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Sperm quality is defined as the sperm cell ability to successfully fertilize eggs and allow normal embryo development⁠. Few studies explore sperm quality using aquatic invertebrates. Parhyale hawaiensis is a marine amphipod with a circumtropical distribution and considered a model for evolution, development, and ecotoxicological studies. We aimed to develop a methodology to collect sperm cells of P. hawaiensis and evaluate their viability and DNA damage (comet assay). We directly exposed the sperm cells to different mutagenic agents to optimize/develop the protocols. Then, as a proof of concept, we exposed the males to mutagenic compounds (EMS, benzo[a]pyrene (BaP), azo and anthraquinone dyes) at non-lethal concentrations verified by the proposed viability test and analyzed their sperm cells for DNA damage (comet assay). Organisms exposed to EMS presented a clear concentration response in the DNA damage response. We also showed that BaP was able to induce a statistically significant increase in DNA damage of the sperm cells. For the two dyes, although DNA damage increased, statistically differences were not observed. We believe we successfully developed a test to detect genotoxicity of chemicals in sperm cells using an invertebrate model. The protocol for sperm cell viability needs to be further explored with different chemicals to verify its utility as a toxicity endpoint. The developed genotoxicity test has the advantages to employ organisms that are easily cultivated in reduced space, use simple laboratory resources and reduced amount of material and reagents. Positive responses with this model could be used to disclose new germ cell mutagen candidates which could be further confirmed in vertebrates' systems., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Toxicant-induced behavioural changes provide important insights into environmental toxicity, particularly in vulnerable tropical marine habitats. However, ecotoxicological knowledge of organisms in these environments is insufficient. We aimed to develop innovative and cost-effective ecotoxicology methods using Parhyale hawaiensis as a tropical model organism. Adult P. hawaiensis were exposed to aqueous benzo[a]pyrene (BaP) (2 μM) and dietary BaP (50, 250, or 1250 μg BaP/g diet). Survival (24 to 96 h) and behavioural responses (21d) to foraging, reproduction, and predator avoidance were studied. Aqueous and dietary exposures to benzo[a]pyrene (BaP) did not affect survival but induced significant immobility with effective concentration (EC 50  ± SE, 96 h at 11.89 ± 1.19 μM). Relative to the control group, aqueous exposure to 2 μM and dietary exposure to 250 and 1250 μg BaP/g feed resulted in statistically significant behavioural changes. These included a 55-76 % reduction in feeding rates, 133 % increase in chemosensation time, 60-122 % drop in moulting frequency, 200 % delay in precopulatory activity, 50-83 % decrease in geotactic activity, and 300-400 % increase in phototactic activity (all significant at p ≤ 0.05). The methods developed in this study are cost-effective, sensitive, and readily integrated into other endpoint analyses, reinforcing the potential of P. hawaiensis as a tropical ecotoxicology model for detecting toxicant-induced behavioural responses and enhancing marine risk assessments., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Ibrahim Lawan reports financial support was provided by Petroleum Technology Development Fund (PTDF) of Nigeria. Ibrahim Lawan reports a relationship with Petroleum Technology Development Fund (PTDF) of Nigeria that includes: funding grants. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Emodin is an anthraquinone secondary metabolite produced by several species of plants and fungi. Emodin is known for its pharmacological versatility, and, in the textile industry, for its good dyeing properties. However, its use in the textile industry can result in the formation and disposal of large volumes of wastewater. Emodin mutagenicity has been shown in bacteria and in human cells, but little is known about its possible toxic, genotoxic, or mutagenic effects in aquatic organisms. We have evaluated the eco/genotoxicity of emodin to aquatic organisms. Emodin was toxic to Daphnia similis (EC 50  = 130 μg L -1 ) and zebrafish embryos (LC 50  = 25 μg L -1 ). No toxicity was observed for Raphidocelis subcapitata, Ceriodaphnia dubia, or Parhyale hawaiensis. Additional biochemistry/molecular studies are needed to elucidate the toxic/mutagenic pathways of emodin in aquatic organisms. The PNEC value for emodin was 0.025 μg L -1 . In addition to mutagenicity in the Salmonella/microsome assay, emodin was mutagenic in the micronucleus assay in the amphipod P. hawaiensis. Among the anthraquinone dyes tested to date, natural or synthetic, emodin was the most toxic to aquatic species., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Plastics contain a mixture of chemical additives that can leach into the environment and potentially cause harmful effects on reproduction and the endocrine system. Two of these chemicals, N-butyl benzenesulfonamide (NBBS) and triphenyl phosphate (TPHP), are among the top 30 organic chemicals detected in surface and groundwater and are currently placed on international watchlist for evaluation. Although bans have been placed on legacy pollutants such as diethylhexyl phthalate (DEHP) and dibutyl phthalate (DBP), their persistence remains a concern. This study aimed to examine the impact of plastic additives, including NBBS, TPHP, DBP, and DEHP, on the reproductive behaviour and male fertility of the marine amphipod Echinogammarus marinus. Twenty precopulatory pairs of E. marinus were exposed to varying concentrations of the four test chemicals to assess their pairing behaviour. A high-throughput methodology was developed and optimised to record the contact time and re-pair time within 15 min and additional point observations for 96 h. The study found that low levels of NBBS, TPHP, and DEHP prolonged the contact and re-pairing time of amphipods and the proportion of pairs reduced drastically with re-pairing success ranging from 75% to 100% in the control group and 0%-85% in the exposed groups at 96 h. Sperm count declined by 40% and 60% in the 50 μg/l and 500 μg/l DBP groups, respectively, whereas TPHP resulted in significantly lower sperms in 50 μg/l exposed group. Animals exposed to NBBS and DEHP showed high interindividual variability in all exposed groups. Overall, this study provides evidence that plastic additives can disrupt the reproductive mechanisms and sperm counts of amphipods at environmentally relevant concentrations. Our research also demonstrated the usefulness of the precopulatory pairing mechanism as a sensitive endpoint in ecotoxicity assessments to proactively mitigate population-level effects in the aquatic environment., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The concept of sustainability has gained prominence in recent years, enhancing the need to develop products that are less harmful to the environment. Dyes are used by various industrial sectors and have a lot of market value; they are used on a large scale mainly by the textile industry that uses large volumes of water and is one of the main contributors to the contamination of water bodies. Some natural compounds, especially anthraquinones are re-emerging as possible alternatives to synthetic dyes, some of which are known for their toxic and/or mutagenic effects. The BioColour project (https://biocolour.fi/) which is interested in promoting the development of new alternative molecules to synthetic dyes, provided us highly purified anthraquinone dyes dermocybin and dermorubin (>98% purity) extracted from a specie of fungus Cortinarius sanguineus. Dyes were tested for their acute and chronic toxicity using different aquatic organisms. Dermorubin was not toxic to any of the organisms tested for the highest test concentration of 1 mg L -1 and it was the most promising dye. Dermocybin was toxic to Daphnia similis (EC 50  = 0.51 mg L -1 ), Ceriodaphnia dubia (IC 10  = 0.13 mg L -1 ) and Danio rerio embryos (extrapolated LC 50  = 2.44 mg L -1 ). A safety limit, i.e, predicted no-effect concentration (PNEC) of 0.0026 mg L -1 was derived based on the toxicity of dermocybin. The PNEC value can be used to provide hazard information for future application in commercial dyeing processes. Then, we compared the toxicity of dermocybin and dermorubin with ecotoxicity data available in the literature on other anthraquinone dyes of natural and synthetic origin. Some natural dyes can be as toxic as synthetic ones, or more toxic when chronic effects are considered. Despite natural dyes being used since centuries past, there are few ecotoxicological studies available. This study is designed to help develop a more comprehensive understanding of their toxicological properties., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Chronic toxicity tests with representative organisms are essential for ecological risk assessment. The circumtropical marine amphipod Parhyale hawaiensis is a promising test organism in ecotoxicology. This study aimed to develop a chronic toxicity protocol for liquid samples testing with P. hawaiensis using reproduction and growth as endpoints. In the proposed protocol, organisms (≤52 days old) are placed in 5 replicates each containing 100 mL of solution, 10 organisms, and 5 g of crushed coral for 42 days of exposure. The protocol was successfully developed but reproduction showed better performance than growth rate. NOECs based on reproduction were determined for zinc (0.10 mg Zn L -1 ) and 3,4-DCA (0.50 mg L -1 ), and they are of the same order of magnitude compared with the values of other amphipods. The developed test based on reproduction can be considered a promising tool for hazard characterizations although more tests with different substances are still needed., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

N-(1,3-dimethylbutyl)-N'-phenyl-p-phenylenediamine-quinone (6PPD-quinone), an oxidation product of the tire additive, 6PPD, has been associated with high mortality of salmonids (0.1 μg/L). The objective of this study was to determine the acute toxicity using neonates and mutagenicity (micronuclei in hemolymph of exposed adults) of 6PPD-quinone in the marine amphipod Parhyale hawaiensis. Also, we studied its mutagenicity in the Salmonella/microsome assay using five strains of Salmonella with and without metabolic system (rat liver S9, 5%). 6PPD-quinone did not present acute toxicity to P. hawaiensis from 31.25 to 500 μg/L. Micronuclei frequency increased after 96 h-exposure to 6PPD-quinone (250 and 500 μg/L) when compared to the negative control. 6PPD-quinone also showed a weak mutagenic effect for TA100 only in the presence of S9. We conclude that 6PPD-quinone is mutagenic to P. hawaiensis and weakly mutagenic to bacteria. Our work provides information for future risk assessment of the presence of 6PPD-quinone in the aquatic environment., (© 2023 Environmental Mutagenesis and Genomics Society.)

Near-shore marine/estuarine environments play an important role in the functioning of the marine ecosystem and are extremely vulnerable to the presence of chemical pollution. The ability to investigate the effects of pollution is limited by a lack of model organisms for which sufficient ecotoxicological information is available, and this is particularly true for tropical regions. The circumtropical marine amphipod Parhyale hawaiensis has become an important model organism in various disciplines, and here we summarize the scientific literature regarding the emergence of this model within ecotoxicology. P. hawaiensis is easily cultured in the laboratory and standardized ecotoxicity protocols have been developed and refined (e.g., miniaturized), and effects of toxicants on acute toxicity (Cd, Cu, Zn, Ag, ammonia, dyes, pesticides, environmental samples), genotoxicity as comet assay/micronuclei, and gene expression (Ag ion and Ag nanoparticles) and regeneration (pesticides) have been published. Methods for determination of internal concentrations of metals (Cu and Ag) and organic substances (synthetic dye) in hemolymph were successfully developed providing sources for the establishment of toxicokinetics models in aquatic amphipods. Protocols to evaluate reproduction and growth, for testing immune responses and DNA damage in germ cells are under way. The sensitivity of P. hawaiensis, measured as 50% lethal concentration (LC50), is in the same range as other amphipods. The combination of feasibility to culture P. hawaiensis in laboratory, the recent protocols for ecotoxicity evaluation and the rapidly expanding knowledge on its biology make it especially attractive as a model organism and promising tool for risk assessment evaluations in tropical environments., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

As a part of an ongoing interest in identifying environmentally friendly alternatives to synthetic dyes and in using liquid CO 2 as a waterless medium for applying the resulting colorants to textiles, our attention turned to yellow-to-red biocolorants produced by Cortinarius sanguineus fungus. The three principal target anthraquinone colorants (emodin, dermocybin, and dermorubin) were isolated from the fungal bodies using a liquid-liquid separation method and characterized using 700 MHz NMR and high-resolution mass spectral analyses. Following structure confirmations, the three colorants were examined for dyeing synthetic polyester (PET) textile fibers in supercritical CO 2 . We found that all three biocolorants were suitable for dyeing PET fibers using this technology, and our attention then turned to determining their toxicological properties. As emodin has shown mutagenic potential in previous studies, we concentrated our present toxicity studies on dermocybin and dermorubin. Both colorants were non-mutagenic, presented low cellular toxicity, and did not induce skin sensitization. Taken together, our results indicate that dermocybin and dermorubin possess the technical and toxicological properties needed for consideration as synthetic dye alternatives under conditions that are free of wastewater production.

Silver nanoparticles (AgNP) are widely used in several applications including as antifouling agents; therefore, they can end up in estuarine and marine environments. These nanoparticles tend to aggregate and to deposit in the sediment, where many organisms feed and reproduce. Parhyale hawaiensis is an epibenthic amphipod globally distributed in tropical zones, and has been considered a potential model for ecotoxicology. The aim of this study was to evaluate genotoxic effects of AgNP and Ag salt via feeding, as P. hawaiensis lives in the sediment where nanoparticles tend to accumulate. Organisms were cultivated in the laboratory, and adults were exposed to food containing both AgNP and Ag salt. We collected hemolymph after different times of exposure, and analysed the hemocytes for nuclear abnormalities (including micronuclei) and DNA damage using the standard alkaline comet assay. Conditions of both assays were developed/optimized to allow their successful application in marine invertebrates. Increased frequencies of micronuclei, nuclear buds and total abnormalities were detected in relation to concentration and time in organisms exposed to AgNP and Ag salt in comparison to the controls. No DNA damage was detected when the alkaline comet assay was applied. After 5 days of exposure, we observed higher micronuclei frequencies in Ag salt treatment in comparison with AgNP. After 13 days, micronuclei frequencies were similar for both silver forms. We believe that the Ag, in its ion form, is causing the mutagenic effect; therefore, more time would be needed for the release of the ion from AgNP, explaining the delayed mutagenic effect., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Graphene oxide (GO) production has increased considerably and therefore its presence in the environment is inevitable. When in aquatic environment GO can interact with co-existing compounds, modifying their toxicities for several organisms. However, the toxic effects of co-exposure of GO and organic compounds are rarely reported in the literature. Herein, we studied the behavior of four organic aquatic contaminants found in surface water such as 2-phenylbenzotriazoles (non-Cl PBTA-9 and PBTA-9) and phenoxyphenyl pesticides, pyriproxyfen (PYR) and lambdacyhalothrin (LCT), in the presence of GO. GO reduced 90% and 83% of the toxicity of non-Cl PBTA-9 and PBTA for Daphnia. When PBTAs were adsorbed onto GO surface their interactions caused GO agglomeration (up to 20 mm) and consequent precipitation, making PBTAs less bioavailable. PYR and LCT's toxicities increased up to 83% for PYR and 47% for LCT in the presence of GO, because their adsorption on GO lead to the stabilization of the suspensions (up to 0.5 μm). Those particles were then easily ingested and retained in the digestive tract of the daphnids, triggering the Trojan horse effect. Based on theoretical calculations we observed that PBTA compounds are planar, electron-poorer and more reactive than the studied pesticides, suggesting a better stability of the GO/PBTA complexes. PYR and LCT are nonplanar, electron-richer and less reactive towards GO than PBTAs, forming less stable GO complexes that could facilitate the desorption of pesticides, increasing toxic effects. Our results suggest that the properties of the organic toxicants can influence the stability of graphene oxide suspensions, playing a fundamental role in the modulation of their toxicity. Further research is needed for a deep understanding of the behavior of nanomaterials in the presence of contaminants and their effect in the toxicity of aquatic organisms., Competing Interests: Declaration of competing interest There are no conflicts to declare., (Copyright © 2022 Elsevier B.V. All rights reserved.)

There has been increased interest in the use of natural dyes for textile coloration as alternatives to synthetic dyes, due to the general belief that natural dyes are more environmentally friendly. However, natural dyes have poor affinity for textiles, which can lead to high dye levels in the resultant wastewater. While chlorine treatment has proven to be effective for dye wastewater disinfection and decolorization, this process can also lead to the formation of more toxic degradation products for certain synthetic dyes. On the other hand, little information is available regarding the ecotoxicity of natural dyes and their chlorination products. To advance knowledge in this area, madder was selected due to its historical importance and wide application in the textile industry. Specifically, we sought to characterize the chlorine-induced degradation products of an aqueous madder solution and to assess their ecotoxicity. The main component of the present madder sample was Alizarin (89.8%). Chlorination led to complete decolorization, and 2-hydroxynaphthalene-1,4-dione and phthalic anhydride were identified as key degradation products. Chlorination of madder decreased toxicity to Daphnia similis (microcrustacean) 10-fold and removed the toxicity to Raphidocellis subcapitata (algae), when compared to the parent dye., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The increasing pollution of aquatic environments due to old and emerging contaminants requires the development of integrative methods for exposure assessment. Internal concentrations are a reliable way to estimate total exposure of contaminants originated from different routes (water, sediment, and food). We developed a protocol to evaluate the concentration of a dye, C.I. Disperse Red 1, in the hemolymph of Parhyale hawaiensis, a marine amphipod. LOD and LOQ were satisfactory to detect the dye in all hemolymph samples. The concentration detected in the hemolymph varied related to exposure time and dye concentration (0.003 to 0.086 μg mL -1 ). Polynomial regression model was the best fit. The protocol was reliable to detect and quantify dye exposure in marine amphipods and can be considered for future assessments of estuarine and marine regions under the influence of dye processing plants. The method possibly can be easily adapted to other amphipods and other azo dyes., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

The chemical pollution crisis severely threatens human and environmental health globally. To tackle this challenge the establishment of an overarching international science-policy body has recently been suggested. We strongly support this initiative based on the awareness that humanity has already likely left the safe operating space within planetary boundaries for novel entities including chemical pollution. Immediate action is essential and needs to be informed by sound scientific knowledge and data compiled and critically evaluated by an overarching science-policy interface body. Major challenges for such a body are (i) to foster global knowledge production on exposure, impacts and governance going beyond data-rich regions (e.g., Europe and North America), (ii) to cover the entirety of hazardous chemicals, mixtures and wastes, (iii) to follow a one-health perspective considering the risks posed by chemicals and waste on ecosystem and human health, and (iv) to strive for solution-oriented assessments based on systems thinking. Based on multiple evidence on urgent action on a global scale, we call scientists and practitioners to mobilize their scientific networks and to intensify science-policy interaction with national governments to support the negotiations on the establishment of an intergovernmental body based on scientific knowledge explaining the anticipated benefit for human and environmental health., Competing Interests: Competing interestsThe authors declare that they have no competing interests. HH is Editor-in-Chief of this Journal., (© The Author(s) 2022.)

Previous studies have demonstrated the presence of precursors and coupling agents in wastewater from hair dyeing processes. The complex reaction involved in the oxidation of these compounds can generate extremely hazardous sub-products, leading to an increase in the mutagenicity and toxicity of wastewater. Without proper treatment, this highly toxic wastewater may find its way into the drinking water treatment plant. The present work aimed to investigate the main products generated after the oxidation reaction involving p-toluenediamine (PTD) and p-aminophenol (PAP) - precursors that widely used in the composition of commercial permanent hair dyes, under experimental conditions close to the routine hair dyeing process (in the presence and absence of hydrogen peroxide in ammoniacal medium), using spectroscopic techniques. The study also investigated the mutagenicity and toxicity of the products formed in the hairdressing wash water and conducted detection analysis to determine the presence of the precursors and Bandrowski's Base Derivative (BBD) in samples of wastewater, surface and drinking water using HPLC-DAD and linear voltammetry techniques. Based on this investigation, we identified several PTD and PAP self-oxidation products and eleven sub-products derived from the reaction between PTD and PAP. Assays conducted using Salmonella typhimurium YG1041, with and without activation-induced rat liver metabolism (S9), indicated mutagenicity of the reaction products in concentrations above 10.0 μg μL -1 . The concentrations of PTD, PAP, and several reactions and oxidation products of these precursors were detected in wastewater and water samples., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Organophosphate esters (OPEs) are substances globally used as flame retardants and plasticizers that have been detected in all environmental compartments. This study aimed to evaluate the occurrence and sources of ten OPEs in the Piracicaba River Basin (Brazil). Twelve sampling sites were selected in five rivers with different pollution sources; six sampling campaigns were performed encompassing dry and wet seasons. ΣOPEs ranged from 0.12 to 6.2 μg L -1 ; the levels in urban areas were higher than in rural and non-urban areas, but no overall tendency concerning the seasonal effect on OPEs concentrations was observed. Tris(2-butoxyethyl) phosphate (TBOEP), tris(2-chloroisopropyl) phosphate (TCIPP), and tris(1,3-dichloroisopropyl) phosphate (TDCIPP) were the most abundant and frequently detected compounds. Nine OPEs were detected at higher concentrations in a site affected by effluents from textile industries. An acute toxicity test using Daphnia similis was performed for tris(2-ethylhexyl) phosphate (TEHP) for the calculation of a preliminary predicted no effect concentration (PNEC). The risk quotient (RQ) approach was applied and risk to aquatic environment related to TEHP levels was observed in areas adjacent to textile industries, but more toxicity studies are required for the determination of a more reliable PNEC., (© 2020. Springer-Verlag GmbH Germany, part of Springer Nature.)

Regeneration is a widely spread process across the animal kingdom, including many species of marine crustaceans. It is strongly linked to hormonal cycles and, therefore, a great endpoint candidate for toxicology studies. We selected the amphipod Parhyale hawaiensis as test organism, already used in ecotoxicological studies and able to regenerate its body appendages. We are proposing a protocol to use the antenna regeneration as a toxicity endpoint. First, we evaluated differences in time of completion of regeneration in males and females after the amputation of one antenna of 6 months old animals. Then we compared the influence of different testing volumes in the regeneration process (100 and 5 mL). We used as testing substances, dimethyl sulfoxide (DMSO) and diflubenzuron, a chitin synthesis inhibitor. The most suitable protocol consisted of volumes of 5 mL in 12-well microplates, with 1 organism per well, 12 organisms per concentration (1:1 females/males) and test time duration of around 5 weeks. DMSO accelerated regeneration time with a NOEC of 0.06%. Diflubenzuron inhibited the time necessary to its completion with a NOEC of 0.32 μg L -1 . We conclude that the Parhyale hawaiensis antenna regeneration protocol proposed here is a potential tool in ecotoxicology, but more studies are required for its validation not only to verify its utility for testing chemicals but also environmental samples.

Phenylbenzotriazoles (PBTA) can be generated unintentionally during textile dyeing factories by reduction of dinitrophenylazo dyes and their subsequent chlorination in disinfection process. Eight non-chlorinated PBTAs (non-Cl PBTA) and their related chlorinated PBTAs have been found in rivers and presented mutagenic activity. No data on their aquatic toxicity are available. In this work, two new phenylbenzotriazoles, non-Cl PBTA-9 and PBTA-9, derived from the dye C.I. Disperse Violet 93 (DV93) were synthesized and chemically/toxicologically characterized. Both compounds were more mutagenic than the parental dye in the Salmonella/microsome assay in the presence of metabolic activation (S9). Mutagenicity studies in vivo with mammals would confirm their potential hazard to humans. The two compounds were acutely toxic to Daphnia similis. We developed an analytical method to simultaneously quantify non-Cl PBTA-9, PBTA-9 and DV93 in river waters. Non-Cl PBTA-9 was found in sites under influence of textile effluents but at concentrations that do not pose risk to the aquatic life according to the P-PNEC calculated based on the acute toxicity tests. PBTA-9 was not detected in any samples analyzed. More studies on the aquatic toxicity and water occurrence of PBTAs should be conducted to verify the relevance of this class of compounds as aquatic contaminants., (Copyright © 2020 Elsevier B.V. All rights reserved.)

In this study, a multi-residue method was used to analyze 13 pesticides and 1 degradation product in surface and groundwater in the region with the largest sugar cane production in the world. The potential effects of individual pesticides and their mixtures, for aquatic life and human consumption, were evaluated. For the surface water, 2-hydroxy atrazine, diuron, carbendazim, tebuthiuron, and hexazinone were the most frequently detected (100, 94, 93, 92, and 91%, respectively). Imidacloprid (2579 ng L -1 ), carbendazim (1114 ng L -1 ), ametryn (1101 ng L -1 ), and tebuthiuron (1080 ng L -1 ) were found at the highest concentrations. For groundwater, tebuthiuron was the only quantified pesticide (107 ng L -1 ). Ametryn, atrazine, diuron, hexazinone, carbofuran, imidacloprid, malathion, carbendazim, and their mixtures presented risk for the aquatic life. No risk was observed for the pesticides analyzed in this work, alone or in their mixtures for human consumption.

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., (© 2020 Wiley Periodicals LLC.)

The extractable organic material (EOM) from atmospheric total suspended particles (TSP) contains several organic compounds including non-substituted polycyclic aromatic hydrocarbons (PAHs), alkyl-PAHs, and nitro-PAHs. These chemicals seem to be among the key drivers of TSP genotoxicity. We have shown previously that the mutagenic potencies of the EOM from Limeira, Stockholm, and Kyoto, cities with markedly different meteorological conditions and pollution sources are similar. Here we compare the profiles of non-substituted PAHs (27 congeners), alkyl-PAHs (15 congeners), and nitro-PAHs (7 congeners) from the same EOM samples from these cities. We also compared the genotoxicity profiles using comet and micronucleus assays in human bronchial epithelial cells. The profiles of PAHs, as well as the cytotoxic and genotoxic potencies when expressed in EOM, were quite similar among the studied cities. It seems that despite the differences in meteorological conditions and pollution sources of the cities, removal, mixing, and different atmospheric transformation processes may be contributing to the similarity of the PAHs composition and genotoxicity profiles. More studies are required to verify if this would be a general rule applicable to other cities. Although these profiles were similar for all three cities, the EOM concentration in the atmospheres is markedly different. Thus, the population of Limeira (∼10-fold more EOM/m 3 than Stockholm and ∼6-fold more than Kyoto) is exposed to higher concentrations of genotoxic pollutants, and Kyoto's population is 1.5-fold more exposed than Stockholm's. Therefore, to reduce the risk of human exposure to TSP genotoxins, the volume of emissions needs to be reduced., (© 2020 Wiley Periodicals, Inc.)

This work reports the study of oxidation reaction of p-aminophenol (PAP) in ammoniacal medium in dissolved atmospheric oxygen and hydrogen peroxide, simulating the process of hair dyeing with permanent dyes. The products formed, which included semi-quinoneimine radical, quinoneimine, dimers, trimers and tetramers, were identified by mass spectrometry, infrared spectroscopy, UV-vis spectrophotometry, and nuclear magnetic resonance of hydrogen. The process was found to involve an autoxidation mechanism. The mutagenicity of the products was carried out by Salmonella Typhimurium YG1041 assay, and the results indicated no mutagenic properties. The presence of PAP and its oxidative products in samples of wastewater collected from hairdressing salon effluent (WW), raw river water (RRW), and water inlet and outlet of drinking water treatment plant (DWTP) was analyzed by HPLC-DAD. PAP was detected in the collected samples of WW, water samples from DWTP (before and after treatment), at concentrations of 2.1 ± 0.5 mg L -1 , 1.9 ± 0.3 × 10 -3 mg L -1 and 1.3 ± 0.2 × 10 -3 mg L -1 , respectively. The reaction products, including dimers, trimers and tetramers were identified only in the WW sample; this shows that both the precursor in the sample and its derivatives were released into the wastewater., Competing Interests: Declaration of Competing Interest The authors declare no conflicts of interest., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Effect-based analyses are being recognized as excellent tools to a comprehensive and reliable water quality evaluation to complement physical and chemical parameters. The Salmonella/microsome mutagenicity test was introduced in the São Paulo State water quality-monitoring program in 1999 and waters from 104 sites used to the production of drinking water were analyzed. Samples were tested after organic extraction, using the microsuspension version of the Salmonella/microsome assay with strains TA98 and TA100 with and without S9-mammalian metabolic system. Of the 1720 water samples analyzed in 20 years, 20% were positive; TA98 was the most sensitive strain, detecting alone 99%. Results were presented in hazard categories to facilitate water managers' understanding and general public communication. Hot spots of mutagenicity were identified, and pollution sources investigated. A flow scheme with instructions of how to proceed in case of mutagenic samples was developed and implemented in the monitoring program. Enforcement actions were taken to reduce exposure of humans and aquatic biota to mutagenic compounds. The results presented provide scientific basis for the incorporation of the Salmonella/microsome assay in a regulatory framework, and to guide water-quality managers. The inclusion of a mutagenicity assay using standardized conditions proved to be an opportunity to improve the quality of water, and the strategy presented here could be applied by any environmental agency around the world. Environ. Mol. Mutagen. 61:200-211, 2020. © 2019 Wiley Periodicals, Inc., (© 2019 Wiley Periodicals, Inc.)

The present work investigated the autoxidation reaction of p-toluenediamine (PTD) - a precursor - widely used in permanent hair dyeing formulation, under experimental conditions close to the hair dyeing process (oxygen and/or peroxide in ammoniacal medium), by chromatographic and spectroscopic techniques. In additional, evaluated the mutagenicity of the PTD oxidation products and the presence of PTD and this products in wastewater from beauty salon, as well as in surface water and drinking water using HPLC coupled to a diode array detector and linear scan voltammetry. Through this study, it was possible the identification of semi-quinonediimine, quinonediimine, dimers (derived from toluenediamine), and trimer radical identified as Bandrowski's Base derivative (BBD) formed during autoxidation of PTD. Salmonella Typhimurium YG1041 assay with and without metabolic activation induced rat-liver (S9) indicated mutagenic activity for BBD. Levels of PTD were determined by the standard addition method in samples collected from the wastewater of a beauty salon, as well as from the water before and after treatment in a drinking water treatment plant (DWTP) reached concentrations of 2.08 ± 0.21, 2.36 ± 0.10 × 10 -3 , and 1.77 ± 0.13 × 10 -3  mg L -1 , respectively. In addition, linear sweep voltammetry was used to monitor the BBD found at the concentration of 1.59 ± 0.35 mg L -1 in wastewater collected from the beauty salon., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Behavioural responses to contaminants are an important endpoint in ecotoxicology because they link effects at biochemical or cellular levels to impacts on individual fitness. Due to the increasing use of silver in nanomaterials, studies of its effects on the behaviour of aquatic organisms are important to assess the risks of silver nanoparticles (AgNP) released into the environment. The aim of this work was to evaluate the behavioural effects of silver on the marine amphipod Echinogammarus marinus after exposure to AgNO 3 via water and AgCl or AgNP via food. Swimming activity of the amphipods was tracked during 6 min alternating dark and light conditions. Animals swam slower and responded less to light at higher concentrations of silver in the water. No differences were found in the behaviour of animals exposed via feeding up to 28 days, hence, longer exposure times may be required for the observation of effects. This is the first work to appraise behaviour effects of silver ions and AgNP on marine amphipods. Although the protocol has been successfully developed for this purpose, specimens appeared to habituate to test conditions during the experiments. Therefore, the need for further understanding of baseline behaviours in these model organisms is discussed., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Atmospheric particulate matter (PM) organic fractions from urban centers are frequently mutagenic for the Salmonella/microsome assay. This mutagenicity is related to both primary and secondary pollutants, and meteorological conditions have great influence on the secondary pollutant's formation. Our objective was to compare the mutagenicity of atmospheric total suspended particulates (TSP) from three cities with marked different meteorological conditions and TSP concentrations: Limeira (Brazil) with 99.0 μg/m 3 , Stockholm (Sweden) with 6.2 μg/m 3 , and Kyoto (Japan) with 28.0 μg/m 3 . For comparison, we used the same batch of filters, sample extraction method, and Salmonella/microsome testing protocol with 11 strains of Salmonella with and without metabolic activation. Samples were collected during winter and pooled into one single extract representing each city. All samples were mutagenic for all tested strains, except for TA102. Based on the strain's selectivity, nitroarenes, polycyclic aromatic hydrocarbons, and aromatic amines play a predominant role in the mutagenicity of these samples. The mutagenic potencies expressed by mass of extracted organic material (EOM; revertants/μg EOM) were similar (~twofold difference) among the cities, despite differences in meteorological conditions and pollution sources. In contrast, the mutagenic potencies expressed by air volume (rev/m 3 ) varied ~20-fold, with Limeira > Kyoto ≈ Stockholm. These results are the first systematic assessment of air mutagenicity from cities on three continents using the same protocols. The results confirm that the mutagenic potency expressed by EOM mass is similar regardless of continent of origin, whereas the mutagenic potency expressed by air volume can vary by orders of magnitude. Environ. Mol. Mutagen. 2019. © 2019 Wiley Periodicals, Inc., (© 2019 Wiley Periodicals, Inc.)

Good water quality for livestock is essential for animal health, supply of safe food and food production economy. Few countries have established water quality criteria for livestock for chemical contaminants. For those that have them, the values are quite variable among each other for the same substance due to differences in the approach for the acceptable daily intakes and algorithms for the calculation. In several countries, including Brazil, these standards are based on international or other countries' data, which differ among protected species, exposure scenarios, and levels of protection. The objective of this work was to discuss critical issues to establish chemical water quality criteria for livestock in Brazil. A discussion about the difficulties involved and the alternatives when sufficient data are not available is presented. Using the Canadian framework for reference, we provide recommendations on a revised framework and alternatives regarding the use of chronic studies as mandatory, defaulting to human health drinking water quality guidelines/criteria for livestock, the use of additional safety factors, and alternatives in the absence of toxicological data., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Ayahuasca is a beverage used in religious rituals of indigenous and nonindigenous groups, and its therapeutic potential has been investigated. Ayahuasca is obtained by decoction of the Banisteriopsis caapi that contains β-carbolines (harmine, harmaline, and tetrahydroharmine) plus Psychotria viridis that contains N,N-dimethyltryptamine. Although plants used in folk medicine are recognized as safe, many of them have genotoxic potential. The Salmonella/microsome assay is usually the first line of the mutagenicity evaluation of products intended for therapeutic use. Our objective was to evaluate the mutagenicity of ayahuasca beverage and their constituents using the Salmonella/microsome assay with TA98 and TA100. We analyzed two ayahuasca samples, and also beverage samples prepared each individual plant P. viridis and B. caapi. Harmine and harmaline were also tested. All beverage samples were chemically characterized and both ayahuasca samples could be considered representative of the beverages consumed in religious rituals. Both ayahuasca samples were mutagenic for TA98 and TA100 with and without S9, with similar potencies. The beverage obtained from P. viridis was not mutagenic, and beverage obtained from B. caapi was mutagenic for TA98 with and without S9. Harmine was nonmutagenic and harmaline was mutagenic only for TA98 without S9. Harmaline fully explain the mutagenicity observed with TA98 without S9 of both ayahuasca samples and the B. caapi beverage samples. We conclude that the ayahuasca samples are mutagenic and this effect is partially explained by harmaline, one of the β-carbolines present in the beverage. Other mutagenic compounds seem to be present and need to be further investigated. Environ. Mol. Mutagen. 60:269-276, 2019. © 2018 Wiley Periodicals, Inc., (© 2018 Wiley Periodicals, Inc.)

Color Index (C.I.) Disperse Blue 291 (DB291) is an azo dye used by the textile industry. After yarn dyeing, wastewater containing the dye, released into the aquatic environment, may pollute drinking water sources. We investigated the mutagenicity and genotoxicity of DB291 in male Swiss mice, following oral administration. Micronucleated cells, primary DNA damage (comet assay) in blood, liver, and kidney cells, and BAX, BCL2, SMAD4 and TNFA gene expression in leukocytes were evaluated. An increased frequency of micronucleated polychromatic erythrocytes (MNPCEs) was observed in animals treated with 50 mg/kg bw; no other genetic alteration was detected. Neither primary DNA damage nor changes in gene expression were observed., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Color Index (C.I.) Disperse Red 1 (DR1) is a widely used textile azo dye found in rivers. As it may not be completely removed by conventional treatments, humans can be exposed through drinking water. Studies have supported in vitro toxicity and mutagenicity of commercial DR1. This study aimed to investigate the mutagenic and toxicogenomic effects of commercial DR1 in multiple tissues/organs of Swiss male mice. For that, animals were orally exposed to the dye (by gavage), at single doses of 0.0005, 0.005, 0.5, 50, or 500 mg/kg bw. The two lowest doses were equivalent to the ones found in two Brazilian rivers receiving influx of textile discharges. Cytotoxicity, micronucleated cell frequencies (for all doses tested), primary DNA damage (comet assay), and gene expression profiling of (0.0005 and 0.005 mg/kg of bw) were investigated 24 hr after animal exposure to commercial DR1. Data showed increased frequencies of micronucleated polychromatic erythrocytes in bone marrow cells after treatment with 0.5 and 50 mg/kg bw. At 0.005 mg/kg bw, commercial DR1 induced an increase of primary DNA damage in liver, but not in kidney cells. Additionally, upregulation of genes involved in the inflammatory process (IL1B) (0.0005 and 0.005 mg/kg bw) and cell-cycle control (CDKN1A) in liver cells, and apoptosis (BCL2 and BAX) in leukocytes (0.005 mg/kg bw) were also detected. In conclusion, the commercial DR1 was genotoxic (chromosome aberrations and primary DNA damage) and modulated gene expression in mice, and such effects were dependent on the doses and tissues analyzed. Environ. Mol. Mutagen. 59:822-828, 2018. © 2018 Wiley Periodicals, Inc., (© 2018 Wiley Periodicals, Inc.)

The Salmonella/microsome assay (Ames test) is the most widely used mutagenicity test for the evaluation of pure chemicals and environmental samples. There are several versions of protocols available in the literature, including those that reduce the amount of sample needed for testing with liquid and agar media. The microsuspension version of the Salmonella/microsome assay is more sensitive than the standard protocol. It is performed using 5-times concentrated bacteria and less sample and S9 mixture, but still uses conventional Petri dishes (90 × 15 mm). It has been extensively used for environmental sample testing, including in effect-directed analysis (EDA). The objective of this study was to miniaturize the microsuspension assay using 12-well microplates instead of the conventional plates. For validation of this miniaturization, thirteen mutagenic compounds were tested using three Salmonella strains that were selected based on their different spontaneous reversion frequencies (low, medium, and high). The conditions of the miniaturized procedure were made as similar as possible to the microsuspension protocol, using the same testing design, metabolic activation, and data interpretation, and the tests were conducted in parallel. The miniaturized plate assay (MPA) and microsuspension procedures provided similar sensitivities although MPA is less laborious and require less sample and reagents, thereby reducing overall costs. We conclude that the MPA is a promising tool and can be particularly suitable for environmental studies such as EDA or monitoring programs. Environ. Mol. Mutagen. 59:488-501, 2018. © 2018 Wiley Periodicals, Inc., (Copyright © 2018 Wiley Periodicals, Inc.)

There is a lack of suitable tropical marine species for ecotoxicity tests. An attractive model organism for ecotoxicology is the marine amphipod Parhyale hawaiensis, which is already a model for genetic and developmental studies. This species is widespread, can tolerate changes in salinity, is easy to handle and is representative of circumtropical regions. The aim of this work was to describe standardized procedures for laboratory husbandry, define conditions for acute toxicity tests, and to provide acute toxicity test results for some reference toxicants. Culturing conditions for the organism in the laboratory were established in reconstituted seawater (30 ± 2 salinity), 24 ± 2 °C, photoperiod 12/12 h light/dark. Acute toxicity test procedures were developed for 96 h-exposure time, and organisms at ages <7 days. The miniaturized version of the test, based on 96-well microplates and 200 µL of exposure media provided consistent results compared to larger exposure volumes (80-mL vials protocol). Acute toxicity of Ag, Cd, Cu, Zn and ammonia determined for P. hawaiensis were consistent to previous results for other marine amphipods. We conclude that P. hawaiensis can be successfully cultured in standardized conditions and be effectively used in acute toxicity testing. Further development and use of this model will enable standardized and reproducible ecotoxicology investigations in understudied and vulnerable tropical marine ecosystems.

The combination of chemical analyses and bioassays allows the identification of potentially mutagenic compounds in different types of samples. Dyes can be considered as emergent contaminants and were detected in waters, under the influence of textile activities. The objective of this study was to evaluate the contribution of 9 azo dyes to the mutagenicity of representative environmental samples. Samples were collected along one year in the largest conglomerate of textile industries of Brazil. We analyzed water samples from an important water body, Piracicaba River, upstream and downstream two main discharges, the effluent of a wastewater treatment plant (WWTP) and the tributary Quilombo River, which receives untreated effluent from local industries. Samples were analyzed using a LC-MS/MS and tested for mutagenicity in the Salmonella/microsome microsuspension assay with TA98 and YG1041. Six dyes were detected in the collected samples, Disperse Blue 291, Disperse Blue 373, Disperse Orange 30, Disperse Red 1, Disperse Violet 93, and Disperse Yellow 3. The most sensitive condition for the detection of the mutagenicity was the strain YG1041 with S9. The concentration of dyes and mutagenicity levels varied along time and the dry season represented the worst condition. Disperse Blue 373 and Disperse Violet 93 were the major contributors to the mutagenicity. We conclude that dyes are contributing for the mutagenicity of Piracicaba River water; and both discharges, WWTP effluent and Quilombo River, increase the mutagenicity of Piracicaba River waters in about 10-fold. The combination of chemical analysis and bioassays were key in the identification the main drivers of the water mutagenicity and allows the selection of priority compounds to be included in monitoring programs as well for the enforcing actions required to protect the water quality for multiple uses., (Copyright © 2017 Elsevier B.V. All rights reserved.)

There is ongoing concern about the consequences of mutations in humans and biota arising from environmental exposures to industrial and other chemicals. Genetic toxicity tests have been used to analyze chemicals, foods, drugs, and environmental matrices such as air, water, soil, and wastewaters. This is because the mutagenicity of a substance is highly correlated with its carcinogenicity. However, no less important are the germ cell mutations, because the adverse outcome is related not only to an individual but also to population levels. For environmental analysis the most common choices are in vitro assays, and among them the most widely used is the Ames test (Salmonella/microsome assay). There are several protocols and methodological approaches to be applied when environmental samples are tested and these are discussed in this chapter, along with the meaning and relevance of the obtained responses. Two case studies illustrate the utility of in vitro mutagenicity tests such as the Ames test. It is clear that, although it is not possible to use the outcome of the test directly in risk assessment, the application of the assays provides a great opportunity to monitor the exposure of humans and biota to mutagenic substances for the purpose of reducing or quantifying that exposure.

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., (© 2016 Wiley Periodicals, Inc.)

Water quality criteria to protect aquatic life are not available for most disperse dyes which are often used as commercial mixtures in textile coloration. In this study, the acute and chronic toxicity of the commercial dye Disperse Red 1 (DR1) to eight aquatic organisms from four trophic levels was evaluated. A safety threshold, i.e. Predicted No-Effect Concentration (PNEC), was derived based on the toxicity information of the commercial product and the purified dye. This approach was possible because the toxicity of DR1 was accounting for most of the toxicity of the commercial mixture. A long-term PNEC of 60 ng L(-1) was proposed, based on the most sensitive chronic endpoint for Daphnia similis. A short-term PNEC of 1800 ng L(-1) was proposed based on the most sensitive acute endpoint also for Daphnia similis. Both key studies have been evaluated with the new "Criteria for Reporting and Evaluating ecotoxicity Data" (CRED) methodology, applying more objective criteria to assess the quality of toxicity tests, resulting in two reliable and relevant endpoints with only minor restrictions. HPLC-MS/MS was used to quantify the occurrence of DR1 in river waters of three sites, influenced by textile industry discharges, resulting in a concentration range of 50-500 ng L(-1). The risk quotients for DR1 obtained in this work suggest that this dye can pose a potential risk to freshwater biota. To reduce uncertainty of the derived PNEC, a fish partial or full lifecycle study should be performed., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

The widespread use of pesticides in agriculture can lead to water contamination and cause adverse effects on non-target organisms. Brazil has been the world's top pesticide market consumer since 2008, with 381 approved pesticides for crop use. This study provides a comprehensive literature review on the occurrence of pesticide residues in Brazilian freshwaters. We searched for information in official agency records and peer-reviewed scientific literature. Risk quotients were calculated to assess the potential risk posed to aquatic life by the individual pesticides based on their levels of water contamination. Studies about the occurrence of pesticides in freshwaters in Brazil are scarce and concentrated in few sampling sites in 5 of the 27 states. Herbicides (21) accounted for the majority of the substances investigated, followed by fungicides (11), insecticides (10) and plant growth regulators (1). Insecticides are the class of major concern. Brazil would benefit from the implementation of a nationwide pesticide freshwater monitoring program to support preventive, remediation and enforcement actions.

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 m(3) 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., (© 2015 Wiley Periodicals, Inc.)

Pharmaceutical effluents have recently been recognized as an important contamination source to aquatic environments and the toxicity related to the presence of antibiotics in effluents has attracted great attention. Conventionally, these effluents have been treated using physico-chemical and aerobic biological processes, usually with low rates of pharmaceuticals removal. Due to the complexity of effluents, it is impossible to determine all pharmaceuticals and their degradation products using analytical methods. Ecotoxicity tests with different organisms may be used to determine the effect level of effluents and thus their environmental impacts. The objective of this work was to compare the sensitivities of five ecotoxicity tests using aquatic and terrestrial organisms to evaluate the toxicity of effluents from the production of veterinary medicines before and after treatment. Raw and chemically treated effluent samples were highly toxic to aquatic organisms, achieving 100,000 toxic units, but only few of those samples presented phytotoxicity. We observed a reduction in the toxicity in the biologically treated effluent samples, which were previously chemically pre-treated, however the toxicity was not eliminated. The rank of test organisms' reactions levels was: Daphnia similis > Raphidocelis subcapitata > Aliivibrio fischeri > Allium cepa ~ Lactuca sativa. Effluent treatment employed by the evaluated company was only partially efficient at removing the effluent toxicity, suggesting potential risks to biota. The acute toxicity test with D. similis proved to be the most sensitive for both raw and treated effluents and is a suitable option for further characterization and monitoring of pharmaceutical effluents.

Benz[j]aceanthrylene (B[j]A) is a cyclopenta-fused polycyclic aromatic hydrocarbon with strong mutagenic and carcinogenic effects. We have identified B[j]A in air particulate matter (PM) in samples collected in Stockholm, Sweden and in Limeira, Brazil using LC-GC/MS analysis. Determined concentrations ranged between 1.57 and 12.7 and 19.6-30.2 pg/m(3) in Stockholm and Limeira, respectively, which was 11-30 times less than benzo[a]pyrene (B[a]P) concentrations. Activation of the DNA damage response was evaluated after exposure to B[j]A in HepG2 cells in comparison to B[a]P. We found that significantly lower concentrations of B[j]A were needed for an effect on cell viability compared to B[a]P, and equimolar exposure resulted in significant more DNA damage with B[j]A. Additionally, levels of γH2AX, pChk1, p53, pp53, and p21 proteins were higher in response to B[j]A than B[a]P. On the basis of dose response induction of pChk1 and γH2AX, B[j]A potency was 12.5- and 33.3-fold higher than B[a]P, respectively. Although B[j]A levels in air were low, including B[j]A in the estimation of excess lifetime cancer risk increased the risk up to 2-fold depending on which potency factor for B[j]A was applied. Together, our results show that B[j]A could be an important contributor to the cancer risk of air PM.

Polycyclic aromatic hydrocarbons (PAHs) are common environmental pollutants that occur naturally in complex mixtures. Many of the adverse health effects of PAHs including cancer are linked to the activation of intracellular stress response signaling. This study has investigated intracellular MAPK signaling in response to PAHs in extracts from urban air collected in Stockholm, Sweden and Limeira, Brazil, in comparison to BP in HepG2 cells. Nanomolar concentrations of PAHs in the extracts induced activation of MEK4 signaling with down-stream increased gene expression of several important stress response mediators. Involvement of the MEK4/JNK pathway was confirmed using siRNA and an inhibitor of JNK signaling resulting in significantly reduced MAPK signaling transactivated by the AP-1 transcription factors ATF2 and c-Jun. ATF2 was also identified as a sensitive stress responsive protein with activation observed at extract concentrations equivalent to 0.1 nM BP. We show that exposure to low levels of environmental PAH mixtures more strongly activates these signaling pathways compared to BP alone suggesting effects due to interactions. Taken together, this is the first study showing the involvement of MEK4/JNK/AP-1 pathway in regulating the intracellular stress response after exposure to nanomolar levels of PAHs in environmental mixtures., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

Caffeine has already been used as an indicator of anthropogenic impacts, especially the ones related to the disposal of sewage in water bodies. In this work, the presence of caffeine has been correlated with the estrogenic activity of water samples measured using the BLYES assay. After testing 96 surface water samples, it was concluded that caffeine can be used to prioritize samples to be tested for estrogenic activity in water quality programs evaluating emerging contaminants with endocrine disruptor activity.

This study evaluated the ecotoxicity of five dyes to freshwater organisms before and during their photo-Fenton degradation. EC50 (48h) of the five tested dyes ranged from of 6.9 to >1000mgL(-1) for Daphnia similis. In the chronic tests IC50 (72h) varied from 65 to >100mgL(-1) for Pseudokirchneriella subcapitata and IC50 (8 days) from 0.5 to 410mgL(-1) for Ceriodaphnia dubia. Toxicity tests revealed that although the applied treatment was effective for decolorization of the dye, the partial mineralization may be responsible for the presence of degradation products which can be either more toxic than the original dye, as is the case of Vat Green 3 and Reactive Black 5, lead to initially toxic products which may be further degraded to non toxic products (acid Orange 7 and Food Red 17), or generate non toxic products as in the case of Food Yellow 3. The results highlighted the importance of assessing both acute and chronic toxicity tests of treated sample before effluent discharge., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The Salmonella/microsome assay is the most used assay for the evaluation of air particulate matter (PM) mutagenicity and a positive correlation between strain TA98 responses and benzo[a]pyrene (B[a]P) levels in PM has been found. However, it seems that the major causes of PM mutagenicity in this assay are the nitro and oxy-PAHs. Salmonella YG5161, a 30-times more responsive strain to B[a]P has been developed. To verify if YG5161 strain was sufficiently sensitive to detect mutagenicity associated with B[a]P mutagenicity, PM samples were collected in Brazil and Sweden, extracted with toluene and tested in the Salmonella/microsome microsuspension assay. PAHs and B[a]P were determined and the extracts were tested with YG5161 and its parental strain TA1538. The extracts were also tested with YG1041 and its parental strain TA98. For sensitivity comparisons, we tested B[a]P and 1-nitropyrene (1-NP) using the same conditions. The minimal effective dose of B[a]P was 155 ng/plate for TA1538 and 7 ng/plate for YG5161. Although the maximum tested dose, 10 m(3) /plate containing 9 ng of B[a]P in the case of Brazilian sample, was sufficient to elicit a response in YG5161, mutagenicity was detected at a dose as low as 1 m(3) /plate (0.9 ng). This is probably caused by nitro-compounds that have been shown to be even more potent than B[a]P for YG5161. It seems that the mutagenicity of B[a]P present in PM is not detectable even with the use of YG5161 unless more efficient separation to remove the nitro-compounds from the PAH extract is performed., (Copyright © 2014 Wiley Periodicals, Inc.)

Triclosan (TCS) is a broad-spectrum bactericide, highly toxic to algae, which is released into the environment via wastewater effluents. Predicted no-effect concentrations (PNECs) for aquatic biota have been proposed in the literature, varying from 1.4 to 1,550 ng/L, reflecting contradicting protection goals. In this work, six rivers in the state of São Paulo were monitored for TCS and caffeine, a tracer for untreated sewage disposal, over a period of more than 1 year. From 71 samples analyzed, 32 contained TCS at concentrations above the limit of quantification, ranging from 2.2 to 66 ng/L, corresponding to a frequency of exceedance of the lowest PNEC of 86 % (six out of seven sites). No correlation between TCS and caffeine was observed, and one of the reasons for that could be the different use patterns in the local populations. Given the high values found in the investigated rivers, TCS seems to be a strong candidate in the priority list of compounds that should be regulated in Brazil to preserve the aquatic environment.

Background: Many dyes can be considered emerging contaminants. The most widely used dyes belong to the class of azo compounds, some of which are known to have toxic and genotoxic properties. They are used in great quantities in textile activities and are of environmental concern because of their potential discharge in water. Planarians have been successfully used as test organisms in hazard evaluation of different chemicals, and we demonstrate the suitability of Girardia tigrina for laboratory testing. The aim of this work was to evaluate the suitability of the planarian species G. tigrina to assess the ability of the azo textile dye disperse red 1 to cause acute toxicity and adverse effects in the regeneration and reproduction of newborn and adult specimens., Results: Disperse red 1 presented a median LC 50 of 75 and 152 mg/L, respectively, for newborns and adults of G. tigrina , showing that newborns are twice as susceptible to the dye. Uncoordinated movements, irregular twists, colored skin, increased mucous production, and regenerative delays were observed after dye exposure at sub-lethal concentrations., Conclusions: A no-observed-adverse-effect concentration (NOAEC) of 0.1 mg/L could be determined for disperse red 1 based on the fecundity test. Zinc seems to be a suitable positive control for monitoring the sensitivity in G. tigrina tests within only 24 h of exposure. This study demonstrates the applicability of G. tigrina tests in the hazard evaluation of water contaminants, such as azo dyes.

Currently, there are a large number of products (sunscreen, pigments, cosmetics, plastics, toothpastes and photocatalysts) that use TiO(2) nanoparticles. Due to this large production, these nanoparticles can be released into the aquatic, terrestrial and aerial environments at relative high concentration. TiO(2) in natural water has the capacity to harm aquatic organisms such as the Daphnia (Cladocera) species, mainly because the photocatalytic properties of this semiconductor. However, very few toxicity tests of TiO(2) nanoparticles have been conducted under irradiation. The aim of this study was to evaluate anatase and rutile TiO(2) toxicity to Daphnia similis exploring their photocatalytic properties by incorporating UV A and visible radiation as a parameter in the assays. Anatase and rutile TiO(2) samples at the highest concentration tested (100 mg L(-1)) were not toxic to D. similis, neither in the dark nor under visible light conditions. The anatase form and a mixture of anatase and rutile, when illuminated by a UV A black light with a peak emission wavelength of 360 nm, presented photo-dependent EC50 values of 56.9-7.8 mg L(-1), which indicates a toxicity mechanism caused by ROS (reactive oxygen species) generation., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Azo dyes are of environmental concern due to their degradation products, widespread use, and low-removal rate during conventional treatment. Their toxic properties are related to the nature and position of the substituents with respect to the aromatic rings and amino nitrogen atom. The dyes Disperse Red 1 and Disperse Red 13 were tested for Salmonella mutagenicity, cell viability by annexin V, and propidium iodide in HepG2 and by aquatic toxicity assays using daphnids. Both dyes tested positive in the Salmonella assay, and the suggestion was made that these compounds induce mainly frame-shift mutations and that the enzymes nitroreductase and O-acetyltransferase play an important role in the observed effect. In addition, it was shown that the presence of the chlorine substituent in Disperse Red 13 decreased the mutagenicity about 14 times when compared with Disperse Red 1, which shows the same structure as Disperse Red 13, but without the chlorine substituent. The presence of this substituent did not cause cytotoxicity in HepG2 cells, but toxicity to the water flea Daphnia similis increased in the presence of the chlorine substituent. These data suggest that the insertion of a chlorine substituent could be an alternative in the design of dyes with low-mutagenic potency, although the ecotoxicity should be carefully evaluated., (Copyright © 2010 Wiley Periodicals, Inc.)