Grantee Research Project Results
2007 Progress Report: Ecotoxicology of Underivatized Fullerenes (C60) in Fish
EPA Grant Number: R833333Title: Ecotoxicology of Underivatized Fullerenes (C60) in Fish
Investigators: Henry, Theodore B. , Sayler, Gary S. , Menn, Fu-Min , Compton, Robert N.
Institution: University of Tennessee
EPA Project Officer: Aja, Hayley
Project Period: May 15, 2007 through May 14, 2010
Project Period Covered by this Report: May 15, 2007 through May 14,2008
Project Amount: $396,807
RFA: Exploratory Research: Nanotechnology Research Grants Investigating Environmental and Human Health Effects of Manufactured Nanomaterials: a Joint Research Solicitation-EPA, NSF, NIOSH, NIEHS (2006) RFA Text | Recipients Lists
Research Category: Safer Chemicals , Nanotechnology
Objective:
Objectives: Investigate characteristics of aqueous C60 aggregates and the impact of dissolved organic material on behavior of nanoparticles. Evaluate bioavailability and toxicity of C60 (both aqueous C60 aggregates and dietary C60) in fish by evaluating changes in gene expression, histopathology, and bioaccumulation of C60 in tissues. The species we will consider are zebrafish Danio rerio and channel catfish Ictalurus punctatus.
Hypotheses: 1) Bioavailability of aqueous C60 aggregates is impacted by nanoparticle characteristics and presence of dissolved organic material; 2) Exposure of fish to C60 can be detected by changes in expression of biomarker genes; and 3) Toxic effects of C60 in fish are evident only after long-term chronic exposure.
Progress Summary:
A major objective of this project period was to investigate the characteristics of aqueous aggregates of C60 and the toxicity of these aggregates in larval zebrafish by evaluating changes in global gene expression. Changes in global gene expression in larval zebrafish exposed to aqueous aggregates of C60 generated by tetrahydrofuran (THF) vehicle or by sonication in water initiated in experiments prior to the reporting period were completed and published. Results indicate that decomposition products of the THF vehicle used to generate C60 aggregates can be responsible for the toxic effects reported in previous investigations. Aggregates generated by stirring and sonication in water were not toxic and there was no evidence of induction of genes coding for enzymes involved in protection from oxidative substances.Characterization of aqueous aggregates of C60 is an important objective of this project and was addressed during this period. C60 nanoparticles were prepared by THF vehicle and by water stirring/sonication and were characterized by size, zeta-potential, and absorbance in the 300-900 nanometer range. Particle size and zeta-potential were measured with a Brookhaven ZetaPlus instrument, which uses dynamic light scattering to obtain size distributions of the nanoparticles and electrophoretic measurements to determine the zeta-potential. The absorbance measurements were made with a HP 845x UV-Visible spectrophotometer.
Initial work has shown that C60 aggregates in water typically range from 100 to 500 nm in diameter, with a zeta-potential ranging from –20 to –30 mV. The UV-Vis spectra shows peaks shifted slightly from pure C60, as seen in previously published reports. Current work involves characterizing these particles in the presence of a model organic toxicant. Preparations with different concentrations of C60 aggregates with and without the model toxicant were examined and the presence of the toxicant corresponded to larger particles with smaller zeta-potentials indicating interactions between the nanoparticle and the toxicant. Detection of the presence of an additional absorption peak in the C60/toxicant spectra is presently inconclusive because of the very low concentrations considered to date. Samples were allowed to settle in the dark for 2-6 weeks and subsequent analyses demonstrated particles of smaller size and zeta-potential, which was consistent with the visual observation that particle sedimentation occurred over time. These experiments are ongoing.
A significant component of this project is to evaluate the bioaccumulation and toxicity of C60 in fish and this objective requires use of fish larger than zebrafish to provide sufficient amounts of tissue for analysis. This component of the project was addressed by preparing the wet lab facilities at UT for channel catfish to be exposed to C60 through the diet. Twelve fiberglass tanks (115 cm diameter X 50 cm deep) were plumbed with flow-through water in preparation for experiments. Experimental design includes four conditions (thee treatment and one control) each with three replicate tanks. Experiments to investigate the bioaccumulation and toxicity of C60 in channel catfish exposed through the diet will be initiated during the next reporting period.
Investigation of the toxicity of dietary C60 was added to an experiment conducted at the University of Plymouth designed to examine the effects of dietary exposure to single walled carbon nanotubes (SWCNTs) in rainbow trout. This 61-d dietary exposure included a 500 mg SWCNT/kg food, a 500 mg C60/kg food, and a control (no carbon nanoparticles CNPs). The C60 used in this experiment was obtained from UT, where the characteristics and purity of the material were confirmed, and is the same material that has been used in toxicity tests conducted at UT as part of this project. This represents the first attempt to directly integrate nanotoxicology investigations in fish between laboratories in the U.S. and the UK, and serves as a mechanism for obtaining results that are directly comparable. No fish died during the exposure and no alterations in behavior or gross appearance were observed among treatments. Presently, samples are being analyzed to determine the effects of exposure on whole body systems including: hematology, osmoregulation, tissue histopathology, and oxidative stress.
Future Activities:
Methods to extract and quantify C60 from rainbow trout tissues are underway to allow bioaccumulation among tissues to be investigated. The uptake and bioaccumulation of nanoparticles in organisms remains an important question in nanotoxicology. The techniques under development may be used in the future to assess C60 burdens in fish to provide recommendations regarding human consumption of wild fish.
The focus of this project does not deviate significantly from the original project objectives; however, we do anticipate the addition of specific experiments that have become relevant during the course of the investigations of present reporting period. These include investigating the effects of aqueous C60 aggregates on the bioavailability and toxicity of model contaminants and how these contaminants alter aggregate characteristics. We will continue to integrate our research with international nanotoxicology efforts, and in particular those that are ongoing and under development at the University of Plymouth. We anticipate continuing experiments with zebrafish for most of the next reporting period and beginning experiments with channel catfish towards the end of the reporting period when young juvenile channel catfish become available for experimentation.
Journal Articles on this Report : 1 Displayed | Download in RIS Format
Other project views: | All 25 publications | 6 publications in selected types | All 6 journal articles |
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Type | Citation | ||
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Henry TB, Menn FM, Fleming JT, Wilgus J, Compton RN, Sayler GS. Attributing effects of aqueous C60 nano-aggregates to tetrahydrofuran decomposition products in larval zebrafish by assessment of gene expression. Environmental Health Perspectives. 2007;115(7):1059-65. |
R833333 (2007) R833333 (2008) |
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Supplemental Keywords:
Health, PHYSICAL ASPECTS, Scientific Discipline, ENVIRONMENTAL MANAGEMENT, Health Risk Assessment, Risk Assessments, Physical Processes, Biochemistry, Risk Assessment, biological pathways, bioavailability, exposure, genetic analysis, nanotechnology, carbon fullerene, human exposure, nanomaterials, fish model, toxicologic assessment, histopathology, nanoparticle toxicity, carcinogenic, human health riskProgress and Final Reports:
Original AbstractThe perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Conclusions drawn by the principal investigators have not been reviewed by the Agency.