USEPA, Office of Research and Development, National … Dubey Department of Environmental Health,...
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Current understanding on potential toxicity of engineered nanomaterials on the aquatic
NANOPARTICLES CHARACTERIZATION
microorganisms is limited for risk assessment and management. Here we show
MetPLATE™ bioassay as an effective and rapid screening tool to test for potential
aquatic toxicity of various metal-based nanoparticles (NPs). MetPLATE™ bioassay is a
simple and cost effective test that uses a mutant strain of Escherichia coli assay - the
enzymatic activity of which is measured as the percentage inhibition compared to the
negative control. Toxicity of NPs was also compared to their corresponding ionic
counterparts. The NPs tested were (1) Citrate coated nAg (Citrate-nAg), (2)
polyvinylpyrrolidone coated nAg (PVP-nAg), (3) uncoated nZnO, (4) uncoated nTiO2,
and (5) 1-Octadecylamine coated CdSe Quantum Dots (CdSe QDs). Citrate-nAg was
further fractionated into clean Citrate-nAg, unclean Citrate-nAg and permeate using
tangential flow filtration (TFF) system to eliminate residual impurities including ions
from the stock Citrate-nAg suspension and differentiate between ionic- versus nano-
specific toxicity. Our results demonstrate that nAg, nZnO, and CdSe QDs were
relatively less toxic than their corresponding ionic forms tested, while both NPs and
ions of TiO2 were not toxic at as high as 2.5 g/L to the MetPLATE™ bacteria. Overall,
the toxicity followed the following trend: CdCl2 > AgNO3 > PVP-nAg > unclean Citrate-
nAg > clean Citrate-nAg > ZnSO4 > nZnO > CdSe QDs > nTiO2/TiO2. As previously
recommended, these results also highlight the importance of fractionating metal NPs
for better toxicity assessment. Although we found that the evaluated NPs were
relatively less toxic than their ionic forms, we caveat to disposing NPs into the receiving
waters as physicochemical properties of NPs may change with changeable water
chem
RESEARCH OBJECTIVES
istry which may promote NPs toxicity.
1
ASSESSMENT OF AQUATIC TOXICITY OF METAL NANOPARTICLES USING
METPLATE™ BIOASSAY AS A RAPID SCREENING TOOL
1Lok R. Pokhrel, 1Thilini Silva, 2Amro M. El Badawy, 3Thabet M. Tolaymat, 1Brajesh Dubey Department of Environmental Health, College of Public Health, East Tennessee State University, Johnson City, TN 37614, USA; 2Department of Civil & Environmental Engineering, University of Cincinnati. Cincinnati, OH,
USA; 3USEPA, Office of Research and Development, National Risk Management Laboratory, 26 West Martin Luther King Drive, Cincinnati, OH 45224, USA. Correspondence: Brajesh Dubey, [email protected].
(1) To evaluate the sensitivity of MetPLATE™ bioassay to the toxicity
of various metal-based NPs suspensions;
(2) To compare the toxicity of NPs with their corresponding ionic
counterparts; and
(3) To examine the toxicity of Ag by fractionating it as unclean Citrate-
nAg (as-synthesized), clean Citrate-nAg (cleaned via tangential
flow filtration system), permeate (obtained as a filtrate) and ionic
Ag (as AgNO3) to differentiate ionic- versus nano-specific toxicity.
1. The dilution matrix (i.e., moderately hard water) and all NPs
suspensions had acceptable characteristics (pH = 6 - 7.5) as
required by the MetPLATE protocol.
2. Our results showed that MetPLATE test can be used as an
effective and rapid screening tool for the toxicity assessment of
wide variety of metal-based NPs.
3. PVP-nAg was the most toxic among all the tested NPs types,
while clean Citrate-nAg showed relatively similar toxicity as that
of unclean Citrate-nAg.
4. Both ions and NPs of TiO2 was nontoxic even at a concentration
as high as 2.5 g/L.
5. CdSe Quantum Dots showed only 34.42% inhibition of
MetPLATE bacteria at 100mg/L.
6. Ions of Cd and Ag were most toxic among all the tested materials
including NPs.
7. MetPLATE toxicity showed the following trend:
CdCl2 > AgNO3 > PVP-nAg > unclean Citrate-nAg > clean
Citrate-nAg > ZnSO4 > nZnO > CdSe QDs > nTiO2/TiO2.
8. Residual Ag ions in unclean Citrate-nAg slightly enhanced the
toxicity as revealed by fractionating Citrate-nAg using Tangential
Flow Filtrtaion (TFF) process.
9. Although particle size and surface charge were not adequate to
explain the nanotoxicity observed, these results however clearly
indicate towards lower nano-specific toxicity than ionic toxicity
of the tested materials.
Conclusion
1. As understanding on the mechanisms by which nanoparticles induce
toxicity is limited, our study indicates that inhibition of β-
galactosidase enzyme activity using MetPLATE bioassay can be an
important consideration for rapid toxicity assessment.
2. Toxicity varied with coatings for nAg, but overall toxicity could not
be explained by particle size, surface charge and coatings.
3. Ionic toxicity was higher than nanotoxicity.
4. Fractionating nanoparticles from ions using diafiltration or field
flow fractionation is crucial to distinguish nano- versus ionic
toxicity.
Wavelength (nm)
300 400 500 600 700
Ab
so
rban
ce
(a.u
.)
0
1
2
3
4
nTiO2
Permeate
AgNO3
Clean Citrate-nAg
Unclean Citrate-nAg
nZnO
Quantum Dots
PVP-nAg
ABSTRACT
β- galactosidase mediated conversion of CPRG into Chlorophenol Red
MetPLATE Procedure
Acknowledgments: We would like to thank the Gordon Research Conference; Office of Research
and Sponsored Programs, ETSU; and School of Graduate Studies, ETSU for providing financial support to Lok R. Pokhrel to attend the Gordon Research Conference.
RESULTS
Fig. 1. Kros Flo Tangential Flow Filtration process for nanoparticles purification
PVP-nAg
nTiO2
Citrate-nAg
Fig. 4. Representative UV-Vis spectra of tested nanoparticles Fig. 5. Particle size distribution of tested nanoparticles
Fig. 2. Representative UV-Vis spectra of tested nanoparticles Fig. 3. TEM images of nanoparticles
Fig. 7. MetPLATE test showing color variability in CPRG as impacted by different
nanoparticles and ionic salts tested
Fig. 6. Comparison of EC50 values showing variability in MetPLATE toxicity for
various nanoparticles and their ionic counterparts
METPLATE BIOASSAY
+ve Control
-ve Control