In vitro assays are broadly used tools to evaluate the estrogenic activity in Waste Water Treatment Plant (WWTP) effluents and their receiving rivers. Since potencies of individual estrogens to induce in vitro and in vivo responses can differ it is not possible to directly evaluate risks based on in vitro measures of estrogenic activity. Estrone, 17beta-estradiol, 17alfa-ethinylestradiol and to some extent, estriol have been shown to be responsible for the majority of in vitro estrogenic activity of municipal WWTP effluents. Therefore, in the present study safe concentrations of Estrogenic Equivalents (EEQs-SSE) in municipal WWTP effluents were derived based on simplified assumption that the steroid estrogens are responsible for all estrogenicity determined with particular in vitro assays. EEQs-SSEs were derived using the bioassay and testing protocol-specific in vitro potencies of steroid estrogens, in vivo predicted no effect concentration (PNECs) of these compounds, and their relative contributions to the overall estrogenicity detected in municipal WWTP effluents. EEQs-SSEs for 15 individual bioassays varied from 0.1 to 0.4ng EEQ/L. The EEQs-SSEs are supposed to be increased by use of location-specific dilution factors of WWTP effluents entering receiving rivers. They are applicable to municipal wastewater and rivers close to their discharges, but not to industrial waste waters.

N-heterocyclic derivatives of polycyclic aromatic hydrocarbons (NPAHs) are widespread concomitantly with their parent analogues and have been detected in air, water, sediments, and soil. Although they were shown to be highly toxic to some organisms, our understanding of their occurrence, environmental fate, biological metabolism, and effects is limited. This study evaluated toxic effects of three homocyclic aromatic hydrocarbons (PAHs—phenanthrene, anthracene, fluorene) and their seven N-heterocyclic derivates on higher terrestrial plants Sinapis alba, Triticum aestivum, and Phaseolus vulgaris. Germinability, morphological end-points, parameters of detoxification, and antioxidant components of plant metabolism as well as lipid peroxidation were studied in acute phytotoxicity tests. Phytotoxicity of NPAHs was generally more pronounced than the effects of parent PAHs, and it significantly differed with respect to the structure of individual NPAHs. Sinapis alba and T. aestivum were more sensitive plant species than P. vulgaris. Chemicals with the strongest inhibition effect on germination and growth of plants were phenanthridine, acridine, benzo[h]quinoline, and 1,10- and 1,7-phenanthroline. All tested chemicals significantly induced activities of detoxification and antioxidant enzymes (glutathione reductase, glutathione peroxidase, and glutathione-S-transferase) at nanomolar to low micromolar concentrations. Levels of reduced glutathione were induced by all tested chemicals except 1,10- and 4,7-phenanthroline. Furthermore, fluorene, carbazole, acridine, phenanthrene, phenanthridine, benzo[h]quinoline, and 1,7-phenanthroline significantly increased lipid peroxidation. The results of our study newly demonstrate significant toxicity of NPAHs to plants and demonstrate suitability of multiple biomarker assessment to characterize mechanisms of oxidative stress and to serve as an early warning of phytotoxicity in vivo.

Air pollution, which represents a major environmental risk to human health, comprises a complex mixture of compounds where only little is known about its specific toxicities. This study examined the specific toxicities associated with ambient air pollutant mixtures with respect to gas/particle partitioning, particulate matter (PM) size, pollutant polarity and bioaccessibility from PM, and evaluated the contribution of PAHs and their oxygenated and nitrated derivatives (OPAHs, NPAHs). Air samples (gas phase, PM₁₀ and size-segregated PM), were collected at urban (in winter and summer) and background (winter) sites in the Czech Republic. The total and bioaccessible concentrations were addressed using organic solvent extraction and simulated lung fluid extraction, respectively. Organic extracts were also further fractionated according to polarity. Aryl hydrocarbon receptor (AhR)-mediated activity, anti-/estrogenicity, anti-/androgenicity, thyroid receptor (TR)-mediated activity and cytotoxicity for bronchial cells were determined by human cell-based in vitro bioassays. The contribution of studied compounds to observed effects was assessed by both modelling and reconstructing the mixtures. Significant effects were detected in the sub-micrometre size fraction of PM (estrogenicity, androgenicity, TR- and AhR-mediated activities) and in the gas phase (TR-mediated activity, antiandrogenicity). Compounds interacting with TR showed high bioaccessibility to simulated lung fluid. Relatively lower bioaccessibility was observed for estrogenicity and AhR-mediated activity. However, the toxicity testing of reconstructed mixtures revealed that the targeted pollutants are not the main contributors, except for urban PM air pollution in winter, where they accounted for 5-88% of several effects detected in the original complex environmental samples. Studied toxicities were mostly driven by polar compounds largely attributed to the easily inhalable PM₁, which is of high relevance for human health risk assessment. Except of parent PAHs in some cases, the targeted compounds contributed to the detected effects mostly to a relatively low extent implying huge data gaps in terms of endocrine disruptive potencies of targeted substances and the significance of other polar compounds present in ambient air.