Grantee Research Project Results
2006 Progress Report: The Bioavailability, Toxicity, and Trophic Transfer of Manufactured ZnO Nanoparticles: A View from the Bottom
EPA Grant Number: R832530Title: The Bioavailability, Toxicity, and Trophic Transfer of Manufactured ZnO Nanoparticles: A View from the Bottom
Investigators: Bertsch, Paul M. , Neal, Andrew , Jackson, Brian , Williams, Phillip , Glenn, Travis
Institution: University of Kentucky , Dartmouth College
Current Institution: Savannah River Ecology Laboratory , Dartmouth College , University of Georgia
EPA Project Officer: Hahn, Intaek
Project Period: October 1, 2005 through September 30, 2008 (Extended to September 30, 2009)
Project Period Covered by this Report: October 1, 2005 through September 30, 2006
Project Amount: $363,680
RFA: Exploratory Research: Nanotechnology Research Grants Investigating Environmental and Human Health Effects of Manufactured Nanomaterials: A Joint Research Solicitation - EPA, NSF, NIOSH (2005) RFA Text | Recipients Lists
Research Category: Nanotechnology , Safer Chemicals
Objective:
The overall objectives of our research are to evaluate: (1) the bioavailability and toxicity of manufactured nanoparticles (ZnO-np) as a function of particle size to the model bacteria, Burkholderia vietnamiensis PR1301 and the model detritivore Caenorhabditis elegans as referenced against aqueous Zinc (Zn2+(aq)); (2) the ability of manufactured ZnO–np to be transferred from one trophic level to the next as assessed in the simple food chain consisting of pre-exposed PR1 and C. elegans;and (3) the synergistic or antagonistic effects of manufactured ZnO-np on the toxicity of Cu2+ to PR1 and C. elegans. These three overall objectives will be approached in the context of the following four hypotheses:
Hypothesis 1: The bioavailability and toxicity of manufactured ZnO-np increases with decreasing particle size (i.e., 3 nm vs. 80 nm).
Hypothesis 2: The toxicity of ZnO-np to PR1 and C. elegans is lower than an equivalent concentration of Zn2+(aq).
Hypothesis 3: The bioavailability and toxicity of ZnO-np introduced via trophic transfer differs from direct exposure.
Hypothesis 4: ZnO-np alter the bioavailability and toxicity of dissolved metals.
Progress Summary:
First year investigations have focused on the characterization of the ZnO-np under physicochemical conditions of the exposure experiments, in parallel with initial experiments examining the bioavailability and toxicity of ZnO-np to B. vietnamensis and C. elegans as referenced to aqueous Zn2+. In agreement with the aim of the project to evaluate trophic transfer, research on detritivores has been expanded to include the model earthworm Eisenia fetida.
The major findings are summarized as follows:
Characterization
- The sole counter anion used to stabilize the suspensions was acetate. Stability of the ZnO-np suspensions depends on acetate concentration and pH.
- There are multiple populations of acetate—free, exchangeable, and structural—associated with the ZnO-np.
- Acetate controls the ZnO-np structure and reactivity.
- Removal of one or more of the acetate populations leads to flocculation/aggregation of
ZnO-np primary particles and irreversible structural transformations that promote surface reactivity.
Nanoparticles-bacteria interactions
- The presence of acetate in ZnO-np suspension was shown to affect Burkholderia vietnamiensis PR1301 (PR1) growth.
- The acetate anion is cytotoxic at pH 5 but can serve as a non-specific C-source at higher pH (6-7).
- PR1301 exhibited a similar pH-dependent toxicity with Zn2+(aq) and ZnO-np.
- The EC50 values of Zn2+(aq) and ZnO-np derived from growth curves are different at pH 6, with ZnO-np less toxic than Zn2+(aq). There were no differences at pH 7.
- While the overall trends of toxicity were very similar, the underlying mechanisms are likely different.
- The microbial acetate utilization influences the stability of ZnO-np over time; there was significant flocculation of primary particles at 24 hours associated with cell growth.
- Aggregates do not form in the absence of PR1, suggesting a biologically controlled induction. Aggregates could result from acetate degradation and/or exopolymer secretion. Ongoing studies are examining both mechanisms.
Nanoparticles-nematode interactions
- Acetate was not toxic to C. elegans.
- There was no significant difference in general toxicity to C. elegans between ZnO-np and Zn2+(aq); yet, as with the bacteria, the mechanism of toxicity is likely different.
- Zn spatial distribution in ZnO-np and Zn2+(aq) exposed nematodes showed similar patterns with the areas of maximum intensity spreading out through the nematode as exposure concentration increases.
- The maximum intensities in Zn2+(aq) exposed nematode are approximately twice as high as in ZnO-np exposed nematode, which may suggest their different bioavailability.
- Preliminary imaging experiments demonstrated that exposure to either Zn2+(aq) or ZnO-np results in greater distribution and co-localization of Zn and mtl2::GFP expression.
Nanoparticles-earthworm interactions
- After 14 days of exposure to artificial soil containing 1000 μg g-1 Zn, there was significantly more Zn accumulated in ZnO-np exposed worms.
- Zn-rich granules were detected in control worms but not in exposed worms. The Zn-rich granules in control worms were accompanied by an elevated concentration of Zn-bound metallothienin in earthworm tissues and are believed to represent a Zn storage mechanism in response to Zn deficient conditions in the artificial soil (85-95% of the total Zn was removed from control soils).
- The zinc tissue and molecular distribution in ZnO-np and Zn2+(aq) exposed earthworms were similar, with high intensity in the gut area likely to be associated with ingested sediments. The absence of granules and Zn-MT in ZnO-np treatment suggests the bioavailability of Zn.
Future Activities:
These results provided a framework for the second year of the project. Although characterization of the ZnO-np will continue, research efforts will be geared toward investigating differences in toxicity mechanisms of ZnCl2 and ZnO-np to B. vietnamensis and C. elegans. Experiments with the 80 nm ZnO nanoparticles will be initiated in addition to studies on the bioavailability and toxicity of ZnO nanoparticles introduced via trophic transfer as opposed to direct exposure. Exposure experiments with Cu2+ will also be started.
Journal Articles:
No journal articles submitted with this report: View all 24 publications for this projectSupplemental Keywords:
nanotechnology, metal oxides, nanoparticles, adsorption, absorption, exposure, ecological effects, bioavailability, trophic transfer, organism, cellular, population, enzymes, metals, nanoparticles, environmental chemistry, analytical, ecosystem protection/environmental exposure & risk, RFA, ecological effects-environmental exposure & risk, ecological monitoring, ecological risk assessment, ecology and ecosystems, ecosystem protection, ecosystem/assessment/indicators, environmental microbiology, risk assessment, exploratory research environmental biology, ecological assessment, ecological impacts, ecosystem assessment, environmental fate,, Health, Scientific Discipline, Water, Environmental Chemistry, Health Risk Assessment, Risk Assessments, Biochemistry, Engineering, Chemistry, & Physics, fate and transport, food chain, trophic transfer, nanotechnology, carbon fullerene, bioavailability, particle exposure, environmental contaminants, nanomaterials, ambient particle health effects, human exposure, engineered nanomaterials, biochemical researchRelevant Websites:
Progress 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.