Rikke Brix

Environmental risk assessment of pharmaceuticals in rivers: Relationships between hazard indexes and aquatic macroinvertebrate diversity indexes in the Llobregat River (NE Spain)

Continuous input of pharmaceuticals into rivers, through wastewater treatment systems, may cause adverse effects on the aquatic ecosystems of the receiving waterbodies, due to the intrinsic biological activity of these compounds. To investigate this issue, we have carried out an Environmental Risk Assessment in the lower part of the Llobregat River basin (NE Spain). The survey was carried out along three campaigns in 7 sampling points, located in the main river and in one of its tributaries (Anoia River). In each sample, 29 commonly used pharmaceuticals, belonging to different therapeutical classes (analgesics and non-steroidal anti-inflammatories (NSAIDs), lipid regulators, psychiatric drugs, anti-histamines, anti-ulcer agents, antibiotics and beta-blockers) have been determined. Simultaneously, the macroinvertebrate community status of the same points has been also studied. Hazard quotient indexes have been estimated for the most representative compounds as the ratio between concentrations and EC(50) reported values, for three bioassays commonly used in environmental toxicology, namely, fish, Daphnia and algae. Hazard indexes are obtained for each sample by summing up the hazard quotients of all the compounds present, and taking its average along the three sampling campaigns. In general, hazard quotients tend to increase when going downstream. Only those points located most upstream of the two rivers can be qualified under low risk for the three bioassays. The most sensitive bioassay seems to be algae, followed by Daphnia and fish. Log-transformed hazard indexes show fairly good inverse correlations (r=-0.58 to -0.93, p<0.05) with Shannon diversity indexes of macroinvertebrates, determined from both densities and biomasses. Best correlations are obtained for Daphnia based hazard indexes, as expected from its taxonomical proximity to macroinvertebrates. The abnormal correlation behaviour found in one point located in the Anoia River is explained by the presence of other previously reported pollutants of industrial origin, generated by the nearby existing industry.

Bridging levels of pharmaceuticals in river water with biological community structure in the Llobregat River basin (northeast Spain).

A wide range of human pharmaceuticals are present at low concentrations in freshwater systems, particularly in sections of polluted river. These compounds show high biological activity, often associated with a high stability. These characteristics imply a potential impact of these substances on aquatic biota even when present at low environmental concentrations. Low flow conditions in Mediterranean rivers, most of which flow through densely populated areas and are subjected to intensive water use, increase the environmental risk of these emergent compounds. Here, we studied whether pharmaceuticals in river water affect the local benthic community structure (diatoms and invertebrates). For this purpose, we analyzed the occurrence of pharmaceuticals along the Llobregat River and examined the benthic community structure (diatoms and invertebrates) of this system. Some pharmaceutical products in the Llobregat River registered concentrations greater than those cited in the literature. Multivariate analyses revealed a potential causal association between the concentrations of some anti-inflammatories and beta-blockers and the abundance and biomass of several benthic invertebrates (Chironomus spp. and Tubifex tubifex). Further interpretation in terms of cause-and-effect relationships is discussed; however, it must be always taken with caution because other pollutants also may have significant contributions. Combined with further community experiments in the laboratory, our approach could be a desirable way to proceed in future risk management decisions.

How to confirm identified toxicants in effect-directed analysis

AbstractDue to the production and use of a multitude of chemicals in modern society, waters, sediments, soils and biota may be contaminated with numerous known and unknown chemicals that may cause adverse effects on ecosystems and human health. Effect-directed analysis (EDA), combining biotesting, fractionation and chemical analysis, helps to identify hazardous compounds in complex environmental mixtures. Confirmation of tentatively identified toxicants will help to avoid artefacts and to establish reliable cause–effect relationships. A tiered approach to confirmation is suggested in the present paper. The first tier focuses on the analytical confirmation of tentatively identified structures. If straightforward confirmation with neat standards for GC–MS or LC–MS is not available, it is suggested that a lines-of-evidence approach is used that combines spectral library information with computer-based structure generation and prediction of retention behaviour in different chromatographic systems using quantitative structure–retention relationships (QSRR). In the second tier, the identified toxicants need to be confirmed as being the cause of the measured effects. Candidate components of toxic fractions may be selected based, for example, on structural alerts. Quantitative effect confirmation is based on joint effect models. Joint effect prediction on the basis of full concentration–response plots and careful selection of the appropriate model are suggested as a means to improve confirmation quality. Confirmation according to the Toxicity Identification Evaluation (TIE) concept of the US EPA and novel tools of hazard identification help to confirm the relevance of identified compounds to populations and communities under realistic exposure conditions. Promising tools include bioavailability-directed extraction and dosing techniques, biomarker approaches and the concept of pollution-induced community tolerance (PICT). FigureToxicity confirmation in EDA as a tiered approach

Toxicity assessment of sediments from three European river basins using a sediment contact test battery

The toxicity of four polluted sediments and their corresponding reference sediments from three European river basins were investigated using a battery of six sediment contact tests representing three different trophic levels. The tests included were chronic tests with the oligochaete Lumbriculus variegatus, the nematode Caenorhabditis elegans and the mudsnail Potamopyrgus antipodarum, a sub-chronic test with the midge Chironomus riparius, an early life stage test with the zebra fish Danio rerio, and an acute test with the luminescent bacterium Vibrio fischeri. The endpoints, namely survival, growth, reproduction, embryo development and light inhibition, differed between tests. The measured effects were compared to sediment contamination translated into toxic units (TU) on the basis of acute toxicity to Daphnia magna and Pimephales promelas, and multi-substance Potentially Affected Fractions of species (msPAF) as an estimate for expected community effects. The test battery could clearly detect toxicity of the polluted sediments with test-specific responses to the different sediments. The msPAF and TU-based toxicity estimations confirmed the results of the biotests by predicting a higher toxic risk for the polluted sediments compared to the corresponding reference sediments, but partly having a different emphasis from the biotests. The results demonstrate differences in the sensitivities of species and emphasize the need for data on multiple species, when estimating the effects of sediment pollution on the benthic community.

Wastewater reuse in Mediterranean semi-arid areas: The impact of discharges of tertiary treated sewage on the load of polar micro pollutants in the Llobregat river (NE Spain).

The presence of sewage-borne micro contaminants in environmental waters is directly related to the discharge of treated effluents from wastewater treatment plants (WWTP) and the flow rate of the receiving river waters. Mediterranean rivers, in particular, are characterized by important fluctuations in the flow rates and heavy pollution pressures resulting from extensive urban, industrial and agricultural activities. This translates into contamination levels in these rivers often higher than those in other larger European basins. The present work provides an overview of the occurrence of five groups of organic contaminants (131 compounds) namely pharmaceuticals, illicit drugs, polar pesticides, estrogens, alkylphenols and related ethoxylates in WWTP tertiary treatment effluents. Data gathered during a period of water reuse carried out in the lower stretch of the Llobregat river (NE Spain), in the surroundings of the town of Barcelona as a consequence of the severe drought that took place along the years 2007-2008 are presented as illustrative example. In general, measured concentrations of the target compounds were in the low to mid ngL(-1) range. The total concentration of each compound class downstream to the discharge point was similar or slightly higher than that found upstream. Regarding the loads calculated for each compound, the relative contribution from the river upstream and the tertiary effluent were highly compound depending with no apparent trend. However, estimation of the overall bulk loads for each compound class determined in the Llobregat river showed the following rank order: pharmaceuticals>alkylphenols>pesticides>illicit drugs≫estrogens.

Solubility of nonylphenol and nonylphenol ethoxylates. On the possible role of micelles

The water solubility of nonylphenol (NP) has been estimated to be 4.9 +/- 0.4 mg/l corresponding to (2.22 +/- 0.18) x 10(-5) mol/l at 25 degrees C using shake flask and surface tension techniques. The low solubility in combination with an observed rather slow dissolution process will limit the leachability of NP in the terrestrial environment. Based on indirect evidence, it is suggested that NP, in contrast to nonylphenol ethoxylate (NPEO) with, e.g., 12 ethoxylate moieties, is not subject to micelle formation, and as such does not constitute a potential vehicle for the transport of hydrophobic pollutants in the environment. For NPEOs with a very high number of ethoxy moieties, e.g., 100, the compounds appear water soluble without micelle formation.

Are pharmaceuticals more harmful than other pollutants to aquatic invertebrate species: A hypothesis tested using multi-biomarker and multi-species responses in field collected and transplanted organisms

The aim of this study was to test if pharmaceuticals could explain observed responses of field collected and transplanted invertebrate species (Hydropsyche exocellata, Echinogammarus longisetosus, and Daphnia magna). The study was performed in the middle and lower course of Llobregat river basin, which is affected by pharmaceuticals and other pollutants coming from sewage treated effluents. Up to 10 different endpoints including enzyme activities related with detoxication mechanisms (i.e. glutathione S transferase, catalase, esterases), the oxidative stress damage marker (lipid peroxidation), and individual responses (mortality, post-exposure feeding rates) were assessed. Biological responses were complemented with a detailed chemical analysis of metals, detergents, pesticides, pharmaceuticals and other general water quality variables to allow identifying causal abiotic factors. Estimated hazard indexes of measured pollutants indicated that pesticides and metals accounted for most of the predicted toxicity (>95%) in the most contaminated site and that the predicted toxicity of pharmaceuticals was marginal (<5%). The three species showed a clear impact across the studied gradient indicated by higher levels of feeding inhibition and of mortality towards lower reaches. Specific responses such as inhibition of cholinesterase activities were closely related to high and presumable toxic levels of diazinon, whereas unspecific responses such as enhanced levels of antioxidant defensive mechanism and of lipid peroxidation levels were associated with most pollutant classes as well as with high and presumable toxic levels of salt and ammonia. These results indicate that pesticides, salinity, ammonia probably had greater effects on the studied species than pharmaceuticals.

Characterizing field sediments from three European river basins with special emphasis on endocrine effects - A recommendation for Potamopyrgus antipodarum as test organism

The assessment of endocrine disrupting potentials of field sediments has until now been mostly limited to classical chemical analysis, in vitro assays and in vivo bioassays performed with vertebrates. There is an urgent need for easy, cheap and reproducible invertebrate tests which may be applied in certain monitoring activities. Since the mudsnail Potamopyrgus antipodarum is known to be tolerant to natural stressors, but also sensitive to endocrine disrupting chemicals, it is very likely that this organism could be suitable for the assessment of endocrine effects of e.g. field sediments. Within this study the endocrine potential of sediments in three European river basins was assessed. The yeast estrogen screen (YES) and a sediment contact test with P. antipodarum were performed. Furthermore, analyses of physico-chemical properties and concentrations of heavy metals, PAHs, organotins, natural steroids and alkylphenols were done. In the sediment contact test, the reproduction of the snail was promoted by a part of the sediments. This phenomenon could not be explained by their physico-chemical properties. However, at some of those sites a high estrogenic activity was detected in the YES, leading to the assumption that endocrine disrupting compounds could be responsible for those effects. This assumption could be confirmed to some extent with partially high concentrations of xeno-estrogens (e.g. nonylphenol) at the certain sites. Our study demonstrates the applicability of the test with P. antipodarum for a variety of sediments and once again points out the need of suitable in vivo biotests for the risk assessment of field sediments.

Identification of disinfection by-products of selected triazines in drinking water by LC-Q-ToF-MS/MS and evaluation of their toxicity

During the development of an on-line solid phase extraction-liquid chromatography-ultraviolet detection (SPE-LC-UV) analytical method for determination of eight selected triazines; ametryn, atrazine, cyanazine, metrybuzine, prometryn, propazin, simazine, and terbutryn, in drinking water, it was observed that the retention times of three of them (ametryn, prometryn, and terbutryn) in Milli-Q water were different from those in chlorinated Milli-Q water, indicating the formation of new products. The cause of this change was found in the oxidation of the molecules as a result of chlorination with sodium hypochlorite. Experiments performed at varying concentrations of triazines and hypochlorite showed that the extent of the reaction depended on their relative concentrations. At the maximum admissible level of 100 ng/l for individual pesticides in drinking water, no apparent transformation was observed in the absence or at low concentrations (0.05 mg/l) of hypochlorite; however, on increasing the concentration of hypochlorite to the level typically present in drinking water (0.9 mg/l) the transformation was complete. The reaction is quite fast; within 1 h the parent compound is completely degraded and after 22 h the concentrations of the by-products are constant. Investigation of the by-products by ultra performance liquid chromatography-quadrupole-time of flight- tandem mass spectrometry (UPLC-Q-ToF-MS/MS) has shown that all three triazines follow a similar transformation pathway, forming four new molecules whose structure have been elucidated. The acute toxicity of the new products was investigated using a standard method based on the bioluminescence inhibition of Vibrio fischeri, and the by-products showed a higher toxicity than that of the parent compounds.

In situ cage experiments with Potamopyrgus antipodarum - a novel tool for real life exposure assessment in freshwater ecosystems.

In situ experiments are an important tool within ecotoxicological research but there is a lack of suitable methodologies especially for freshwater invertebrate species. Within this study, a novel in situ methodology with Potamopyrgus antipodarum was developed. Snails were inserted into cages, made of Plexiglas measuring 7 × 9 × 7 cm(3) and fixed with stainless steel pins into the sediment at the relevant sampling sites. During the experiment physico-chemical properties of the water and concentrations of metals, PAHs and PCBs were measured in the sediment. The growth and survival of the snails was not affected, but the reproduction increased significantly at one of the most polluted sites. The increase in reproduction was neither correlated with physico-chemical parameters, nor with the concentrations of the different compounds, but maybe related to certain groups of estrogenic compounds. The study demonstrates the excellent applicability of this novel in situ test.