Fish Embryo Acute Toxicity Research Articles

Impact of the glyphosate-based commercial herbicide, its components and its metabolite AMPA on non-target aquatic organisms

Glyphosate (GLY) is the active ingredient of several herbicide formulations widely used to control weeds in agricultural and non-agricultural areas. Due to the intensive use of GLY-based herbicides and their direct application on soils, some of their components, including the active ingredient, may reach the aquatic environment through direct run-off and leaching. The present study assessed the acute toxicity and genotoxicity of the GLY-based formulation Atanor 48 (ATN) and its major constituents GLY, surfactant polyethoxylated tallow amine (POEA), as well as the main metabolite of GLY aminomethylphosphonic acid (AMPA) on non-target aquatic organisms. The toxic effects of these chemicals were evaluated in the fish embryo acute toxicity test with zebrafish (Danio rerio), while genotoxic effects were investigated in the comet assays with cells from zebrafish larvae and rainbow trout gonad-2 (RTG-2). GLY and AMPA caused no acute toxic effect, while ATN and POEA induced significant lethal effects in zebrafish (LC50-96 h 76.50 mg/L and 5.49 mg/L, respectively). All compounds were genotoxic in comet experiments with zebrafish larvae (LOEC 1.7 mg/L for GLY, ATN, AMPA and 0.4 mg/L for POEA). Unlike in vivo, only POEA induced DNA damage in RTG-2 cells (LOEC 1.6 mg/L), suggesting that it is a direct acting genotoxic agent. In summary, these data indicate that the lethal effects on zebrafish early-life stages can be ranked in the following order from most to least toxic: surfactant POEA > formulation ATN > active ingredient GLY ≈ metabolite AMPA. Genotoxic effects were observed in both RTG-2 cells (only POEA) and zebrafish (all test compounds) with the lowest tested concentrations. Therefore, it is important to evaluate different toxicological endpoints as well as use different non-target organisms to predict the hazards of GLY-based formulations and their components and breakdown product to aquatic biota.

Ecotoxicological assessment and electrochemical remediation of doxorubicin

Doxorubicin (DOX) is an anthracycline widely used in treatments of several cancers, so it has found in hospital effluents with a significant concentration (above 1 μg L−1). Electrochemical remediation is an alternative to promote its degradation. The aim of this work was to evaluate the ability of nanostructured graphite electrodes with metallic oxides to degrade DOX by electro-oxidation (EO). Graphite, TiO2@graphite and AuO-TiO2@graphite electrodes were used in medium with tap water or 10 mmol L−1 NaCl. DOX treatments at concentrations of 1.25–5 mg L−1 were carried out in a voltage source with 1.5–5 V. The cathode used was the platinum electrode. The treatment of DOX 1.25 mg L−1 with 10 mmol L−1 NaCl electrolyte using the AuO-TiO2@graphite electrode at 5 V and 1 mA was the best methodology to promote its degradation. Also, the modified electrode was efficient to DOX degradation after 17 cycles of reuse. An energy expenditure of 1.11 and 0.2 kWh m−3 were obtained for 3 and 50 mL of treatment, respectively. Fish embryo acute toxicity test with zebrafish (Danio rerio) were performed before and after treatment by EO using NaCl. This treatment caused no effect on embryo-larval development, however it induced significant damage in the DNA of the zebrafish larvae after 96 h of exposure, which emphasizes the importance of a depth ecotoxicological evaluation during the development of EO methodologies.

Zebrafish embryo and acute fish toxicity test show similar sensitivity for narcotic compounds_suppl2

The zebrafish embryo test has been discussed as an alternative test system to provide data on acute fish toxicity required by diverse regulations. A meta-analysis of zebrafish embryo acute toxicity (ZFET) data has revealed conflicting evidence that narcotic compounds (i.e., compounds with baseline toxicity) may exhibit weaker sensitivity in the ZFET compared to the acute (adult) fish toxicity test (AFT). Therefore, six compounds with presumably narcotic or unknown mode of action, and for which a previous meta-analysis had indicated weaker sensitivity, were experimentally analyzed for their fish embryo acute toxicity and exposure concentrations were monitored. The data indicated that ZFET and AFT for the selected compounds had similar sensitivity and differences were in the range of species differences of the AFT.

Identifying sublethal endpoints for evaluating neurotoxic compounds utilizing the fish embryo toxicity test

Fish embryos are increasingly being utilized in aquatic toxicity testing, as evidenced by the Organisation for Economic Co-operation and Development's approval of the fish embryo acute toxicity (FET) test. However, the FET test only allows for the estimation of acute toxicity, whereas other test methods such as the larval growth and survival (LGS) test allow for the estimation of both acute and chronic toxicity. Additionally, it has been demonstrated that the FET test is less sensitive than other test methods for some neurotoxic compounds. To address these limitations, efforts to identify sublethal endpoints that increase FET test sensitivity and allow for the prediction of sublethal adverse effects have begun. As such, the objectives of the current study were 1) to compare estimated LC50 values from the FET and LGS test for three known neurotoxicants: fluoride (F), nickel (Ni), and cadmium (Cd) and 2) to evaluate the responsiveness of potential sublethal endpoints for the FET test related to growth (i.e., wet weight and snout-vent length), neurological development (i.e., spontaneous contraction frequency and eye size), and cardiovascular function (i.e., heart rate and pericardial area). The calculated LC50 values from the F and Cd FET test were significantly higher than those from the LGS test, demonstrating that the FET test is less sensitive than the LGS test for neurotoxic compounds. Only Cd exposure resulted in alterations in any of the sublethal endpoints investigated. Embryos/eleutheroembryos exposed to Cd displayed alterations in length, eye size, and pericardial area at concentrations five-fold less than the estimated LC50 value, suggesting that for Cd the inclusion of these sublethal endpoints would improve the sensitivity of the FET test. Overall, these results provide evidence that for some neurotoxicants, the inclusion of sublehtal endpoints may improve the utility of the FET test; however, further research utilizing a broader range of neurotoxicants with differing mechanisms of action, is needed to fully establish such endpoints in the context of routine FET test.

283 - Mapping glutathione utilization in the developing zebrafish embryo

Glutathione (GSH), the most abundant endogenous redox buffer in vertebrates, plays a key role in organogenesis and embryonic development. Toxic insults during development are commonly neutralized by glutathionylation and excretion. Despite its importance, scant information exists regarding organ-specific GSH utilization during development. Monochlorobimane (MCB) is a dye that is conjugated with GSH by glutathione-s-transferase (GST) to form a fluorescent adduct. Here, we use MCB staining to visualize organ-specific location of GSH utilization in live developing zebrafish embryos at developmental timepoints commonly used in the OECD fish embryo acute toxicity test. At 24 hours post-fertilization (hpf), GSH conjugation with MCB was highest in the heart, brain and yolk, increasing in the gut and otolith at 48 hpf. By 72 hpf, MCB staining was higher in the endoderm-derived lineage, and at 96hpf it was brightest in the liver, gut, and gill arches. We then tested the sensitivity and specificity of MCB staining with known GSH modulators . A 10-minute treatment at 48 hpf with the prooxidant tert-butyl hydroperoxide, caused organ-specific reductions in MCB staining, with the heart and brain losing 30% fluorescence, and, the gut and myotome losing 50%. A 24 hour treatment from 24-48 hpf with the GSH precursor N-Acetylcysteine resulted in significantly increased fluorescence, with the brain showing a 640% increase and the heart showing a 350% increase. The gut and myotome showed a 100% increase. Ethacrynic acid, a highly specific GST inhibitor caused decreased fluorescence; the heart, brain and myotome lost 30% fluorescence and the gut lost 50%. 2-AAPA, an inhibitor of Glutathione Reductase, caused similar organ specific losses in fluorescence with the gut being more susceptible than the heart and brain. Our data indicate highly specific spatio-temporal GSH utilization, with different organs having differing resilience against GSH system disruptions during embryonic development.

Zebrafish embryo and acute fish toxicity test show similar sensitivity for narcotic compounds.

The zebrafish embryo test has been discussed as an alternative test system to provide data on acute fish toxicity required by diverse regulations. A meta-analysis of zebrafish embryo acute toxicity (ZFET) data has revealed conflicting evidence that narcotic compounds (i.e. compounds with baseline toxicity) may exhibit weaker sensitivity in the ZFET if compared to the acute (adult) fish toxicity test (AFT). Therefore, six compounds with presumably narcotic or unknown mode of action, and for which a previous meta-analysis has indicated weaker sensitivity were experimentally analysed for their fish embryo acute toxicity and exposure concentrations were monitored. The data indicated that ZFET and AFT for the selected compounds revealed similar sensitivity and differences were in the range of species differences of the AFT.

RuII( p-cymene) Compounds as Effective and Selective Anticancer Candidates with No Toxicity in Vivo.

Ruthenium(II) complexes are currently considered a viable alternative to the widely used platinum complexes as efficient anticancer agents. We herein present the synthesis and characterization of half-sandwich ruthenium compounds with the general formula [Ru( p-cymene)(L-N,N)Cl][CF3SO3] (L = 3,6-di-2-pyridyl-1,2,4,5-tetrazine (1) 6,7-dimethyl-2,3-bis(pyridin-2-yl)quinoxaline (2)), which have been synthesized by substitution reactions from the precursor dimer [Ru( p-cymene)(Cl)(μ-Cl)]2 and were characterized by elemental analysis, mass spectrometry, 1H NMR, UV-vis, and IR spectroscopy, conductivity measurements, and cyclic voltammetry. The molecular structure for complex 2 was determined by single-crystal X-ray diffraction. The cytotoxic activity of these compounds was evaluated against human tumor cells, namely ovarian carcinoma A2780 and breast MCF7 and MDAMB231 adenocarcinoma cells, and against normal primary fibroblasts. Whereas the cytotoxic activity of 1 is moderate, IC50 values found for 2 are among the lowest previously reported for Ru( p-cymene) complexes. Both compounds present no cytotoxic effect in normal human primary fibroblasts when they are used at the IC50 concentration in A2780 and MCF7 cancer cells. Their antiproliferative capacity is associated with a combined mechanism of apoptosis and autophagy. A strong interaction with DNA was observed for both with a binding constant value of the same magnitude as that of the classical intercalator [Ru(phen)2(dppz)]2+. Both complexes bind to human serum albumin with moderate to strong affinity, with conditional binding constants (log Kb) of 4.88 for complex 2 and 5.18 for complex 1 in 2% DMSO/10 mM Hepes pH7.0 medium. The acute toxicity was evaluated in zebrafish embryo model using the fish embryo acute toxicity test (FET). Remarkably, our results show that compounds 1 and 2 are not toxic/lethal even at extremely high concentrations. The novel compounds reported herein are highly relevant antitumor metallodrug candidates, given their in vitro cytotoxicity toward cancer cells and the lack of in vivo toxicity.

Comparative ecotoxicity of potential biofuels to water flea (Daphnia magna), zebrafish (Danio rerio) and Chinese hamster (Cricetulus griseus) V79 cells

The ecotoxicity of two biofuel candidates (1‑octanol and 2‑butanone) was investigated by an integrative test strategy using three bioassays: the acute immobilisation test with water flea (D. magna), the fish embryo acute toxicity test with zebrafish (Danio rerio) and the in vitro micronucleus assay with Chinese hamster (Cricetulus griseus) V79 cells. The median effective concentration (EC50) values were 14.9±0.66mgL−1 for 1‑octanol, and 2152.1±44.6mgL−1 for 2‑butanone in the D. magna test. Both 1‑octanol and 2‑butanone caused teratogenic and lethal effects on zebrafish embryos, while exposure to 1‑octanol significantly induced these effects at concentrations ≥2.0mgL−1. These results indicate that 1‑octanol exert much higher ecotoxicity than 2‑butanone to D. magna and zebrafish embryos. Moreover, both 1‑octanol and 2‑butanone did not cause significant genotoxic effects, while their metabolites significantly induced micronuclei in V79 cells. The present study proposed an integrative test approach to evaluate the potential ecotoxicity of biofuels using simple, quick and inexpensive bioassays.

Applicability of the fish embryo acute toxicity (FET) test (OECD 236) in the regulatory context of Registration, Evaluation, Authorisation, and Restriction of Chemicals (REACH).

In 2013 the Organisation for Economic Co-operation and Development (OECD) test guideline (236) for fish embryo acute toxicity (FET) was adopted. It determines the acute toxicity of chemicals to embryonic fish. Previous studies show a good correlation of FET with the standard acute fish toxicity (AFT) test; however, the potential of the FET test to predict AFT, which is required by the Registration, Evaluation, Authorisation, and Restriction of Chemicals (REACH) regulation (EC 1907/2006) and the Classification, Labelling and Packaging (CLP) Regulation (EC 1272/2008), has not yet been fully clarified. In 2015 the European Chemicals Agency (ECHA) requested that a consultant perform a scientific analysis of the applicability of FET to predict AFT. The purpose was to compare the toxicity of substances to fish embryos and to adult fish, and to investigate whether certain factors (e.g., physicochemical properties, modes of action, or chemical structures) could be used to define the applicability boundaries of the FET test. Given the limited data availability, the analysis focused on organic substances. The present critical review summarizes the main findings and discusses regulatory application of the FET test under REACH. Given some limitations (e.g., neurotoxic mode of action) and/or remaining uncertainties (e.g., deviation of some narcotic substances), it has been found that the FET test alone is currently not sufficient to meet the essential information on AFT as required by the REACH regulation. However, the test may be used within weight-of-evidence approaches together with other independent, relevant, and reliable sources of information. The present review also discusses further research needs that may overcome the remaining uncertainties and help to increase acceptance of FET as a replacement for AFT in the future. For example, an increase in the availability of data generated according to OECD test guideline 236 may provide evidence of a higher predictive power of the test. Environ Toxicol Chem 2018;37:657-670. © 2017 SETAC.

Preclinical safety and immunological efficacy of Alternaria alternata polymerized extracts.

Introduction Alternaria alternata is a widespread fungi whose allergy is a risk factor for asthma development. The use of a polymerized allergen extract (allergoid) may be safer than native extract based treatments while maintaining efficacy. The objective of this study was to characterize biochemically and immunochemically a new Alternaria alternata allergoid.MethodsCharacterization of native and allergoid extracts was performed by determination of protein content, protein and allergenic profile, biological potency, identification of Alternaria allergens, and Alt a 1 quantification. Safety was evaluated in toxicological assays (Ames test, limit test, and fish embryo acute toxicity test in zebrafish, and maximum tolerated dose and Dose‐range finding study in rats). Efficacy was evaluated as the capacity to induce IgG antibodies that block IgE‐binding to the allergen and cytokine induction (IFN‐γ, IL‐4, IL‐6, IL‐10, and TNF‐α) in PBMC from atopic donors.ResultsProtein and antigenic profiles showed significant modification of the depigmented allergoid with respect to the native extract, inducing a lower IgE binding capacity. Alt a 1, Alt a 3, Alt a 6, and Alt a 8 allergen sequences were identified in the polymer. No toxicological nor genotoxicity effects were observed. The polymer induced IgG antibodies that blocked human IgE binding epitopes, and it induced higher IL‐10 levels and similar levels of the other cytokines than native extract in PBMC.ConclusionsThis new A. alternata allergoid could be an effective immunotherapy treatment leading to cytokine stimulation and inducing synthesis of IgG antibodies able to block IgE binding to the allergen. In addition, no toxicological effect was observed, and it may be safer than native extract due to its lower IgE binding capacity and cytokine induction that suggest tolerance induction via T cell shift to Treg (IL‐10).

Developmental toxicity and inhibition of the fungicide hymexazol to melanin biosynthesis in zebrafish embryos

Hymexazol is an efficacious and widely used fungicide. However, its environmental toxicological assessment has not been well documented. It had no report of its toxicity to fish embryo. Fish embryo acute toxicity tests are highly predictive of aquatic embryotoxicity outcome. In this study, zebrafish (Danio rerio) embryos were exposed to hymexazol at varying concentrations for the study of the developmental toxicity, melanin biosynthesis, biochemical and transcriptional endpoints. The embryotoxicity tests indicated that the 96h LC50 value of hymexazol was 649mg/L with a 95% confidence interval range of 632–667mg/L. Hymexazol at concentrations of 417–738mg/L decreased the heart rate and increased the voluntary swing. Hymexazol inhibited normal development at concentrations above 554mg/L. the 96h EC50 was 411mg/L. Hymexazol in a concentration range of 417–738mg/L induced cardiac edema and yolk sac edema. Exposure of hymexazol at such concentrations to zebrafish embryos for 48h decreased the pigment area density compared with the no hymexazol control. Tyrosinase activity was inhibited by hymexazol relative to the untreated control. The P53 mRNA expression level in embryos upon exposure to 480mg/L or greater of hymexazol was significantly higher than that of the control. The results indicated that hymexazol has quite low acute toxicity and low embryotoxicity to zebrafish

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

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

Assessment of the permeability and toxicity of polymeric nanocapsules using the zebrafish model.

To assess the capacity of a new drug delivery nanocapsule (NC) with a double shell of hyaluronic acid and protamine to overcome biological barriers using the zebrafish model. NCs were prepared by the solvent displacement method, tagged with fluorescent makers and physicochemically characterized. Toxicity was evaluated according to the Fish Embryo Acute Toxicity test, and permeability was tested by exposing zebrafish, with and without chorion, to the fluorescent NCs. Toxicity of NCs was very low as compared with that of a control nanoemulsion. Double-shell NCs were able to cross chorion and skin. Beyond the potential value of hyaluronic acid:protamine NCs for overcoming epithelial barriers, this works highlights the utility of zebrafish for fast screening of nanocarriers.

Uranium mining wastes: The use of the Fish Embryo Acute Toxicity Test (FET) test to evaluate toxicity and risk of environmental discharge

Active and abandoned uranium mining sites often create environmentally problematic situations, since they cause the contamination of all environmental matrices (air, soil and water) with stable metals and radionuclides. Due to their cytotoxic, genotoxic and teratogenic properties, the exposure to these contaminants may cause several harmful effects in living organisms. The Fish Embryo Acute Toxicity Test (FET) test was employed to evaluate the genotoxic and teratogenic potential of mine liquid effluents and sludge elutriates from a deactivated uranium mine. The aims were: a) to determine the risk of discharge of such wastes in the environment; b) the effectiveness of the chemical treatment applied to the uranium mine water, which is a standard procedure generally applied to liquid effluents from uranium mines and mills, to reduce its toxicological potential; c) the suitability of the FET test for the evaluation the toxicity of such wastes and the added value of including the evaluation of genotoxicity. Results showed that through the FET test it was possible to determine that both elutriates and effluents are genotoxic and also that the mine effluent is teratogenic at low concentrations. Additionally, liquid effluents and sludge elutriates affect other parameters namely, growth and hatching and that water pH alone played an important role in the hatching process. The inclusion of genotoxicity evaluation in the FET test was crucial to prevent the underestimation of the risks posed by some of the tested effluents/elutriates. Finally, it was possible to conclude that care should be taken when using benchmark values calculated for specific stressors to evaluate the risk posed by uranium mining wastes to freshwater ecosystems, due to their chemical complexity.

Sub-lethal pharmaceutical hazard tracking in adult zebrafish using untargeted LC–MS environmental metabolomics

Antibiotics in the aquatic environment are dispersed through anthropogenic activities at low concentrations. Despite their sub lethal concentration, these biologically active compounds may still have adverse effects to non-target species. This study examined the response of adult zebrafish to 0.1mg/L concentration of clarithromycin, florfenicol, sulfamethazine, and their mixture using environmental metabolomics. Embryo and larvae of the fish were also used to assess fish embryo acute toxicity and behavior tests respectively. The fish embryo toxicity test did not show any inhibition of growth and development of the embryos after 96h of exposure to the antibiotics. Changes in swimming activity were seen in 5-dpf larvae which is believed to be correlated with the length of exposure to the compounds. Meanwhile, environmental metabolomics revealed diverse metabolites and pathways that were affected after 72h of exposure of the adult fish to sub-lethal concentration of the compounds. We found that even at low concentration of the antibiotics, behavioral and metabolic effects were still observed despite the lack of visible morphological changes. Further studies involving other aquatic organisms and bioactive compounds are encouraged to strengthen the findings presented in this novel research.

Evaluating the zebrafish embryo toxicity test for pesticide hazard screening.

Given the numerous chemicals used in society, it is critical to develop tools for accurate and efficient evaluation of potential risks to human and ecological receptors. Fish embryo acute toxicity tests are 1 tool that has been shown to be highly predictive of standard, more resource-intensive, juvenile fish acute toxicity tests. However, there is also evidence that fish embryos are less sensitive than juvenile fish for certain types of chemicals, including neurotoxicants. The utility of fish embryos for pesticide hazard assessment was investigated by comparing published zebrafish embryo toxicity data from pesticides with median lethal concentration 50% (LC50) data for juveniles of 3 commonly tested fish species: rainbow trout, bluegill sunfish, and sheepshead minnow. A poor, albeit significant, relationship (r2 = 0.28; p < 0.05) was found between zebrafish embryo and juvenile fish toxicity when pesticides were considered as a single group, but a much better relationship (r2 = 0.64; p < 0.05) when pesticide mode of action was factored into an analysis of covariance. This discrepancy is partly explained by the large number of neurotoxic pesticides in the dataset, supporting previous findings that commonly used fish embryo toxicity test endpoints are particularly insensitive to neurotoxicants. These results indicate that it is still premature to replace juvenile fish toxicity tests with embryo-based tests such as the Organisation for Economic Co-operation and Development Fish Embryo Acute Toxicity Test for routine pesticide hazard assessment, although embryo testing could be used with other screening tools for testing prioritization. Environ Toxicol Chem 2017;36:1221-1226. © 2016 SETAC.

Development of a general baseline toxicity QSAR model for the fish embryo acute toxicity test

Fish embryos have become a popular model in ecotoxicology and toxicology. The fish embryo acute toxicity test (FET) with the zebrafish embryo was recently adopted by the OECD as technical guideline TG 236 and a large database of concentrations causing 50% lethality (LC50) is available in the literature. Quantitative Structure-Activity Relationships (QSARs) of baseline toxicity (also called narcosis) are helpful to estimate the minimum toxicity of chemicals to be tested and to identify excess toxicity in existing data sets. Here, we analyzed an existing fish embryo toxicity database and established a QSAR for fish embryo LC50 using chemicals that were independently classified to act according to the non-specific mode of action of baseline toxicity. The octanol-water partition coefficient Kow is commonly applied to discriminate between non-polar and polar narcotics. Replacing the Kow by the liposome-water partition coefficient Klipw yielded a common QSAR for polar and non-polar baseline toxicants. This developed baseline toxicity QSAR was applied to compare the final mode of action (MOA) assignment of 132 chemicals. Further, we included the analysis of internal lethal concentration (ILC50) and chemical activity (La50) as complementary approaches to evaluate the robustness of the FET baseline toxicity. The analysis of the FET dataset revealed that specifically acting and reactive chemicals converged towards the baseline toxicity QSAR with increasing hydrophobicity. The developed FET baseline toxicity QSAR can be used to identify specifically acting or reactive compounds by determination of the toxic ratio and in combination with appropriate endpoints to infer the MOA for chemicals.

A new supported TiO2 film deposited on stainless steel for the photocatalytic degradation of contaminants of emerging concern

A new supported catalyst composed of a nanostructured TiO2 film deposited on a stainless steel mesh (nanoTiO2-SS) using the Metal Organic Chemical Vapour Deposition (MOCVD) technique was evaluated for the photocatalytic degradation of a mixture of contaminants of emerging concern. Results showed that under the oxidative conditions tested, the nanoTiO2-SS catalyst demonstrated an efficiency in degrading the target contaminants higher than that observed under direct photolysis and photocatalysis using the conventional TiO2 Degussa P25 catalyst at the same amount of TiO2 participating to the photocatalysis. Specifically, the rate of removal of warfarin and trimethoprim obtained with the new catalyst was found twice the one observed by using TiO2 Degussa P25 and approximately 1.6 times faster for metoprolol, carbamazepine and gemfibrozil. An evaluation of the electrical energy per order magnitude of removal (EE/O) confirmed the enhanced performance of the new catalyst (24.3–31.8kWhm−3 rather than 32.8–39.3kWhm−3 for conventional TiO2) and that the performance is compound-dependent. Toxicity testing revealed that some assays are suitable for the investigation of bioactivity of treated waters containing contaminants of emerging concern at μgL−1 level. Specifically, the AMES Fluctuation Test, Fish Embryo Acute Toxicity Test and Green alga Selenastrum capricornutum test provided valuable results for an environmental impact assessment. On the other hand, the Daphnia magna and Vibrio fischeri acute toxicity tests were not sensitive enough to detect bioactivity in the samples analysed without prior pre-concentration.

The effect of tramadol hydrochloride on early life stages of fish.

The aim of this study was to perform the fish embryo acute toxicity test (FET) on zebrafish (Danio rerio) and the early-life stage toxicity test on common carp (Cyprinus carpio) with tramadol hydrochloride. The FET was performed using the method inspired by the OECD guideline 236. Newly fertilized zebrafish eggs were exposed to tramadol hydrochloride at concentrations of 10; 50; 100 and 200μg/l for a period of 144h. An embryo-larval toxicity test on C. carpio was performed according to OECD guideline 210 also with tramadol hydrochloride at concentrations 10; 50; 100 and 200μg/l for a period of 32 days. Hatching was significantly influenced in both acute and subchronic toxicity assays. Subchronic exposure also influenced early ontogeny, both morphometric and condition characteristics and caused changes in antioxidant enzyme activity. The LOEC value was found to be 10μg/l tramadol hydrochloride.

A high throughput passive dosing format for the Fish Embryo Acute Toxicity test

High throughput testing according to the Fish Embryo Acute Toxicity (FET) test (OECD Testing Guideline 236) is usually conducted in well plates. In the case of hydrophobic test substances, sorptive and evaporative losses often result in declining and poorly controlled exposure conditions. Therefore, our objective was to improve exposure conditions in FET tests by evaluating a passive dosing format using silicone O-rings in standard 24-well polystyrene plates. We exposed zebrafish embryos to a series of phenanthrene concentrations until 120h post fertilization (hpf), and obtained a linear dilution series. We report effect values for both mortality and sublethal morphological effects based on (1) measured exposure concentrations, (2) (lipid normalized) body residues and (3) chemical activity. The LC50 for 120hpf was 310μg/L, CBR50 (critical body residue) was 2.72mmol/kg fresh wt and La50 (lethal chemical activity) was 0.047. All values were within ranges expected for baseline toxicity. Impaired swim bladder inflation was the most pronounced morphological effect and swimming activity was reduced in all exposure concentrations. Further analysis showed that the effect on swimming activity was not attributed to impaired swim bladder inflation, but rather to baseline toxicity. We conclude that silicone O-rings (1) produce a linear dilution series of phenanthrene in the 120hpf FET test, (2) generate and maintain aqueous concentrations for reliable determination of effect concentrations, and allow for obtaining mechanistic toxicity information, and (3) cause no toxicity, demonstrating its potential as an extension of the FET test when testing hydrophobic chemicals.