1. 18.03.2013Eivind Farmen Giftighet av nanosølv for fisk I norske vann og vassdrag 1

2. Eivind Farmen2 Silver - the most common material used in commercial nano-products 18.03.2013

3. Nanosilver from a washing machine Concentration in effluent: 2,7 µg/L to 25 µg/L Average: 10,522 µg/L Large variability! Particles found with TEM: Size range: 6 – 11 nm Farkas (2011) Environment international 37 (6), 1057-1062

4. Objectives Determine acute and sublethal toxicity of AgNP to fish & fish cells Determine AgNP characteristics in various matrixes Compare toxic potential of AgNP to ionic silver and AuNP Assess risk to fish under ecologicaly relevant exposure scenarios 18.03.2013Eivind Farmen

5. In vitro experimental model What are the possible effects of nanoparticles on fish cells used as an in vitro test system? Liver Gill Exposure (48h) Cytotoxicity Reactive oxygen species (ROS) production Epithelial integrity Primary hepatocytes Primary gill epithelium cells 18.03.2013Eivind Farmen

6. Particle characteristics by TEM AgNPs Water cell media cell media +DOC Roundish to oval particle shape AuNPs In–house synthesized citrate-coated NPs

7. ROS production & Cytotoxicity (hepatocytes) Silver NPs Gold NPs ROS Cytotoxicity Farkas et al. 2009. Effects of silver and gold nanoparticles on rainbow trout (Oncorhynchus mykiss) hepatocytes. Aquatic toxicol. 18.03.2013Eivind Farmen

8. Effects on primary gill cells Ag is less cytotoxic to gill cells than to hepatocytes AuNPs were not cytotoxic 18.03.2013Eivind Farmen

9. 18.10.2012Eivind Farmen What happens in natural water? Commercial AgNP 1:10 in natural lake water In-house AgNP 1:10 in natural lake water 4 nm 220 nm 9

10. Effects Ag nanoparticles ? ? Control 1  g/L AgNP 20  g/L AgNP 100  g/L AgNP 20  g/L Ag+ 100  g/L AgNP2 18.03.2013Eivind Farmen10

11. 18.03.2013Eivind Farmen GO analysis AgNP 20 µg/L 11

12. 18.10.2012Eivind Farmen GO analysis 12

13. Transcriptional UP-regulation (Ag) 100 ug/L AgNP20 ug/L Ag+ 20 ug/L AgNP Heme-binding protein 2 Protein-Glutamine  -glutamyltransferase GTPase IMAP family member 4 Na/H exchanger 9 18.03.2013Eivind Farmen13

14. Transcriptional Down-regulation (Ag) 100 ug/L AgNP20 ug/L Ag+ 20 ug/L AgNP Carbonic anhydrase 7 Na/K-ATPase Interferon gamma MHC Class II 18.03.2013Eivind Farmen14

15. Mode of action Ag BloodGillWater HCO 3 - Cl - NaK- ATPase H + (NH 4 + ) Na + K+K+ Carbonic anhydrase CO 2 + H 2 O => H + + HCO 3 - CO 2 Na + K+K+ Cl - CO 2 18.03.2013Eivind Farmen15

16. Hazard/risk assessment 100 ug/L AgNP 20 ug/L Ag + 20 ug/L AgNP Control Toxicity of AgNP due to acting as reservoir for Ag + ? Toxicity of AgNP higher in soft waters ! 18.03.2013Eivind Farmen16

17. Effects Ag nanoparticles Control 1  g/L AgNP 20  g/L AgNP 100  g/L AgNP 20  g/L AgNP 100  g/L AgNP2 18.03.2013Eivind Farmen17 Osmo- regulation Necrosis/ apoptosis

18. Work in progress 18.03.2013Eivind Farmen18

19. Differences Ag-NPs 18.03.2013Eivind Farmen19

20. 18.03.2013Eivind Farmen Conclusion High toxicity of Ag-NPs in natural Norwegian waters Ag-NP effect mediated by Ag+ ions? Effects in gills/osmoregulation Modulation of immune system Effect in gills > liver 20

21. 18.03.2013Eivind Farmen Acknowledgements Farkas J, Mikkelsen HN, Evensen Ø, Heier LS, Einset J, Salbu B, Rosseland BO, Oughton DH, Tollefsen KE, Paul Christian, Julián Alberto Gallego-Urrea, Norbert Roos, Martin Hassellöv, Kevin V Thomas, Mie Jareid, Hanne Fossnes, Hans-Christian Teien Funding: – Norwegian Research Council » Nanotrace Project » NIVA, SIP project 24024 NEWPOLL Thank you for listening! 21