TW2004

1. Are current ecotoxicological testing approaches appropriate to assess the hazard of emerging contaminants ? Helmut Segner, University of Bern TW2004

2. What concerns us on „emerging contaminants“ ? • According to the invitation, „emerging contaminants“ include • Pharmaceuticals, and their metabolites • Personal care products, and their metabolites • Nanomaterials • Mixtures • concern on appearance of „new“ compounds in the environment ? • concern on non-anticipated toxic potencies of contaminants ?

3. What concerns us on „emerging contaminants“ ? GHS classification of acute toxicity of pharmaceuticals and industrial chemicals Sanderson and Thomas 2009, Tox Lett 187:84 • Nothing special on the „emerging contaminants • No specific needs for hazard assessment • talk finished

4. What concerns us on „emerging contaminants“ ? One concern on these compounds is that they may cause specific toxicities at (very) low concentrations long-term exposure which are not predicted by current hazard assessment schemes Stealth bombers flying under the radar of hazard assessment

5. Initial steps in aquatic toxicity assessement; what „effects“ are measured ? Substance degradation time < 3 d ? Chemical X yes no Procedure under Reach/ EMEA phaase II tier A Mitigating factors ? yes no PNEC Acute toxicity data available for algae ? (Scenedesmus) no AF 1000 yes Production volume < 10 T ? Acute toxicity data available for fish ? yes no Acute toxicity data available for invertebrates ? (Daphnia) no Water solubility < 1 mg/L) ? yes yes

6. Hypothetical example: assessing aquatic toxicity of EE2 Acute algal toxicity: 72 h EC50 = 0.84 mg/L Acute Daphnia toxicity: 48 h EC50 = 5.7 mg/L Acute fish toxicity: 96 h EC50 = 1.6 mg/L AF 1000 Reproductive NOEC of EE2 in fish: 0. 3 ng/L  PNEC = 0.03 ng/L PNEC= 0.84 µg/L A posteriori, we know that EE2 can cause reproductive toxicity through estrogen receptor binding, and would subject this compound to chronic testing, however, this kind of information is not routinely asked for in environmental hazard assessment

7. Example azole fungicides Frische and Kotschik, SETAC Milano 2011

8. Emerging contaminants may show low acute but increased chronic toxicity due to change in MoA acute toxicity follows baseline, chronic toxicity specific MoA acute and chronic toxicity are baseline MoA

9. Due to specific toxicity, emerging contaminants may have specific ecologcial receptor groups Species sensitivity distribution for EE2

10. Due to specific toxicities, emerging contaminants may have specific ecological receptor groups

11. „Ecological receptors“ refers not only to interspecies but also intra-species differences: life history The contaminant effects are present in specific life stages only: estrogenic impact on zebrafish sex ratio ovary testis 100 80 % sex ratio 60 40 20 0 control, exposed control exposed developmental exposure adult exposure

12. Shaded: acute LC50 range of all test com-pounds So, does current hazard assessment scheme fail for contaminants with specific toxicities ? For the majority of chemicals, it may perform well Vertical lines: LC10 of the most sensitive life stage Ratio of acute LC50 to reproductive EC50 1 500 000 13 500 32  Even for certain estrogenic chemicals, the elevated chronic toxicity of the other EDCs would be well covered by the Assessment Factors

13. How to identify those contaminants that are likely to cause specific hazards ? • Alerts • to go for more detailed hazard characterization • to search for specific receptor groups • Effect • MoA classification • Human pharmacolo-gical/toxicological information • Screening infor-mation • Exposure • For instance, pharmaceuticals dis- playing „critical en-vironmental concen-trations“ (Fick et 2010)

14. Mode of action alerts • Mode of action describes selected key events that lead to toxicity (Vonk et al. 2009) • MoAs used in ecotoxicology group chemicals on the basis of the relation-ship between (acute) effects and physicochemical/structural descriptors • Verhaar scheme: • MoA 1: baseline toxicity, inert „narcotic“ chemical • MoA 2: less inert polar narcotics • MoA 3: unselectively reactive chemicals • MoA 4: specifically reactive chemicals, e.g., receptor ligands • ECETOC Report No 102: • MoA 4 alerts for extended hazard characterization

15. Mode of action alerts • Caveats to the MoA approach: • There exist significant uncertainties in MoA assignment • Current procedures work mainly with acute MoAs, what, however, appears to be not the critical point with respect to „emerging contaminants“ • Current MoA approaches rely on the standard ecotoxicological test endpoints (growth, survival, lethality) measured with standard test organisms, and thus may fail to reveal unexpected toxic qualities or unexpected ecological receptor groups

16. Use of pharmacological/toxicological information from mammalian toxicology Conservation of molecular targets across taxonomic groups Owen et al., 2007, Aquat Tox 82.145

17. „Conservation of targets“ may extend well beyond vertebrates Ibuprofen toxicity in Daphnia magna Ibuprofen interferes with eoicosanoid synthesis in mammals and with juvenile hormone synthesis in Daphnia Heclmann et al. 2007

18. Human pharmacological data may not only provide qualitative but also quantitative information Berninger and Brooks, 2010, Tox lett 193:69

19. Alerts from screening assay and non-conventional endpoints For instance, -omics technologies • they can group chemicals • they can reveal similarities and dissimilarities compared to benchmark compounds • as open approach, they can indicate unexpected effects • they can indicate species at risk

20. Are such alerts toxicologically relevant ? Relation between VTG induction and reproductive success of zebrafish Similar relations exist for other (xeno)estrogens: octylphenol, ethynlestradiol, genistein

21. Are such alerts toxicologically relevant ? no conventional (sub) chronic test data available only non-conventional data on sublethal endpoints available (3 independent studies): histopathology: NOEC/LOEC in the range of 1 g/L Immunomodulation: NOEC/LOEC in the range of 0.5 g/L  Can these data be used in risk assessment ? Non-steroideal anti-inflammatory drug Use in Germany (2009): 91 583 kg Environmental concentration: 0.71 g/L (DG ENV, UBA) Maximum concentration 1.2 g/L (Boxall, 2008)

22. Even if we accept such alerts, definitive confirma-tion of chronic, low dose toxicities can be difficult • Low-dose, chronic effect data are not easy to reproduce • For instance, propanol: • Low concentrations (ng/L) inhibited egg laying in medaka • Higher concentrations (0.5 and 1 g/L) did not inhibit egg laying of medaka ? •  artefact, reality, or the famous U-shaped concentration-response curve ? Huggett et al. 2002, Arch Env Contam Toxicol 43:229;

23. This was all lab-based prediction of potential hazards – but what about field assessment ?

24. Mixture issue: would specifically acting contaminants change the overall toxic pressure at a field site ? Chemical status relates to ecological status Chemical status does not relate to ecological status Data from EU IP „MODELKEY“

25. Multiple stressor issue: would specifically acting con-taminants increase vulnerability to other stresors ? Estrogenic compounds alone do not induce mortality but enhance susceptibility towards pathogenic bacteria bacteria + EDC bacteria alone EDC alone

26. Ecological issue: effects of specifically acting contaminants on ecological/selection processes ? A3A 1.0 Ibuprofen Pharmaceuticals and temperature explain 71 % of the species variance in the Llobregat river (MODELKEY case study; redundancy analysis) propanolol LL4C LL4A LL3A LL2C LL1A LL1C LL3C LL2A A2C LL2B Propranolol A3B A3C LL3B A1A A1C A1B Indomethacine LL1B A2A LL4B -0.6 A2B -0.4 1.2 Muñoz et al.; Env.Tox.Chem., 28: 2706-2714 (2009)

27. THANK YOU VERY MUCH FOR YOUR ATTENTION