Effect-based and chemical analytical monitoring of steroidal estrogens

Effect-based and chemical analytical monitoring of steroidal estrogens

Project coordination: Robert Kase, Mario Carere (ISS‐Italian Institute of Health, IT)

Endocrine disrupting chemicals may pose a risk to the quality of European water bodies. Three steroidal estrogens, 17α-ethinylestradiol (EE2), 17β-estradiol (E2) and estrone (E1), are currently included in the so-called “watch list” of the Water Framework Directive (WFD) and are therefore recommended for monitoring. However, this may be difficult because the detection limits of most existing routine analytical methods are above the biological effect concentrations, and high-end analytical methods are very costly. However, effect-based methods can measure the estrogenic activity of environmental samples in a cost-efficient way at very low concentrations. Therefore, we will compare seven specific effect-based methods with three sensitive chemical analysis methods to measure EE2, E2 and E1. For this purpose, 35 surface water and wastewater samples are collected and analysed. 24 institutions from 12 countries are involved in this activity (see below). The project is based on the Science-to-Policy-Interface and Chemical Monitoring of Emerging Pollutants activities under the Common Implementation Strategy for the EU WFD, in which the Working Group Chemicals identified a strong need for research on effect-based monitoring tools in support of implementing the relevant legislation. 

In this project we will provide harmonised methods for sample collection, sample extraction and data evaluation, and screening and risk assessment. We aim at: a) Promoting reliable screening methods to support the monitoring of endocrine disrupting activity in wastewater and surface water b) Harmonizing monitoring options across Europe c) Linking reliable effect-based tools with regulatory needs d) Linking effect-based tools with chemical analysis for estrogen monitoring e) Supporting national and EU monitoring for endocrine disruptors f) Supporting the watch-list mechanism within the WFD 

Applied test systems

Estrogenic substances are biologically active at very low concentrations at which they can be poorly analysed chemically. As an alternative, bioassays can be used to detect these substances in surface waters. Bioassays are also able to measure the total estrogenic activity of all active substances in the sample. Various in vitro assays are available that can, however, provide different results for the same sample. Therefore, the variability and reproducibility of five different assays with artificially generated and natural water samples are analyzed and the suitability of the assays for environmental monitoring is assessed.


Kase, R., Javurkova, B., Simon, E., Swart, K., Buchinger, S., Könemann, S., Escher, B., Carere, M., Dulio, V., Ait-Aissa, S., Hollert, H., Valsecchi, S., Polesello, S., Behnisch, P., di Paolo, C., Olbrich, D., Sychrova, E., Gundlach, M., Schlichting, R., Leborgne, L., Clara, M., Scheffknecht, C., Marneffe, Y., Chalon, C., Tusil, P., Soldan, P., von Danwitz, B., Schwaiger, J., Moran, A., Bersani, F., Perceval, O., Kienle, C., Vermeirssen, E., Hilscherova, K., Reifferscheid, G., Werner, I. (2018) Screening and risk management solutions for steroidal estrogens in surface and wastewater. Trends in Analytical Chemistry https://doi.org/10.1016/j.trac.2018.02.013

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Könemann, S., Kase, R., Simon, E., Swart, K., Buchinger, S., Schlüsener, M. Hollert, H., Escher, B.I., Werner, I., Ait-Aissa, S., Vermeirssen, E., Dulio, V., Valsecchi, S., Polesello, S., Behnisch, P., Javurkova, B., Perceval, O., di Paolo, C., Olbrich, D. … Carere, M. (2018) Effect-based and chemical analytical methods to monitor estrogens under the European Water Framework Directive. Trends in Analytical Chemistry 102, 225-235

Project results

Project related references

[1] Carere Mario, Polesello Stefano, Kase Robert, Gawlik Bernd (2015): The emerging contaminants in the context of the EU Water Framework Directive. Bookchapter Springer-Verlag. Part of the series The Handbook of Environmental Chemistry pp 1-19. Avaiable on request from Marion Carere mario.carere@iss.it or at: http://link.springer.com/chapter/10.1007/698_2015_5011

[2] Wernersson Ann-Sofie; Carere Mario, et al. (2015): The European technical report on aquatic effect-based monitoring tools under the water framework directive. Environmental Sciences Europe, 2015; 27 (1) DOI: 10.1186/s12302-015-0039-4. http://www.enveurope.com/content/pdf/s12302-015-0039-4.pdf

Press release at:http://www.sciencedaily.com/releases/2015/03/150313083449.htm

[3] Kunz Petra, Kienle Cornelia, Carere Mario, Homazava Nadzeya, Kase Robert (2014): In vitro bioassays to screen for endocrine active pharmaceuticals in surface and waste waters, Journal of Pharmaceutical and Biomedical Analysis,http://dx.doi.org/10.1016/j.jpba.2014.11.018 .

[4] Jarošová Barbora, Bláha Luděk, Giesy John P, Hilscherová Klára (2014): What level of estrogenic activity determined by in vitro assays in municipal waste waters can be considered as safe? Environment International 64: 98–109

[5] Wernersson A-S , Maggi C, Carere M. 2014. EU technical report on aquatic effect-based monitoring tools. Report available at:https://circabc.europa.eu/w/browse/80c5932e-8e8b-4cf8-b34e-db18ba127e95

[6] Heiss Christiane et al. 2013. Recommendation for a monitoring strategy for estrogens in coastal and continental surface waters. Available at :http://www.bafg.de/DE/05_Wissen/02_Veranst/2013/2013_02_27_votum_en.pdf?__blob=publicationFile

[7] Johnson A C, Dumont E, Williams R J, Oldenkamp R, Cisowska I, Sumpter J. 2013: Do concentrations of ethinylestradiol, estradiol and diclofenac in European rivers exceed proposed EU environmental quality standards?. Environmental Science & Technology47(21) 12297-12304.

[8] Loos Robert in EU JRC report. 2012: “Analytical methods relevant to the European Commission's 2012 proposal on Priority Substances under the Water Framework Directive. Including: Kase R, Kunz P, Hollert H, Werner I (2012): Contribution on bioanalytical assays for steroidal estrogens”. ISBN 978-92-79-26642-3. Publications Office of the European Union, 2012. Available at CIRCABC orhttp://publications.jrc.ec.europa.eu/repository/handle/111111111/26936.

[9] Kase Robert, Clayton Helen, Martini Frederique. 2012. Science-Policy Interface (SPI) activity on prioritisation of research needs, knowledge availability and dissemination for the Working Group E (Chemical Aspects) 2010-2012. Open available at CIRCABC at: https://circabc.europa.eu/w/browse/5bf63ff3-b24b-4365-8a57-38e4d56b941c

[10] Hecker Markus and Hollert Henner. 2011. Endocrine disruptor screening: regulatory perspectives and needs. Environmental Sciences Europe 2011. 23:15. http://www.enveurope.com/content/23/1/5

[10] European Commission. 2011. 4th Report Commission Staff Working Paper on the implementation of the “Community Strategy for Endocrine Disruptors” a range of substances suspected of interfering with the hormone systems of humans and wildlife (COM (1999) 706). SEC(2011) 1001 final. Brussels, 10.08.2011