Passive sampling in effect-based monitoring of two European rivers - explicability of in vitro toxic potentials by detected chemicals

EU commission Water Framework Directive considers employment of passive sampling and use of effect-based tools in the monitoring of aquatic pollution. A combination of both approaches was used for monitoring of two rivers differing significantly in pollution levels. The Bosna, moderate-sized river in Bosnia-Herzegovina, which is burdened by untreated wastewaters, was sampled by semipermeable passive sampling devices (SPMD) and POCIS samplers. The Danube, the largest river in the EU with relatively low pollution level, was sampled using a mobile dynamic passive sampling device with silicone rubber (SR) and SDB-RPS Empore™ (ED) disc samplers. Both sampler sets consisted of partitioning sampler for non-polar chemicals (SPMD, SR) and adsorption sampler for the polar-ones (POCIS, ED). For the partitioning samplers, concentrations of collected chemicals in river water were derived using dissipation of performance reference compounds. For the adsorption samplers, the sampling rates were either taken from literature (POCIS) or calculated from correlated levels of chemicals that were detected both in adsorption (ED) and partitioning samplers (SR). The samples were analyzed for aryl hydrocarbon-, estrogen- and androgen receptor-mediated effects using in vitro bioassays. The effects were expressed as bioanalytical equivalents (BEQbio) of respective model compounds in water. The BEQbio levels were significantly higher in extracts from POCIS and ED samplers showing that the polar chemicals were responsible for most of the detected effects. Chemical analyses detected 103 and 209 chemicals in the Bosna and the Danube samples, respectively. The passive sampling allowed detection of chemicals at pg/L concentrations. The levels of chemicals with known biological potency for the studied endpoints were used for modeling of bioanalytical equivalents (BEQchem). The comparison of bioassay- and chemical analysis-derived equivalents showed that the detected chemicals explained mostly a low fraction of the BEQbio. Only in the case of estrogenicity in extracts of the samplers collecting polar chemicals, the BEQchem was comparable with the BEQbio levels. Both sampler combinations proved to be suitable for the detection of a large set of chemicals even at trace levels and for the complementary assessment of the biological potentials of the environmental mixtures. The SOLUTIONS Project was supported by the 7th Framework Programme EU (FP7-ENV-2013) with grant agreement no. 603437

Pull-down assay coupled to non-target mass spectrometry analysis as a tool to identify unknown endocrine disruptive transthyretin ligands in waste and surface water

Surface and treated wastewaters are contaminated with highly complex mixtures of micropollutants, which may cause adverse effects on aquatic biota or humans, often mediated by endocrine disruption. However, there is very limited knowledge regarding some important modes of action, such as interference with thyroid hormone (TH) regulation and the compounds driving these effects. The effects of environmental samples observed in bioassays addressing various endpoints in the endocrine, namely thyroid hormone pathways, remain largely unexplained with known active chemicals. Transthyretin (TTR) is a serum transport protein distributing thyroid hormones to target tissues in vertebrates; its binding inhibition by xenobiotics may lead to adverse effects such as impaired (neuro)development. In this study, we describe a novel approach for the identification of compounds with the potential to bind to TTR, based on the specific separation of these compounds in a pull-down assay followed by non-target analysis (NTA). The pull-down assay using purified TTR protein was established and optimized with known TTR ligands. The method was applied to separate and identify compounds responsible for TH displacing activity in highly complex wastewater and surface water samples. The samples after the pull-down assay elicited TH displacing activity and the specific separation of TTR ligands provided a substantial reduction of chromatographic features from the original complex water extract. The applied non-target screening workflow resulted in the identification of 34 structures. Thirteen identified compounds with available analytical standards were quantified in the original water extracts and their TH-displacement potency was confirmed. Twelve compounds were discovered as TTR binders for the first time and linear alkylbenzene sulfonates (LAS) were highlighted as contaminants of concern regarding the TH-displacement activity. Pull-down assay combined with NTA proved to be a well-functioning approach for the identification of bioactive compounds in complex environmental mixtures with great application potential across various biological target endpoints and environmental compartments

Identification of algal growth inhibitors in treated waste water using effect-directed analysis based on non-target screening techniques

Growth inhibition of freshwater microalga Pseudokirchneriella subcapitata caused by a waste water treatment plant (WWTP) effluent extract was investigated using an effect directed analysis (EDA) approach. The objective was to identify compounds responsible for the toxicity by combining state-of-the-art sampling, bioanalytical, fractionation and non-target screening techniques. Three fractionation steps of the whole extract were performed and bioactive fractions were analysed with GC (xGC)-MS and LC-HRMS. In total, 383 compounds were tentatively identified, and their toxicity was characterized using US EPA Ecotox database, open scientific literature or modelled by ECOSAR. Among the top-ranking drivers of toxicity were pesticides and their transformation products, pharmaceuticals (barbiturate derivatives and macrolide antibiotics e.g. azithromycin), industrial compounds or caffeine and its metabolites. Several of the top-ranking pesticides are no longer registered for use in plant protection products or biocides in the Czech Republic (e.g. prometryn, atrazine, acetochlor, resmethrin) and some are approved only for use in biocides (e.g. terbutryn, carbendazim, phenothrin), which indicates that their non-agricultural input into aquatic environment via WWTPs should be carefully considered. The study demonstrated a functional strategy of combining biotesting, fractionation and non-target screening techniques in the EDA study focused on the identification of algal growth inhibitors in WWTP effluent

Assessment of a novel device for onsite integrative large-volume solid phase extraction of water samples to enable a comprehensive chemical and effect-based analysis

The implementation of targeted and nontargeted chemical screening analysis in combination with in vitro and organism-level bioassays is a prerequisite for a more holistic monitoring ofwater quality in the future. For chemical analysis, little or no sample enrichment is often sufficient, while bioanalysis often requires larger sample volumes at a certain enrichment factor for conducting comprehensive bioassays on different endpoints or further effect-directed analysis (EDA). To avoid logistic and technical issues related to the storage and transport of large volumes ofwater, samplingwould benefit greatly from onsite extraction. This study presents a novel onsite large volume solid phase extraction (LVSPE) device tailored to fulfill the requirements for the successful effect-based and chemical screening of water resources and complies with available international standards for automated sampling devices. Laboratory recovery experiments using 251 organic compounds in the log D range from −3.6 to 9.4 (at pH 7.0) spiked into pristine water resulted in acceptable recoveries and from 60 to 123% for 159 out of 251 substances.Within a European-wide demonstration program, the LVSPE was able to enrich compounds in concentration ranges over three orders of magnitude (1 ng L−1 to 2400 ng L−1). Itwas possible to discriminate responsive samples from samples with no or only low effects in a set of six different bioassays (i.e. acetylcholinesterase and algal growth inhibition, androgenicity, estrogenicity, fish embryo toxicity, glucocorticoid activity). The LVSPE thus proved applicable for onsite extraction of sufficient amounts ofwater to investigate water quality thoroughly by means of chemical analysis and effect-based tools without the common limitations due to small sample volumes.publishedVersio

Corrigendum to “European demonstration program on the effect-based and chemical identification and monitoring of organic pollutants in European surface waters” [Sci. Total Environ. 601–602 (2017) 1849–1868]

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