Ecotoxicology of engineered nanoparticles
Carmen Gil, Ksenia Groh, Carl Isaacson, Alexandra Kroll, Xiaomei Li, Ursula Lindauer, Niksa Odzak, Flavio Piccapietra, Lena Röhder, Irene Schwyzer, Ahmed Tlili, Bettina Wagner, Yang Yue.
Collaboration between Renata Behra, Kristin Schirmer, Laura Sigg, Marc Suter, Ralf Kägi, Bernd Nowack (Empa, St. Gallen), Marc Gessner (IGB, Stechlin, Germany), Enrique Navarro (CSIC, Zaragoza, Spain), Ruth Sofield (Western Washington University, USA).
Developments of nanotechnology lead to a rapid proliferation of nanomaterials (1) that are likely to become a source of many different nanoparticles for the environment. Currently, a general lack of information on the ecotoxicology of engineered nanoparticles (NP) prevents an accurate evaluation of their environmental risks (2, 3). We are currently examining several types of NP with regard to their fate in aquatic systems and their effects on aquatic organisms and ecological processes.
Projects
Interactions of silver and cerium oxide nanoparticles with the alga Chlamydomonas reinhardtii: influence of particle physico-chemical characteristics. Flavio Piccapietra and Lena Röhder. SNSF 200020_134750.
The present project aims at gathering scientific information on the environmental fate and effects of silver- and cerium oxide nanoparticles. Silver NPs belong to the most important materials incorporated in consumer products and used because of their toxicity to microorganisms. Cerium oxide is industrially exploited because of its catalytic properties also as diesel fuel additive. Due to the wide application of Ag and CeO2 nanoparticles and likelihood of release in the environment, investigations of their fate and effects is required to assist in evaluating their role.
Specific objectives of the project are:
• to examine how physicochemical characteristics of CeO2NPs are influenced by factors relevant for natural waters;
• to examine the toxicity of AgNP and CeO2NP s to algae;
• to assess which physicochemical characteristics of the NPs are determinant for toxicity.
The project adopts a multidisciplinary approach combining chemical, biological and particle-analysis expertise. The ultimate purpose of the project is to contribute to laying a solid foundation that supports environmental risk assessment.
Influence of chemical coatings on the short-term toxicity of silver nanoparticles on photosynthesis. In collaboration with E. Navarro (CSIC, Zaragoza). Link: http://www.enriquenavarro.com.
We are currently examining the influence of various coatings on the toxicity of silver NP to photosynthesis in the alga Chlamydomonas reinhardtii. Considered coatings include polyvinylpyrrolidone, citrate, lactate, gelatin, chitosan and carbonate. Toxicity data are compared as function of the total silver mass as well as function of the concentration of dissolved silver, which is also present in AgNP suspensions. The results indicate a difference in toxicity among the various AgNP when examined as function of the total Ag mass, though they display similar toxicity when analyzed as function of the dissolved silver concentration. Furthermore, since toxicity of all types of AgNP was abolished in presence of the strong silver ligand cysteine, we conclude that toxicity to photosynthesis is determined by the dissolved silver fraction and that none of the coatings are toxic to algal photosynthesis.
Influence of fulvic and humic acids on silver nanoparticle toxicity to algae. Ruth Sofield (Western Washington University, USA). Link: http://faculty.wwu.edu/harperr3/.
During a sabbatical stay at Eawag, Ruth Sofield has been investigating the influence of fulvic and humic acids on Ag speciation and on the effects of AgNPto algae. Ag binding to humic and fulvic acids was examined using an ion-specific electrode for Ag+ and a cation-exchange resin technique. Effects of Ag nanoparticles on photosynthesis of algae were studied in various media in presence and absence of humic and fulvic acids.
Silver nanoparticle effects on simple stream food webs and ecosystem processes (SNEP). SNSF, NRP 64. 406440_131272. Carmen Gil, Ahmed Tlili (IGB, Stechlin, Germany).
With the aim of developing an understanding of the impact of AgNP on food web interactions and ecological processes, we are currently using the fungal litter-decomposition and algal biofilm system in streams as models to assess effects of nanomaterials in ecosystems. Emphasis is being placed on testing for effects on interactions in simplified food webs (fungi-bacteria-detritivores and algae-bacteria-herbivores, respectively) and on key ecosystem processes (leaf litter decomposition and primary production). The experiments are carried out in microcosms and indoor experimental stream channels. A range of state-of-the-art analytical techniques are used to characterize nanoparticles throughout the experiments. Assessed responses include growth and activity parameters of fungi, bacteria, algae and invertebrates as well as rates of litter decomposition and primary production. The outcomes of the proposed project will provide essential baseline information for environmental risk assessment. Because of the direct relevance of the measured endpoints for ecosystem functioning, the results of our experiments will be immediately useful for industries developing nanoparticles, regulators, environmental managers and other stakeholders.
Interaction of metal nanoparticles with aquatic organisms (MeNanoqa). SNSF, NRP 64. 406440_131240. Xiaomei Li, Yang Yue.
The rapid development of nanotechnology spurs the societal and economic need to use this technology in a sustainable way. Regulations regarding production, use and disposal of synthetic nanomaterials are clearly needed to maximize opportunities and minimize potential human and environmental health risks. Rational implementation of such regulations, however, requires comprehensive knowledge of the interaction of nanomaterials with their surrounding environment. The overall aim of MeNanoqa is to develop a mechanistic understanding of the interactions of metal nanoparticles (NPs) with aquatic organisms.
The main goals of the project are:
(1) to gain an understanding of metal NP/organism interactions upon contact, in particular with regard to uptake, elimination, intracellular transformations and interaction with biological molecules. Our focus will be on algae and fish cells.
(2) to provide a framework to aid optimized metal NP design by taking the mechanisms of interactions of the NPs with aquatic organisms into account.
The work proposed is divided into four parts. Firstly, we will quantify how particle characteristics are modified by aging in different media and interactions with biomolecules. We will secondly explore the uptake and intracellular fate of NPs. This research will provide information in terms of NP exposure and potential long-term behavior and effects and guide protein identification in part three. Part three will focus on identifying and characterizing proteins bound to NPs and on learning how NPs alter structure and function of proteins. Part four of the project will synthesize the gained knowledge to build a framework to serve as a guide for future particle design and for hazard assessment.
Project Nanotrack: Study of life cycle of nanoparticles using [45Ti]TiO2 and [105Ag]Ag0 (funded by BMBF, Germany)
Link zu Project Nanotrack: http://www.hzdr.de/db/Cms?pOid=32171&pNid=2058
In collaboration with : Institut für Radiochemie, HZDR Leipzig; Institut für Oberflächenmodifizierung, Leipzig; Cetelon Nanotechnik GmbH, Eilenburg, D.
Eawag participants: Carl Isaacson, Kristin Schirmer, Laura Sigg, Adrian Ammann.
Behavior of TiO2 nanoparticles in a simple food chain. Carl Isaacson.
Very little is known about how exposure to TiO2 nanoparticles at one level of a food chain may effect higher levels of the food chain. In order to provide valuable data to address this gap, toxicology studies must be conducted in a manner which builds on current knowledge.
To date, many studies have shown that once TiO2 nanoparticles enter aquatic environments, solution forces acting on the nanoparticles result in aggregation, then deposition of these particles from the water column to the sediment. This aggregation and deposition will likely result in TiO2 nanoparticles entering the sediment where nanoparticle exposure to benthic organisms is likely to occur.
As the interface between sediment and many water bodies, biofilms are likely to be a prime recipient of TiO2 nanoparticles as they aggregate and deposit from the water column. While many studies have reported on the interactions of TiO2 nanoparticles and pure cultures of planktonic microorganisms and the potential for toxic effects as a result of these interaction, very little is known about the interactions of TiO2 nanoparticles with biofilms, if nanoparticles can be taken up by biofilms or become trapped in the extracellular matrix of biofilms, and if interactions between nanoparticles and biofilms can result in toxic effects to biofilms. Additionally, as a primary producer in many aquatic systems, biofilms serve as a vital food source for many higher level predators and consumption of biofilms and associated TiO2 nanoparticles by organisms at higher trophic levels may adversely impact these organisms. Rotifers, protozoa, and nematodes are three types of predators that are known to feed on biofilms and determining the ability for trophic transfer of nanoparticles from biofilms to these organisms, will provide much needed information for assessing the implications food chain transfer of nanoparticles.
Interactions of engineered nanoparticles with periphyton. Alexandra Kroll.
We are currently investigating the distribution and bioavailability,of engineered nanoparticles (silver NP, cerium dioxide NP, titaniumdioxide NP) in freshwater periphyton as well as potential biological effects. In particular, we focus on the distribution of engineered nanoparticles between water body and periphyton, the interaction with extracellular components of periphytic organisms, and material properties such as size, charge, and dissolution. The impact on biological parameters such as particle uptake, species composition, and photosynthetic activity will be quantified. To this end, periphyton is colonized on glass slides in small indoor channels in a flow-through system fed by natural river water taken directly on campus from river Chriesbach. Chriesbach water quality is regularly monitored by the local water authority. Exposure to nanoparticles is then realized in closed recirculation systems. Methods applied in this project include dynamic light scattering (DLS), nanoparticle tracking analysis (NTA), electron microscopy, inductively coupled plasma mass spectrometry (ICP-MS), flow cytometry, confocal laser scanning microscopy (CLSM), photosynthesis measurements, and algae culturing techniques.
The study will provide valuable insides in the fate of engineered nanoparticles in the aquatic environment and help to evaluate the suitability of algal biofilms and the biological parameters tested for the assessment of nanoparticle effects on freshwater algae.
A proteomics view on the effects of silver nanoparticles in zebrafish embryos. Ksenia Groh.
The current project aims to investigate the mechanisms of toxicity of silver nanoparticles (AgNP) to zebrafish embryos. AgNP were shown to be toxic to zebrafish embryos. However, the question whether obseved effects are due to the silver ions present in the nanoparticle solution, or to other properties of the particles themselves, has not been properly addressed so far. We expose zebrafish embryos to silver nitrate (a source of silver ions) or nanoparticles, and look at the effects exerted by corresponding concentrations. To avoid an uncontrolled loss of silver ions from the solution caused by precipitation in form of AgCl, we use a specially designed low chloride exposure medium. Whenever needed, L-cysteine, a strong silver ligand, is added as a means for complete abolishment of silver ions bioavailability. Further, we use multidimensional Protein Identification Technology to characterize the proteomes of embryos exposed to silver nitrate or nanoparticles, with or without cysteine. The observed alterations in protein profiles may provide the insights into the molecular mechanisms of AgNP toxicity in zebrafish embryos.
Solubilization of carbon nanotubes in natural waters. Irene Schwyzer, in collaboration with Bernd Nowack, Empa
Link to: project description at Empa (http://www.empa.ch/plugin/template/empa/*/69978)
Carbon nanotubes (CNTs) are an important new class of engineered nanoparticles, which consist of graphene sheets and may be single-walled or multi-walled. Fate and effects of CNTs depend on their aggregation state in natural waters. CNTs are virtually insoluble in water, but may be solubilized in presence of natural polymers such as humic and fulvic acids. The aim of this project is to analyze the behavior of CNTs under conditions similar as those of natural waters. The results indicate that the CNT properties (size, functional groups), as well as their initial state (dry vs. predispersed) strongly influence the stability of CNT suspensions(8).
Dissolution of metal and metal oxide nanoparticles in natural aqueous media (Bafu project No. 10.0007.KP / J115-274). Niksa Odzak, Ursula Lindauer.
Engineered metal and metal oxide nanoparticles will likely reach natural waters, either directly from use of consumer products, or after those products are discharged into the environment. Dissolution of these nanoparticles under environmental conditions is thus an important process for their effects on aquatic organisms. Toxic effects of some metal ions (e.g. Ag+, Cd2+, Cu2+, Zn2+, etc.) are very well documented. Some nanoparticulate forms of those metals show similar characteristics (e.g. Ag nanoparticles), but it is still not clear whether observed toxic effects are caused by the particulate or dissolved form, or both. It is therefore of extrem importance to accurately measure dissolved metal and metal oxide nanoparticle fractions under different aquatic conditions.
Dissolution of Ag nanoparticles with various coatings, of ZnO nanoparticles and of Cu and CuO nanoparticles is examined in various media. Dissolution of nanoparticles in simple artificial aqueous media was highly dependent on the particle type and coating, but in general only small amounts of Ag nanoparticles were dissolved, while ZnO nanoparticles dissolved almost completely within a time period of several days. We have used for dissolution experiments the DGT technique together with two other, better known techniques (ultrafiltration and dialysis membrane). The results of these techniques were comparable in simple aqueous media.
It is planned to further investigate dissolution of metal and metal oxide nanoparticles in more complex, natural aqueous media. The results of previous study in defined media will be compared with the results achieved using different types of natural water, which are differing in important factors for the dissolution, such as dissolved organic matter, pH, ionic strength.
This project will contribute to the risk assessment of nanoparticles in the environment and thus of nanotechnology.
References
1. Project on Emerging Nanotechnologies. The Nanotechnology Consumer Products Inventory 2006. www.nanotechproject.org/consumer-products.
2. Behra R & Krug H (2008) Nanoecotoxicology - Nanoparticles at large. Nature Nanotechnology 3: 253-254.
3. Navarro E, Baun A, Behra R, Hartmann NB, Filser J, Miao AJ, Quigg A, Santschi PH & Sigg L (2008a) Environmental behavior and ecotoxicity of engineered nanoparticles to algae, plants, and fungi. Ecotoxicology 17: 372-386C.
4. Navarro E, Piccapietra F, Wagner B, Marconi F, Kaegi R, Odzak N, Sigg L & Behra R (2008b) Toxicity of Silver Nanoparticles to Chlamydomonas reinhardtii. Environmental Science & Technology 42: 8959-896.
5. Kühnel D, Busch W, Meißner T, Springer A, Potthoff A, Richter V, Gelinsky M, Scholz S, Schirmer K. (2009). Agglomeration of tungsten carbide nanoparticles in exposure medium does not prevent uptake and toxicity toward a rainbow trout gill cell line. Aquatic Toxicology, 93, 91-99.
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7. Busch, W., Kuhnel, D., Schirmer, K., and Scholz, S. Tungsten carbide cobalt nanoparticles exert hypoxia-like effects on the gene expression level in human keratinocytes. BMC Genomics 11:65.
8. Schwyzer, I., Kaegi, R., Sigg, L., Magrez, A., and Nowack, B., 2011. Influence of the initial state of carbon nanotubes on their colloidal stability under natural conditions. Environ. Poll. 159, 1641-1648.
9. Schwab, F., Bucheli, T. D., Lukhele, L. P., Magrez, A., Nowack, B., Sigg, L., and Knauer, K., 2011. Are carbon nanotube effects on green algae caused by shading and agglomeration? Environ. Sci Technol. 45, 6136-6144.
10. Busch W, Bastian S, Trahorsch U, Iwe M, Kühnel D, Meissner T, Springer A, Gelinsky M, Richter V, Ikonomidou C, Potthoff A, Lehmann I, Schirmer K. (2010). Internalisation of engineered nanoparticles into mammalian cells in vitro: influence of cell type and particle properties. Journal of Nanoparticle Research 13(1), 209-310.
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12. Busch W, Bastian S, Trahorsch U, Iwe M, Kühnel D, Meissner T, Springer A, Gelinsky M, Richter V, Ikonomidou C, Potthoff A, Lehmann I, Schirmer K. (2010). Internalisation of engineered nanoparticles into mammalian cells in vitro: influence of cell type and particle properties. J. Nanoparticle Res. 13(1), 209-310.
13. Meissner T, Kühnel D, Busch W, Oswald S, Richter V, Schirmer K, Potthoff A. (2010). Physical-chemical characterization of tungsten carbide nanoparticles as a basis for toxicological investigations. Nanotoxicology 4(2), 196-206.
14. Piccapietra, F., Sigg, L., and Behra, R., 2012. Colloidal stability of carbonate-coated silver nanoparticles in synthetic and natural freshwater Environ. Sci Technol. 46, 818-825.
15. Scheringer, M., MacLeod, M., Behra, R., Sigg, L. & Hungerbühler, K. Environmental risks associated with nanoparticulate silver used as biocide. Household and Personal Care today 1, 34-37 (2010).
16. Piccapietra, F., Gil-Allué, C., Sigg, L., and Behra, R., 2012. Intracellular silver accumulation in Chlamydomonas reinhardtii upon exposure to carbonate coated silver nanoparticles and silver nitrate. Environ. Sci Technol. 46, 7390-7397.
17. Schwyzer, I., Kaegi, R., Sigg, L., Smajda, R., Magrez, A., and Nowack, B., 2012. Long-term colloidal stability of 10 carbon nanotube types in the absence/presence of humic acid and calcium. Environ. Poll. 169, 64-73.