Upload
ylpkaso
View
216
Download
0
Embed Size (px)
Citation preview
8/3/2019 Leonid Vigderman et al- Quantitative Replacement of Cetyl Trimethylammonium Bromide by Cationic Thiol Ligands
1/15
Gold NanorodsDOI: 10.1002/anie.201107304
Quantitative Replacement of Cetyl Trimethylammonium Bromide byCationic Thiol Ligands on the Surface of Gold Nanorods and TheirExtremely Large Uptake by Cancer Cells**
Leonid Vigderman, Pramit Manna, and Eugene R. Zubarev*
Over the past decade, gold nanorods (Au NRs) have received
broad attention as possible therapeutic and diagnostic agents
because of their anisotropic physical properties.[18] However,
as-synthesized Au NRs based on the most common synthetic
approach, the seed-mediated synthesis,[9, 10] are not directly
useable for these applications. They are synthesized in a
concentrated cetyl trimethylammonium bromide (CTAB)
solution and are noncovalently coated with a CTAB
bilayer.[11] For CTAB-capped rods to remain soluble, the
concentration of free CTAB in solution must remain above acertain value and there is a constant dynamic exchange of
CTAB molecules between the solution and NR surfaces.[12]
This feature limits the usefulness of these rods for many
biological applications because free CTAB is known to be
highly cytotoxic.[13, 14] Various strategies have been developed
to functionalize gold nanorods with a variety of different
ligands to reduce their cytotoxicity.[12, 1520] Further reports
have dealt with the problems of stability and biocompatibility
of Au NRs by using various polymer shells,[21] polyelectro-
lytes,[22, 23] peptides,[24] surfactants,[25] and lipids[14,26] to modify
the Au NR surface. Partial replacement of CTAB was
qualitatively confirmed in many of these cases, but the exact
surface composition of nanorods and the amount of residualCTAB was either unknown or not possible to determine. For
that reason, there is always some ambiguity in interpretation
of in vitro and in vivo experiments involving such nanorods.[27]
Beyond just biocompatibility and low toxicity, there has
been great interest in understanding the cellular uptake of Au
NRs.[2832] For example, if CTAB is replaced by nonionic
polyethylene glycol, the cytotoxicity is substantially reduced,
but the cellular uptake drops as much as 94%, and virtually
no nanorods can enter the cells.[30] When polyelectrolytes are
randomly wrapped around the CTAB bilayer, the uptake
increases and the average number of rods that enter each cell
ranges from hundreds[31] to one hundred thousand.[29] How-
ever, even polyelectrolyte-coated Au NRs are taken by cellsin picogram quantities, which is only 0.1% of a typical cell
mass. This small weight fraction may explain a somewhat
limited success of photothermal oblation of tumor cells,[33]
which is strongly dependent on the actual number of nano-
rods present inside the cells. Therefore, an obvious goal is to
design stable and nontoxic (CTAB free) nanorods that could
enter cancer cells in much larger quantities.
Here we report a strategy for complete exchange of
CTAB for its thiolated analogue (16-mercaptohexadecyl)tri-
methylammonium bromide (MTAB) and we directly deter-
mine the chemical composition of the surface coating.
Through
1
H NMR analysis, we are able to prove that CTABis fully replaced by a covalent MTAB monolayer. By
combining thermogravimetric analysis (TGA) with TEM
size analysis of the nanorods, we are able to accurately
determine the packing density of the self-assembled thiol
monolayer on the surface of Au NRs. Cytotoxicity of these
novel nanorods was studied by MTT assay on breast cancer
cells (MCF-7), which were imaged by correlated optical and
scanning electron microscopy (SEM). Extremely large
number of nanorods (about 2 106 NRs per cell) were
found inside the viable cells as confirmed by transmission
electron microscopy (TEM) and quantified by inductively
coupled plasma-optical emission spectrometry (ICP-OES).
Several reports have shown that even with systems thathave successfully functionalized gold nanorods, CTAB is still
observed to be present on the NRs.[28] We hypothesized that
we could use a molecule with a structure very similar to
CTAB to completely exchange the CTAB, but with the ability
to bind more strongly to the NR surface. Thus, we chose to
synthesize a thiolated CTAB analogue (MTAB) whose
synthesis is shown in Scheme 1. Commercially available
1,16-hexadecanediol was converted to a corresponding dibro-
mide under standard bromination conditions, which was
followed by the synthesis of its monothioester. Cleavage of
the acetyl group was carried out in anhydrous methanol using
Scheme 1. Synthesis of (16-mercaptohexadecyl)trimethylammoniumbromide (MTAB; NBS=N-bromosuccinimide, THF= tetrahydrofuran).
[*] L. Vigderman, Dr. P. Manna, Prof. E. R. ZubarevDepartment of Chemistry, Rice UniversityHouston, TX 77005 (USA)E-mail: [email protected]: http://www.owlnet.rice.edu/~zubarev/
[**] This work was supported by the NSF (grant numbers DMR-0547399and DMR-1105878).
Supporting information for this article is available on the WWWunder http://dx.doi.org/10.1002/anie.201107304.
Angew. Chem. Int. Ed. 2011, 50, 1 7 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
These are not the final page numbers!
http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://dx.doi.org/10.1002/anie.201107304http://dx.doi.org/10.1002/anie.201107304http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-8/3/2019 Leonid Vigderman et al- Quantitative Replacement of Cetyl Trimethylammonium Bromide by Cationic Thiol Ligands
2/15
in situ generated hydrogen chloride. The resulting 16-bromo-
1-hexadecanethiol was subjected to exhaustive methylation to
yield the final product (see the Supporting Information).
Importantly, the purified MTAB compound was found to be
water-soluble, which offers an opportunity to perform ligand
exchange in aqueous media. The MTAB ligand contains an
entire CTAB moiety, but installs a pendant thiol group to be
able to strongly anchor it to the gold surface. The cross-
section of this molecule is small enough to form a compact
monolayer on the surface of nanorods, thus providing their
solution stability.
Stabilization of gold nanorods is generally difficult
because of their small surface-to-volume ratio, and finding a
compound that can directly exchange with CTAB is further
complicated by the high-density positive charge and the
amphiphilic nature of the CTAB bilayer (Figure 1). In the
past, researchers have exchanged only the tips of goldnanorods because of relatively weak CTAB binding there
and have noted that it is much stronger on the sides.[3436]
Furthermore, as the CTAB on the sides of the rods begins to
exchange, it appears that the bilayer structure breaks down
and rods tend to aggregate prematurely. However, choosing a
cationic ligand with a chemical structure similar to native
CTAB allows for the direct exchange in water to proceed
smoothly as shown schematically in Figure 1. The noncova-
lent bilayer of CTAB is replaced with a compact thiolate
monolayer of MTAB which is covalently anchored to the
surface through goldsulfur bonds. UV/Vis absorbance spec-
troscopy is often used to assess the stability of gold nanorods
in solution as the intensity, the peak shape, and the peakposition of the longitudinal plasmon resonance (LSPR) are
sensitive to any aggregation that may occur. The UV/Vis
spectrum (see Figure S1A in the Supporting Information)
shows the typical absorbance spectrum for Au NRs and
demonstrates what happens when excess CTAB is removed
from solution through multiple rounds of centrifugation
followed by dispersion in pure water. As expected, after just
four rounds, the NRs have lost all solubility as evidenced by
the lack of any absorbance. In stark contrast, when MTAB-
modified NRs are purified by the same technique, there is no
appreciable change in the absorbance spectrum even after
five rounds of centrifugation and dispersion (see Figure S1B
in the Supporting Information), indicating their much
improved solution stability while also completely purifying
the product.
Additional experiments show that these rods can be
completely dried and kept in the solid state indefinitely
without losing their water solubility. Surprisingly, a standard
lyophilization technique can be applied to aqueous solution of
MTAB NRs to produce a fluffy dark brown powder. Fig-
ure 2a shows a photograph of 15 mg of lyophilized Au NRs
that occupy an area of several square centimeters. Most
importantly, when a small amount of this powder is placed on
the surface of pure water, rapid dissolution takes place as
manifested by the concentration swirls and a continuous
diffusion of the colored solute (Figure 2 b). The nanorods
dissolve in a matter of seconds without any heating or
sonication. It is sufficient to flip the vial only once to form a
homogeneous solution which does not contain any free
organic molecules. The solution remains intact for at least
several months as confirmed by UV/Vis analysis. This data
clearly shows that a dense monolayer of small cationic thiol
MTAB is capable of stabilizing large metallic particles in purewater and preventing their flocculation through entropically
driven depletion interactions.[37] This data also showes for the
first time that the presence of free surfactant in the solution of
gold nanorods is not necessary as long as their surface is
covalently functionalized by a dense organic shell.
Another indirect method that has often been used to
characterize Au NR complexes is the zeta potential measure-
ment, which can qualitatively describe the charge surrounding
a nanoparticle. The zeta potential of the MTAB NRs was
found to be +55 mV, demonstrating the cationic structures as
expected from the high concentration of quaternary ammo-
nium groups positioned around the NR surface. However, the
Figure 1. Exchange of the CTAB bilayer for the MTAB thiol monolayer. Figure 2. a) Photograph of lyophilized powder of MTAB-functionalizedAu NRs and b) its spontaneous dissolution in pure water. The imageswere taken consecutively within intervals of five second.
Communications
2 www.angewandte.org 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim Angew. Chem. Int. Ed. 2011, 50, 1 7
These are not the final page numbers!
http://-/?-http://-/?-http://-/?-http://www.angewandte.org/http://www.angewandte.org/http://-/?-http://-/?-http://-/?-8/3/2019 Leonid Vigderman et al- Quantitative Replacement of Cetyl Trimethylammonium Bromide by Cationic Thiol Ligands
3/15
zeta potential is unable to distinguish between CTAB and
MTAB on the surface of the rods given that they both have
the same cationic headgroup exposed to the solution and so is
useful only as a secondary confirmation of structure. We
chose 1H NMR spectroscopy to analyze the organic compo-
nent of the MTAB NRs which should be able to differentiate
between the CTAB and MTAB molecules. We used KCN to
dissolve the gold NR core and release any surface-bound
organic material into solution[39] after having rigorously
purified the functionalized NRs. Because of the low surface-
to-volume ratio as well as high density of the gold core, we
performed an oxidative dissolution of 105 mg of Au NRs in
one milliliter of D2O to get enough organic material for a
sufficiently strong 1H NMR signal. The dissolution proceeded
very slowly, taking approximately two weeks to dissolve all of
the gold, suggesting a high packing density for the MTAB
surface functionality.[40] On the contrary, CTAB-coated NRs
dissolved in about one hour under similar conditions. The
dissolution procedure caused the organic component to
precipitate from the D2O solution, apparently because of
the formation of a Au/CN/MTAB complex. The NMRspectrum of the D2O supernatant showed no signals, con-
firming that the entire organic component had precipitated
from the solution. The NMR spectrum of this precipitate
dissolved in deuterated methanol, as well as that of pure
CTAB and MTAB thiol, is shown in Figure 3. Comparing the
spectra of CTAB and MTAB, it is clear that the isolated signal
from the terminal methyl group of CTAB at 0.91 ppm is not
present in the MTAB spectrum, making it a convenient
marker for the presence of any CTAB in solution. Analysis of
the NMR spectrum of the dissolved NRs (spectrum C in
Figure 3) clearly shows that there is no signal corresponding
to this methyl group, proving that no CTAB remained on the
surface of the NRs after the exchange and the following
purification. Further analysis of the NMR spectrum of the
dissolved NRs shows the formation of the expected disulfide
of MTAB as evidenced by the triplet at 2.75 ppm correspond-
ing to the a-disulfide protons (instead of 2.5 ppm for a-thiol
protons). Importantly, the NMR technique is capable of
detecting small organic molecules at concentration close to
105m, whereas the concentration of our solution was 102m.
This implies that even if there were some residual CTAB
undetectable by NMR spectroscopy, it would not exceed
0.1%. Therefore, for all practical purposes one can conclude
that the method described hear offers an opportunity to
quantitatively replace CTAB by its thiolated analogue.
Because the exact chemical composition of the NR
surface agents was determined, it was possible to use TGA
to further characterize the MTAB NRs. TGA analysis is a
highly useful analytical technique for hybrid inorganic
organic nanostructures because it allows us to accurately
quantify the weight percentage of a nanostructure that is
organic compared to inorganic core.[39,41, 42] For our study,
more than 100 mg of MTAB NRs were synthesized and
functionalized to be used for TGA and NMR measurements,
which is quite a large amount when it comes to Au NR
synthesis, requiring a synthesis on the four liter scale.
However, even this amount of material is difficult to handle,
forming an almost unusable thin film when dried normally. Tosolve this problem, the MTAB NRs were lyophilized from an
aqueous solution, forming a low density powder which could
be easily handled, as mentioned previously (Figure 2). TGA
analysis of the lyophilized material (see Figure S2 in the
Supporting Information) shows a weight loss of 5.3% in the
range between 200 and 4508C corresponding to the percent-
age of organic material in the structure. Since the NMR
analysis proved that no residual CTAB was left, this weight
must be entirely because of MTAB ligands covalently
attached to the surface of rods.
We then performed a careful TEM analysis by measuring
over 300 nanorods, which determined that the average length
and width of MTAB NRs were 41.88 and 9.87 nm, respec-tively. With this information, we were able to calculate a
grafting density of about 3.7 moleculesnm2 for an MTAB
monolayer and 5013 MTAB molecules residing on each Au
NR (see the Supporting Information for detailed calcula-
tions). This is close to the grafting density of 4.5 mole-
culesnm2 for neutral alkanethiols on flat gold substrates[43]
and may be slightly lower because of repulsive forces between
the positively charged headgroups. We believe this is the first
proof of a dense self-assembled thiol monolayer on gold
nanorods with a calculated grafting density. Estimation of the
extent of thiol functionalization has been carried out on
DNA-functionalized Au NRs by measuring fluorescence
intensity, but such structures contained only about 40 DNAmolecules per nanorod[17] as compared to over 5000 MTAB
molecules.
With the characterization complete, we performed cyto-
toxicity and cell uptake experiments to see if the MTAB NRs
could be useful for biological applications. We first measured
their in vitro cytotoxicity compared to regular CTAB-capped
NRs using the standard MTT assay on MCF-7 breast cancer
cells (see Figure S3 in the Supporting Information). These
studies reaffirmed the cytotoxicity of regular CTAB-capped
gold nanorods and showed the decreased cytotoxicity of the
thiolated rods, which are not cytotoxic even up to concen-
trations of 0.1 gL1. This finding supports previous assertions
Figure 3. 1H NMR spectra in deuterated methanol of CTAB (A), pureMTAB (B), and the organic product released upon oxidative dissolu-tion of MTAB NRs (C).
Angew. Chem. Int. Ed. 2011, 50, 1 7 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim www.angewandte.org
These are not the final page numbers!
http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://www.angewandte.org/http://www.angewandte.org/http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-http://-/?-8/3/2019 Leonid Vigderman et al- Quantitative Replacement of Cetyl Trimethylammonium Bromide by Cationic Thiol Ligands
4/15
that surface charge may not be a key factor for the
cytotoxicity of nanoparticle systems.[31] Although previous
reports have shown that it is mostly free CTAB that
contributes to the cytotoxicity of CTAB NRs in the short
term,[31] it is unknown what may happen to surface-bound
CTAB in the longer term in in vivo experiments. Our system
has the advantage of having quantitative CTAB replacement
such that this is no longer an issue. Low cytotoxicity is
particularly useful if the NRs can be taken up by cells
efficiently. To study the uptake, MCF-7 cells were treated with
a 20 mg mL1 solution of MTAB NRs in Eagles minimum
essential medium for 24 h. The cells were subsequently rinsed
multiple times with PBS solution to remove the excess of NRs
that did not enter the cells. After that, the cells were
trypsinized, redispersed into medium, and carefully plated
onto a new culture slide. This approach allowed us to create a
clear background without any free nanorods, which is
critically important for differentiating between the internal-
ized particles and those which are not associated with the
cells. The dark field optical image (Figure 4A) clearly shows a
large uptake of MTAB NRs and demonstrates their high
scattering efficiency. It also appears that NRs are clustered
together inside the cells rather than being evenly distributed
throughout their interior, suggesting that the MTAB NRs
enter the cells through an endosomal pathway. To perform a
correlated SEM imaging of the exact same group of cells, we
fixed the sample with glutaraldehyde and further cross-linked
the cells with osmium tetroxide. This enabled us to prevent
the collapse of cells under high vacuum conditions required
for the SEM. The SEM imaging correlated with the optical
experiments (Figure 4B) shows areas of increased brightness
inside the cells because of clusters of internalized MTAB
NRs. The locations of the NR clusters match up well with the
bright spots seen in the optical micrograph, although there are
some morphological changes that occurred during the fixation
and dehydration processes. Most importantly, SEM imaging
reveals that there are no MTAB NRs on the surface of the
cells, which would be impossible to confirm by the optical
microscopy alone. In contrast, the visualization of nanorods
residing in thicker cell areas is more efficient by optical
microscopy, displaying their strong potential as imaging
contrast agents.To further characterize the cellular uptake, we performed
transmission electron microscopy (TEM) imaging of cells
treated with the MTAB NRs as shown in Figure 5. Unlike
SEM and optical microscopy, TEM images of the microtomed
cross-sections reveal precise location and spatial distribution
of individual nanorods. We prepared 75 nm thick sections that
were cut through the center of cells. The TEM image of the
entire cell cross-section (Figure 5A) shows an extremely large
amount of NRs, which appear as dark particles. The volume of
this cross-sectional sample is approximately 0.5% of the total
cell volume (20 mm in diameter), which means that the total
number of nanorods inside the cell is approximately 200 times
Figure 4. A) Dark field optical micrograph and B) SEM image of cellstreated with MTAB NRs.
Figure 5. TEM images of microtomed MCF-7 cancer cells treated withthe MTAB NRs (75 nm thick). A) View of an entire cell cross-section.B) Magnified image of the area outlined by the black box in panel A;C) Magnified image of the area outlined by the dashed box in panel B.
Communications
4 www.angewandte.org 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim Angew. Chem. Int. Ed. 2011, 50, 1 7
These are not the final page numbers!
http://-/?-http://-/?-http://www.angewandte.org/http://www.angewandte.org/http://-/?-http://-/?-8/3/2019 Leonid Vigderman et al- Quantitative Replacement of Cetyl Trimethylammonium Bromide by Cationic Thiol Ligands
5/15
greater than that present in this image (about 10000 NRs).
Thus, even on the basis of TEM one can roughly estimate that
there are at least two million nanorods per cell. If confirmed
by quantitative methods, it would be significantly larger than
the numbers reported in the literature for nanorods of similar
size, which range from several hundred[31] to one-hundred-
fifty thousand NRs per cell.[29] Even at low magnification one
can see that NRs do not enter the nucleus and are clustered in
endosomes. Higher magnification of regions outlined by the
boxes (Figure 5B and C, respectively) clearly shows individ-
ual nanorods that do not appear to be free in the cyto-
plasm.[29,4446] The amount of uptake was compared to another
popular NR system, pegylated nanorods (PEG NRs), which
are known to resist cellular uptake and thus form a good
negative control for our system. To quantify the average
number of NRs taken by cells, we performed ICP-OES
analysis on dissolved cells which had been treated with both
MTAB NRs and their pegylated analogues. Based on these
results, while almost no PEG NRs were taken up (less than
1%), about 40% of MTAB NRs in solution were taken up by
the cells. This corresponds to a level of about 2.17 millionMTAB NRs per cell that have been treated with a solution of
nanorods (see the Supporting Information for detailed
calculations). Such a great amount of metallic gold increases
the mass of cancer cell by 0.13 ng, which is approximately
13% of a typical cell mass (1 ng). It is particularly remarkable
that the cells retain their viability and continue to proliferate
with such a significant amount of gold nanostructures inside
them. These results show that the MTAB NRs have a very low
cytotoxicity and suggest that they may be promising for
further biomedical applications such as drug/gene delivery
and photothermal therapy.
In conclusion, we have synthesized highly stable, func-
tionalized gold nanorods as an alternative to CTAB-cappedNRs through a direct quantitative exchange with a thiolated
CTAB analogue. The MTAB NRs showed a highly increased
stability to multiple rounds of purification compared to
CTAB NRs. We were able to quantitatively prove the
complete removal of CTAB through the use of 1H NMR
spectroscopy, and thus determined the exact composition of
the organic component of the hybrid nanostructures. We also
successfully performed a TGA analysis to measure the exact
organic versus inorganic composition of hybrid nanostruc-
tures and determined a grafting density for the MTAB thiol,
proving the formation of a compact self-assembled monolayer
on gold nanorods. The MTAB NRs were not only found to be
nontoxic, but were also efficiently taken up by cancer cellsin vitro in very large amounts as shown by a comprehensive
combination of optical microscopy, SEM, TEM, and ICP-
OES data.
Received: October 17, 2011
Published online:&& &&, &&&&
.Keywords: cancer cells cellular uptake gold nanorods surface chemistry
[1] N. J. Durr, T. Larson, D. K. Smith, B. A. Korgel, K. Sokolov, A.
Ben-Yakar, Nano Lett. 2007, 7, 941 945.
[2] X. Huang, I. H. El-Sayed, W. Qian, M. A. El-Sayed, J. Am.
Chem. Soc. 2006, 128, 2115 2120.
[3] C. Yu, J. Irudayaraj, Anal. Chem. 2007, 79, 572 579.
[4] X. Liu, Q. Dai, L. Austin, J. Coutts, G. Knowles, J. Zou, H. Chen,
Q. Huo, J. Am. Chem. Soc. 2008, 130, 2780 2782.
[5] R. Guo, L. Zhang, H. Qian, R. Li, X. Jiang, B. Liu, Langmuir
2010, 26, 5428 5434.
[6] T. Kawano, Y. Niidome, T. Mori, Y. Katayama, T. Niidome,
Bioconjugate Chem. 2009, 20, 209 212.
[7] H. Wang, T. B. Huff, D. A. Zweifel, W. He, P. S. Low, A. Wei,
J. X. Cheng, Proc. Natl. Acad. Sci. USA 2005, 102, 15752 15756.
[8] a) L. Zhao, X. Pang, R. Adhikary, J. Petrich, M. Jeffries-EL, Z.
Lin, Adv. Mater. 2011, 23, 2844 2849; b) L. Zhao, X. Pang, R.
Adhikary, J. Petrich, Z. Lin, Angew. Chem. 2011, 123, 4044
4048; Angew. Chem. Int. Ed. 2011, 50, 3958 3962.
[9] B. Nikoobakht, M. A. El-Sayed, Chem. Mater. 2003, 15, 1957
1962.
[10] N. R. Jana, L. Gearheart, C. J. Murphy, J. Phys. Chem. B 2001,
105, 4065 4067.
[11] B. Nikoobakht, M. A. El-Sayed, Langmuir2001, 17, 6368 6374.
[12] C. J. Orendorff, T. M. Alam, D. Y. Sasaki, B. C. Bunker, J. A.Voigt, ACS Nano 2009, 3, 971983.
[13] R. Cortesi, E. Esposito, E. Menegatti, R. Gambari, C. Nastruzzi,
Int. J. Pharm. 1996, 139, 69 78.
[14] H. Takahashi, Y. Niidome, T. Niidome, K. Kaneko, H. Kawasaki,
S. Yamada, Langmuir2006, 22, 2 5.
[15] H. W. Liao, J. H. Hafner, Chem. Mater. 2005, 17, 4636 4641.
[16] S. K. Basiruddin, A. Saha, N. Pradhan, N. R. Jana, Langmuir
2010, 26, 7475 7481.
[17] A. Wijaya, K. Hamad-Schifferli, Langmuir2008, 24, 9966 9969.
[18] A. Gole, C. J. Murphy, Chem. Mater. 2005, 17, 1325 1330.
[19] D. Gentili, G. Ori, F. M. Comes, Chem. Commun. 2009, 5874
5876.
[20] I. E. Sendroiu, M. E. Warner, R. M. Corn, Langmuir 2009, 25,
11282 11284.[21] G. Prencipe, S. M. Tabakman, K. Welsher, Z. Liu, A. P. Good-
win, L. Zhang, J. Henry, H. Dai, J. Am. Chem. Soc. 2009, 131,
47834787.
[22] A. P. Leonov, J. Zheng, J. D. Clogston, S. T. Stern, A. K. Patri, A.
Wei, ACS Nano 2008, 2, 2481 2488.
[23] H. C. Huang, S. Barua, D. B. Kay, K. Rege, ACS Nano 2009, 3,
29412952.
[24] N. Chanda, R. Shukla, K. V. Katti, R. Kannan, Nano Lett. 2009,
9, 1798 1805.
[25] A. M. Alkilany, P. K. Nagaria, M. D. Wyatt, C. J. Murphy,
Langmuir 2010, 26, 9328 9333.
[26] S. E. Lee, D. Y. Sasaki, T. D. Perroud, D. Yoo, K. D. Patel, L. P.
Lee, J. Am. Chem. Soc. 2009, 131, 14066 14074.
[27] N. Khlebtsov, L. Dykman, Chem. Soc. Rev. 2011, 40, 1647 1671.
[28] A. K. Oyelere, P. C. Chen, X. Huang, I. H. El-Sayed, M. A. El-Sayed, Bioconjugate Chem. 2007, 18, 1490 1497.
[29] T. S. Hauck, A. A. Ghazani, W. C. Chan, Small2008, 4, 153 159.
[30] H. J. Parab, H. M. Chen, T. C. Lai, J. H. Huang, P. H. Chen, R. S.
Liu, M. Hsiao, C. H. Chen, D. P. Tsai, Y. K. Hwu, J. Phys. Chem.
C 2009, 113, 7574 7578.
[31] A. M. Alkilany, P. K. Nagaria, C. R. Hexel, T. J. Shaw, C. J.
Murphy, M. D. Wyatt, Small2009, 5, 701708.
[32] T. B. Huff, M. N. Hansen, Y. Zhao, J. X. Cheng, A. Wei,
Langmuir 2007, 23, 1596 1599.
[33] a) L. C. Kennedy, L. R. Bickford, N. A. Lewinski, A. J. Cough-
lin, Y. Hu, E. S. Day, J. L. West, R. A. Drezek, Small 2011, 7,
169183; b) R. Choi, J. Yang, J. Choi, E.-K. Lim, E. Kim, J.-S.
Suh, Y.-M. Huh, S. Haam, Langmuir 2009, 25, 17520 17527.
Angew. Chem. Int. Ed. 2011, 50, 1 7 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim www.angewandte.org
These are not the final page numbers!
http://-/?-http://-/?-http://-/?-http://-/?-http://dx.doi.org/10.1021/nl062962vhttp://dx.doi.org/10.1021/nl062962vhttp://dx.doi.org/10.1021/nl062962vhttp://dx.doi.org/10.1021/nl062962vhttp://dx.doi.org/10.1021/nl062962vhttp://dx.doi.org/10.1021/ja057254ahttp://dx.doi.org/10.1021/ja057254ahttp://dx.doi.org/10.1021/ja057254ahttp://dx.doi.org/10.1021/ja057254ahttp://dx.doi.org/10.1021/ja057254ahttp://dx.doi.org/10.1021/ja057254ahttp://dx.doi.org/10.1021/ac061730dhttp://dx.doi.org/10.1021/ac061730dhttp://dx.doi.org/10.1021/ac061730dhttp://dx.doi.org/10.1021/ac061730dhttp://dx.doi.org/10.1021/ac061730dhttp://dx.doi.org/10.1021/ja711298bhttp://dx.doi.org/10.1021/ja711298bhttp://dx.doi.org/10.1021/ja711298bhttp://dx.doi.org/10.1021/ja711298bhttp://dx.doi.org/10.1021/ja711298bhttp://dx.doi.org/10.1021/la903893nhttp://dx.doi.org/10.1021/la903893nhttp://dx.doi.org/10.1021/la903893nhttp://dx.doi.org/10.1021/la903893nhttp://dx.doi.org/10.1021/la903893nhttp://dx.doi.org/10.1021/bc800480khttp://dx.doi.org/10.1021/bc800480khttp://dx.doi.org/10.1021/bc800480khttp://dx.doi.org/10.1021/bc800480khttp://dx.doi.org/10.1021/bc800480khttp://dx.doi.org/10.1073/pnas.0504892102http://dx.doi.org/10.1073/pnas.0504892102http://dx.doi.org/10.1073/pnas.0504892102http://dx.doi.org/10.1073/pnas.0504892102http://dx.doi.org/10.1073/pnas.0504892102http://dx.doi.org/10.1002/adma.201100923http://dx.doi.org/10.1002/adma.201100923http://dx.doi.org/10.1002/adma.201100923http://dx.doi.org/10.1002/adma.201100923http://dx.doi.org/10.1002/adma.201100923http://dx.doi.org/10.1002/ange.201100200http://dx.doi.org/10.1002/ange.201100200http://dx.doi.org/10.1002/ange.201100200http://dx.doi.org/10.1002/ange.201100200http://dx.doi.org/10.1002/ange.201100200http://dx.doi.org/10.1002/ange.201100200http://dx.doi.org/10.1002/anie.201100200http://dx.doi.org/10.1002/anie.201100200http://dx.doi.org/10.1002/anie.201100200http://dx.doi.org/10.1002/anie.201100200http://dx.doi.org/10.1002/anie.201100200http://dx.doi.org/10.1021/cm020732lhttp://dx.doi.org/10.1021/cm020732lhttp://dx.doi.org/10.1021/cm020732lhttp://dx.doi.org/10.1021/cm020732lhttp://dx.doi.org/10.1021/cm020732lhttp://dx.doi.org/10.1021/cm020732lhttp://dx.doi.org/10.1021/jp0107964http://dx.doi.org/10.1021/jp0107964http://dx.doi.org/10.1021/jp0107964http://dx.doi.org/10.1021/jp0107964http://dx.doi.org/10.1021/jp0107964http://dx.doi.org/10.1021/la010530ohttp://dx.doi.org/10.1021/la010530ohttp://dx.doi.org/10.1021/la010530ohttp://dx.doi.org/10.1021/la010530ohttp://dx.doi.org/10.1021/la010530ohttp://dx.doi.org/10.1021/nn900037khttp://dx.doi.org/10.1021/nn900037khttp://dx.doi.org/10.1021/nn900037khttp://dx.doi.org/10.1021/nn900037khttp://dx.doi.org/10.1021/nn900037khttp://dx.doi.org/10.1016/0378-5173(96)04574-7http://dx.doi.org/10.1016/0378-5173(96)04574-7http://dx.doi.org/10.1016/0378-5173(96)04574-7http://dx.doi.org/10.1016/0378-5173(96)04574-7http://dx.doi.org/10.1016/0378-5173(96)04574-7http://dx.doi.org/10.1021/la0520029http://dx.doi.org/10.1021/la0520029http://dx.doi.org/10.1021/la0520029http://dx.doi.org/10.1021/la0520029http://dx.doi.org/10.1021/la0520029http://dx.doi.org/10.1021/cm050935khttp://dx.doi.org/10.1021/cm050935khttp://dx.doi.org/10.1021/cm050935khttp://dx.doi.org/10.1021/cm050935khttp://dx.doi.org/10.1021/cm050935khttp://dx.doi.org/10.1021/la904189ahttp://dx.doi.org/10.1021/la904189ahttp://dx.doi.org/10.1021/la904189ahttp://dx.doi.org/10.1021/la904189ahttp://dx.doi.org/10.1021/la904189ahttp://dx.doi.org/10.1021/la8019205http://dx.doi.org/10.1021/la8019205http://dx.doi.org/10.1021/la8019205http://dx.doi.org/10.1021/la8019205http://dx.doi.org/10.1021/la8019205http://dx.doi.org/10.1021/cm048297dhttp://dx.doi.org/10.1021/cm048297dhttp://dx.doi.org/10.1021/cm048297dhttp://dx.doi.org/10.1021/cm048297dhttp://dx.doi.org/10.1021/cm048297dhttp://dx.doi.org/10.1039/b911582jhttp://dx.doi.org/10.1039/b911582jhttp://dx.doi.org/10.1039/b911582jhttp://dx.doi.org/10.1039/b911582jhttp://dx.doi.org/10.1021/la902675shttp://dx.doi.org/10.1021/la902675shttp://dx.doi.org/10.1021/la902675shttp://dx.doi.org/10.1021/la902675shttp://dx.doi.org/10.1021/la902675shttp://dx.doi.org/10.1021/la902675shttp://dx.doi.org/10.1021/ja809086qhttp://dx.doi.org/10.1021/ja809086qhttp://dx.doi.org/10.1021/ja809086qhttp://dx.doi.org/10.1021/ja809086qhttp://dx.doi.org/10.1021/ja809086qhttp://dx.doi.org/10.1021/ja809086qhttp://dx.doi.org/10.1021/nn800466chttp://dx.doi.org/10.1021/nn800466chttp://dx.doi.org/10.1021/nn800466chttp://dx.doi.org/10.1021/nn800466chttp://dx.doi.org/10.1021/nn800466chttp://dx.doi.org/10.1021/nn900947ahttp://dx.doi.org/10.1021/nn900947ahttp://dx.doi.org/10.1021/nn900947ahttp://dx.doi.org/10.1021/nn900947ahttp://dx.doi.org/10.1021/nn900947ahttp://dx.doi.org/10.1021/nn900947ahttp://dx.doi.org/10.1021/nl8037147http://dx.doi.org/10.1021/nl8037147http://dx.doi.org/10.1021/nl8037147http://dx.doi.org/10.1021/nl8037147http://dx.doi.org/10.1021/nl8037147http://dx.doi.org/10.1021/la100253khttp://dx.doi.org/10.1021/la100253khttp://dx.doi.org/10.1021/la100253khttp://dx.doi.org/10.1021/la100253khttp://dx.doi.org/10.1021/la100253khttp://dx.doi.org/10.1021/ja904326jhttp://dx.doi.org/10.1021/ja904326jhttp://dx.doi.org/10.1021/ja904326jhttp://dx.doi.org/10.1021/ja904326jhttp://dx.doi.org/10.1021/ja904326jhttp://dx.doi.org/10.1039/c0cs00018chttp://dx.doi.org/10.1039/c0cs00018chttp://dx.doi.org/10.1039/c0cs00018chttp://dx.doi.org/10.1039/c0cs00018chttp://dx.doi.org/10.1039/c0cs00018chttp://dx.doi.org/10.1021/bc070132ihttp://dx.doi.org/10.1021/bc070132ihttp://dx.doi.org/10.1021/bc070132ihttp://dx.doi.org/10.1021/bc070132ihttp://dx.doi.org/10.1021/bc070132ihttp://dx.doi.org/10.1002/smll.200700217http://dx.doi.org/10.1002/smll.200700217http://dx.doi.org/10.1002/smll.200700217http://dx.doi.org/10.1002/smll.200700217http://dx.doi.org/10.1002/smll.200700217http://dx.doi.org/10.1021/jp9000169http://dx.doi.org/10.1021/jp9000169http://dx.doi.org/10.1021/jp9000169http://dx.doi.org/10.1021/jp9000169http://dx.doi.org/10.1021/jp9000169http://dx.doi.org/10.1021/jp9000169http://dx.doi.org/10.1002/smll.200801546http://dx.doi.org/10.1002/smll.200801546http://dx.doi.org/10.1002/smll.200801546http://dx.doi.org/10.1002/smll.200801546http://dx.doi.org/10.1002/smll.200801546http://dx.doi.org/10.1021/la062642rhttp://dx.doi.org/10.1021/la062642rhttp://dx.doi.org/10.1021/la062642rhttp://dx.doi.org/10.1021/la062642rhttp://dx.doi.org/10.1021/la062642rhttp://dx.doi.org/10.1002/smll.201000134http://dx.doi.org/10.1002/smll.201000134http://dx.doi.org/10.1002/smll.201000134http://dx.doi.org/10.1002/smll.201000134http://dx.doi.org/10.1002/smll.201000134http://dx.doi.org/10.1002/smll.201000134http://www.angewandte.org/http://www.angewandte.org/http://dx.doi.org/10.1002/smll.201000134http://dx.doi.org/10.1002/smll.201000134http://dx.doi.org/10.1021/la062642rhttp://dx.doi.org/10.1002/smll.200801546http://dx.doi.org/10.1021/jp9000169http://dx.doi.org/10.1021/jp9000169http://dx.doi.org/10.1002/smll.200700217http://dx.doi.org/10.1021/bc070132ihttp://dx.doi.org/10.1039/c0cs00018chttp://dx.doi.org/10.1021/ja904326jhttp://dx.doi.org/10.1021/la100253khttp://dx.doi.org/10.1021/nl8037147http://dx.doi.org/10.1021/nl8037147http://dx.doi.org/10.1021/nn900947ahttp://dx.doi.org/10.1021/nn900947ahttp://dx.doi.org/10.1021/nn800466chttp://dx.doi.org/10.1021/ja809086qhttp://dx.doi.org/10.1021/ja809086qhttp://dx.doi.org/10.1021/la902675shttp://dx.doi.org/10.1021/la902675shttp://dx.doi.org/10.1039/b911582jhttp://dx.doi.org/10.1039/b911582jhttp://dx.doi.org/10.1021/cm048297dhttp://dx.doi.org/10.1021/la8019205http://dx.doi.org/10.1021/la904189ahttp://dx.doi.org/10.1021/la904189ahttp://dx.doi.org/10.1021/cm050935khttp://dx.doi.org/10.1021/la0520029http://dx.doi.org/10.1016/0378-5173(96)04574-7http://dx.doi.org/10.1021/nn900037khttp://dx.doi.org/10.1021/la010530ohttp://dx.doi.org/10.1021/jp0107964http://dx.doi.org/10.1021/jp0107964http://dx.doi.org/10.1021/cm020732lhttp://dx.doi.org/10.1021/cm020732lhttp://dx.doi.org/10.1002/anie.201100200http://dx.doi.org/10.1002/ange.201100200http://dx.doi.org/10.1002/ange.201100200http://dx.doi.org/10.1002/adma.201100923http://dx.doi.org/10.1073/pnas.0504892102http://dx.doi.org/10.1021/bc800480khttp://dx.doi.org/10.1021/la903893nhttp://dx.doi.org/10.1021/la903893nhttp://dx.doi.org/10.1021/ja711298bhttp://dx.doi.org/10.1021/ac061730dhttp://dx.doi.org/10.1021/ja057254ahttp://dx.doi.org/10.1021/ja057254ahttp://dx.doi.org/10.1021/nl062962vhttp://-/?-http://-/?-http://-/?-http://-/?-http://-/?-8/3/2019 Leonid Vigderman et al- Quantitative Replacement of Cetyl Trimethylammonium Bromide by Cationic Thiol Ligands
6/15
[34] Z. Nie, D. Fava, E. Kumacheva, S. Zou, G. C. Walker, M.
Rubinstein, Nat. Mater. 2007, 6, 609 614.
[35] K. K. Caswell, J. N. Wilson, U. H. Bunz, C. J. Murphy, J. Am.
Chem. Soc. 2003, 125, 1391413915.
[36] M. Sethi, G. Joung, M. R. Knecht, Langmuir 2009, 25, 1572
1581.
[37] a) R. Tuinier, J. Rieger, C. G. de Kruif, Adv. Colloid Interface
Sci. 2003, 103, 1 31; b) M. C. LeMieux, Y.-H. Lin, P. D. Cuong,
H.-S. Ahn, E. R. Zubarev, V. V. Tsukruk, Adv. Funct. Mater.
2005, 15, 1529 1540.
[38] R. H. Terrill et al, J. Am. Chem. Soc. 1995, 117, 12537 12548.
[39] S. S. Agasti, C. C. You, P. Arumugam, V. M. Rotello, J. Mater.
Chem. 2008, 18, 70 73.
[40] A. C. Templeton, M. J. Hostetler, C. T. Kraft, R. W. Murray, J.
Am. Chem. Soc. 1998, 120, 1906 1911.
[41] a) B. P. Khanal, E. R. Zubarev, Angew. Chem. 2009, 121, 7020
7023; Angew. Chem. Int. Ed. 2009, 48, 68886891; b) W.-S.
Chang, L. S. Slaughter, B. P. Khanal, P. Manna, E. R. Zubarev, S.
Link, Nano Lett. 2009, 9, 1152 1157.
[42] Z. Li, P. Huang, X. Zhang, J. Lin, S. Yang, B. Liu, F. Gao, P. Xi, Q.
Ren, D. Cui, Mol. Pharm. 2010, 7, 94104.
[43] L. H. Dubois, R. G. Nuzzo, Annu. Rev. Phys. Chem. 1992, 43,
437463.
[44] R. Shukla, V. Bansal, M. Chaudhary, A. Basu, R. R. Bhonde, M.
Sastry, Langmuir 2005, 21, 10644 10654.
[45] A. Verma, O. Uzun, Y. Hu, Y. Hu, H. S. Han, N. Watson, S. Chen,
D. J. Irvine, F. Stellacci, Nat. Mater. 2008, 7, 588 595.
[46] P. Nativo, I. A. Prior, M. Brust, ACS Nano 2008, 2, 1639 1644.
Communications
6 www.angewandte.org 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim Angew. Chem. Int. Ed. 2011, 50, 1 7
These are not the final page numbers!
http://dx.doi.org/10.1038/nmat1954http://dx.doi.org/10.1038/nmat1954http://dx.doi.org/10.1038/nmat1954http://dx.doi.org/10.1038/nmat1954http://dx.doi.org/10.1038/nmat1954http://dx.doi.org/10.1021/ja037969ihttp://dx.doi.org/10.1021/ja037969ihttp://dx.doi.org/10.1021/ja037969ihttp://dx.doi.org/10.1021/ja037969ihttp://dx.doi.org/10.1021/ja037969ihttp://dx.doi.org/10.1021/ja037969ihttp://dx.doi.org/10.1021/la802845bhttp://dx.doi.org/10.1021/la802845bhttp://dx.doi.org/10.1021/la802845bhttp://dx.doi.org/10.1021/la802845bhttp://dx.doi.org/10.1021/la802845bhttp://dx.doi.org/10.1021/la802845bhttp://dx.doi.org/10.1016/S0001-8686(02)00081-7http://dx.doi.org/10.1016/S0001-8686(02)00081-7http://dx.doi.org/10.1016/S0001-8686(02)00081-7http://dx.doi.org/10.1016/S0001-8686(02)00081-7http://dx.doi.org/10.1016/S0001-8686(02)00081-7http://dx.doi.org/10.1016/S0001-8686(02)00081-7http://dx.doi.org/10.1002/adfm.200500088http://dx.doi.org/10.1002/adfm.200500088http://dx.doi.org/10.1002/adfm.200500088http://dx.doi.org/10.1002/adfm.200500088http://dx.doi.org/10.1002/adfm.200500088http://dx.doi.org/10.1021/ja00155a017http://dx.doi.org/10.1021/ja00155a017http://dx.doi.org/10.1021/ja00155a017http://dx.doi.org/10.1021/ja00155a017http://dx.doi.org/10.1021/ja00155a017http://dx.doi.org/10.1039/b711434fhttp://dx.doi.org/10.1039/b711434fhttp://dx.doi.org/10.1039/b711434fhttp://dx.doi.org/10.1039/b711434fhttp://dx.doi.org/10.1039/b711434fhttp://dx.doi.org/10.1039/b711434fhttp://dx.doi.org/10.1021/ja973863+http://dx.doi.org/10.1021/ja973863+http://dx.doi.org/10.1021/ja973863+http://dx.doi.org/10.1021/ja973863+http://dx.doi.org/10.1021/ja973863+http://dx.doi.org/10.1021/ja973863+http://dx.doi.org/10.1002/ange.200903524http://dx.doi.org/10.1002/ange.200903524http://dx.doi.org/10.1002/ange.200903524http://dx.doi.org/10.1002/ange.200903524http://dx.doi.org/10.1002/ange.200903524http://dx.doi.org/10.1002/ange.200903524http://dx.doi.org/10.1002/anie.200903524http://dx.doi.org/10.1002/anie.200903524http://dx.doi.org/10.1002/anie.200903524http://dx.doi.org/10.1002/anie.200903524http://dx.doi.org/10.1002/anie.200903524http://dx.doi.org/10.1021/nl803796dhttp://dx.doi.org/10.1021/nl803796dhttp://dx.doi.org/10.1021/nl803796dhttp://dx.doi.org/10.1021/nl803796dhttp://dx.doi.org/10.1021/nl803796dhttp://dx.doi.org/10.1021/mp9001415http://dx.doi.org/10.1021/mp9001415http://dx.doi.org/10.1021/mp9001415http://dx.doi.org/10.1021/mp9001415http://dx.doi.org/10.1021/mp9001415http://dx.doi.org/10.1146/annurev.pc.43.100192.002253http://dx.doi.org/10.1146/annurev.pc.43.100192.002253http://dx.doi.org/10.1146/annurev.pc.43.100192.002253http://dx.doi.org/10.1146/annurev.pc.43.100192.002253http://dx.doi.org/10.1146/annurev.pc.43.100192.002253http://dx.doi.org/10.1146/annurev.pc.43.100192.002253http://dx.doi.org/10.1021/la0513712http://dx.doi.org/10.1021/la0513712http://dx.doi.org/10.1021/la0513712http://dx.doi.org/10.1021/la0513712http://dx.doi.org/10.1021/la0513712http://dx.doi.org/10.1038/nmat2202http://dx.doi.org/10.1038/nmat2202http://dx.doi.org/10.1038/nmat2202http://dx.doi.org/10.1038/nmat2202http://dx.doi.org/10.1038/nmat2202http://dx.doi.org/10.1021/nn800330ahttp://dx.doi.org/10.1021/nn800330ahttp://dx.doi.org/10.1021/nn800330ahttp://dx.doi.org/10.1021/nn800330ahttp://dx.doi.org/10.1021/nn800330ahttp://www.angewandte.org/http://www.angewandte.org/http://dx.doi.org/10.1021/nn800330ahttp://dx.doi.org/10.1038/nmat2202http://dx.doi.org/10.1021/la0513712http://dx.doi.org/10.1146/annurev.pc.43.100192.002253http://dx.doi.org/10.1146/annurev.pc.43.100192.002253http://dx.doi.org/10.1021/mp9001415http://dx.doi.org/10.1021/nl803796dhttp://dx.doi.org/10.1002/anie.200903524http://dx.doi.org/10.1002/ange.200903524http://dx.doi.org/10.1002/ange.200903524http://dx.doi.org/10.1021/ja973863+http://dx.doi.org/10.1021/ja973863+http://dx.doi.org/10.1039/b711434fhttp://dx.doi.org/10.1039/b711434fhttp://dx.doi.org/10.1021/ja00155a017http://dx.doi.org/10.1002/adfm.200500088http://dx.doi.org/10.1002/adfm.200500088http://dx.doi.org/10.1016/S0001-8686(02)00081-7http://dx.doi.org/10.1016/S0001-8686(02)00081-7http://dx.doi.org/10.1021/la802845bhttp://dx.doi.org/10.1021/la802845bhttp://dx.doi.org/10.1021/ja037969ihttp://dx.doi.org/10.1021/ja037969ihttp://dx.doi.org/10.1038/nmat19548/3/2019 Leonid Vigderman et al- Quantitative Replacement of Cetyl Trimethylammonium Bromide by Cationic Thiol Ligands
7/15
Communications
Gold Nanorods
L. Vigderman, P. Manna,
E. R. Zubarev* &&&&&&&&
Quantitative Replacement of Cetyl
Trimethylammonium Bromide by
Cationic Thiol Ligands on the Surface of
Gold Nanorods and Their Extremely
Large Uptake by Cancer Cells
Stable and biocompatible : The chemical
composition of synthesized cationic thi-
olate-monolayer-protected gold nanorods
was precisely determined. In vitro cell
culture experiments showed no cytotox-
icity of these nanorods, and the number
of nanorods that were taken up by each
cancer cell exceeded two million particles
(see picture).
Angew. Chem. Int. Ed. 2011, 50, 1 7 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim www.angewandte.org
These are not the final page numbers!
http://www.angewandte.org/http://www.angewandte.org/8/3/2019 Leonid Vigderman et al- Quantitative Replacement of Cetyl Trimethylammonium Bromide by Cationic Thiol Ligands
8/15
Supporting Information
Wiley-VCH 2011
69451 Weinheim, Germany
Quantitative Replacement of Cetyl Trimethylammonium Bromide byCationic Thiol Ligands on the Surface of Gold Nanorods and TheirExtremely Large Uptake by Cancer Cells**
Leonid Vigderman, Pramit Manna, and Eugene R. Zubarev*
anie_201107304_sm_miscellaneous_information.pdf
8/3/2019 Leonid Vigderman et al- Quantitative Replacement of Cetyl Trimethylammonium Bromide by Cationic Thiol Ligands
9/15
1
Supporting Information
Synthesis of MTAB. Synthesis of 1,16-dibromohexadecane, 1. A 50 mL solution of
triphenylphosphine (3.93 g, 15 mmol) in anhydrous THF was added to a stirred solution ofN-
bromosuccinamide (2.67 g, 15 mmol) in 50 mL THF at 0C. Upon vigorous stirring, a solution of
hexadecane-1,16-diol (1 g, 3.9 mmol) in 25 mL THF was slowly added to the mixture of NBS and
Ph3P. The resulting solution was warmed to room temperature and then heated at 60C for 3.5 hours.
THF was removed by rotary evaporation and the residue was re-crystallized form ethanol to obtain 1.1
g of2 as white powder (70 % isolated yield).1H NMR (CDCl3, 400 MHz): 1.26-1.46 (m, 24 H), 1.85
(q, 4H), 3.41 (t, 4H).
Synthesis of 16-bromo-1-hexadecanethioacetate, 2. One gram (2.60 mmol) of 1 was dissolved in 40
mL methanol in a three-neck flask and the solution was degassed for one hour. Separately, 124 mg of
sodium methoxide was dissolved in 12 mL dry, ice-cold methanol and mixed with 204 mg (2.6 mmol)
of thioacetic acid. This mixture was transferred into a funnel. The solution in the three-neck flask was
refluxed under argon atmosphere and the content in the funnel was slowly added to the solution over
the course of 4 hours. After the reaction was complete, the content of the flask was cooled to room
temperature and methanol was removed under reduced pressure. The yellow oil was further purified by
column chromatography (20 % ethyl acetate in hexane) to obtain 480 mg of3 (50 % yield).
Synthesis of 16-bromo-1-hexadecanethiol 3. To a stirred solution of2 (400 mg, 1.05 mmol) in 10 mL
of methanol, 4 mL of acetyl chloride was added drop-wise and the reaction mixture was kept at 50
C
for 4 hours. After the reaction was complete, 200 mL of CH2Cl2 was added to the reaction mixture and
excess acetyl chloride and HCl were removed by multiple extractions with DI water. The mixture was
8/3/2019 Leonid Vigderman et al- Quantitative Replacement of Cetyl Trimethylammonium Bromide by Cationic Thiol Ligands
10/15
2
dried over sodium sulfate. Methylene chloride was evaporated under reduced pressure to obtain 284
mg of3 as colorless oil (80 % isolated yield).1H NMR (CDCl3, 400 MHz): 1.26-1.46 (m, 25H), 1.60
(m, 2H), 1.85 (q, 2H), 2.52 (q, 2H), 3.41 (t, 2H).
Synthesis of 16-mercaptohexadecyltrimethylammonium bromide, MTAB. To a solution of3 (284 mg,
0.85 mmol) in 5 mL of ethyl acetate, 3 mL of 4.2 M ethanolic solution of trimethylamine was added.
The mixture was vigorously stirred under argon for 4 days. The resulting white precipitate was filtered
and the washed several times with ethyl acetate to remove excess trimethylamine. The residue was
vacuum dried to obtain 270 mg of MTAB (80 % isolated yield).1H NMR (CDCl3, 400 MHz): 1.26-
1.46 (m, 25H), 1.60 (m, 2H), 1.85 (m, 2H), 2.52 (q, 2H), 3.5 (s, 9H), 3.55-3.7 (m, 2H).
Gold nanorods synthesis. Gold nanorods were synthesized according to procedure described
elsewhere.[41]
Briefly, a seed solution was created by adding 60 L of a 0.1 M NaBH4 solution to 10
ml of 5.0x10-4
M HAuCl4 in 0.1 M CTAB solution upon rigorous stirring. Separately, the growth
solution was prepared by adding 2 mL of 0.1 M AgNO3 to 2 liters of 5.0x10-4
M HAuCl4 in 0.1 M
CTAB solution. To this solution was added 11.5 mL of 0.1 M ascorbic acid solution and hand-stirred
until the solution became clear, followed immediately by the addition of 3.2 mL of seed solution. A
dark brown color indicating the presence of gold nanorods was visible after about 30 min and the
solution was allowed to sit overnight before functionalization.
Gold nanorods functionalization with MTAB thiol. Four liters of Au NRs solution was
concentrated to about 9 mL using centrifugation at 13,000 rpm. The concentrated NRs solution was
purified by two cycles of centrifuging at 13,000 rpm, removing the supernatant and redispersing the
precipitate in pure water. To this solution was added 60 mg of MTAB and the solution was stirred for
8/3/2019 Leonid Vigderman et al- Quantitative Replacement of Cetyl Trimethylammonium Bromide by Cationic Thiol Ligands
11/15
3
2 days. The MTAB-NRs were purified by five cycles of centrifugation as described earlier and finally
dispersed in 4 mL of water to a concentration of 15 mg/mL as determined by ICP-OES measurements.
Figure S1.UV-vis absorbance spectrum of CTAB-NRs (A) and MTAB-NRs (B) after purification steps by
high-speed centrifugation and redispersion into pure water.
Analysis of MTAB-NRs. For TGA analysis (TA Q-600 TGA/DSC), MTAB-NRs were first
lyophilized to obtain a useable powder. 1 mL of 15 mg/mL NR solution was placed into an Eppendorf
tube, flash frozen with liquid nitrogen, and lyophilized. In the TGA measurement, the temperature was
held at 120 C for 1 hour to remove any residual water and then ramped at 10 C/min to 850 C. For
1H-NMR analysis (Bruker 400 MHz), 1.5 mL of 15 mg/mL NR solution was transferred into D2O by
two rounds of centrifugation at 13,000 rpm and redispersion into D2O. To the D2O solution was added
50 mg KCN and the solution stirred for two days until the solution was colorless and a white
precipitate had formed. The precipitate was isolated by centrifugation and dissolved in deuterated
methanol. UV-Vis analysis was carried out on a Varian Cary 500 UV-Vis-NIR Spectrophotometer.
Zeta Potential measurements were carried out on a Malvern Instruments Zetasizer Nano-ZS.
8/3/2019 Leonid Vigderman et al- Quantitative Replacement of Cetyl Trimethylammonium Bromide by Cationic Thiol Ligands
12/15
4
Figure S2.TGA curve of MTAB-NRs. Residual water is lost at 100 C. Main weight loss of about 5.3 %
from organic component occurs from 200 to 450 C.
Calculation of MTAB binding parameters from TGA data. From TEM size analysis of ~200 NRs,
average length and width are 41.88 nm and 9.87 nm, respectively. Assume NR = cylindrical middle
with radius (r) and height (h) plus 2 spherical caps with base radius (r) and height (a). Based on TEM,
the spherical caps extend for about 2 nm on each side so h = 37.88 nm, r = is 4.935 nm, and a = 2 nm.
Thus, the surface area of the rod = (r2
+ a2) + 2rh = 1352.6 nm
2and the volume =( /3) a (3r
2+ a
2) +
r2h = 3059.6 nm
3. Mass of Au per NR = 3059.6 nm
3 59 (atoms/nm
3) 197 = 3.5610
7Da. Mass
MTAB = 0.053 (mass Au per NR + mass MTAB). Mass MTAB = 1.99106
Da. Number of MTAB
per NR = 1.99106
Da/ 397 (Da/molecule) = 5,013 MTAB molecules per NR. Molecular footprint =
1352.6 nm2
/ 5,013 molecules = 0.270 nm2
per molecule of MTAB or 3.7 MTAB molecules per nm2.
8/3/2019 Leonid Vigderman et al- Quantitative Replacement of Cetyl Trimethylammonium Bromide by Cationic Thiol Ligands
13/15
5
Figure S3. Cytotoxicity plot based on MTT analysis, which shows the percent survival of MCF-7 cells when
treated with different concentrations of CTAB-NRs (blue) and MTAB-NRs (red).
Calculation of amount of NR uptaken per cell. 500,000 cells were treated with 2 mL of medium
containing 80 g/mL MTAB-NR in medium. From ICP-OES, the total amount of up-taken gold was
determined to be 40% of 160 g or 64 g Au / 500,000 cells = 1.2810-4
(g Au/cell). Given the
density of gold and the previously calculated volume per NR of 3060 nm3
1.9310-14
(g Au/nm3) =
5.9110-11 (g Au/NR). So, 1.2810-4 (g Au/cell) / 5.9110-11 (g Au/NR) = 2.17106 NR per cell.
TEM imaging of MTAB-NR uptake into cells. MCF-7 cells were plated into a 6-well plate at
500,000 cells per well and treated with 2 mL of a 20 g/mL solution of NRs per well in medium for 24
8/3/2019 Leonid Vigderman et al- Quantitative Replacement of Cetyl Trimethylammonium Bromide by Cationic Thiol Ligands
14/15
6
hours. Cells were washed several times with PBS and fixed with a solution containing 3%
glutaraldehyde plus 2% paraformaldehyde in 0.1 M cacodylate buffer, pH 7.3, for 1 hour. After
fixation, the samples were washed and treated with 0.1% Millipore-filtered cacodylate buffered tannic
acid, postfixed with 1% buffered osmium tetroxide for 30 min, and stained en bloc with 1% Millipore-
filtered uranyl acetate. The samples were dehydrated in increasing concentrations of ethanol,
infiltrated, and embedded in LX-112 medium. The samples were polymerized in a 70oC oven for 2
days. Ultrathin sections were cut in a Leica Ultracut, stained with uranyl acetate and lead citrate in a
Leica EM Stainer, and examined in a JEM 1010 transmission electron microscope (JEOL) at an
accelerating voltage of 80 kV. Digital images were obtained using AMT Imaging System (Advanced
Microscopy Techniques Corp).
Cytotoxicity and cell uptake. MCF-7 cells were cultured in Eagles Minimum Essential Medium with
non-essential amino acids, 2 mM L-glutamine, 1 mM sodium pyruvate, and fetal bovine serum to a
final concentration of 10 %. MCF-7 cells were plated into a 96-well plate at 15,000 cells per well and
6 wells per each concentration. Cells were treated with an appropriate amount MTAB-NRs or CTAB-
NRs in medium for 24 hours, followed by the addition of 25 L of a 5 mg/mL MTT solution in PBS.
After incubating for 4 hours, the medium was removed, the cells were lysed with DMSO, and the
absorbance measured on a plate reader at 570 nm. For cell uptake imaging experiments, MCF-7 cells
were plated into a 6-well plate at 500,000 cells per well and treated with 2 ml of a 20 g/mL solution
of NRs in medium for 24 hours, trypsinized, and then plated onto a culture slide. For quantification of
NR uptake by ICP-OES, cells were treated with 2 mL of an 80 g/mL solution of NRs in medium for
24 hours, washed several times with PBS, trypsinized, pelleted, dissolved with aqua regia, and the gold
concentration measured by ICP. For SEM imaging cells were first fixed by using 2.5 % glutaraldehyde
in PBS for an hour followed by 0.4 % osmium tetroxide solution. Next, cells were dehydrated by
8/3/2019 Leonid Vigderman et al- Quantitative Replacement of Cetyl Trimethylammonium Bromide by Cationic Thiol Ligands
15/15
7
consecutive treatment with 25 %, 50 %, 70 %, 95 %, and 100 % ethanol followed by 50 % and 100%
hexamethyldisilazane and allowed to dry overnight.
Figure S4. TEM image of MTAB-functionalized gold nanorods.