Grantee Research Project Results
2007 Progress Report: Microbial Impacts of Engineered Nanoparticles
EPA Grant Number: R832534Title: Microbial Impacts of Engineered Nanoparticles
Investigators: Alvarez, Pedro J. , Wiesner, Mark R.
Institution: Rice University
Current Institution: Rice University , Duke University
EPA Project Officer: Hahn, Intaek
Project Period: December 15, 2005 through December 14, 2008
Project Period Covered by this Report: December 15, 2006 through December 14, 2007
Project Amount: $375,000
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:
Responsible usage of nanomaterials in commercial products and environmental applications, and prudent management of the associated risks, require an understanding of nanoparticle mobility, bioavailability and ecotoxicology. This project seeks to elucidate processes governing the transport and microbial impacts of two classes of catalytic nanomaterials in soil-water systems: fullerenes and metallic nanoparticles (e.g., TiO2). Specific tasks include to
- characterize nanomaterials size, shape, functionality, reactivity, aggregation, deposition potential, and bioavailability;
- screen nanomaterials of varying sizes and properties for bactericidal activity;
- discern bacterial physiologic characteristics that confer resistance (or susceptibility) to catalytic nanomaterials;
- evaluate the potential for fullerene biotransformation by reference bacteria and fungi; and
- assess the impact of simulated nanomaterial releases on microbial diversity and community structure.
The relevance of this work to the EPA mission is related to the fact that microorganisms are the foundation of all ecosystems and are often the basis for food chains and the main agents of biogeochemical cycles. Thus, understanding their interactions with engineered nanomaterials is important to ensure that nanotechnology improves material and social conditions without exceeding the ecological capabilities that support them. At the conclusion of this project, we will have an improved understanding of the chemical and physical factors that control nanoparticle mobility and bioavailability, and their impacts on microbial activities, diversity and community structure. This will benefit risk assessment and management efforts, and may contribute indirectly to the development of nanotechnology-based disinfection and biofouling control strategies.
Progress Summary:
Antibacterial activity of fullerene water suspensions (nC60) is not due to ROS-mediated damage
The cytotoxic and antibacterial properties of nC60, a fullerene water suspension, have been attributed to photocatalytically-generated reactive oxygen species (ROS). However, in this work, neither ROS production nor ROS-mediated damage are found in nC60-exposed bacteria. Furthermore, the colorimetric methods used to evaluate ROS production and damage are confounded by interactions between nC60 and the reagents, yielding false positives. On the contrary, we propose that nC60 exerts ROS-independent oxidative stress, with evidence of protein oxidation, changes in bacterial membrane potential, and interruption of cellular respiration. thus These findings reconcile conflicting results in the literature. Unlike previous reported nanomaterial antibacterial mechanisms of ROS generation from metal oxides or leaching of toxic ions from nanosilver and quantum dots, this nanoparticle directly oxidizes the cell.
Comparative photoactivity and microbial toxicity of fullerenes and titanium dioxide
Nanosize anatase (TiO2) and fullerenes are two nanomaterials whose photoactivity has been scrutinized for its possible involvement in cytotoxicity as well as its potential applications. We compared the singlet oxygen (1O2), superoxide (O2-), and hydroxyl radical (OH•) generation capacity of different fullerene water suspensions to that of a nano-TiO2 (Degussa P25) in both ultra-pure water and a microbial growth medium. In ultra-pure water, nano-TiO2 produced primarily OH•, and fullerol produced only 1O2. In the mineral medium, nano-TiO2’s OH• were quenched, but O2- production was enhanced. More 1O2 was generated by PVP/C60 and fullerol than by nano-TiO2, whereas O2- was generated by nano-TiO2, PVP/C60 and fullerol, in order of decreasing activity. However, in none of the media, the C60 aggregates obtained by extensive stirring (Aq/nC60) or exchange of solvent (THF/nC60) were able to produce reactive oxygen species (ROS). Additional tests showed that nano-TiO2 exhibited antibacterial activity in presence of light but not in the dark. Other nanoparticles were either not antibacterial at all (fullerol, Aq/nC60) or had similar antibacterial activity (THF/nC60, and PVP/C60) regardless of light. These results demonstrate the lack of correlation between ROS production and toxicity for the fullerenes, and suggest that O2- contributed to nano-TiO2 phototoxicity. Based on ROS speciation in the two media tested and the oxidation potential of each ROS, we recommend that fullerol and encapsulated fullerenes be applied in biomedical applications and to accompany water treatment targeting more specifically pollutants or microorganisms more sensitive to either superoxide or singlet oxygen, whereas nano-TiO2 would be more efficient for water treatment in general, involving UV or solar energy, to enhance disinfection or contaminant oxidation.
Mechanisms of reactive oxygen production by fullerene suspensions in water.
Buckminsterfullerene (C60) is a known photosensitizer that produces reactive oxygen species (ROS) in the presence of light. However, its properties in relevant environmental systems are still not well understood or modeled. This is of the utmost importance if a predictive knowledge of the impact of released nanomaterials is ever to be obtained. In this study, production of singlet oxygen and superoxide by UV photosensitization of colloidal aggregates of C60 in water was measured by electron paramagnetic resonance (EPR) and reduction followed by spectrophotometric detection of the compound XTT. Both singlet oxygen and superoxide were generated by fullerol suspensions while neither was detected in the aqu/nC60 suspensions. These trends can be mechanistically described when taken in the context of those observed in organic solvents and predicted by quantum yields. A mechanism for the behavior or fullerene based aggregates in water was proposed based on these considerations.
Inactivation of Bacteriophages via Photosensitization of Fullerol Nanoparticles
The production of reactive oxygen species through UV photosensitization of polyhydroxylated fullerene (fullerol) is shown to enhance viral inactivation rates. The production of both singlet oxygen and superoxide by fullerol in the presence of UV light is confirmed via two unique methods: electron paramagnetic resonance (EPR) and reduction of nitro blue tetrazolium (NBT). These findings build on previous results both in the area of fullerene photosensitization and fullerene impact on micro-fauna. Results showed that the first-order MS2 bacteriophage inactivation rate nearly doubled due to the presence of singlet oxygen, and increased by 125% due to singlet oxygen and superoxide as compared with UV-A illumination alone. When fullerol and NADH are present in solution, dark inactivation of viruses occurs at nearly the same rate as that produced by UV-A illumination without nanoparticles. These results suggest a potential for fullerenes to impact microbial populations in both natural and engineered systems ranging from surface waters to disinfection technologies for water and wastewater treatment.
Educational; Achievements:
This project has served as the basis for one PhD dissertation:
E. Matt Hotze (2008). “Mechanisms and applications of reactive oxygen production by fullerene suspensions in water.” PhD Thesis, Duke University.
Journal Articles on this Report : 7 Displayed | Download in RIS Format
Other project views: | All 16 publications | 15 publications in selected types | All 13 journal articles |
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Badireddy AR, Hotze EM, Chellam S, Alvarez P, Wiesner MR. Inactivation of bacteriophages via photosensitization of fullerol nanoparticles. Environmental Science & Technology 2007;41(18):6627-6632. |
R832534 (2007) R832534 (Final) |
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Brunet L, Lyon DY, Zodrow K, Rouch J-C, Caussat B, Serp P, Remigy J-C, Wiesner MR, Alvarez PJJ. Properties of membranes containing semi-dispersed carbon nanotubes. Environmental Engineering Science 2008;25(4):565-576. |
R832534 (2006) R832534 (2007) R832534 (Final) |
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Fang J, Lyon DY, Wiesner MR, Dong J, Alvarez PJJ. Effect of a fullerene water suspension on bacterial phospholipids and membrane phase behavior. Environmental Science & Technology 2007;41(7):2636-2642. |
R832534 (2006) R832534 (2007) R832534 (Final) |
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Hotze EM, Labille J, Alvarez P, Wiesner MR. Mechanisms of photochemistry and reactive oxygen production by fullerene suspensions in water. Environmental Science & Technology 2008;42(11):4175-4180. |
R832534 (2007) R832534 (Final) |
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Klaine SJ, Alvarez PJJ, Batley GE, Fernandes TF, Handy RD, Lyon DY, Mahendra S, McLaughlin MJ, Lead JR. Nanomaterials in the environment: behavior, fate, bioavailability, and effects. Environmental Toxicology and Chemistry 2008;27(9):1825-1851. |
R832534 (2007) R832534 (Final) |
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Lyon DY, Adams LK, Falkner JC, Alvarez PJJ. Antibacterial activity of fullerene water suspensions: effects of preparation method and particle size. Environmental Science & Technology 2006;40(14):4360-4366. |
R832534 (2006) R832534 (2007) R832534 (Final) |
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Lyon DY, Brunet L, Hinkal GW, Wiesner MR, Alvarez PJJ. Antibacterial activity of fullerene water suspensions (nC60) is not due to ROS-mediated damage. Nano Letters 2008;8(5):1539-1543. |
R832534 (2007) R832534 (Final) |
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Supplemental Keywords:
Health, Scientific Discipline, Health Risk Assessment, Environmental Chemistry, Risk Assessments, Ecological Risk Assessment, anthropogenic stress, ecotoxicogenomics, bioavailability, nanotechnology, nanomaterials, microbial 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.