g. solubility and reactive functional groups, which significantly affect their toxicological and ecological risks. In this paper, we used in vitro models to evaluate the metabolism of selected textile dyes: Disperse Red 73 (DR 73), Disperse Red 78 (DR 78) and Disperse Red 167 (DR 167). To evaluate the mutagenic potential of the textile dyes, the Salmonella mutagenicity assay (Ames test) with strains TA 98 and TA 100 in the presence and absence of the exogenous metabolic system (S9) was used. DR73 was considered the most mutagenic compound, inducing both replacement base pairs (TA 100) and also changing frameshift (TA 98) mutations that are reduced in the presence of the S9 mixture. Furthermore, we used rat liver microsomes in the same experimental conditions of the S9 mixture to metabolize the dyes and the resultant solutions were analyzed using a liquid chromatography coupled to a quadrupole linear ion trap mass spectrometry (LC-MS/MS) to investigate the metabolites formed by the in vitro biotransformation. Based on this experiment, we detected and identified two biotransformation products for each textile dye substrate analyzed. Furthermore, to evaluate the interaction and reactivity of these compounds with DNA, theoretical calculations were also carried out. The results showed that the chemical reaction occurred preferentially at the azo group and the nitro group, indicating that there was a reduction in these groups by the CYP P450 enzymes presented in the rat microsomal medium. Our results clearly demonstrated that the reduction of these dyes by biological systems is a great environmental concern due to increased genotoxicity for the body of living beings, especially for humans.

Allergens of permanent hair dyes induces epidermal damage, skin barrier loss and IL-1 α increase in epidermal in vitro model.

Allergic and irritant skin reactions caused by topical exposure to permanent hair dyes are a common problem. For regulatory and ethnical purposes, it is required to perform chemical safety assessment following the replacement, reduction, and refinement of animal testing (3Rs). Permanent hair dyes are formed by a mixture of ingredients that vary from low to extreme skin sensitizing potency and that inter-react to form unknown by-products. Because of the complex reaction, this cytotoxic mechanism has not yet been elucidated and is the subject of this study. Here, we topically exposed p-phenylenediamine (PPD), Resorcinol (RES), Hydrogen Peroxide (H2O2) alone or as a mixture to RhE and evaluated parameters related to skin irritation such as epidermal viability, keratinocytes damage, barrier loss and IL-1 α. Our data indicates that ingredients tested alone did not lead to an increase of cytotoxic parameters related to skin irritation. However, when the mixture of PPD/H2O2/RES and PPD/H2O2 was applied to the RhE, some of the parameters such as morphological changes including the presence of apoptotic cells, barrier loss and increased IL- 1 α release were observed. The results indicate that the mixture of ingredients used in permanent hair dyes have an irritant effect in RhE while the ingredients alone not.

Investigation of a Brazilian tannery effluent by means of zebra fish (Danio rerio) embryo acute toxicity (FET) test.

Tannery effluents consist of a complex chemical composition not only limited to primary pollutants, which also require biological detection as these compounds may produce adverse effects. The fish embryo toxicity (FET) test with Danio rerio is an alternative method in hazard and risk assessment for determination of chemical-mediated effects. The aim of this investigation was to use the FET test to detect compounds and consequent effects in Brazilian tannery effluents. Samples were collected from the inlet and outlet of the effluent treatment plant at a tannery located in Restinga, São Paulo, Brazil. The toxicological effects were assessed using FET assay for a period of 144 hr using indices such as (1) coagulation of fertilized eggs, (2) lack of detachment of tail-bud from yolk sac, (3) absence of spontaneous movement, (4) yolk sack edema, (5) malformation of the tail, (6) scoliosis, and (7) deformation of swim bladder in the embryos. Data showed that effluent treatment plant exposure produced acute toxicity in D. rerio embryos as evidenced by coagulation of fertilized eggs in up to 5% of all diluted samples 24 hr post fertilization for inlet effluent samples compared to 100% coagulation for outlet samples. Results demonstrated that these effects may not be attributed to metals, but to other non-detected components, such as dyes, pigments, biocides, carriers, surfactants, or other organic compounds that might be present in these complex mixtures. The use of D. rerio embryos was found to be useful as an additional tool for ecotoxicity testing to assess the potential environmental acute toxicity influence of tannery effluents.

The herbicides trifluralin and tebuthiuron have no genotoxic or mutagenic potential as evidenced by genetic tests.

Brazil has been the largest world consumer of pesticides since 2008, followed by the USA. The herbicides trifluralin and tebuthiuron have been widely applied in agriculture. These herbicides are selective for some plant species, and their use brings various benefits. However, the genotoxic and mutagenic effects of tebuthiuron on non-target organisms are poorly known, and in addition, the effects of trifluralin must be better investigated. Therefore, this study employed genetic tests including the comet assay and micronucleus test to evaluate the genotoxic effects of trifluralin and tebuthiuron on HepG2 cells. In addition, we have used the Ames test to assess the mutagenic effects of the herbicides on the TA97a, TA98, TA100, and TA1535 strains of Salmonella typhimurium. On the basis of the comet assay and the micronucleus test, trifluralin did not cause genetic damage to HepG2 cells. In addition, trifluralin did not impact the tested S. typhimurium strains. Regarding tebuthiuron, literature has shown that this herbicide damaged DNA in Oreochromis niloticus. Nevertheless, we have found that tebuthiuron was not genotoxic to either HepG2 cells or the S. typhimurium strains. Therefore, neither trifluralin nor tebuthiuron exerted genotoxic or mutagenic potential at the tested conditions.

Ecotoxicological assessment of glyphosate-based herbicides: Effects on different organisms.

Glyphosate-based herbicides are the most commonly used worldwide because they are effective and relatively nontoxic to nontarget species. Unlimited and uncontrolled use of such pesticides can have serious consequences for human health and ecological balance. The present study evaluated the acute toxicity and genotoxicity of 2 glyphosate-based formulations, Roundup Original (Roundup) and Glyphosate AKB 480 (AKB), on different organisms: cucumber (Cucumis sativus), lettuce (Lactuca sativa), and tomato (Lycopersicon esculentum) seeds, and microcrustacean Artemia salina and zebrafish (Danio rerio) early life stages. For the germination endpoint, only L. esculentum presented significant sensitivity to AKB and L. sativa to Roundup, whereas both formulations significantly inhibited the root growth of all species tested. Both AKB and Roundup induced significant toxicity to A. salina; both are classified as category 3, which indicates a hazard for the aquatic environment, according to criteria of the Globally Harmonized Classification System. However, Roundup was more toxic than AKB, with 48-h median lethal concentration (LC50) values of 14.19 mg/L and 37.53 mg/L, respectively. For the embryo-larval toxicity test, Roundup proved more toxic than AKB for the mortality endpoint (96-h LC50 values of 10.17 mg/L and 27.13 mg/L, respectively), whereas for the hatching parameter, AKB was more toxic than Roundup. No significant genotoxicity to zebrafish larvae was found. We concluded that AKB and Roundup glyphosate-based formulations are phytotoxic and induce toxic effects in nontarget organisms such as A. salina and zebrafish early life stages. Environ Toxicol Chem 2017;36:1755-1763. © 2016 SETAC.

Ecotoxicological risk assessment of the "Acid Black 210" dye.

The "Acid Black 210" dye is one of the most used black dyes by the leather industry. This compound contains three azo groups in its chemical structure, and has been quoted as a non-regulated dye with toxicological concern, since it could generate carcinogenic aromatic amines. The objective of this study was to perform the ecotoxicological risk assessment of this dye through testing its toxicity in vitro and in vivo with the Ames test, the Comet assay, the Daphnia similis test, and the zebrafish embryo acute toxicity test. Moreover, we evaluated the presence of this dye in environmental samples related with a tannery industry. All the tests performed were negative, with the exception of the Ames test with the Salmonella typhimurium TA98 strain, which resulted in a low mutagenic potency. Due to the low concentrations of the "Acid Black 210" dye found in tannery effluents, and the high concentrations where any toxic activity is occasionally described, we concluded that this dye is safe from the ecotoxicological point of view in the areas evaluated and in the light of the current knowledge.

Comparative Study of Genotoxicity Induced by Six Different PBDEs.

Indiscriminate use of synthetic substances has led to environmental contamination and increasing human and animal exposure to harmful chemicals. Polybrominated flame retardants (PBDEs), which serve as non-covalent additives that enhance the safety of a variety of commercial and consumer goods, are an important class among potentially damaging synthetic substances. Its use is very common in developing countries, including Brazil. In theory, 209 different PBDE congeners exist, and many are currently being used during the manufacture of several products. Unfortunately, PBDEs are easily released from the original products, promptly reaching the environment. Knowledge about the toxicological power of these substances is still limited, which has prevented environmental and regulatory authorities from conducting adequate risk assessments. This research addresses the genotoxic and mutagenic potential of PBDEs. The effects of HepG2 cells and Salmonella typhimurium exposure to six main representatives of PBDEs, namely tetrabromodiphenyl ether (BDE-47), pentabromodiphenyl ether (BDE-99 and BDE-100), hexabromodiphenyl ether (BDE-153 and BDE-154) and decabromodiphenyl ether (BDE-209), were evaluated. The comet assay revealed that all the assessed BDEs exerted genotoxic effects but induced no micronuclei formation in HepG2 cells. These BDEs had no significant mutagenic effects on the Salmonella typhimurium strains TA98 and TA100. Taken together, the results of the genomic instability assays showed that PBDEs can represent a risk to the health of directly and indirectly exposed population, because the assessed BDEs induce genotoxic effects in the HepG2 cell line.

Textile dyes induce toxicity on zebrafish early life stages.

Textile manufacturing is one of the most polluting industrial sectors because of the release of potentially toxic compounds, such as synthetic dyes, into the environment. Depending on the class of the dyes, their loss in wastewaters can range from 2% to 50% of the original dye concentration. Consequently, uncontrolled use of such dyes can negatively affect human health and the ecological balance. The present study assessed the toxicity of the textile dyes Direct Black 38 (DB38), Reactive Blue 15 (RB15), Reactive Orange 16 (RO16), and Vat Green 3 (VG3) using zebrafish (Danio rerio) embryos for 144 h postfertilization (hpf). At the tested conditions, none of the dyes caused significant mortality. The highest RO16 dose significantly delayed or inhibited the ability of zebrafish embryos to hatch from the chorion after 96 hpf. From 120 hpf to 144 hpf, all the dyes impaired the gas bladder inflation of zebrafish larvae, DB38 also induced curved tail, and VG3 led to yolk sac edema in zebrafish larvae. Based on these data, DB38, RB15, RO16, and VG3 can induce malformations during embryonic and larval development of zebrafish. Therefore, it is essential to remove these compounds from wastewater or reduce their concentrations to safe levels before discharging textile industry effluents into the aquatic environment.

The oxidation of p-phenylenediamine, an ingredient used for permanent hair dyeing purposes, leads to the formation of hydroxyl radicals: Oxidative stress and DNA damage in human immortalized keratinocytes.

The hair-dyeing ingredient, p-phenylenediamine (PPD), was previously reported to be mutagenic, possibly by inducing oxidative stress. However, the exact mechanism of PPD in inducing oxidative stress upon skin exposure during hair-dyeing in human keratinocytes remains unknown. The aim of our studies was therefore to investigate the toxicity of PPD and its by-products in human immortalized keratinocytes (HaCaT) after auto-oxidation and after reaction with hydrogen peroxide (H2O2). We found that the PPD half maximal effective cytotoxic concentration (EC50) to HaCaT is 39.37 and 35.63 µg/mL after 24 and 48 h, respectively, without addition of H2O2 to induce oxidation. When PPD (10 or 100 µg/mL) is combined with 10.5 µg/mL of H2O2, intracellular ROS production by HaCaT after 1 h was significantly increased and enhanced levels of DNA damage were observed after 4 h of exposure. After 24 h incubations, 20 µg/mL of PPD increased the level of DNA oxidation in HaCaT. Also, we found that the in vitro reaction between PPD and H2O2, even below the maximum allowance by cosmetic industries, released hydroxyl radicals which can damage DNA. Taken together, we conclude that PPD alone and when combined with H2O2 increases the formation of reactive oxygen species in human keratinocytes, leading to oxidative stress and subsequent DNA damage. These alterations suggest that the mechanism by which PPD exposure, alone or combined with H2O2, damages keratinocytes by the formation of the high reactive HO∙ radicals.

The azo dye Disperse Red 13 and its oxidation and reduction products showed mutagenic potential.

Common water pollutants, azo dyes and their degradation products have frequently shown toxicity, including carcinogenic and mutagenic effects, and can induce serious damage in aquatic organisms and humans. In the present study, the mutagenic potential of the azo dye Disperse Red 13 (DR13) was first evaluated using the Micronucleus Assay in human lymphocytes. Subsequently, in order to mimic hepatic biotransformation, controlled potential electrolysis was carried out with a DR13 solution using a Potentiostat/Galvanostat. In addition, a DR13 solution was oxidized using S9 (homogenate of rat liver cells). DR13 oxidation and the reduction products were identified using HPLC-DAD and GC/MS, and their mutagenic potential investigated by way of a Salmonella/microsome assay using TA98 and YG1041 strains, with no S9. The original azo dye DR13 induced chromosomal damage in human lymphocytes, and the respective oxidation and reduction products also showed mutagenic activity, as detected by the Salmonella/microsome assay. Furthermore sulfate 2-[(4-aminophenyl)ethylamino]-ethanol monohydrate, 2-chloro-4-nitro-benzamine, 4-nitro-benzamine and 2-(ethylphenylamine)-ethanol were identified as products of the DR13 reduction/oxidation reactions. Thus it was concluded that the contamination of water effluents with DR13 is a health risk not only due to the dye itself, but also due to the possibility of drinking contaminated water, considering the harmful compounds that can be produced after hepatic biotransformation.