Aquatic testing guidelines insufficiently control the influence of dilution water toc and hardness on cationic polymer toxicity - A proposal to improve standardized test procedures.

Cationic polymers (CPs) are widely used chemicals for wastewater treatment applications and in various "down-the-drain" household products. The aquatic toxicity of CPs results from an electrostatic interaction with negatively charged cell surfaces. These effects are greatly mitigated by the binding affinity of CPs to total organic carbon (TOC) in surface water. Consequently, baseline aquatic toxicity tests of CPs using clean lab water (TOC < 2 mg/L) typically overestimate toxicity and risk which is greatly mitigated at higher environmentally relevant OC levels. However, the point at which mitigation begins is not well defined and low-level TOC in lab water may influence the baseline toxicity outcome. Similarly, divalent cations, quantified as water hardness, may modulate the electrostatic binding between OC and CP. Although standard guidelines define limits for lab water hardness and TOC, the consequences of variability within those limits on test outcome is unknown. We investigated the impact of part-per-billion (ppb) additions of TOC to lab water at different hardness levels on CP acute toxicity to Daphnia magna and Raphidocelis subcapitata. In both species, the acute toxicities of CPs with different molecular weight and charge density varied by > 10-fold in response to slight changes in TOC and water hardness, although parameters were maintained within guideline limits. When determining the baseline aquatic toxicity of CPs, the lab water should be standardized at the lowest biologically tolerable hardness and TOC at a reliably measurable level (>1 - < 2 mg/L) to reduce variability and increase the reliability of the toxicity estimate.

Evaluation of the bioaccumulation of octocrylene after dietary and aqueous exposure.

Octocrylene is used as UV filter in personal care products with a high production volume and can be detected in surface water and biota. It is liquid at ambient temperature, highly lipophilic, has a high adsorption capacity to organic material and is considered as persistent in the environment. The very low water solubility complicates the evaluation of potential long-term effects in aquatic toxicity testing, since effect thresholds are often above the water solubility limit. Thus, the evaluation of the bioaccumulation potential becomes highly relevant for the assessment of long-term environmental effects. However, even the determination of the water solubility limit for a substance with such difficult properties is challenging. The following experiments are described, and results compared to available environmental monitoring data: A bioconcentration study with aqueous exposure (BCF) in zebrafish and a biomagnification study with dietary exposure (BMF) in rainbow trout, as well as supporting experiments to evaluate the water solubility. The growth and lipid corrected BCF determined by aqueous exposure was 858 L kg-1 while the corrected BMF was 0.0335. The model-based estimation of the BCF from BMF (152-1182 L kg-1) is in good agreement with the measured BCF value. Environmental monitoring data provide only limited information on the bioaccumulation potential of octocrylene, as only few investigations were made in biota and water in parallel and concentrations of octocrylene vary by several orders of magnitude during seasons. Based on the determined fish BCF data, we conclude that OCR is not bioaccumulative according to the criteria as laid down by ECHA, 2017. Furthermore, the low BMF value indicates no accumulation along the food chain.

Multi-criteria decision analysis of test endpoints for detecting the effects of endocrine active substances in fish full life cycle tests.

Fish full life cycle (FFLC) tests are increasingly required in the ecotoxicological assessment of endocrine active substances. However, FFLC tests have not been internationally standardized or validated, and it is currently unclear how such tests should best be designed to provide statistically sound and ecologically relevant results. This study describes how the technique of multi-criteria decision analysis (MCDA) was used to elicit the views of fish ecologists, aquatic ecotoxicologists and statisticians on optimal experimental designs for assessing the effects of endocrine active chemicals on fish. In MCDA qualitative criteria (that can be valued, but not quantified) and quantitative criteria can be used in a structured decision-making process. The aim of the present application of MCDA is to present a logical means of collating both data and expert opinions on the best way to focus FFLC tests on endocrine active substances. The analyses are presented to demonstrate how MCDA can be used in this context. Each of 3 workgroups focused on 1 of 3 species: fathead minnow (Pimephales promelas), Japanese medaka (Oryzias latipes), and zebrafish (Danio rerio). Test endpoints (e.g., fecundity, growth, gonadal histopathology) were scored for each species for various desirable features such as statistical power and ecological relevance, with the importance of these features determined by assigning weights to them, using a swing weighting procedure. The endpoint F1 fertilization success consistently emerged as a preferred option for all species. In addition, some endpoints scored highly in particular species, such as development of secondary sexual characteristics (fathead minnow) and sex ratio (zebrafish). Other endpoints such as hatching success ranked relatively highly and should be considered as useful endpoints to measure in tests with any of the fish species. MCDA also indicated relatively less preferred endpoints in fish life cycle tests. For example, intensive histopathology consistently ranked low, as did measurement of diagnostic biomarkers, such as vitellogenin, most likely due to the high costs of these methods or their limited ecological relevance. Life cycle tests typically do not focus on identifying toxic modes and/or mechanisms of action, but rather, single chemical concentration-response relationships for endpoints (e.g., survival, growth, reproduction) that can be translated into evaluation of risk. It is, therefore, likely to be an inefficient use of limited resources to measure these mechanism-specific endpoints in life cycle tests, unless the value of such endpoints for answering particular questions justifies their integration in specific case studies.

Environmental effect assessment for sexual endocrine-disrupting chemicals: Fish testing strategy.

Current standard testing and assessment tools are not designed to identify specific and biologically highly sensitive modes of action of chemicals, such as endocrine disruption. This information, however, can be important to define the relevant endpoints for an assessment and to characterize thresholds of their sublethal, population-relevant effects. Starting a decade ago, compound-specific risk assessment procedures were amended by specifically addressing endocrine-disrupting properties of substances. In 2002, the Conceptual Framework, agreed upon by OECD's Task Force on Endocrine Disrupters Testing and Assessment, did not propose specific testing strategies, and appropriate testing methods had not yet been developed and approved. In the meantime, the OECD Test Guidelines Programme has undertaken important steps to revise established and to develop new test methods, which can be used to identify and quantify effects of endocrine-disrupting chemicals on mammals, birds, amphibians, fish, and invertebrates. For fish testing of endocrine-disrupting chemicals, the first Test Guidelines have recently been adopted by the OECD and validation of further test systems is under progress. Based on these test systems and the experience gained during their validation procedures, we propose a 3-step fish testing strategy: 1) Weight-of-evidence approach for identifying potential sexual endocrine-disrupting chemicals; even after advanced specification of systematic criteria, this step of establishing initial suspicion will still require expert judgment; 2) in vivo evaluation of sexual endocrine-disrupting activity in fish by applying in vivo fish screening assays; sufficient data are available to diagnose the aromatase-inhibition and estrogen-receptor agonist mechanisms of action by indicative endpoints (biomarkers), whereas the ability of the respective biomarkers in the screening assay to identify the estrogen-receptor antagonists and androgen-receptor agonists and antagonists requires further validation; 3) characterization of sexual endocrine-mediated adverse effects including threshold concentrations; in cases when the most sensitive population-relevant endpoints and the most sensitive time window for exposure are known for the mechanisms of action, the fish full life-cycle or 2-generation test, which are the normal definitive tests, might be abbreviated to, e.g., the fish sexual development test. In the European Union, the measurement of indicative endpoints in the definitive test might be crucial for the authorization procedure under REACH and plant-protection products. The results of the definitive tests can be used in existing schemes of compound-specific environmental risk assessments.

Acute and chronic effects of nano- and non-nano-scale TiO(2) and ZnO particles on mobility and reproduction of the freshwater invertebrate Daphnia magna.

Among the emerging literature addressing the biological effects of nanoparticles, very little information exists, particularly on aquatic organisms, that evaluates nanoparticles in comparison to non-nanocounterparts. Therefore, the potential effects of nano-scale and non-nano-scale TiO(2) and ZnO on the water flea, Daphnia magna, were examined in 48-h acute toxicity tests using three different test media, several pigment formulations--including coated nanoparticles--and a variety of preparation steps. In addition, a 21-d chronic Daphnia reproduction study was performed using coated TiO(2) nanoparticles. Analytical ultracentrifugation analyses provided evidence that the nanoparticles were present in a wide range of differently sized aggregates in the tested dispersions. While no pronounced effects on D. magna were observed for nano-scale and non-nano-scale TiO(2) pigments in 19 of 25 acute (48-h) toxicity tests (EC50>100 mg L(-1)), six acute tests with both nano- and non-nano-scale TiO(2) pigments showed slight effects (EC10, 0.5-91.2 mg L(-1)). For the nano-scale and non-nano-scale ZnO pigments, the acute 48-h EC50 values were close to the 1 mg L(-1) level, which is within the reported range of zinc toxicity to Daphnia. In general, the toxicity in the acute tests was independent of particle size (non-nano-scale or nano-scale), coating of particles, aggregation of particles, the type of medium or the applied pre-treatment of the test dispersions. The chronic Daphnia test with coated TiO(2) nanoparticles demonstrated that reproduction was a more sensitive endpoint than adult mortality. After 21d, the NOEC for adult mortality was 30 mg L(-1) and the NOEC for offspring production was 3 mg L(-1). The 21-d EC10 and EC50 values for reproductive effects were 5 and 26.6 mg L(-1), respectively. This study demonstrates the utility of evaluating nanoparticle effects relative to non-nano-scale counterparts and presents the first report of chronic exposure to TiO(2) nanoparticles in D. magna.

Aerobic biodegradation of polyethylene glycols of different molecular weights in wastewater and seawater.

In order to distinguish between aerobic biodegradation of synthetic polymers in fresh and seawater, polyethylene glycols (PEGs) were systematically and comparatively investigated in inocula from municipal wastewater and seawater aquarium filters for the first time. The molecular weight (MW) of the PEGs, (HO(CH(2)CH(2)O)(n)H, n=3-1350) as representatives of water-soluble polymers, ranged from 250 to 57,800Da. The biodegradation was observed by removal of dissolved organic carbon and carbon dioxide production by applying standardized ISO and OECD test methods. Specific analyses using liquid chromatography mass spectrometry (LC-MS) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) were performed. All PEGs selected were completely biodegradable in freshwater media within 65d. PEGs with an MW up to 14,600Da have a similar degradation pathway which is characterized by gradual splitting of C(2)-units off the chain resulting in formation of short-chain PEGs. In artificial seawater media, full biodegradation of PEGs up to 7400Da required more time than in freshwater. PEGs with MW 10,300 and 14,600Da were only partially degraded whereas PEGs with MW 26,600 and 57,800Da were not degraded for a period of 135d. The biodegradation pathway of PEG 250 and PEG 970 in seawater is similar to that for freshwater. For PEGs having an MW from 2000 to 10,300Da, the degradation pathway in seawater differs from the pathway of the shorter PEGs.