Journal of Colloid and Interface Science 348 (2010) 329–334

Contents lists available at ScienceDirect

Journal of Colloid and Interface Science

www.elsevier.com/locate/jcis

Stresses exerted by ZnO, CeO2 and anatase TiO2 nanoparticles

on the Nitrosomonas europaea
Xiaohua Fang, Ran Yu, Bingquan Li, Ponisseril Somasundaran *, Kartik Chandran
Department of Earth and Environmental Engineering, Columbia University, NY 10027, USA

a r t i c l e i n f o a b s t r a c t

Article history: Recent studies have shown that nano-bio interfaces are the most complex and the least understood.
Received 27 January 2010 Notably, nanotoxicity of these nanoparticles is not even well recognized. In this work, we examined
Accepted 24 April 2010 the toxic effects of different nanoparticles on bacteria cells (Nitrosomonas europaea). The four nanoparti-
Available online 23 May 2010
cles involved are: 25 nm anatase TiO2, 200 nm anatase TiO2, ZnO and CeO2 particles. These particles will
have different electrical charges in the cell cultivating media. It has been observed that even with only 4 h
Keywords: of dosing, all of the particles caused apparent morphological damage to the cells. Experimental results
Nanoparticles
suggest that ZnO particles exert the stress on cells by its dissolution and releasing of Zn2+ ions. The
Bacteria
TEM and AUC (analytical ultracentrifuge) result suggest that cells become heavier in presence of CeO2
and TiO2 particles. No visible clear intrusions of bulk nanoparticles were observed. However, both the
analytical ultracentrifuge and TEM results show that cells are heavier when being damaged.
Ó 2010 Elsevier Inc. All rights reserved.

1. Introduction nano/bio interfacial region. It is to be further noted that the entire

Nanoparticles (NPs) in the range of 1–100 nm are increasingly
used for a variety of clinical and commercial purposes due to their
large surface-to-volume ratio and special physico-chemical and
electronic properties. The increasing application of nanoparticles
(NPs) in commercial and consumer product industry, biological sci-
ences and medical diagnostics have brought attention on the haz-
ards NPs pose to human health. The toxicity of NPs is dependent to
a large extent on their dispersion which in turn is controlled by the
NP properties such as size, charge and the surface groups [1–10].
For example, it has been found very recently that the nanoparticles
with different surface properties would cause different responses
in the culture medium, which leads to a different degree of toxicity
[11,12].
The potential adverse effects of the inorganic nanoparticle–cell
have attracted attention of the scientific community [5,13–17].
Some studies found intracellular reactive oxygen species (ROS)
by CuO nanoparticles. DNA lesions were observed in presence of
ZnO, TiO2 (mixture of rutile and anatase) as well as super-para-
magnetic iron oxide nanoparticles (SPIONs) [18,19]. As in the case
of engineered composite nanoparticles, cytotoxicity of polymeric
nanogels can be driven by surface and morphological effects, by
mechanisms that have not yet been clearly addressed [20]. The
effects of the heterogeneity of the nanoparticles are further com-
plicated by many other interactions that take place in the complex
nano-biointerface is a dynamic system where many short and long
term interactions occur simultaneously as well as sequentially.
In addition, since the safety of existing nanomaterials that have
been marketed for decades does not appear to be adequately ad-
dressed, experimental results are urgently needed to screen haz-
ardous effects of current and future nanomaterials. As of today,
most of the research on nanotoxicity has been put on the eukary-
otic cells. We report in this work cytotoxicity of ZnO, CeO2, and
anatase TiO2 nanoparticles on Nitrosomonas europaea cultures.
The model system studied here should provide useful references
for helping in setting parameters of future studies on the effect
of nanoparticles/nanogels on other bacterial cells [11,20,21].
2. Materials and methods
N. europaea is a microbe whose growth rely on the oxidation of
ammonia to nitrite [22]. Therefore, it is used in general for waste
water treatment. The presence of nanoparticles could possibly
damage the activity of the N. europaea cells, leading to their poor
performance in the treatment of industrial and sewage waste of
oxidizing ammonia to nitrate. In this study, this toxic effect by
the different NPs on N. europaea cells is evaluated from the per-
spect of physical chemistry.
2.1. Cell-culture and loading of nanoparticles

* Corresponding author. Fax: +1 212 854 8362. N. europaea bacteria in chemostat state (the American true type
E-mail address: ps24@columbia.edu (P. Somasundaran). culture collection) were cultured in a media containing (NH4)2SO4

0021-9797/$ - see front matter Ó 2010 Elsevier Inc. All rights reserved.
doi:10.1016/j.jcis.2010.04.075
330 X. Fang et al. / Journal of Colloid and Interface Science 348 (2010) 329–334
(1.32 g/L), MgSO4.7H2O (0.2 g/L), CaCl22H2O (0.02 g/L), K2HPO4
(0.087 g/L), 3-(4-(2-Hydroxyethyl)-1-piperzine) propane sulfonic
acid (2.52 g/L), and 13% EDTA-Fe3+ (1 ml/l). ZnO, CeO2 and anatase
TiO2 nanoparticles were loaded into the media at concentrations of
20 ppm and 200 ppm. After being cultured for 4 h in the media
containing those particles, the cells were collected for further
characterization.
2.2. Nanoparticle and media characterizations
2.2.1. Size and morphology
Four types of NPs were applied in the study: 25 nm and 200 nm
anatase TiO2, (Sigma–Aldrich, St. Louis, MO, USA), ZnO (meliorum
tech.), and CeO2 (meliorum tech.). Their size and morphology were
characterized with a Transmission Electron Microscope (Philips
430 transmission electron microscope). The particles were dis-
persed in 0.1 mM Nonyl phenol-7 ethoxylate solutions. After over-
night mixing and equilibration, a droplet of the suspension was
dried onto a 300 mesh gilder copper TEM grid (Electron Micros-
copy Sciences, Inc., Hatfield, PA, USA) before TEM analysis with
an accelerating voltage of 100 kV.
2.2.2. Surface area
The surface area of the particle was determined with a Quanta-
sorb Surface Area Analyzer (Quantachrome Instruments, Monosorb
(The name of the instrument), Boynton Beach, FL, USA). The parti-
cles were preheated at 70 °C for 30 min to remove the pre-ad-
sorbed water before analysis.
2.2.3. Charge
A type of back titration method was adopted for particle’s PZC
measurement. In this method, the particles were loaded into a ser-
ies of solutions (at 0.1 mg/ml) at different pH values prepared with
hydrochloric acid or sodium hydroxide. The amount of base (Dn)
loaded for both the blank and the particles suspensions were then
plotted as a function of pH. The crossing point of the two curves
gives the PZC, which is 8.4, 7.5, 6.5 and 6.9 for ZnO, CeO2,
25 nm and 200 nm TiO2 particles, respectively.
2.3. Bacteria characterizations
Transmission Electron microscopy: The cells after 4 h of culture
were fixed with 2.5% glutaraldehyde in 0.1 M Sorenson’s buffer (PH
7.2). The pellet obtained is then postfixed with 1% OsO4 in Soren-
son’s buffer for 1 h. After dehydration the samples were embedded
in Lx-112 (Ladd Research Industries, Inc.). Thin sections were cut
on the MT-7000 ultramicrotome at 60 nm thickness. The sections
were stained with uranyl acetate and lead citrate. The bacteria
samples were examined under a JEOL JEM-1200 EXII electron
microscope. Pictures were taken on an ORCA-HR digital camera
(Hamamatsu) and recorded with an AMT Image Capture Engine.
The charges (the zeta potential) as well as size conditions for
the cell/particle complex in the medium were measured using a
Malvern nano ZS zetasizer (Malvern, Worcestershire, United King-
dom). Cell suspensions were put in a DTS0012 sample holder. The
scattered light signals from the suspended cells in the cultivation
medium were collected by the instrument. It is assumed that the
particles in the medium do not participate in the scattering since
they tend to settle down to the bottom of the sample cell quickly.
A Beckman Optima XL-1 analytical ultracentrifuge with scan-
ning optics in an interference system was employed to investigate
the complex distribution in solution. Sedimentation velocity meth-
od was performed. A two-channel cell was used to accommodate
the sample and reference solvent (buffer). As sedimentation pro-
gresses, the solutes will redistribute along the cell channel accord-
ing to their sedimentation coefficients. The total concentration will
be detected by the interference optical system through measuring
the refractive index difference between the sample channel and
the reference channel at each radial position as indicated by the
vertical displacement of a set of evenly spaced horizontal fringe.
When the vacuum was reached at 5e 3 Torr, the centrifugation
was set at a motor speed 3000 rpm with the temperature at
25 °C. Software Sedfit92 developed by Peter Shuck was used to ana-
lyze the sedimentation data [23].
3. Results
3.1. Properties of nanoparticles
Fig. 1 shows the transmission electron microscopic images of the
particles tested in this work. The median size of the elliptical
shaped TiO2 particles are 25 nm and 200 nm (Fig. 1a and b). The
elliptically shaped ZnO particles had a median size of 30 nm
(Fig. 1c). CeO2 particles are needle-shaped with an average dimen-
sion of 60 nm in length and 20 nm in width (Fig. 1d). The surface
areas from the BET measurement for 25 nm anatase TiO2, anatase
200 nm TiO2, ZnO and CeO2 particles were 117.7 m2/g, 9.2 m2/g,
42.1 m2/g, and 93.8 m2/g, respectively.
When the particles are placed into a solution, dissolution of io-
nic groups or preferential adsorption of ions can induce surface
charge on their solid surfaces. For general metal oxides in acid/base
solutions, protonation/deprotonation controls the sign and value of
the surface charge density. The pH at which the net charge on the
particles surface equals to zero is called the point of zero charge
(PZC). Below PZC, the particles are positively charged, and above
PZC, the particles are negatively charged. The PZC values are
6.5, 6.8, 8.4, and 7.5 for 25 nm TiO2, 200 nm TiO2, ZnO, and
CeO2 particles. The pH of the cell culture media is 7.5 ± 0.1. There-
fore, 25 nm TiO2, 200 nm TiO2, ZnO, and CeO2 particles were orig-
inally expected to be negative, slightly negative, positive, and
neutral, respectively. On the other hand, the media is a very com-
plex system containing multi-valence ionic species. It is thus very
possible that the charges on the particle as a whole being reversed
after ionic adsorption. The zeta potential result shows 21 mv for
25 nm TiO2, 22 mv for 200 nm TiO2, 20 mv for ZnO, and 19 mv
for CeO2 particles, indicating the existence of strong selective
adsorption of negative ionic species or the dissolution of positive
ionic species in the media-particle suspension.
3.2. Average size for particle–cell complexes under NP stress
The dynamic light scattering results on statistical particle–cell
complex size change in N. europaea cultivating medium is shown
in Fig. 2. It is assumed that light scattering measures the size of
the cell-particle complex suspended in the media. Pure N. europaea
cells show an averaged size of 1050 ± 27 nm.
From Fig. 2 it can be seen that cells in media with 20 ppm TiO2-
25 nm, TiO2-200 nm and ZnO NPs show a smaller size as compared
with control cells. Increase of particle content to 200 ppm NPs in
the media did not reduce the particle–cell complex size further.
On the contrary, the cells in 200 ppm NPs media either remained
their size level at 20 ppm as in the zinc oxide case, or slightly
recovered a bit as the 25 nm or 200 nm titania cases. The loading
of 20 ppm CeO2 NPs did not affect cell size significantly. When
the loading was increased to 200 ppm, an apparent decrease in size
was observed.
It is also seen from Fig. 2 that 20 nm and 200 nm TiO2 NPs have
similar impact in terms of cell size. However, the larger size ana-
tase particles seem to have created a little more shrinkage of cells.
Thus, NP type and loading concentration are seen to exert impact
differently on the size of the particle–cell complex. The observed
 X. Fang et al. / Journal of Colloid and Interface Science 348 (2010) 329–334
Fig. 1. Transmission electron microscopy images of (a) 25 nm anatase TiO2, (b) 200 nm anatase TiO2, (c) ZnO, and (d) CeO2 particles studied in this research.
Fig. 2. Size of the cell–nanoparticle complex in the cultivating media.
phenomena implied the underlying diverse cell morphology varia-
tion under NPs stress and were confirmed by TEM analysis dis-
cussed as shown below.
3.3. Statistical charge for particle–cell complexes under NP stress
Pure N. europaea culture showed an averaged zeta potential of
22.4 ± 0.9 mv in the cultivating media, which moved slightly to-
wards the positive side upon the loading of 20 ppm, 200 nm,
25 nm TiO2 anatase and ZnO particles Fig. 3. ZnO NPs have the
most effect to reverse the charge of the cells, while CeO2 does
not show any significant effect at this loading level (20 ppm). Fur-
ther increment in NPs concentration to 200 ppm converged the
charge for all the particles (25 nm TiO2, 200 nm TiO2, ZnO and
CeO2 particles) to similar zeta potential levels of around 20.5 mv.
Compared with the statistical size results shown in Fig. 2, it can
be seen that 20 ppm loading of NPs show the most significant and
consistent effect on cells irrespective of the type of the NP. For ZnO
and CeO2 particles, a size decrease of cell–NP complexes correlates
331
Fig. 3. Zeta potential of the cell–nanoparticle complex in the cultivating media.
with an increment in the zeta potential. Cells in media with 25 nm
and 200 nm TiO2 nanoparticles, however, only shows the rise in
zeta potentials at low dose of NPs (20 ppm).
3.4. Morphological stress exerted by nanoparticles on N. europaea
The morphological changes of bacteria were captured by the
transmission electron microscope (Fig. 4a–h). The intercellular par-
ticle accumulation was found for cells treated with all three types
of NPs except for the 200 nm anatase TiO2.
After an exposure to 25 nm anatase TiO2, the morphology of N.
europaea changed mostly from the regular elliptical shape for the
control (Fig. 4a) to an irregular and distorted form (Fig. 4b). Most
of the cells that remained in their elliptical shapes contained a cav-
ity. Those cells that are distorted still kept their cytosolic composi-
tions. Hardly any dividing cells were observed in the cells with
particles. Also, it was hard to distinguish the clear multilayered cell
membranes under NPs stress, especially when 200 ppm of NPs
were added (Fig. 4c). At the same loading concentration, i.e.,
332 X. Fang et al. / Journal of Colloid and Interface Science 348 (2010) 329–334
Fig. 4. N. europaea cells grown in media containing (a) control (no anatase TiO2 nanoparticles (NPs)), (b) 20 ppm of 25 nm anatase TiO2 NPs, (c) 200 ppm of 25 nm anatase
TiO2 NPs, (d) 200 ppm of 200 nm anatase TiO2 NPs for 4 h, (e) 20 ppm of ZnO NPs, (f) 20 ppm CeO2 NPs for 4 h, (g) 20 ppm CeO2 NPs, showing the particle align themselves
longitudinally on the cell wall, (h) 20 ppm CeO2 NPs, the cells close to the particle aggregates shows more cavities in them.
20 ppm, smaller particles distorted the cells more than the larger
particles. Cells in the media with 200 nm of particles remained
spherical in shape with their membrane less perturbed (Fig. 4d)
than the ones in media with 25 nm particles. Empty cavities, how-
ever, are still observed in the cell. The examined NPs normally
aggregated in the medium and were seldom observed attached
onto cell walls. At similar total surface area, 200 ppm, 200 nm ana-
tase TiO2 nanoparticles show less damage to cell shape as com-
pared to 20 ppm 25 nm anatase TiO2 NPs. Empty vacuoles,
however, were still observed inside of the cell for both the sizes.
No visible particles intrusion was observed, indicating that the par-
ticles that enter the cells might have sizes on the sub-nano level,
which is hard to distinguish by the TEM.
The cells show little morphological distortion in shape under
the stress of ZnO NPs (Fig. 4e) as compared with cells cultured
in media with 25 nm anatase TiO2 particles. However, most of
the cell inner leaflet of the membrane wrapped the interior cell
structures and separated from the rest of outer membrane bilayer
compartments. The outer cell membrane bilayers become un-dis-
tinguishable too. No broken membrane debris was observed in
the fixed cell sample. Neither was empty cavities/vacuoles seen
in the cell. On the other hand, a few dark sparkles can be ob-
served having entered the cells and gathered within the center
of the cells.
Similar morphological responses were found for CeO2 treated
N. europaea (Fig. 4f). No particles were observed to intrude the cell,
although some can be observed to adhere to the cell wall along
their longitudinal needle axis (Fig. 4g). Much significant damages
were found in cells that were close to CeO2 aggregates, which dis-
play large cavities and bubbles in them (Fig. 4h). The cell walls/
membranes, however, remain undamaged. Considering the fact
that during TEM sample preparation heavy particles as well as cells
precipitate first due to centrifugation, it can be stated that the cells
near the CeO2 aggregates would be heavier and show more damage
with vacuoles than the cells far away from them.
4. Discussion
The stresses exerted by the same type of nanoparticles on
eukaryotic cells have been reported by Xia et al. [16]. It has been
concluded that ZnO particles induce cytotoxicity by dissolution
of Zn2+ ions as well as the uptake of ZnO particle remnants. The
nano-ZnO exposure on cells could increase the membrane perme-
ability via Na+ and K+ fluxes through cells and ionic homeostasis
disturbance based on rat tests [24]. Since the N. europaea has a cell
wall and multilayered membrane bilayer shell, it would very unli-
kely that uptake the particles via endosomal ways. Moreover, a
close contact between cells and particles is also prevented by the
electric static interaction since both of them are negative in sign.
This is proposed to be the reason that very few particles were ob-
served inside of the cells as shown by the TEM images. The dark
sparkles we observed could be the agglomerate of sub-nano parti-
cles that got into the cells by diffusion. To test whether Zn2+ causes
the damage, we also cultured the cells in the ZnSO4 loaded media
at the same level of Zn2+ (2.6 ppm) as in the 4-h 20 ppm and
200 ppm particles suspensions. The Zn2+ levels were decided by
an atomic absorption spectrometry (Atomic Absorption Spectro-
photometer, Model 200A, Buck Scientific, East Norwalk, CT, USA).
After centrifugation of 20 ml cultures at 16,100g for 10 min, the
supernatant was filtered through the 0.22 lm filter (Fisherbrand,
Waltham, MA, USA) followed by an immediate Zn measurement.
It is then assumed that very little particles and cells exist in the
media at this stage. Compared with results from ZnO particles,
the size-particle concentration and charge-particle concentration
curves shown in Figs. 2 and 3 for ZnSO4 salt showed similar trends,
supporting the validity of Xia’s statement. On the other hand, our
results also show that the Zn2+ ions in the salt are a bit more toxic
than the particle suspension by decreasing the cell size more sig-
nificantly and making the zeta potential more positive. This obser-
vation, however, is different from that of Xia’s et al. We propose
that this difference is due to the dissolution kinetics of ZnO and
ZnSO4 in the cell culturing media during the initial stage of
experiments.
As from previous references on eukaryotic cells, TiO2 NPs would
be toxic under UV energy in the cells [14]. Therefore, the TiO2 NPs
that did not get into the cells would not produce as much damage.
The CeO2 particles, on the other hand, have been proven to be safe
and can protect the cells against secondary oxidative stress stimu-
lus [16]. Our results of stress exerted by the anatase TiO2 and CeO2
nanoparticles on the prokaryotic N. europaea, however, showed
toxic observations. From the morphological observation, all
 X. Fang et al. / Journal of Colloid and Interface Science 348 (2010) 329–334 333

a b

Fig. 5. The analytical ultracentrifuge result on N. europaea cells grown in media loaded with 0 ppm, 20 ppm and 200 ppm of (a) 25 nm anatase TiO2 NPs, (b) CeO2 NPs and (c)
ZnO NPs. Cells under the stress of NPs have larger sedimentation coefficient than the control.

particles were observed to have caused damage irrespective of
their sizes and charges. The shrinkage of the inner cell membrane
against the cell wall and the release of cytosolic compartments
indicate possible high osmotic pressure in and out of the cell mem-
brane under the stress of nanoparticles. TiO2 cause cell membrane
dissociation even when possible UV-radiation was screened to
some degree under dark culturing conditions. It is also seen from
the TEM images that there are many fibrous structures surround-
ing the cells, which can be considered to belong to the cell walls
or membrane compartments. Since the charges on the cell-com-
plex shifted to the positive side and the original zeta potential
for the particles are negative, we propose that only some positive
ionic species from the media adsorbed on the cell walls. This state-
ment is validated by the results from three separate analytical
ultracentrifuge measurements in Fig. 5, which shows in general
when under the NP stress, the cells become heavier than the con-
trol and thus sediment faster (Fig. 5a–c). Cells in 200 ppm TiO2 and
CeO2 NPs media have a sedimentation peak position close to that of
the cells with lower NP loading, while apparent right-shift is seen
between the 200 ppm and 20 ppm peak positions of cells in ZnO
NPs media (Fig. 5c). This is not unexpected considering the lower
dissolution power of the TiO2 and CeO2 NPs as compared with
ZnO NPs. These results correlate well with the light scattering
and zeta potential data.
5. Conclusion
The possible toxicity effects of ZnO, CeO2, 25 nm and 200 nm
anatase TiO2 nanoparticles on waste water treatment N. europaea
bacteria are probed. Within 4 h of cultivating time, cells were
found to be distorted with cell wall/membrane damage as from
TEM, light scattering, and zeta potential results. Very few particles
get into the cells. Toxicity of ZnO NPs is due to the releasing of Zn2+
ions from the particles. Anatase TiO2 and CeO2 particles are also
toxic, which are different from previous observations on eukaryotic
cells by the literature. Preferential dissolution of specific ions of the
particles is proposed to contribute to the toxicity.
Acknowledgments
This material is based upon work supported by the National Sci-
ence Foundation and the Environmental Protection Agency under
Cooperative Agreement Number EF 0830117. Any opinions, find-
ings, and conclusions or recommendations expressed in this mate-
rial are those of the author(s) and do not necessarily reflect the
views of the National Science Foundation or the Environmental
Protection Agency. This work has not been subjected to EPA review
and no official endorsement should be inferred.
References
[1] W. Noohom, K.S. Jack, D. Martin, M. Trau, Biomed. Mater. 4 (1) (2009).
[2] A.S. Mistry, S.H. Cheng, T. Yeh, E. Christenson, J.A. Jansen, A.G. Mikos, J. Biomed.
Mater. Res., Part A 89A (1) (2009) 68–79.
[3] C.R. Kothapalli, C.R. Kothapalli, Acta Biomater. 5 (2) (2009) 541–553.
[4] D. Depan, A.P. Kumar, R.P. Singh, Acta Biomater. 5 (1) (2009) 93–100.
[5] J.T. Seil, T.J. Webster, Int. J. Nanomed. 3 (4) (2008) 523–531.
[6] C. Erisken, D.M. Kalyon, H.J. Wang, Biomaterials 29 (30) (2008) 4065–4073.
[7] T. Crouzier, A. Nimmagadda, M.U. Nollert, P.S. McFerridge, Langmuir 24 (22)
(2008) 13173–13181.
[8] S.X. Bi, X.Y. Wei, N. Li, L. Lei, Mater. Lett. 62 (17–18) (2008) 2963–2966.
334 X. Fang et al. / Journal of Colloid and Interface Science 348 (2010) 329–334
[9] A. Nel, T. Xia, L. Madler, N. Li, Science 311 (2006) 622–627.
[10] P. Somasundaran, S.C. Mehta, L. Rhein, S. Chakraborty, MRS Bull. 32 (10) (2007)
779–786.
[11] M. Mahmoudi, M.A. Shokrgozar, A.Simchi;M. Imani, A.S. Milani, P. Stroeve, H.
Vali, U.O. Hafeli, S. Bonakdar, J. Phys. Chem. C 113 (6) (2009) 2322–2331.
[12] M. Mahmoudi, A. Simchi, M. Imani, A. S. Milani, P. Stroeve, Nanotechnology 20
(2009) (22) 8.
[13] T. Xia, N. Li, A.E. Nel, Annu. Rev. Public Health 30 (2009) 137–150.
[14] Z. Pan, W. Lee, L. Slutsky, R.A.F. Clark, N. Pernodet, M.H. Rafailovich, Small 5 (4)
(2009) 511–520.
[15] T. Xia, M. Kovochich, M. Liong, J.I. Zink, A.E. Nel, ACS Nano 2 (1) (2008) 85–96.
[16] T. Xia, M. Kovochich, M. Liong, L. Madler, B. Gilbert, H.B. Shi, J.I. Yeh, J.I. Zink,
A.E. Nel, ACS Nano 2 (10) (2008) 2121–2134.
[17] A. Nel, T. Xia, L. Madler, N. Li, Science 311 (5761) (2006) 622–627.
[18] H.L. Karlsson, P. Cronholm, J. Gustafsson, L. Moller, Chem. Res. Toxicol. 21 (9)
(2008) 1726–1732.
[19] M. Mahmoudi, A. Simchi, H. Vali, M. Imani, M.A. Shokrgozar, K. Azadmanesh, F.
Azari, Adv. Eng. Mater. 11 (12) (2009) B243–B250.
[20] A.E. Nel, L. Madler, D. Velegol, T. Xia, E.M.V. Hoek, P. Somasundaran, F. Klaessig,
V. Castranova, M. Thompson, Nat. Mater. 8 (7) (2009) 543–557.
[21] T. Cedervall, I. Lynch, S. Lindman, T. Berggard, E. Thulin, H. Nilsson, K.A.
Dawson, S. Linse, Proc. Natl. Acad. Sci. U. S. A. 104 (7) (2007) 2050–2055.
[22] P. Chain, J. Lamerdin, F. Larimer, W. Regala, V. Lao, M. Land, L. Hauser, A.
Hooper, M. Klotz, J. Norton, L. Sayavedra-Soto, D. Arciero, N. Hommes, M.
Whittaker, D. Arp, J. Bacteriol. 185 (21) (2003) 6496.
[23] P. Schuck, Biophys. J. 78 (3) (2000) 1606–1619.
[24] J. Zhao, L. Xu, T. Zhang, G. Ren, Z. Yang, NeuroToxicology 30 (2009) 220–
230.