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Bisphenols and phthalates, chemicals frequently used in plastic products, promote obesity in cell and animal models. However, these well-known metabolism disrupting chemicals (MDCs) represent only a minute fraction of all compounds found in plastics. To gain a comprehensive understanding of plastics as a source of exposure to MDCs, we characterized all chemicals present in 34 everyday products using nontarget high-resolution mass spectrometry and analyzed their joint adipogenic activities by high-content imaging. We detected 55,300 chemical features and tentatively identified 629 unique compounds, including 11 known MDCs. Importantly, chemicals that induced proliferation, growth, and triglyceride accumulation in 3T3-L1 adipocytes were found in one third of the products. Since the majority did not target peroxisome proliferator-activated receptor γ, the effects are likely to be caused by unknown MDCs. Our study demonstrates that daily-use plastics contain potent mixtures of MDCs and can, therefore, be a relevant yet underestimated environmental factor contributing to obesity.
Teaser Plastics contain a potent mixture of chemicals promoting adipogenesis, a key process in developing obesity.
Bisphenols and phthalates, chemicals frequently used in plastic products, promote obesity in cell and animal models. However, these well-known metabolism-disrupting chemicals (MDCs) represent only a minute fraction of all compounds found in plastics. To gain a comprehensive understanding of plastics as a source of exposure to MDCs, we characterized the chemicals present in 34 everyday products using nontarget high-resolution mass spectrometry and analyzed their joint adipogenic activities by high-content imaging. We detected 55,300 chemical features and tentatively identified 629 unique compounds, including 11 known MDCs. Importantly, the chemicals extracted from one-third of the products caused murine 3T3-L1 preadipocytes to proliferate, and differentiate into adipocytes, which were larger and contained more triglycerides than those treated with the reference compound rosiglitazone. Because the majority of plastic extracts did not activate the peroxisome proliferator-activated receptor γ and the glucocorticoid receptor, the adipogenic effects are mediated via other mechanisms and, thus, likely to be caused by unknown MDCs. Our study demonstrates that daily-use plastics contain potent mixtures of MDCs and can, therefore, be a relevant yet underestimated environmental factor contributing to obesity.
Surface water can contain a complex mixture of organic micropollutants (i.e. residues of pharmaceuticals or biocides). Conventional wastewater treatment plants (WWTPs) do not completely remove a broad range of anthropogenic chemicals and therefore represent a leading point source. To upgrade WWTPs, technical solutions based on oxidative and sorptive processes have been developed and successfully implemented. Acknowledging these substantial advances, this thesis focuses on another key topic and aims to investigate whether improved biological treatment processes likewise effectively remove anthropogenic micropollutants from wastewater. The work conducted on this topic was part of two European research projects (ATHENE, ENDETECH).
The ATHENE project aimed to go beyond the state-of-the-art by developing biological wastewater treatment processes that exploit the full potential of biodegradation. With the objective to explore the potential of complementary strictly anaerobic conditions within the biological wastewater treatment, combinations of aerobic and anaerobic treatments on site of a WWTP were implemented. Based on pre-experiments, two promising treatment combinations were selected for a more comprehensive evaluation. An aerobic treatment was paired with an anaerobic pre-treatment under iron-reducing conditions, and an activated sludge treatment was combined with an anaerobic post-treatment under substrate-limiting conditions. For the evaluation of these processes, an effect-based assessment was applied and combined with chemical data of 31 selected target organic micropollutants as well as ten metabolites. To assess the removal of endocrine disrupting chemicals (EDCs), yeast based reporter gene assays covering seven receptor-mediated mechanisms of action including (anti-)estrogenicity, (anti-) androgenicity, retinoid-like, and dioxin-like activity were conducted. Furthermore, the removal of unspecific toxicity (Microtox assay) and oxidative stress response as a marker for reactive toxicity (AREc32 assay) were analyzed to cover micropollutants acting via a non-specific mechanism of action. Moreover, to assess toxicity of the whole effluent in vivo, standardized in vivo bioassays with four aquatic model species (Desmodesmus subspicatus, Daphnia magna, Lumbriculus variegatus, Potamopyrgus antipodarum) were performed.
The combination of aerobic and anaerobic treatments resulted in a low additional removal of the selected target organic micropollutants (by 14-17%). In contrast, the removal of endocrine and dioxin-like activities (by 17-75%) and non-specific in vitro toxicities (by 27-60%) was significantly enhanced. Compared to technical solutions (i.e. ozonation), the combination with an anaerobic pre-treatment under iron-reducing conditions was likewise effective in removing the estrogenic activity as well as the unspecific toxicity, whereas anti-androgenic activity and dioxin-like activity were less effectively removed. Exposure to effluents of the conventional activated sludge treatment did not induce adverse in vivo effects in the investigated aquatic model species. Accordingly, no further improvement in water quality could be observed. In conclusion, the combination of aerobic and anaerobic treatment processes significantly enhanced the removal of specific and non-specific in vitro toxicities. Thus, an optimization of the biological wastewater treatment can lead to a substantially improved detoxification. These capacities of a treatment technology can only be uncovered by complementary effect-based measurements.
The global objective of the ENDETECH project was to develop a biotechnological solution to eliminate recalcitrant pharmaceuticals in wastewater direct from sites, where high loads are expected (i.e. hospitals). For this purpose, laccase, an enzyme mainly found in wood decaying fungi, was immobilized on ceramic membranes for application in bioreactors. In a proof of principle experiment, the performance of immobilized laccase in removing a mixture of 38 antibiotics without and in combination with a natural mediator (syringaldehyde; SYR) was investigated. For the evaluation of the enzymatic membrane bioreactors, chemical data on the elimination of the selected target antibiotics was combined with the outcomes of two in vitro bioassays. Growth inhibition tests with an antibiotic sensitive Bacillus subtilis strain were conducted to assess the residual antibiotic activity of the effluents, and Microtox assays were performed to detect a potential formation of toxic by-products.
The treatment by laccase without SYR did not reduce the load of antibiotics significantly. In contrast, in combination with a SYR concentration of 10 µmol L-1, 26 out of 38 antibiotics were removed by >50% after 24 h treatment. Moreover, increasing the SYR concentration to 1000 µmol L-1 resulted in a further improvement of the antibiotic removal. 32 out of 38 antibiotics were removed by over 50%, whereby 17 were almost completely eliminated (>90%). However, the treatment with laccase in combination with SYR resulted in a time-dependent increase of unspecific toxicity. While SYR alone did not affect B. subtilis, the combination of laccase with SYR led to a strong time-dependent growth inhibition up to 100%. Similar to that, a time-dependent increase of unspecific toxicity in the Microtox assay was observed. In conclusion, the laccase-mediator process successfully degrades a broad spectrum of antibiotics and thus represents a promising technology to treat wastewater from sites, where high loads are expected. However, further research is required to reduce the formation of unspecific toxicity before an implementation of this technology can be considered.
Microplastics (MP) are contaminants of emerging concern in aquatic ecosystems. While the number of studies is rapidly increasing, a comparison of the toxicity of MP and natural particulate matter is largely missing. In addition, research focusses on the impacts of hydrophobic chemicals sorbed to plastics. However, the interactive effects of MP and hydrophilic, dissolved chemicals remain largely unknown. Therefore, we conducted chronic toxicity studies with larvae of the freshwater dipteran Chironomus riparius exposed to unplasticised polyvinyl chloride MP (PVC-MP) as well as kaolin and diatomite as reference materials for 28 days. In addition, we investigated the effects of particles in combination with the neonicotinoid imidacloprid in a multiple-stressor experiment. High concentrations of kaolin positively affected the chironomids. In contrast, exposure to diatomite and PVC-MP reduced the emergence and mass of C. riparius. Likewise, the toxicity of imidacloprid was enhanced in the presence of PVC-MP and slightly decreased in the co-exposure with kaolin. Overall, parallel experiments and chemical analysis indicate that the toxicity of PVC-MP was not caused by leached or sorbed chemicals. Our study demonstrates that PVC-MP induce more severe effects than both natural particulate materials. However, the latter are not benign per se, as the case of diatomite highlights. Considering the high, environmentally irrelevant concentrations needed to induce adverse effects, C. riparius is insensitive to exposures to PVC-MP.
Ecological speciation assumes reproductive isolation to be the product of ecologically based divergent selection. Beside natural selection, sexual selection via phenotype-assortative mating is thought to promote reproductive isolation. Using the neotropical fish Poecilia mexicana from a system that has been described to undergo incipient ecological speciation in adjacent, but ecologically divergent habitats characterized by the presence or absence of toxic H2S and darkness in cave habitats, we demonstrate a gradual change in male body colouration along the gradient of light/darkness, including a reduction of ornaments that are under both inter- and intrasexual selection in surface populations. In dichotomous choice tests using video-animated stimuli, we found surface females to prefer males from their own population over the cave phenotype. However, female cave fish, observed on site via infrared techniques, preferred to associate with surface males rather than size-matched cave males, likely reflecting the female preference for better-nourished (in this case: surface) males. Hence, divergent selection on body colouration indeed translates into phenotype-assortative mating in the surface ecotype, by selecting against potential migrant males. Female cave fish, by contrast, do not have a preference for the resident male phenotype, identifying natural selection against migrants imposed by the cave environment as the major driver of the observed reproductive isolation.