Ce

l
View Online

10 eb
Journal of ra

Volume 10 | Number 6 | 2008

ye tin
a g

&
Environmental rs

NE RO
IM
Analytical Abstracts…

W VED
P
Analytical Abstracts covers all
Monitoring
Cutting-Edge Research on Environmental Processes & Impacts
areas of analytical and
bioanalytical science, including www.rsc.org/jem Volume 10 | Number 6 | June 2008 | Pages 685–784
Downloaded by University of Massachusetts - Amherst on 01 December 2010

Journal of Environmental Monitoring

the latest applications and
Published on 13 May 2008 on http://pubs.rsc.org | doi:10.1039/B805998E

cutting edge techniques.

The database is updated
weekly and spans 25 years of
research sourced from over 100
international journals.

…the first stop for analytical scientists

New and improved features include:

Improved search features with a basic search and optional

advanced searches by index term and bibliographic data.
You can also browse by subject area.

Results can be sorted and displayed in different formats

and have additional information embedded in the text
accessed via pop up boxes.

Results can be exported to reference

management software and to email.
06020716

www.rsc.org/aa ISSN 1464-0325

Pages 685–784

Registered Charity Number 207890

Editorial Hot Article
Stephen M. Mudge Jin et al.
Is the use of biofuels environmentally Nanoparticle uptake, translocation,
sound or ethical? and accumulation in pumpkin plants 1464-0325(2008)10:6;1-7
 COMMUNICATION
Uptake, translocation, and accumulation of manufactured iron oxide
nanoparticles by pumpkin plants
Hao Zhu,a Jie Han,b John Q. Xiaoa and Yan Jin*b
Received 9th April 2008, Accepted 29th April 2008
First published as an Advance Article on the web 13th May 2008
DOI: 10.1039/b805998e
Rapid development and application of nanomaterials and
nanotechnology make assessment of their potential health and
environmental impacts on humans, non-human biota, and ecosys-
Downloaded by University of Massachusetts - Amherst on 01 December 2010
tems imperative. Here we show that pumpkin plants (Cucurbita
Published on 13 May 2008 on http://pubs.rsc.org | doi:10.1039/B805998E
maxima), grown in an aqueous medium containing magnetite
(Fe3O4) nanoparticles, can absorb, translocate, and accumulate the
particles in the plant tissues. These results suggest that plants, as an
important component of the environmental and ecological systems,
need to be included when evaluating the overall fate, transport and
exposure pathways of nanoparticles in the environment.
Nanotechnology is becoming a major force of the future economy
and continues to attract increasing investment from governments and
industry worldwide. The potential impacts of nanotechnology have
been predicted to far exceed the Industrial Revolution, generating
a $1 trillion market by 2015 and employing 2 million workers
worldwide.1,2 Promising applications in a wide variety of areas using
newly engineered or enhanced materials in the form of nanosized
particles (typically defined as particles smaller than 100 nm) are being
developed rapidly. More than 300 nanotechnology products have
already appeared on the market.3 Increasing production and use of
nanoparticles (NPs) in commercial products has instigated growing
regulatory concerns and public debates4–10 on the potential environ-
mental and health implications of nanotechnology.
To support sustainable development of nanotechnology, potential
risks, whether real or imagined, have to be evaluated based on sound,
risk-focused research that addresses all relevant aspects of the
concern. In a recent Nature Commentary, Maynard and colleagues
proposed five grand challenges with a goal to stimulate collaborative
and integrated research that is imaginative, innovative and relevant to
the safe development of nanotechnology.11 Research efforts are being
funded by the governments of various countries, most notably the
US, the European Union, and Japan, to develop priorities for risk-
based safety evaluations of nanomaterials and nanotechnology. In
the US, the Environmental Protection Agency (EPA) and other
federal agencies have tentatively identified the following research
needs for developing comprehensive risk assessments: (1) charac-
terization, (2) transformation and interaction, (3) environmental fate
and transport, (4) exposure assessment, (5) ecological effects, (6)
human health effects assessment, and (7) life-cycle analysis.12
To date, toxicological effects of manufactured NPs have attracted
a majority of the attention. It has been reported that carbon nanotubes
a
University of Delaware, Department of Physics and Astronomy, Newark,
Delaware, USA
b
University of Delaware, Department of Plant and Soil Sciences, Newark,
Delaware, USA. E-mail: yjin@udel.edu; Fax: +1-302-831-0605; Tel: +1-
302-831-6962
This journal is ª The Royal Society of Chemistry 2008
www.rsc.org/jem | Journal of Environmental Monitoring
could damage lung tissues in mice, PTFE NPs killed laboratory rats in
15 min after exposure, and inhaled carbon-13 and manganese NPs
reached rats’ olfactory bulbs and then migrated through the brain.13 A
recent review14 of the studies on biokinetics and toxicology reports that
when inhaled, NPs of specific sizes are effectively deposited in all
regions of the respiratory tract. The small size of NPs facilitate their
uptake into cells, therefore can reach potentially sensitive organs (e.g.,
bone marrow, lymph nodes, spleen, and heart), access the central
nervous system and ganglia, and penetrate skin via uptake into
lymphatic channels.14 While there has been an increasing amount of
research on the toxicity of NPs to human cells, bacteria, and rodents
recently,15–18 very few studies have been conducted to assess the effects
of NPs on ecological terrestrial species such as plants.19
A the end of their life cycle, most nanotechnology products are
disposed in landfills. As a result, NPs may be released into the soil–
water system. In addition, NPs may be intentionally put into the soil to
facilitate soil and groundwater remediation.2 Information on the
potential exposure through soil and water and whether NPs will
bioaccumulate through the food chain and end up in higher-level
organisms is extremely limited.20 Plants are an important component
in the ecological system and may serve as an potential pathway for NP
transport and a route forbioaccumulation into the food chain.
However, the few studies conducted to date on the effects of NPs on
plants have focused mainly on phytotoxicity and how certain plant
metabolic functions are affected. The reported effects vary depending
on the type of NP, as well as plant species, and are not always
consistent between different studies. For example, titania NPs were
shown to improve the growth of spinach seeds,10 whereas alumina NPs
were reported to inhibit root elongation of several plants (corn,
cucumber, soybean, cabbage, and carrot).21 A more recent study22
investigated the effects of five types of NPs (multi-walled carbon
nanotube, aluminium, alumina, zinc and zinc oxide) on seed germi-
nation and root growth of six higher plant species (radish, rape,
ryegrass, lettuce, corn, and cucumber) and showed that seed genera-
tion was in general not affected in most cases while root elongation was
inhibited. The 50% inhibitory concentration (IC50) varied among the
particles and plants species tested. An evaluation on the effect of
magnetic NPs coated with tetramethylammonium hydroxide (TMA-
OH) on the growth of young popcorn (Zea mays) plants found that
low concentrations of aqueous ferrofluid stimulated plant growth
while high concentrations induced inhibitory or toxic effects.23 The
authors proposed bio-membrane penetration as one of the possible
reasons for their observed effects of TMA-OH coated magnetic NPs
(8 nm average size) on popcorn plants but this hypothesis was not
verified.
Here we report the results from a recent study where we demon-
strated significant uptake of magnetite (Fe3O4) NPs by pumpkin
plants and their subsequent translocation and accumulation in
various tissues. To the best of our knowledge, this is the first study to
J. Environ. Monit., 2008, 10, 713–717 | 713
 show that manufactured NPs can be taken up by plants as well as
undergo translocation and accumulation within plant tissues.
Pumpkin (Cucurbita maxima) was selected as a model plant
because of its large water uptake capacity, and was grown hydro-
ponically in a growth medium (Scotts-Sierra Horticultural Products
Co., Marysville, OH, USA). We chose Fe3O4 (NanoAmor Inc.,
manufacture reported diameter of 20 nm) as representative NPs
because of its biological compatibility and structural stability. More
importantly, we selected Fe3O4 because of its magnetic properties,
which allowed non-intrusive tracking and quantification of the
particles via magnetometry measurement. Dynamic light scattering
measurements were made to determine particle size distribution and
zeta potential of suspended Fe3O4 particles using a Zetasizer Nano
Downloaded by University of Massachusetts - Amherst on 01 December 2010
ZS (Malvern Instruments Ltd.). As shown in Fig. 1, both individual
Published on 13 May 2008 on http://pubs.rsc.org | doi:10.1039/B805998E
Fe3O4 particles of 20 nm as well as agglomerates of various sizes (up
to 2 mm) existed in the suspension, and particle surface possessed
a slightly negative charge (
7 to
9 meV).
Pumpkin seeds were germinated using root cubes in a greenhouse
where temperature and relative humidity were controlled at 22  C
and 60%, respectively. After germination, the seedlings were trans-
ferred into separate containers (A and B) filled with the growth
medium and allowed to grow till the appearance of the 3rd leaf.
Suspended Fe3O4 particles were then added into container A (treat-
ment) at a concentration of 0.5 g L
1 while container B received the
growth medium free of NPs (control). Water consumed by the
pumpkin plants was replenished: container A with the growth
Fig. 1 Dynamic light scattering measurements of (a) particle size
distribution by intensity and (b) zeta potential for Fe3O4 nanoparticles
dispersed in growth medium. The different lines represent replicate
measurements.
714 | J. Environ. Monit., 2008, 10, 713–717
medium containing the same concentration of Fe3O4 particles
(0.5 g L
1) and B with the growth medium without particles. When
the pumpkin plants reached an average height of 27 cm (20 d of
growth), they were removed from the containers, rinsed with water,
cut into short pieces, and subsequently dried in a vacuum desiccator
for 3 d. The concentrations of the Fe3O4 particles were measured with
a vibrating sample magnetometer (VSM, LakeShore 7400) at room
temperature (23  C). VSM is a non-destructive technique that can
be used to detect very weak magnetic signals. Briefly, the specimen is
vibrated in an external magnetic field. The total magnetization can
thus be determined from the voltage induced in a pick-up coil. Fig. 2
shows selected VSM measurements of control plants (Fig. 2(a)) and
various tissues of the treatment plants (Fig. 2(b)–2(f)).
After 20 d of growth, there were no apparent visual differences in
the plants grown with and without Fe3O4 particles, indicating that the
particles did not pose any toxicological effects to the plants at the
concentration level tested. No magnetization signal was detected
from the control plants (VSM detection limit was 0.5 memu),
whereas most of the tissue samples of the treatment plants showed
magnetization of various strengths. Fig. 3(a) shows a schematic
pumpkin plant and VSM measured magnetization at various
sampling locations, marked as numbers in the unit of 10
3 emu g
1
(or memu g
1). The magnetization of pure Fe3O4 particles is
53.19 emu g
1, and 1 memu correlates to 8.48  1011 particles
(assuming Fe3O4 particles have a density of 5.17 g cm
3). Strong
magnetic signals (>1.0 memu g
1) were detected in all leaf specimens
regardless of their distances from the roots, while the signals were
much weaker from the stem tissue samples except those close to the
roots. Although large numbers of the suspended particles accumu-
lated on root surfaces (Fig 3(b)), it is clear that Fe3O4 particles were
taken up by the pumpkin plants and moved into the various tissues.
The strongest magnetization (3.26 memu g
1) was detected right
above the roots, which might be due to NP agglomeration.
SEM micrographs of the major tissue components are also shown
in Fig. 3, which include horizontal and vertical (near the surface)
cross sections of the stem tissue, respectively in (f) and (g), magnified
xylem wall (h), and a stomata located on the back of a leaf (d). As
indicated by the strong ferromagnetic signals detected in the leaf
samples, large numbers of Fe3O4 particles were transported to and
accumulated in the leaves, where transport pathways are much
narrower compared to those in the stems.
At the end of the experiment, concentrations of Fe3O4 particles
remaining in the growth medium and accumulated in plant tissues
were measured to obtain mass balance. The concentration in
suspension was determined by VSM. Plant tissues were dried at
70  C, ground to pass a 20-mesh sieve, and each 0.5 g of subsample
was digested with HNO3 and H2O2 in a CEM Mars5 Microwave
Digestion System (CEM Corporation, Matthews, NC, USA). Total
Fe content was then measured by inductively coupled plasma (ICP)
optical emission spectroscopy using a Thermo Electron IRIS Intrepid
II XSP-Duo View ICP (Thermo Electron Corporation, Madison WI,
USA). A mass balance of 67.4% was obtained based on total Fe
recovered and is the sum of the following components: (1) remaining
in suspension 21.4%, (2) leaf tissue 0.6%, and (3) root tissue 45.4%. As
noted previously, many Fe3O4 particles accumulated on the root
surfaces so the 45.4% includes both the particles inside of the roots
and those on the outside that were not rinsed off by water. The low
mass recovery indicates that a portion of the particles was lost during
the experiment and/or during sample handling. We believe that most
This journal is ª The Royal Society of Chemistry 2008
Downloaded by University of Massachusetts - Amherst on 01 December 2010
Published on 13 May 2008 on http://pubs.rsc.org | doi:10.1039/B805998E

Fig. 2 Selected VSM measurements of pumpkin plant tissues for Fe3O4 nanoparticles. (a) control plant (root), (b) treatment plant stem at height of 23–
27 cm, (c) treatment plant root, (d) treatment plant stem at height of 0–6 cm, (e) treatment plant leaf at height of 22–23 cm, and (f) treatment plant leaf
above 27 cm. The background diamagnetic signal is linear with magnetic field. The magnetization at H ¼ 10 000 Oe is determined after the background
signal is removed.

of the unaccounted particles were lost with the xylem sap when the
plants were cut and separated into different tissue fractions. A small
portion could also be lost through leaves via controlled openings of
the stomata (8 mm  3 mm in size) that allow water evaporation and
gas exchange.
It is also worth mentioning that at the beginning of this study, we
conducted an initial screening test, growing pumpkin plants as well as
lima bean plants (Phaseolus limensis) hydroponically in Fe3O4
particle suspension. However, we did not detect any magnetic signals
in any parts of the lima bean plants while strong signals were
measured in pumpkin plant tissues. This suggests that different plants
have varied responses to nanosized particles and therefore warrants
further investigation with more plant species.
We have also grown pumpkin plants in sand and soil media,
respectively, by irrigating the plants periodically with Fe3O4 particle
suspensions. While the level of uptake and accumulation were
significantly reduced in plants grown in sand as compared to those
grown in liquid media, measurable levels of magnetic signals were
detected in all tissues, ranging from <0.1 memu g
1 in most tissues to
0.4–1.2 memu g
1 in leaves and stems (close to roots), respectively.
However, magnetic signals were not detectable in the plants grown in
soil. Clearly, the availability of Fe3O4 particles in the aqueous phase
was significantly reduced, likely due to their attachment to the
surfaces of sand or soil grains.
In summary, we demonstrated that a significant amount of Fe3O4
particles suspended in a liquid medium can be taken up by pumpkin

This journal is ª The Royal Society of Chemistry 2008 J. Environ. Monit., 2008, 10, 713–717 | 715
Downloaded by University of Massachusetts - Amherst on 01 December 2010
Published on 13 May 2008 on http://pubs.rsc.org | doi:10.1039/B805998E
Fig. 3 Schematic of Fe3O4 particle distribution in various tissues of a pumpkin plant.
plants and be translocated throughout the plant tissues. Particles tend
to accumulate near the roots as well as in leaves. Although this study
likely represents a worst-case scenario (plants grown in a liquid
medium containing high particle concentrations), it nevertheless
provides convincing evidence that plant uptake is a potential
transport pathway of nanoparticles in the environment. Clearly,
interactions between plants and nanoparticles deserve more in depth
investigation on many fronts, such as uptake potential of different
plant species, mechanisms of uptake and translocation and the
interactions between the particles with plant tissue at the cellular level.
Values inside the circles indicate magnetization in unit of memu g
1
with 1 memu g
1 corresponding to 8.48  1011 particles g
1. These
future studies will help answer the question on how significant plant
uptake of nanoparticles is as a potential transport pathway and its
role in bioaccumulation through the food chain. Such studies will
play an important role in assessing the impacts of nanomaterials and
nanotechnology on public health and ecological systems.
Acknowledgements
We acknowledge the financial support provided by the US National
Science Foundation’s Delaware EPSCoR grant EPS-0447610
716 | J. Environ. Monit., 2008, 10, 713–717
through Delaware Biotechnology Institute. We also would like to
express our gratitude to Dr Jung-Youn Lee at the University of
Delaware, for her suggestion to use pumpkin as the model plant for
this study and to Dr Harsh Bais, also at the University of Delaware,
for helpful discussions.
Notes and references
1 A. Nel, T. Xia, L. Madler and N. Li, Science, 2006, 311, 622–627.
2 W. X. Zhang and B. Karn, Environ. Sci. Technol., 2005, 39, 94A–95A.
3 The Nanotechnology Consumer Products Inventory, Woodrow Wilson
International Center for Scholars, Washington DC, 2006, http://
www.nanotechproject.org/consumerproducts.
4 V. L. Colvin, Nat. Biotechnol., 2003, 21, 1166–1170.
5 W. W. Center, The Hill, http://www.thehill.com, October 11, 2006.
6 R. Liddle, The Guardian, London, July 30, 2003.
7 A. Broache, CNET, http://www.news.com, January 11, 2006.
8 A. Press, MSNBC, http://www.msnbc.com, January 11, 2006.
9 S. R. Morrissey, ACS, C&EN, 2005, 83, 46–48.
10 L. Zheng, F. S. Hong, S. P. Lu and C. Liu, Biol. Trace Elem. Res.,
2005, 104, 83–91.
11 A. D. Maynard, R. J. Aitken, T. Butz, V. Colvin, K. Donaldson,
G. Oberdorster, M. A. Philbert, J. Ryan, A. Seaton, V. Stone,
S. S. Tinkle, L. Tran, N. J. Walker and D. B. Warheit, Nature,
2006, 444, 267–269.
This journal is ª The Royal Society of Chemistry 2008
 12 K. Thomas, P. Aguar, H. Kawasaki, J. Morris, J. Nakanishi and
N. Savage, Toxicol. Sci., 2006, 92, 23–32.
13 R. F. Service, Science, 2003, 300, 243–243.
14 G. Oberdorster, E. Oberdorster and J. Oberdorster, Environ. Health
Perspect., 2005, 113, 823–839.
15 T. J. Brunner, P. Wick, P. Manser, P. Spohn, R. N. Grass, L. K. Limbach,
A. Bruinink and W. J. Stark, Environ. Sci. Technol., 2006, 40, 4374–4381.
16 G. Jia, H. F. Wang, L. Yan, X. Wang, R. J. Pei, T. Yan, Y. L. Zhao
and X. B. Guo, Environ. Sci. Technol., 2005, 39, 1378–1383.
17 C. W. Lam, J. T. James, R. McCluskey, S. Arepalli and R. L. Hunter,
Crit. Rev. Toxicol., 2006, 36, 189–217.
Downloaded by University of Massachusetts - Amherst on 01 December 2010
Published on 13 May 2008 on http://pubs.rsc.org | doi:10.1039/B805998E
This journal is ª The Royal Society of Chemistry 2008
18 K. F. Soto, A. Carrasco, T. G. Powell, L. E. Murr and K. M. Garza,
Mater. Sci. Eng., C, 2006, 26, 1421–1427.
19 Science Policy Council Agency E P United States, Nanotechnology
White Paper External Review Draft, US Environmental Protection
Agency, Feb. 2005.
20 The Royal Society and the Royal Academy of Engineering, London,
http://www.royalsoc.ac.uk, 2004.
21 L. Yang and D. J. Watts, Toxicol. Lett., 2005, 158, 122–132.
22 D. Lin and B. Xing, Environ. Pollut., 2007, 150, 243–250.
23 M. Racuciu and D. Creanga, Rom. J. Phys., 2007, 52,
395–402.
J. Environ. Monit., 2008, 10, 713–717 | 717