High resolution effect-directed analysis of steroid hormone (ant)agonists in surface and wastewater quality monitoring.

Monitoring of chemical water quality is extremely challenging due to the large variety of compounds and the presence of biologically active compounds with unknown chemical identity. Previously, we developed a high resolution Effect-Directed Analysis (EDA) platform that combines liquid chromatography with high resolution mass spectrometry and parallel bioassay detection. In this study, the platform is combined with CALUX bioassays for (anti)androgenic, estrogenic and glucocorticoid activities, and the performance of the platform is evaluated. It appeared to render very repeatable results, with high recoveries of spiked compounds and high consistency between the mass spectrometric and bioassay results. Application of the platform to wastewater treatment plant effluent and surface water samples led to the identification of several compounds contributing to the measured activities. Eventually, a workflow is proposed for the application of the platform in a routine monitoring context. The workflow divides the platform into four phases, of which one to all can be performed depending on the research question and the results obtained. This allows one to make a balance between the effort put into the platform and the certainty and depth by which active compounds will be identified. The EDA platform is a valuable tool to identify unknown bioactive compounds, both in an academic setting as in the context of legislative, governmental or routine monitoring.

Advancements in effect-based surface water quality assessment.

Legally-prescribed chemical monitoring is unfit for determining the pollution status of surface waters, and there is a need for improved assessment methods that consider the aggregated risk of all bioavailable micropollutants present in the aquatic environment. Therefore, the present study aimed to advance effect-based water quality assessment by implementing methodological improvements and to gain insight into contamination source-specific bioanalytical responses. Passive sampling of non-polar and polar organic compounds and metals was applied at 14 surface water locations that were characterized by two major anthropogenic contamination sources, agriculture and wastewater treatment plant (WWTP) effluent, as well as reference locations with a low expected impact from micropollutants. Departing from the experience gained in previous studies, a battery of 20 in vivo and in vitro bioassays was composed and subsequently exposed to the passive sampler extracts. Next, the bioanalytical responses were divided by their respective effect-based trigger values to obtain effect-based risk quotients, which were summed per location. These cumulative ecotoxicological risks were lowest for reference locations (4.3-10.9), followed by agriculture locations (11.3-27.2) and the highest for WWTP locations (12.8-47.7), and were mainly driven by polar organic contaminants. The bioanalytical assessment of the joint risks of metals and (non-)polar organic compounds resulted in the successful identification of pollution source-specific ecotoxicological risk profiles: none of the bioassays were significantly associated with reference locations nor with multiple location types, while horticulture locations were significantly characterized by anti-AR and anti-PR activity and cytotoxicity, and WWTP sites by ERα activity and toxicity in the in vivo bioassays. It is concluded that the presently employed advanced effect-based methods can readily be applied in surface water quality assessment and that the integration of chemical- and effect-based monitoring approaches will foster future-proof water quality assessment strategies on the road to a non-toxic environment.

Effect-based nationwide surface water quality assessment to identify ecotoxicological risks.

A large portion of the toxic effects observed in surface waters cannot be attributed to compounds regularly measured by water authorities. Hence, there is an urgent need for an effect-based monitoring strategy that employs bioassays to identify environmental risks. The aim of the present study was to perform an effect-based nationwide water quality assessment to identify ecotoxicological risks in a wide variety of surface waters. At 45 locations silicone rubbers and polar organic chemical integrative samplers were exposed to surface water for 6 weeks. Alongside the passive samplers an in-situ daphnid test was performed. Subsequent to field exposure, accumulated compounds were extracted from the passive samplers after which a battery of in vivo and in vitro bioassays was exposed to the extracts. The bioassay battery was selected such that it could identify the risks posed by a wide range of chemical pollutants and their transformation products, while simultaneously allowing for targeted identification of groups of compounds that cause specific effects. Bioassay responses were compared to effect-based trigger values to identify potential ecotoxicological risks at the investigated locations. Responses were observed in all bioassays, and trigger values were exceeded in 9 out of the 21 applied assays, allowing for ranking of the investigated locations based on ecotoxicological risks. No relationship between land use and the identification of ecotoxicological risks was observed. Based on the results, considerations regarding future improvements of effect-based monitoring are given. It is concluded that effect-based water quality assessment allowed prioritization of sites based on ecotoxicological risks, identified the presence of hazardous compounds regardless of being listed as priority substances, and meanwhile could prevent costly chemical analysis at sites with low ecotoxicological risks.

An ecotoxicological view on neurotoxicity assessment.

The numbers of potential neurotoxicants in the environment are raising and pose a great risk for humans and the environment. Currently neurotoxicity assessment is mostly performed to predict and prevent harm to human populations. Despite all the efforts invested in the last years in developing novel in vitro or in silico test systems, in vivo tests with rodents are still the only accepted test for neurotoxicity risk assessment in Europe. Despite an increasing number of reports of species showing altered behaviour, neurotoxicity assessment for species in the environment is not required and therefore mostly not performed. Considering the increasing numbers of environmental contaminants with potential neurotoxic potential, eco-neurotoxicity should be also considered in risk assessment. In order to do so novel test systems are needed that can cope with species differences within ecosystems. In the field, online-biomonitoring systems using behavioural information could be used to detect neurotoxic effects and effect-directed analyses could be applied to identify the neurotoxicants causing the effect. Additionally, toxic pressure calculations in combination with mixture modelling could use environmental chemical monitoring data to predict adverse effects and prioritize pollutants for laboratory testing. Cheminformatics based on computational toxicological data from in vitro and in vivo studies could help to identify potential neurotoxicants. An array of in vitro assays covering different modes of action could be applied to screen compounds for neurotoxicity. The selection of in vitro assays could be guided by AOPs relevant for eco-neurotoxicity. In order to be able to perform risk assessment for eco-neurotoxicity, methods need to focus on the most sensitive species in an ecosystem. A test battery using species from different trophic levels might be the best approach. To implement eco-neurotoxicity assessment into European risk assessment, cheminformatics and in vitro screening tests could be used as first approach to identify eco-neurotoxic pollutants. In a second step, a small species test battery could be applied to assess the risks of ecosystems.

Complex physiological traits as biomarkers of the sub-lethal toxicological effects of pollutant exposure in fishes.

Complex physiological traits, such as routine aerobic metabolic rate or exercise performance, are indicators of the functional integrity of fish that can reveal sub-lethal toxicological effects of aquatic pollutants. These traits have proved valuable in laboratory investigations of the sub-lethal effects of heavy metals, ammonia and various xenobiotics. It is not known, however, whether they can also function as biomarkers of the complex potential range of effects upon overall functional integrity caused by exposure to mixtures of chemicals in polluted natural environments. The current study used portable swimming respirometers to compare exercise performance and respiratory metabolism of fish exposed in cages for three weeks to either clean or polluted sites on three urban European river systems: the river Lambro, Milan, Italy; the rivers Blythe, Cole and Tame, Birmingham, UK; and the river Amstel, Amsterdam, The Netherlands. The UK and Italian rivers were variously polluted with high levels of both bioavailable heavy metals and organics, and the Amstel by mixtures of bioavailable organics at high concentrations. In both the UK and Italy, indigenous chub (Leuciscus cephalus) exposed to clean or polluted sites swam equally well in an initial performance test, but the chub from polluted sites could not repeat this performance after a brief recovery interval. These animals were unable to raise the metabolic rate and allocate oxygen towards exercise in the second trial, an effect confirmed in successive campaigns in Italy. Swimming performance was therefore a biomarker indicator of pollutant exposure in chub exposed at these sites. Exposure to polluted sites on the river Amstel did not affect the repeat swimming performance of cultured cloned carp (Cyprinus carpio), indicating either a species-specific tolerance or relative absence of heavy metals. However, measurements of oxygen uptake during swimming revealed increased rates of routine aerobic metabolism in both chub and carp at polluted sites in all of the rivers studied, indicating a sub-lethal metabolic loading effect. Therefore, the physiological traits of exercise performance and metabolic rate have potential as biomarkers of the overall sub-lethal toxic effects of exposure to complex mixtures of pollutants in rivers, and may also provide insight into why fish do not colonize some polluted environments.

Interrelationships between bioaccumulation of organic trace pollutants (PCBs, organochlorine pesticides and PAHs), and MFO-induction in fish.

1. Hydrophobic compounds that are more easily biotransformed (e.g. PAHs) generally show less bioaccumulation in fish than the more persistent PCBs and DDTs. 2. In lake Nieuwe Meer, with the highest levels of organic micropollutants, the hepatic MFO activity was elevated in three fish species. This indicates that MFO activity might be a sensitive indicator for organic micropollutants in the aquatic environment. 3. Activities of the 3-MC type inducible isozymes were most pronounced in all three fish species studied. Though further research is required, indications for the existence of a PB-type inducible enzyme system have been demonstrated for two fish species, i.e. pike and eel. 4. Fish liver enlargement as a consequence of MFO induction could not be demonstrated in fish from the most polluted lake. 5. In fish, a correlation was observed between PCB or OCP tissue concentrations on the one hand, and hepatic MFO activities on the other. The low PAH fish/sediment ratios are further reduced when MFO systems are induced due to organic micropollution. The interrelationship between bioaccumulation and enzyme induction demonstrates the importance of an integrated study of these phenomena in field research.

DNA dosimetry in biological indicator species living on PAH-contaminated soils and sediments.

A large variety of environmental carcinogens are metabolically activated to electrophilic metabolites that can bind to nucleic acids, forming covalent adducts. In organisms possessing active metabolic systems for a particular carcinogen, DNA adducts generally have longer biological half-lives than the substrate carcinogens. Thus, measurement of specific DNA adduct concentrations in terrestrial and water organisms may provide a relevant biological indicator of prior exposure to environmental carcinogens. Analysis of carcinogen load in indicator species with specific behavioral patterns may indicate human exposure risk to environmental carcinogens. Recently, sensitive assays have been developed to measure carcinogen-DNA adducts in organisms exposed to complex mixtures such as polycyclic aromatic hydrocarbons (PAH). At first instance, the nuclease P1 version of the 32P-postlabeling assay was used to examine the liver of eel (Anguilla anguilla) for the presence of aromatic DNA adducts. The fish were collected from six freshwater sites in the Amsterdam area with different levels of PAH contamination in their sediments. Chromatograms derived from DNA of fish from polluted sites revealed a broad diagonal zone indicating the presence of DNA adducts containing aromatic or bulky hydrophobic moieties not present in DNA of fish from an unpolluted reference site. Significant correlations were found between the aromatic DNA adducts levels and the levels of PAH in sediments (P < 0.001). To examine the validity of DNA adduct dosimetry in terrestrial organisms earthworms (Lumbricus terrestris) were kept on industrially contaminated PAH soils for several weeks. Several aromatic DNA adducts could be detected in DNA from the exposed earthworms; adduct levels were significantly increased with increasing exposure time.(ABSTRACT TRUNCATED AT 250 WORDS)