Chemical and toxicological assessment of leachates from UV-degraded plastic materials using in-vitro bioassays.

The broad use of plastics and the persistence of the material results in plastic residues being found practically everywhere in the environment. If plastics remain in the (aquatic) environment, natural weathering leads to degradation processes and compounds may leach from plastic into the environment. To investigate the impact of degradation process on toxicity of leachates, different types of UV irradiation (UV-C, UV-A/B) were used to simulate weathering processes of different plastic material containing virgin as well as recyclate material and biodegradable polymers. The leached substances were investigated toxicologically using in-vitro bioassays. Cytotoxicity was determined by the MTT-assay, genotoxicity by using the p53-CALUX and Umu-assay, and estrogenic effects by the ERα-CALUX. Genotoxic as well as estrogenic effects were detected in different samples depending on the material and the irradiation type. In four leachates of 12 plastic species estrogenic effects were detected above the recommended safety level of 0.4 ng 17ß-estradiol equivalents/L for surface water samples. In the p53-CALUX and in the Umu-assay leachates from three and two, respectively, of 12 plastic species were found to be genotoxic. The results of the chemical analysis show that plastic material releases a variety of known and unknown substances especially under UV radiation, leading to a complex mixture with potentially harmful effects. In order to investigate these aspects further and to be able to give recommendations for the use of additives in plastics, further effect-related investigations are advisable.

Effect-based evaluation of ozone treatment for removal of micropollutants and their transformation products in waste water.

The aim of this interdisciplinary research project in North Rhine-Westphalia (NRW), Germany, entitled "Elimination of pharmaceuticals and organic micropollutants from waste water" involved the conception of cost-effective and innovative waste-water cleaning methods. In this project in vitro assays, in vivo assays and chemical analyses were performed on three municipal waste-water treatment plants (WWTP). This publication focuses on the study of the in vitro bioassays. Cytotoxic, estrogenic, genotoxic and mutagenic effects of the original as well as enriched water samples were monitored before and after wastewater treatment steps using MTT and PAN I, ER Calux and A-YES, micronucleus and Comet assays as well as AMES test. In most cases, the measured effects were reduced after ozonation, but in general, the biological response depended upon the water composition of the WWTP, in particular on the formed by-products and concentration of micropollutants. In order to be able to assess the genotoxic and/or mutagenic potential of waste-water samples using bioassays like Ames test, Comet assay or micronucleus test an enrichment of the water sample via solid-phase extraction is recommended. This is in agreement with previous studies such as the "ToxBox"-Project of the Environmental Agency in Germany.

Evaluation of mixture effects of endocrine active substances in wastewater using CALUX reporter-gene assays.

Endocrine active substances (EAS), which are commonly used in pharmaceuticals and personal care products, are released into surface water mainly through WWTP effluents and have been shown to cause adverse effects in aquatic organisms. In wastewater, a variety of EAS with different hormonal activities is present, which can lead to additive effects or mask an endocrine activity. To investigate hormonal combination effects, with a focus on estrogen and androgen-modulators, influent samples from municipal and hospital wastewater treatmenr plants were spiked with 17α-ethinylestradiol, toremifene, 17α-methyltestosterone and bicalutamide and analyzed using in vitro reporter gene CALUX assays. All wastewaters caused endocrine activities in human cells, which were modified by adding one or several endocrine active substances. As expected, estrogenic activity was reduced in presence of the anti-estrogenic toremifene and androgenic activity decreased with the anti-androgen bicalutamide. In general, substance addition caused a similar trend in altered endocrine activities; however, their intensities differed between the wastewaters. Our results indicate that masking effects, leading to a suppressed biological signal, are of significant importance in the assessment of complex water samples, and combination effects rather than single substances determine the final biological effect. This emphasizes the need of effect-based tools in the assessment of water samples.

Effect-based methods in combination with state-of-the-art chemical analysis for assessment of water quality as integrated approach.

Complex mixtures of chemicals in waste and finally in surface water may pose a risk to the environment and also to human health. This contamination of surface water cannot be addressed with chemical analysis alone. Tools are required to detect and assess these micropollutants which might cause adverse effects. Effect-directed analysis (EDA) with effect-based methods in combination with state-of-the-art chemical analysis can meet this challenge. The present paper summarizes and outlines current experiences with analytical tools and bioassays as integrated approach for assessment of water quality. The need for a holistic and solution-oriented procedure of water quality monitoring is described. To integrate and evaluate existing information about toxicity pathways, which are essential for the EDA approach, the adverse outcome pathway (AOP) concept is useful and recommended. An integration of AOP concept in water quality assessment and further requirements are discussed.