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Systems-based approach for assessing hazard and risk of manufactured nanomaterials and non-human species and ecosystems


This task focuses on the ecotoxicology of manufactured nanomaterials (particles or fibers that have at least one dimension between 1 and 100 nm). These materials are of immediate concern to regulatory offices because they exhibit properties not observed in their traditional bulk form, are being developed and incorporated into products at a rapid rate, and require novel and non-standardized test approaches due to their particulate nature. In addition, due to their small size alone, these particles may be taken up and translocated via mechanisms very different from those typical of truly soluble substances. The inherent properties that determine toxicity might also differ significantly from traditional, soluble chemicals, for example surface plasmon activity that occurs only at the nanoscale and allows surface electrons to behave in unpredictably. These unique properties have the potential to elicit system-level responses at every level of organization, from the molecular/sub-cellular to the ecosystem (for example carbon or nutrient cycling). Many of these responses could be mitigated by variation in size or other material attributes that can be manipulated during production (e.g., using green chemistry principles).

This task is designed to serve two broad purposes: 1) to develop methods for working with a broad range of nanomaterials with the immediate goal of providing test guidance to regulatory Offices, primarily OCSPP; 2) to characterize and quantify toxicity of various nanomaterials in freshwater, marine, and terrestrial systems to provide Offices with basic toxicity information, but also to develop a basis for investigating chronic toxicity, mechanisms of action, toxic pathways, and initiating events, and ultimately to develop predictive tools to preclude extensive plant and animal testing.

Rationale and Research Approach:

Understanding how nanomaterial exposure and toxicity differ from soluble chemicals is critical for elucidating their ecological hazard and risk, and for determining whether their behaviors and effects across levels of biocomplexity (individuals, populations, communities, intact ecosystems), are inherently different from traditional chemicals. It has been well established that, because of their particulate and fibrous nature, nanomaterial exposure occurs via colloidal suspensions of insoluble particles rather than true solutions, and that these suspensions typically behave in a manner that is not addressed in standardized test methods. The particles agglomerate (change size), settle, and exhibit continuously-changing surface properties. This level of variability is also seen in dry testing (e.g., terrestrial systems) even where wet suspensions are not used in development of exposure media. Preliminary research suggests that it is generally not possible to limit variation in critical nanomaterial properties over the duration of most tests and that characterization at relatively frequent intervals (relative to soluble chemicals) is necessary to fully quantify exposure. For example, titanium dioxide nanoparticles tend to agglomerate over 24-hour renewal periods in aquatic media, changing in average particle size from 200 nm to 2000 nm. This behavior results in settling of material from the water column and an order-of-magnitude reduction in the total number of particles present, both factors that greatly reduce exposure.

Understanding these behaviors allow us to identify susceptible targets in ecosystems, for example benthic communities in aquatic and marine systems. In addition to presenting profound challenges for regulatory testing, these behaviors are also not easily documented and typically require methods and approaches that have yet to be developed or standardized. These challenges are made more complex by the nearly unlimited variation in the as-produced form of nanomaterials, including particle size (and size distribution), surface coating, material combinations (e.g. CdSe quantum dots), functionalization (e.g. addition of hydroxyl or carboxyl groups), and pre-application "packaging" (e.g. stabilization in matrices that will be removed during product incorporation). All of these modifications can affect the behaviors described above.

Our research approach will be to focus initially on developing methods for preparing test media and approaches for characterizing nanomaterials in media during testing. This effort will address media development for freshwater, marine, and terrestrial test systems, each of which presents novel problems, including highly variable agglomeration behavior due to varying salinity and the lack of methods for detecting nanomaterials in terrestrial and other complex media (e.g. sediments). Initially, these efforts will focus on TiO2, nano-silver, and carbon nanotubes. These materials are representative of metal oxides, metals, and carbon-based materials, respectively, and may provide a basis for broad application of developed methods across these material classes. Tested media preparation approaches will include stirring, sonication, and use of solvents, dispersion, and stabilizing agents. In the latter case, naturally-occurring stabilizing agents such as dissolved organic carbon will be tested, as this will also provide insight into nanomaterial behavior in natural environments. Characterization approaches will include particle sizing (dynamic laser-light scattering), fractionation, near infrared fluorescence detection, visible and electron microscopy (with back-scattering detection for material identification), as well as material extraction and isolation methods.

The hypothesis underlying this effort is that nanomaterials will require novel testing approaches. Our aim is to develop these approaches and to provide guidance to Offices on how nanomaterial testing should be undertaken to accurately represent the potential hazard of nanomaterials and their applications. Concurrent with our media-development efforts, we will characterize and quantify the toxicity of the materials being studied. These efforts are closely aligned, as nanomaterial behavior in media is changed by the presence of organisms (due to alteration of ionic strength, pH, and other critical parameters that are affected by or derived from organism exudates). The necessity of developing test media with organisms present allows for immediate preliminary assessment of toxicity and identification of potential test endpoints. The refinement of media preparation methods, material characterization approaches, and exposure-response relationships will occur simultaneously. This approach will also provide the opportunity to continuously identify and evaluate material characteristics (inherent properties) that are most closely related to organism responses, e.g. particle size effects on the surface area available for material-target interactions or release of soluble, toxic species. The responses of organisms will be used to infer, and guide the further investigation of modes and mechanisms of action and the toxic events that initiate adverse outcome pathways. For a subset of tested materials and systems, ingestion, uptake, and food-chain exposure will be investigated, and where possible, other effects on ecological processes such as benthic community interactions or nutrient cycling (in simple, model exposure systems) will be identified and characterized. A likely focus will be investigating the role of reactive oxygen species that are likely to be produced on the surface of many nano-scale materials, and are well-understood relative to initiating events and AOPs. The hypothesis underlying this effort is that nanomaterials will exhibit some level of toxicity and that organism responses might be unique relative to soluble chemicals.

Our aim is to provide data to regulatory Offices on toxicity and eco-toxicity of selected nanomaterials, and to identify and characterize modes and mechanisms of action where possible. These near-term efforts (FY12/13) will focus on a limited number of nanomaterials (carbon nanotubes, nano-silver, and nano-TiO2). These materials are the focus of ongoing research and were selected based on Office input, development of the ORD Nanomaterials Research Strategy and ORD involvement in Organisation for Economic Cooperation and Development efforts. A goal of this effort to develop a suite of techniques, approaches, and methods that allow ORD to rapidly address immediate Office needs across a broad range of nanomaterials. These efforts will be undertaken in close collaboration and integration with CSS, Systems Task 2.6.1 and Tasks in the CSS Inherency Topic. These efforts will also provide the basis for out-year (FY14/15/16) deeper investigation of chronic and indirect effects, material and environmental properties that mitigate toxicity or other effects (and inform green and sustainable nanomaterial development), and full modeling of dosimetry, and identification and development of alternate assays that might accelerate testing and avoid material-by-material plant or animal testing. The scope of these out-year efforts is dependent on levels of funding and other forms of support and the results of the near-term efforts described above.

MED Scientists:

Dale Hoff
Terry Highland
Teresa Norberg-King
Larry Heinis


Li, S., L.K. Wallis, H. Ma, and S.A. Diamond. 2014. Phototoxicity of TiO2 nanoparticles to a freshwater benthic amphipod: Are benthic systems at risk? Science of the Total Environment 466-467:800-808.

Li, S., F. Irin, F.O. Atore, M.J. Green, and J.E. Canas-Carrell. 2013. Determination of multi-walled carbon nanotube bioaccumulation in earthworms measured by a microwave-based detection technique. Science of the Total Environment 445:9-13.

Ma, H. and S.A. Diamond. 2013. Phototoxicity of TiO2 nanoparticles to zebrafish (Danio rerio) is dependent on life stage. Environmental Toxicology and Chemistry 32:2139-2143.

Hoheisel, S.M., S.A. Diamond, and D.R. Mount. 2012. Comparison of nanosilver and ionic silver toxicity in Daphnia magna and Pimephales promelas. Environmental Toxicology and Chemistry 31:2557-2563.

Ma, H., A. Brennan, and S.A. Diamond. 2012. Photocatalytic ROS production and phototoxicity of titanium dioxide nanoparticles is dependent on solar UV radiation spectrum. Environmental Toxicology and Chemistry 31: 2099-2107.

Ma, H., A. Brennan, and S.A. Diamond. 2012. Phototoxicity of TiO2 nanoparticles under solar radiation to two aquatic species: Daphnia magna and Japanese Medaka. Environmental Toxicology and Chemistry 31:1621-1629.

Nowack, B., J.F. Ranville, S. Diamond, J.A. Gallego-Urrea, C. Metcalfe, J. Rose, N. Horne, A.A. Koelmans, and S.J. Klaine. 2012. Potential scenarios for nanomaterial release and subsequent alteration in the environment. Environmental Toxicology and Chemistry 31:50-59.

Diamond, S.A., D. Utterback, C.P. Andersen, R. Burgess, S. Hirano, K. Ho, C. Ingersoll, M.G. Johnson, A.J. Kennedy, D.R. Mount, J.W. Nichols, P. Pandard, P. Rygiewicz, J.J. Scott-Fordsmand, and K. Stewart. 2009. Review of OECD/OPPTS-harmonized and OPPTS ecotoxicity test guidelines for their applicability to manufactured nanomaterials. EPA Report, EPA/600/R-09/065, 13 pp.

Morris, J., R. Wentsel, M. Conlon, M.J. Davis, S.A. Diamond, K. Dreher, M. Gwinn, T.J. Holdsworth, K. Houck, E. Hubal, D. McKinney, D.R. Mount, C.M. Nunez, N. Savage, C.R. Shoaf, B. Walton, and E. Weber. 2009. Nanomaterial research strategy. EPA Report, EPA/620/K-09/011, 44 pp.

Expected Products:




Sep 30, 2013

(1) Guidance on best available methods and approaches for eco-testing and characterization of selected nanomaterials.

Dale Hoff

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