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An abstract of the article "Membrane Dialysis Extraction (MDE): A Novel Approach for Extracting Toxicologically Relevant Hydrophobic Organic Compounds from Soils and Sediments for Assessment in Biotests" is presented. It was found that MDE is useful for the extraction of toxicologically organic compounds from soil. This compound is extracted from wet and dry sediments. The efficacy of the MDE is evaluated trough testing the extracts.

Chemicals in the environment occur in mixtures rather than as individual entities. Environmental quality monitoring thus faces the challenge to comprehensively assess a multitude of contaminants and potential adverse effects. Effect-based methods have been suggested as complements to chemical analytical characterisation of complex pollution patterns. The regularly observed discrepancy between chemical and biological assessments of adverse effects due to contaminants in the field may be either due to unidentified contaminants or result from interactions of compounds in mixtures. Here, we present an interlaboratory study where individual compounds and their mixtures were investigated by extensive concentration-effect analysis using 19 different bioassays. The assay panel consisted of 5 whole organism assays measuring apical effects and 14 cell- and organism-based bioassays with more specific effect observations. Twelve organic water pollutants of diverse structure and unique known modes of action were studied individually and as mixtures mirroring exposure scenarios in freshwaters. We compared the observed mixture effects against component-based mixture effect predictions derived from additivity expectations (assumption of non-interaction). Most of the assays detected the mixture response of the active components as predicted even against a background of other inactive contaminants. When none of the mixture components showed any activity by themselves then the mixture also was without effects. The mixture effects observed using apical endpoints fell in the middle of a prediction window defined by the additivity predictions for concentration addition and independent action, reflecting well the diversity of the anticipated modes of action. In one case, an unexpectedly reduced solubility of one of the mixture components led to mixture responses that fell short of the predictions of both additivity mixture models. The majority of the specific cell- and organism-based endpoints produced mixture responses in agreement with the additivity expectation of concentration addition. Exceptionally, expected (additive) mixture response did not occur due to masking effects such as general toxicity from other compounds. Generally, deviations from an additivity expectation could be explained due to experimental factors, specific limitations of the effect endpoint or masking side effects such as cytotoxicity in in vitro assays. The majority of bioassays were able to quantitatively detect the predicted non-interactive, additive combined effect of the specifically bioactive compounds against a background of complex mixture of other chemicals in the sample. This supports the use of a combination of chemical and bioanalytical monitoring tools for the identification of chemicals that drive a specific mixture effect. Furthermore, we demonstrated that a panel of bioassays can provide a diverse profile of effect responses to a complex contaminated sample. This could be extended towards representing mixture adverse outcome pathways. Our findings support the ongoing development of bioanalytical tools for (i) compiling comprehensive effect-based batteries for water quality assessment, (ii) designing tailored surveillance methods to safeguard specific water uses, and (iii) devising strategies for effect-based diagnosis of complex contamination. [ABSTRACT FROM AUTHOR]

Surface waters can contain a diverse range of organic pollutants, including pesticides, pharmaceuticals and industrial compounds. While bioassays have been used for water quality monitoring, there is limited knowledge regarding the effects of individual micropollutants and their relationship to the overall mixture effect in water samples. In this study, a battery of in vitro bioassays based on human and fish cell lines and whole organism assays using bacteria, algae, daphnids and fish embryos was assembled for use in water quality monitoring. The selection of bioassays was guided by the principles of adverse outcome pathways in order to cover relevant steps in toxicity pathways known to be triggered by environmental water samples. The effects of 34 water pollutants, which were selected based on hazard quotients, available environmental quality standards and mode of action information, were fingerprinted in the bioassay test battery. There was a relatively good agreement between the experimental results and available literature effect data. The majority of the chemicals were active in the assays indicative of apical effects, while fewer chemicals had a response in the specific reporter gene assays, but these effects were typically triggered at lower concentrations. The single chemical effect data were used to improve published mixture toxicity modeling of water samples from the Danube River. While there was a slight increase in the fraction of the bioanalytical equivalents explained for the Danube River samples, for some endpoints less than 1% of the observed effect could be explained by the studied chemicals. The new mixture models essentially confirmed previous findings from many studies monitoring water quality using both chemical analysis and bioanalytical tools. In short, our results indicate that many more chemicals contribute to the biological effect than those that are typically quantified by chemical monitoring programs or those regulated by environmental quality standards. This study not only demonstrates the utility of fingerprinting single chemicals for an improved understanding of the biological effect of pollutants, but also highlights the need to apply bioassays for water quality monitoring in order to prevent underestimation of the overall biological effect. [ABSTRACT FROM AUTHOR]

A key aspect of the transformation of the economic sector towards a sustainable bioeconomy is the development of environmentally friendly alternatives for hitherto used chemicals, which have negative impacts on environmental health. However, the implementation of an ecotoxicological hazard assessment at early steps of product development to elaborate the most promising candidates of lowest harm is scarce in industry practice. The present article introduces the interdisciplinary proof-of-concept project GreenToxiConomy, which shows the successful application of a Green Toxicology strategy for biosurfactants and a novel microgel-based pesticide release system. Both groups are promising candidates for industrial and agricultural applications and the ecotoxicological characterization is yet missing important information. An iterative substance- and application-oriented bioassay battery for acute and mechanism-specific toxicity within aquatic and terrestrial model species is introduced for both potentially hazardous materials getting into contact with humans and ending up in the environment. By applying in silico QSAR-based models on genotoxicity, endocrine disruption, skin sensitization and acute toxicity to algae, daphnids and fish, individual biosurfactants resulted in deviating toxicity, suggesting a pre-ranking of the compounds. Experimental toxicity assessment will further complement the predicted toxicity to elaborate the most promising candidates in an efficient pre-screening of new substances. [Display omitted] • Interdisciplinary project (GreenToxiConomy) on new bioeconomic high-value products. • Proof-of-concept study on the applicability of Green Toxicology in bioeconomy. • Development of an application-oriented bioassay battery for early hazard assessment. • QSAR-based toxicity prediction differentiates compounds of deviating toxicity. • Experimental toxicity assessment will identify most promising candidates. [ABSTRACT FROM AUTHOR]

Abstract: To evaluate the effect of crop residue burning of agriculturally tilled soils, the soil''s sorption of polycyclic aromatic hydrocarbons (PAHs) is studied using naphthalene as a representative model contaminant in the soil. Soils from agriculturally tilled fields (Dorobantu farm, Calarasi, Romania), either burned or unburned for clearance, were examined for their toxicity and sorption capacity (fields being burned for cleansing experience an increase in nanoparticles because soot particles enter the soil). The burned and unburned soils were separated into several fractions on the basis of size using membranes with different pore sizes. Batch experiments were conducted to test the impact of the following operating parameters: the initial concentration of naphthalene, initial concentration of soil, pH, soil structure and composition and soil characterization. The toxic effectiveness of the various soils was also investigated. [Copyright &y& Elsevier]

Petroleum products including crude oils and refined distillates are unique environmental pollutants consisting of thousands of compounds with varying physical-chemical properties and resulting toxicity for aquatic biota. Hence, for a reliable risk assessment individual petroleum product toxicity profiles are needed. Furthermore, the influence of oil spill response strategies like the application of chemical dispersants has to be implemented. The present study addressed the toxicity of water-accommodated fractions (WAFs) of two different oil types on fish early life stages on different biological organization levels in the laboratory model species Danio rerio. Experiments with a 3rd generation dispersant used in loading rated resembling the exposure in experiments with chemically dispersed oils were included, enabling a direct comparability of results. This approach is of high importance as especially the investigation of dispersant toxicity in relevant exposure concentrations is rather scarce. Zebrafish embryos were exposed to different WAFs shortly after and up to 120 hour post fertilization (hpf). Besides phenotypic effects including edema and spine deformations, reduced responses to dark stimuli, increased CYP1A activity and marginal AChE inhibition were observed in sublethal effect concentrations. Both oil types had varying strength of toxicity, which did not correlate with corresponding chemical analysis of target PAHs. Chemically dispersed oils induced stronger acute toxicity in zebrafish embryos compared to native (initial) oil exposure, which was further reflected by very low exposure concentrations for biomarker endpoints. Based on a comparison to the dispersant alone, a higher toxicity of dispersed oils was related to a combination of dispersant toxicity and an elevated crude oil compound bioavailability, due to dispersion-related partitioning kinetics. In contrast to LEWAF and CEWAF neither typical morphological effects nor mechanism-specific toxicity were observed for the dispersant alone, indicating narcosis as the responsible cause of effects. Unlabelled Image • Oil-specific (sub-)lethal effect concentrations for oil spill risk assessment were established. • Oil types induced different strength of embryo toxicity not correlated with common PAHs. • Sublethal exposure concentrations resulted in CYP1A induction but no AChE inhibition. • Larvae showed strong behavioral impairment already at low exposure concentrations. • Dispersant contributes to higher toxicity of chemically dispersed oils compared to native oils. [ABSTRACT FROM AUTHOR]

Ecotoxicological assays have traditionally focused on the effects of chemicals at the individual level by exploiting mortality and reproduction as endpoints. Although these two parameters are ecologically relevant, they rarely provide information regarding the elemental toxic mechanisms. Obviously, the number of xenobiotics used has been rapidly increased. Thus, any established measurement that shortens the actual outcome and, simultaneously provides information about toxic mechanisms is desirable. This research focused on the study of oxidative stress response as a biomarker in the eukaryotic model organism, Saccharomyces cerevisiae. For this, yeast cells were exposed to a set of selected environmentally relevant chemicals via different approaches, including cellular diagnostics, gene expression analysis and chemo-genetic screening. The results demonstrated that at the cellular level, model organisms reacted to different chemicals in distinct manner. For each xenobiotic, the correlation between toxic response of molecular and cellular levels are presented. Namely, the expression of target genes after chemical exposure affected the cellular alteration as evidenced by an elevated level of superoxide dismutase and a reduced amount of glutathione. Furthermore, the results derived from chemo-genetic screening, in which mutants lacking of gene of interest were employed, exhibited more susceptibility to test chemicals in comparison to the wildtype. The response of oxidative stress upon chemical exposure in budding yeast from this study is potentially useful for an establishment of a proper bio-test system which can eventually be linked to adverse effects at an individual level in higher eukaryotes., 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 Inc. All rights reserved.)

The pollution state in the German Bight was investigated by determination of pollutant concentrations of sediment samples using equilibrium passive sampling. Polychlorinated biphenyls (PCBs) and polycyclic aromatic hydrocarbons (PAH) were determined in the pore water of North Sea sediment. The freely dissolved pore water concentration (C free ) was measured applying Solid Phase Microextraction (SPME) by using PDMS-coated glass fibers. The obtained results show that the North Sea contamination level with the investigated pollutants is relatively low. However, the stations close to the sediment-dumping site were higher contaminated. A macrofauna analysis showed that bioturbation activities were mostly present in the upper sediment layers, but a direct bioturbation influence on the sediment concentration distribution could not be shown. Overall, the contamination load was below baseline toxicity, but considering that several other priority pollutants will also make a contribution to the baseline toxicity, it can be counted as relatively high., 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.)

Around the world, many ambitious environmental conventions and regulations have been implemented over recent decades. Despite this, the environment is still deteriorating. An increase in the volume and diversity of chemicals is one of the main drivers of this deterioration, of which biodiversity loss is a telling indicator. In response to this situation, in October 2020, a chemicals strategy for sustainability (CSS) was published in the EU. The CSS is the first regional framework aiming to address chemical pollution in a holistic manner. The CSS covers the complete lifecycle of a chemical, including the design of better substances and remediation options, to remove chemicals from the environment. The strategy contains terms, such as a "toxic-free environment," for which no clear definition exists, potentially hampering the implementation of the CSS. In this paper, a definition for a "toxic-free environment" is proposed on the basis of a survey and a discussion held at the 2020 SETAC Europe Annual Meeting. In addition, key issues that are absent from the CSS but are considered to be key for the realization of a toxic-free environment are identified. To achieve the policy goals, it is recommended to align the definition of risk across the different chemical legislations, to establish a platform for open data and data sharing, and to increase the utility and use of novel scientific findings in policymaking, through the development of a strong science to regulation feedback mechanism and vice versa. The paper concludes that environmental scientists have the tools to address the key challenges presented in the CSS. However, an extra step is needed by both policymakers and scientists to develop methods, processes and tools, to increase the robustness and transparency of deliberation processes, and the utility of science. Integr Environ Assess Manag 2021;17:1105-1113. © 2021 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals LLC on behalf of Society of Environmental Toxicology & Chemistry (SETAC)., (© 2021 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals LLC on behalf of Society of Environmental Toxicology & Chemistry (SETAC).)

Green toxicology is a novel approach increasingly applied for the development of materials and chemicals that are more benign to the environment and human health than their conventional counterparts. It includes predictive eco-toxicological assessments of chemicals during the early developmental process to exclude adverse effects. In the present study, two guanidine zinc catalysts for the ring-opening polymerization of lactide were investigated using eco-toxicological tools. Namely, the fish embryo toxicity assay for teratogenic effects, the ER (α) CALUX assay for endocrine activity and the Ames fluctuation assay for mutagenic potential were applied. Both complexes showed no endocrine activity, mutagenicity or acute aquatic toxicity, however a delayed hatch could be observed, therefore suggesting potential effects on a molecular level. This proof-of-concept study aims to assess the toxicity of guanidine zinc catalysts and is a first step towards the incorporation of toxicological assessments into chemical developmental processes to achieve a sustainable and safe production of catalysts., (Copyright © 2021 Elsevier B.V. All rights reserved.)

In situ burning (ISB) is discussed to be one of the most suitable response strategies to combat oil spills in extreme conditions. After burning, a highly viscous and sticky residue is left and may over time pose a risk of exposing aquatic biota to toxic oil compounds. Scientific information about the impact of burn residues on the environment is scarce. In this context, a comprehensive ISB field experiment with approx. 1000L IFO 180 was conducted in a fjord in Greenland. The present study investigated the toxicity of collected ISB residues to early life stages of zebrafish (Danio rerio) as a model for potentially exposed pelagic organisms. The toxicity of ISB residues on zebrafish embryos was compared with the toxicity of the initial (unweathered) IFO 180 and chemically dispersed IFO 180. Morphological malformations, hatching success, swimming behavior, and biomarkers for exposure (CYP1A activity, AChE inhibition) were evaluated in order to cover the toxic response on different biological organization levels. Across all endpoints, ISB residues did not induce greater toxicity in zebrafish embryos compared with the initial oil. The application of a chemical dispersant increased the acute toxicity most likely due to a higher bioavailability of dissolved and particulate oil components. The results provide insight into the adverse effects of ISB residues on sensitive life stages of fish in comparison with chemical dispersant application.

The leaf-shredding crustacean Hyalella azteca, which is indigenous to Northern and Central America, is used to assess environmental risks associated with (metal-)contaminated sediments and to propose sediment quality standards also in Europe. Yet, it is unknown if H. azteca is protective for European crustacean shredders. We thus compared the sensitivity of H. azteca with that of the European species Asellus aquaticus and Gammarus fossarum towards copper- and cadmium-contaminated sediments (prepared according to OECD 218) under laboratory conditions employing mortality and leaf consumption as endpoints. H. azteca either reacted approximately fourfold more sensitive than the most tolerant tested species (as for cadmium) or its sensitivity was only 1.6 times lower than the highest sensitivity determined (as for copper), which should be covered by safety factors applied during risk assessments. Therefore, the results for the sediment type and the two heavy metals tested during the present study in combination with the existence of standardized testing protocols, their ease of culture, and short generation time, suggest H. azteca as suitable crustacean model shredder for assessing the toxicity of sediment-associated metals in Europe.

Water pollution risks to human health and the environment are emerging as serious concerns in the European Union and worldwide. With the aim to achieve good ecological and chemical status of all European water bodies, the "European Water Framework Directive" (WFD) was enacted. With the framework, bioanalytical techniques have been recognized as an important aspect. However, there are limitations to the application of bioassays directly for water quality assessment. Such approaches often fail to identify pollutants of concern, since the defined priority and monitored pollutants often fail to explain the observed toxicity. In this study, we integrated an effect-based risk assessment with a zebrafish-based investigation strategy to evaluate water sample extracts and fractions collected from the Danube. Four tiered bioassays were implemented, namely RNA-level gene expression assay, protein-level ethoxyresorufin-O-deethylase (EROD) assay, cell-level micronucleus assay and organism-level fish embryo test (FET). The results show that teratogenicity and lethality during embryonic development might be induced by molecular or cellular damages mediated by the aryl hydrocarbon receptor (AhR) -mediated activity, estrogenic activity and genotoxic activity. With the combination of high-throughput fractionation, this effect-based strategy elucidated the major responsible mixtures of each specific toxic response. In particularly, the most toxic mixture in faction F4, covering a log Kow range from 2.83 to 3.42, was composed by 12 chemicals, which were then evaluated as a designed mixture. Our study applied tiered bioassays with zebrafish to avoid interspecies differences and highlights effect-based approaches to address toxic mixtures in water samples. This strategy can be applied for large throughput screenings to support the main toxic compounds identification in water quality assessment., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Investigations of deleterious effects on non-target species, including earthworms, have been conducted for a number of pesticides, but there is a need for additional assessments of potential adverse effects. In the present study, the acute toxicity of eight pesticides to the earthworm Eisenia andrei was assessed and compared. The exposures were conducted using the filter paper contact toxicity method. Based on the 48-h LC 50 values, one pesticide was classified as supertoxic (combined fungicide containing difenoconazole and fludioxonil), four as extremely toxic (combined herbicide containing pethoxamide and terbuthylazine, combined fungicide containing fluopyram and tebuconazole, fungicide containing pyrimethanil, and combined fungicide containing thiram and carboxin), two as very toxic (combined fungicide containing flutriafol and thiabendazole, and herbicide containing fluroxypyr-meptyl), and one as moderately toxic (insecticide containing thiamethoxam). Additionally, effects of pesticides on the multixenobiotic resistance (MXR) activity were measured. Results showed that four pesticides caused significant effects with a recorded inhibition of the activity, which can consequently lead to a higher toxicity due to longer retention of the pesticides in the cells. Finally, for three chosen pesticides, gene expression of cat, sod, and gst was measured, and significant changes were observed. The obtained results show that earthworms could be significantly affected by pesticides commonly used in agriculture.

In Germany, micropollutants that (may) occur in drinking water are assessed by means of the health-related indicator value (HRIV concept), developed by the German Federal Environment Agency. This concept offers five threshold values (≤ 0.01 to ≤ 3 μg l -1 ) depending on availability and completeness of data regarding genotoxicity, neurotoxicity, and germ cell-damaging potential. However, the HRIV concept is yet lacking integration of endocrine disruptors as one of the most prominent toxicological concerns in water bodies, including drinking water. Thresholds and proposed bioassays hence urgently need to be defined. Since endocrine disruption of ubiquitary chemicals as pharmaceuticals, industrial by-products, or pesticides is a big issue in current ecotoxicology, the aim of this study was to explore endocrine effects, i.e., estrogenic and androgenic effects, as an important, additional toxicological mode of action for the HRIV concept using a hierarchical set of well-known but improved bioassays. Results indicate that all of the 13 tested substances, industrial chemicals and combustion products (5), pharmaceuticals and medical agents (4), and pesticides and metabolites (4), have no affinity to the estrogen and androgen receptor in human U2OS cells without metabolic activation, even when dosed at their water solubility limit, while in contrast some of these substances showed estrogenic effects in the RYES assay, as predicted in pre-test QSAR analysis. Using a specifically developed S9-mix with the U2OS cells, those micropollutants, i.e., Benzo[a]pyrene, 2,4-Dichlorophenol, 3,3-Dichlorbenzidin, 3,4-Dichloranilin, and diclofenac, they show estrogenic effects at the same concentration range as for the yeast cells. Three of the drinking water-relevant chemicals, i.e., atrazine, tributyltin oxide, and diclofenac, caused effects on hormone production in the H295R assay, which can be correlated with changes in the expression of steroidogenic genes. One chemical, 17α-Ethinylestradiol, caused an estrogenic or anti-androgenic effect in the reproduction test with Potamopyrgus antipodarum. Considering these results, a proposal for a test strategy for micropollutants in drinking water regarding potential endocrine effects (hormonal effects on reproduction and sexual development) will be presented to enhance the existing HRIV concept.

The binding of the p53 tumor suppression protein to DNA response elements after genotoxic stress can be quantified by cell-based reporter gene assays as a DNA damage endpoint. Currently, bioassay evaluation of environmental samples requires further knowledge on p53 induction by chemical mixtures and on cytotoxicity interference with p53 induction analysis for proper interpretation of results. We investigated the effects of genotoxic pharmaceuticals (actinomycin D, cyclophosphamide) and nitroaromatic compounds (4-nitroquinoline 1-oxide, 3-nitrobenzanthrone) on p53 induction and cell viability using a reporter gene and a colorimetric assay, respectively. Individual exposures were conducted in the absence or presence of metabolic activation system, while binary and tertiary mixtures were tested in its absence only. Cell viability reduction tended to present direct correlation with p53 induction, and induction peaks occurred mainly at chemical concentrations causing cell viability below 80%. Mixtures presented in general good agreement between predicted and measured p53 induction factors at lower concentrations, while higher chemical concentrations gave lower values than expected. Cytotoxicity evaluation supported the selection of concentration ranges for the p53 assay and the interpretation of its results. The often used 80% viability threshold as a basis to select the maximum test concentration for cell-based assays was not adequate for p53 induction assessment. Instead, concentrations causing up to 50% cell viability reduction should be evaluated in order to identify the lowest observed effect concentration and peak values following meaningful p53 induction.

Bioassays are particularly useful tools to link the chemical and ecological assessments in water quality monitoring. Different methods cover a broad range of toxicity mechanisms in diverse organisms, and account for risks posed by non-target compounds and mixtures. Many tests are already applied in chemical and waste assessments, and stakeholders from the science-police interface have recommended their integration in regulatory water quality monitoring. Still, there is a need to address bioassay suitability to evaluate water samples containing emerging pollutants, which are a current priority in water quality monitoring. The presented interlaboratory study (ILS) verified whether a battery of miniaturized bioassays, conducted in 11 different laboratories following their own protocols, would produce comparable results when applied to evaluate blinded samples consisting of a pristine water extract spiked with four emerging pollutants as single chemicals or mixtures, i.e. triclosan, acridine, 17α-ethinylestradiol (EE2) and 3-nitrobenzanthrone (3-NBA). Assays evaluated effects on aquatic organisms from three different trophic levels (algae, daphnids, zebrafish embryos) and mechanism-specific effects using in vitro estrogenicity (ER-Luc, YES) and mutagenicity (Ames fluctuation) assays. The test battery presented complementary sensitivity and specificity to evaluate the different blinded water extract spikes. Aquatic organisms differed in terms of sensitivity to triclosan (algae > daphnids > fish) and acridine (fish > daphnids > algae) spikes, confirming the complementary role of the three taxa for water quality assessment. Estrogenicity and mutagenicity assays identified with high precision the respective mechanism-specific effects of spikes even when non-specific toxicity occurred in mixture. For estrogenicity, although differences were observed between assays and models, EE2 spike relative induction EC 50 values were comparable to the literature, and E2/EE2 equivalency factors reliably reflected the sample content. In the Ames, strong revertant induction occurred following 3-NBA spike incubation with the TA98 strain, which was of lower magnitude after metabolic transformation and when compared to TA100. Differences in experimental protocols, model organisms, and data analysis can be sources of variation, indicating that respective harmonized standard procedures should be followed when implementing bioassays in water monitoring. Together with other ongoing activities for the validation of a basic bioassay battery, the present study is an important step towards the implementation of bioanalytical monitoring tools in water quality assessment and monitoring., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Effluents from wastewater treatment plants (WWTPs) are a major source of estrogenic compounds to the aquatic environment. In the present work, estrogenic activities of effluents from eight municipal WWTPs in Finland were studied. The main objectives of the study were to quantify the concentrations of selected estrogenic compounds, to evaluate their contribution to estrogenic potency and to test the feasibility of the commercial bioassays for wastewater analysis. The effluent samples were analyzed by two in vitro tests, i.e. ERα-CALUX(®) and ELISA-E2, and by liquid chromatography mass spectrometry for six estrogenic compounds: estrone (E1), 17β-estradiol (E2), estriol (E3), 17α-ethinylestradiol (EE2), 17α-estradiol and bisphenol A (BPA). Estrogenic effects were found in all of the effluent samples with both of the bioassays. The concentrations measured with ELISA-E2 (8.6-61.6 ng/L) were clearly higher but exhibited a similar pattern than those with chemical analysis (E2

This protocol describes a quantitative and robust 96-well-plate-reader-based assay for the measurement of ethoxyresorufin-O-deethylase (EROD) activity using the rat hepatoma cell line H4IIE. The assay can be used to determine the cytochrome P450 subfamily 1A (CYP1A)-inducing potential of single substances, as well as of mixtures and extracts of samples. It is based on the aryl hydrocarbon receptor (AhR)-mediated induction of cytochrome P450 enzymes (subfamily 1A) in cells after exposure to dioxins and dioxin-like compounds. One enzymatic reaction catalyzed by CYP1A is the deethylation of the exogenous substrate 7-ethoxyresorufin to the fluorescent product resorufin, which is measured as EROD activity in the assay. The CYP1A-inducing potential of a sample can be reliably quantified by comparing the EROD activity with the concentration-response curve of the standard substance 2,3,7,8-tetrachlorodibenzo-p-dioxin, which can be detected at concentrations down to the picogram per liter range. A researcher familiar with the procedure can process up to 160 samples with four wells each within 3 d. The series described uses four plates with three concentrations per sample, which can be easily scaled to accommodate different sample sizes.

A wide variety of organic contaminants including pesticides, polycyclic aromatic hydrocarbons (PAHs) and polychlorinated biphenyls (PCBs) have previously been detected in surface waters in the river Ythan catchment, North East Scotland UK. While the concentrations detected were below Water Framework Directive Environmental Quality Standards (WFD-EQSs) environmental exposures to the diverse mixtures of contaminants, known and unknown, may pose chronic and/or sublethal effects to non target organisms. The present study assessed the embryo and algal toxicity potential of freely dissolved organic contaminants from the Ythan catchment using silicone rubber passive sampling devices (SR-PSDs) and miniaturised bioassay techniques. Zebrafish (Danio rerio) embryos and marine phytoplankton species (Diacronema lutheri) were exposed to extracts from SR-PSDs deployed at different locations along the river Ythan and an undeployed procedural blank. Statistically significant developmental and algal toxicities were measured in all tests of extracts from deployed samples compared with the procedural blanks. This indicates environmental exposure to, and the combined toxicity potential of, freely dissolved organic contaminants in the catchment. The present and previous studies in the Ythan catchment, coupling SR-PSDs and bioassay techniques, have both helped to understand the interactions and combined effects of dissolved organic contaminants in the catchment. They have further revealed the need for improvement in the techniques currently used to assess environmental impact.

The zebrafish embryo test is a widely used bioassay for the testing of chemicals, effluents and other types of environmental samples. Oxygen depletion in the testing of sediments and effluents is especially important and may be a confounding factor in the interpretation of apparent toxicity. In order to identify oxygen levels critical to early developmental stages of zebrafish, oxygen consumption of zebrafish embryos between 0 and 96h post-fertilization, minimum oxygen levels required by the embryos for survival as well as the effects of oxygen depletion following exposure to model sediments were determined. No significant effects on zebrafish embryo development were observed for oxygen concentrations between 7.15 and 3.33mg/L, whereas at concentrations between 3.0and 2.0mg/L minor developmental retardations were observed, yet without any pathological consequences. Oxygen concentrations lower than 0.88mg/L were 100% lethal. In the sediment contact tests with zebrafish embryos, native sediments rich in organic materials rapidly developed strongly hypoxic conditions, particularly at the sediment-water interface (0 to 500μm distance to the sediment)., (Copyright © 2010 Elsevier Inc. All rights reserved.)