Functionalized gold nanoparticles as an approach to the direct colorimetric detection of DCNP nerve agent simulant

Article abstract of DOI:10.1002/ejoc.201300339

New functionalized gold nanoparticles have been synthesized and their ability to act as colorimetric molecular probes for the naked-eye detection of nerve agent simulant DCNP has been studied. The detection process is based on the compensation of charges

Full text of DOI:10.1002/ejoc.201300339

FULL PAPER
DOI: 10.1002/ejoc.201300339
Functionalized Gold Nanoparticles as an Approach to the Direct Colorimetric
Detection of DCNP Nerve Agent Simulant
Almudena Martí,[a,b] Ana M. Costero,[a,b] Pablo Gaviña,*[a,b] Salvador Gil,[a,b]
Margarita Parra,*[a,b] Mauro Brotons-Gisbert,[c] and Juan Francisco Sánchez-Royo[c]
Keywords: Gold / Nanoparticles / Sensors / Nerve agents / Colorimetry
New functionalized gold nanoparticles have been synthe-
sized and their ability to act as colorimetric molecular probes
for the naked-eye detection of nerve agent simulant DCNP
has been studied. The detection process is based on the com-
pensation of charges at the surfaces of the functionalized
AuNPs, which triggers their aggregation with the resulting
change in the color of the solution.
ylamino)phenyl]ethanol reactive groups that are part of the
conjugated π system of donor–acceptor dyes.[5] The nucleo-
philic hydroxy group of this moiety is easily phosphorylated
by nerve agent simulants to give the intermediate II
(Scheme 1, a), which undergoes rapid intramolecular N-alk-
ylation to afford the quaternary ammonium salt III. The
change in the electronic distribution promoted by these pro-
cesses allows the colorimetric detection of DCP, DFP and
DCNP, whereas the reagents remain silent in the presence
of other organophosphorus derivatives. On the other hand,
the nucleophilic reactivity of the pyridine moiety of a push–
pull azo dye towards nerve gases has allowed us to develop
off–on colorimetric sensors (Scheme 1, b).[6]
Introduction
The use of chemical warfare (CW) agents in terrorist at-
tacks has proven the need for the development of reliable
and accurate methods for detecting these lethal com-
pounds.[1] Among CW weapons, nerve agents are especially
dangerous and the United Nations classifies them as weap-
ons of mass destruction. Nerve agents are capable of inter-
fering with the action of the nervous system. Their primary
mode of action is the inhibition of acetylcholinesterase, re-
sulting in an accumulation of acetylcholine in the synaptic
junctions, which hinders the relaxing of muscles.[2]
Analytical methods based on enzymatic assays and phys-
ical measurements have generally been used to detect these
agents.[3] However, these protocols usually have limitations,
such as low selectivity, poor portability and a certain level
of complexity. In recent years several chromo- and fluo-
rogenic sensors and probes for nerve agents have been de-
scribed.[4] Colorimetric detection is particularly appealing
because it uses low-cost, widely available instruments and
allows assays to be detected by the naked eye. Our research
group has developed a new family of reagents for the chro-
mogenic detection of the nerve agent simulants DFP, DCP
and DCNP (Figure 1) based on the use of 2-[2-(dimeth-
Figure 1. Chemical structures of different organophosphorus nerve
reagents and their simulants.
In both cases, the reaction between the probe and the
simulant generates a positive charge on the molecule. Based
on this fact, we focused on the study of the behaviour of
gold nanoparticles (AuNPs) functionalized with this type
of moiety towards nerve agents. The functionalization of
AuNPs has attracted research interest, especially in recep-
tor-based sensor applications.[7] Gold nanoparticles have
unique optoelectronic properties that can easily be tuned
by changing their size, shape or chemical environment.[7–9]
Colorimetric assays based on gold nanoparticles have re-
cently become useful for many types of analytes without the
need for advanced instrumentation because the molecular
[a] Centro de Reconocimiento Molecular y Desarrollo Tecnológico
(IDM), Unidad Mixta Universidad de Valencia-Universidad
Politécnica de Valencia,
Valencia, Spain
Fax: +34-963543151
E-mail: pablo.gavina@uv.es
margarita.parra@uv.es
Homepage: http://idm.webs.upv.es
[b] Departamento de Química Orgánica, Universidad de Valencia,
c/ Dr. Moliner 50, 46100 Burjassot, Valencia, Spain
Homepage: http://www.uv.es/
[c] Instituto de Ciencia de los Materiales, Universidad de Valencia,
c/ Dr. Moliner 50, 46100 Burjassot, Valencia, Spain
Homepage: http://www.uv.es
Supporting information for this article is available on the
WWW under http://dx.doi.org/10.1002/ejoc.201300339.
4770
© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Eur. J. Org. Chem. 2013, 4770–4779

Gold Nanoparticles for the Detection of Nerve Agent Simulants
Scheme 1. Colorimetric sensing paradigm developed by our group.
recognition events can be transformed into color the reactions of the terminal nucleophilic ligands with mo-
changes.[10] The selective and sensitive strategy for sensing is lecules of the nerve agent simulant. These positive charges
based on the color change that arises from the interparticle partially neutralize the original negative charges of the
plasmon coupling that occurs during aggregation of AuNPs nanoparticles, inducing their aggregation and thus a change
or the dispersion of AuNP aggregates. The red color of dis- in their surface plasmon resonance absorption and conse-
persed nanoparticles turns to dark blue upon aggregation, quently a change in their color.
and this color change can be observed by the naked eye
even at low concentrations.[11] Thus, the generation of posi-
tive charges on the surfaces of dispersed functionalized an-
ionic AuNPs as a result of their reaction with nerve agents
will induce their aggregation with the concomitant color
change.
Recently, the detection of highly toxic organophosphate
pesticides by using thioctic acid capped AuNPs as colori-
metric probes in an indirect strategy with very low detection
limits has been described.[7c,12,13] In that research sensors
were developed for the detection of thiocholine obtained by
catalytic hydrolysis of acetylthiocholine by acetylcho-
linesterase. The ligand-exchange reaction between cationic
thiocholine and the negatively charged thioctic carboxylate
induces the aggregation of Au nanoparticles. The irrevers-
ible inhibition of acetylcholinesterase by the organophos-
phorus pesticides prevents aggregation and the red color of
the Au nanoparticles remains.
Scheme 2. Paradigm of the sensing mechanism. Reaction of the
terminal ligands with the simulant produces positive charges that
can compensate the negative charges of the AuNPs and induce ag-
gregation processes.
Synthesis of the Ligands
Following a completely different approach, we report
herein the results obtained in the direct detection of DCNP
(as simulant of tabun) by using thioctic acid capped AuNPs
functionalized with pyridines or 2-[2-(dimethylamino)phen-
yl]ethanol derivatives.
AuNPs have affinity for both reduced (thiols) and oxid-
ized (disulfide) sulfur groups through Au–S bonds, and thus
both groups can be used as anchors for the functionaliza-
tion of nanoparticles. The different ligands used in this
study are presented in Figure 2.
Ligands 1 and 2 are commercially available, whereas 3
and 4 were obtained by the esterification of the correspond-
ing alcohol or disulfide derivatives with thioctic acid (TA)
under Steghich[14] or Mitsunobu[15] conditions. Ligands 5–
Results and Discussion
The sensing paradigm of the detection process, which is 7 were synthesized by an azo-coupling reaction between
presented in Scheme 2, involves the generation of positive pyridine-4-diazonium (obtained from 4-aminopyridine by
charges on the surfaces of the gold nanoparticles through diazotization) and phenol, N-phenyl-N-methyl-2-amino-
Eur. J. Org. Chem. 2013, 4770–4779
© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
4771

P. Gaviña, M. Parra et al.
FULL PAPER
Figure 2. Ligands 1–8 synthesized for the functionalization of AuNPs.
ethanol or 3-(dimethylamino)phenol, respectively, followed which was lithiated with BuLi and then transformed into
by esterification with TA. Ligand 8 required a seven-step the formyl derivative 13 by reaction with DMF. Compound
synthesis and is depicted in Scheme 3. Thus, 2-(2-ni- 13 was subsequently reduced to the corresponding benzylic
trophenyl)ethanol (9) was reduced in the presence of form- alcohol 14. After esterification with thioctic acid and depro-
aldehyde to the corresponding dimethylamino derivative 10. tection of the silyl-protected hydroxy moiety with tert-
This amino compound was transformed into the 4-bromo butylammonium fluoride (TBAF), ligand 8 was obtained in
derivative 11 by reaction with N-bromosuccinimide (NBS). a 20% overall yield. The chemical structures and purity of
Protection of the hydroxy group as a silyl ether yielded 12, ligands 1–8 were confirmed by spectroscopic techniques.
Scheme 3. Synthesis of ligand 8.
4772
www.eurjoc.org
© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Eur. J. Org. Chem. 2013, 4770–4779

Gold Nanoparticles for the Detection of Nerve Agent Simulants
Synthesis of the Functionalized Gold Nanoparticles
In this study, the nanoparticles were centrifuged and re-
dissolved in a buffered aqueous solution (MOPS) at pH 7.
In this medium, no obvious change either in color nor in
the characteristic peak at 526 nm was observed.
The resulting functionalized gold nanoparticles were
characterized by UV/Vis and zeta-potential measurements,
and the results are summarized in Table 1. In some cases
the AuNPs were also characterized by TEM or XPS.
AuNPs can be synthesized by different procedures,[16–18]
the size, stability and optical properties of the resulting
nanoparticles being strongly dependent upon the method
used. The functionalized gold nanoparticles AuNP1-
AuNP8 were synthesized by a two-step procedure. First, cit-
rate-stabilized nanoparticles were prepared by reducing tet-
rachloroauric acid with trisodium citrate in boiling
water.[8,19] In this step we found that both the relative con-
centration of the reagents and thorough cleaning of the
glassware with aqua regia were crucial to avoid either nu-
cleation during the synthesis or aggregation of the gold col-
loid solutions. Monodisperse citrate-stabilized nanopar-
ticles were thus obtained with an average size of 13 nm, as
Table 1. Characterization of the gold nanoparticles.[a]
AuNP Optimized TA:L SPR peak
c [m]
ζ-potential
molar ratio
λmax [nm]
[mV][b]
AuNP2
AuNP3
AuNP4
1:1
526
526
526
524
520
525
524
1.29ϫ10–8
9.47ϫ10–8
9.47ϫ10–8
1.18ϫ10–8
5.79ϫ10–9
8.32ϫ10–9
1.13ϫ10–8
–32.5
–35.6
1.2:1
1.2:1
1.2:1
1.2:1
4:1
determined by TEM. The surface plasmon peak appears at AuNP5
–37.3
–35.9
AuNP6
526 nm, in perfect agreement with the experimental data for
AuNP7
particle sizes smaller than 25 nm.[8] The molar extinction
AuNP8
–36.2
1.5:1
–24.0[c]
coefficient was estimated to be ε = 2.47ϫ108 m–1 cm–1 (ob-
[a] Dispersed in buffered aqueous solution (MOPS, pH 7). [b] ZP =
–37.5 mV for TA-capped AuNPs. [c] Suspended in deionized water.
tained from the plot of ε vs. nanoparticle size reported by
Huo and co-workers[20]). The initial concentration of the
citrate-capped AuNPs was calculated from the Beer–Lam-
bert law to be 5.3ϫ10–9 m.
In the second step, in a ligand-exchange reaction, the cit-
rate was displaced from the surface of the nanoparticles by
a mixture of thioctic acid and the different ligands 1–8. In
each case it was necessary to optimize the ligand/TA molar
ratio (from 1:1 to 1:4) to improve the stabilization of the
nanoparticles as well as the response with respect to the
analyte.
AuNP1 experienced aggregation during the functionali-
zation reaction and this prevented its use in the detection
studies. The other gold nanoparticles remained stable in
cold, buffered aqueous solution for at least a month, which
is in agreement with their high zeta-potential values in this
medium.
Figure 3 (a) shows the Au 4f core level spectra measured
by XPS for the samples AuNP2, AuNP5 and AuNP7 as
As an alternative, a three-step protocol was tested for the well as for TA- and citrate-capped AuNPs. The presence of
functionalization of AuNPs in which the citrate was first Au was detected in all of them, although the Au 4f doublets
displaced by TA and the resulting TA-capped AuNPs were measured in the functionalized samples tend to shift to
stirred in the presence of ligands for a second ligand-ex- higher binding energies with respect to the unfunctionalized
change reaction. However, the degree of functionalization sample. This fact can be attributed to charge-transfer effects
was much lower by this method, as evidenced by a weaker between the Au nanoparticles and the incorporated ligands,
response towards the war agent simulant.
indicating that Au tends to donate electrons.
Figure 3. XPS spectra for Au, N and S atoms in AuNP2, AuNP5, AuNP7, TA-capped and citrate-capped (reference) AuNPs.
Eur. J. Org. Chem. 2013, 4770–4779
© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
4773

P. Gaviña, M. Parra et al.
FULL PAPER
Figure 3 (b,c) show the N 1s and S 2p core level spectra
measured by XPS, respectively. In most of the samples a
weak but distinguishable feature attributable to a signal of
the N 1s core level can be observed at binding energies of
around 400 eV, but is not detectable in the TA-function-
alized AuNP nor in the reference sample. In the binding
energy region corresponding to the S 2p core level, a clear
structure emerges between 160 and 172 eV in the function-
alized samples. In these samples, the S 2p spectra appear to
be dominated by photoelectrons with binding energies of
around 162–163 eV, which can be attributed to sulfur atoms
bound and unbound to the gold surface, accompanied by a
weaker structure appearing at around 168 eV attributable
to oxidized S atoms.
Sensing Studies
To verify the reactivity of the pyridine ligands with the
DCNP simulant, a preliminary study was conducted with
free ligands 5–7 in acetonitrile (the lack of absorption of
ligands 1–4 in the UV/Vis region precluded their study).
The UV/Vis spectra of ligands 5 and 6 in acetonitrile
(5ϫ10–5 m) show a band centred at 316 and 431 nm (logε
= 4.36 and 4.31 m–1 cm–1), respectively. The addition of
30 equiv. of DCNP to each ligand induced in both cases a
bathochromic shift to give a new band centred at 340 and
532 nm, respectively, corresponding to the phosphorylation
of the pyridine moiety, which enhances the intramolecular
charge-transfer process in the dyes. The UV spectrum of
ligand 7 in acetonitrile (5ϫ10–5 m) shows a band centred
at 454 nm (logε = 4.15 m–1 cm–1). In this case, addition of
30 equiv. of DCNP induced the appearance of two broad
absorption bands, which, upon deconvolution, split into
four well-defined bands at 394, 431, 507 and 537 nm (see
Figure 4). Based on previous studies,[6] these bands were as-
signed to ligand 7 protonated at the dimethylamino group
(band centred at 431 nm, confirmed by the addition of
30 equiv. of HCl to a sample of ligand 7 in acetonitrile) and
ligand 7 phosphorylated in three different places. The band
centred at 537 nm, which corresponds to the phosphoryl-
ation of the pyridine moiety, increases the pull–push char-
acter of the chromophore, which accounts for the batho-
Figure 4. UV/Vis spectrum of ligand 7 (5ϫ10–5 m in acetonitrile)
on addition of 30 equiv. of DCNP. Deconvolution of the two bands
and assigned products.
chromic shift similar to ligand 6. The bands at 394 and aqueous MOPS solutions, a color change from red wine to
507 nm can be assigned to the phosphorylation of the di- purple blue was readily observed by the naked eye. This
methylamino and azo groups of the ligand. The phosphor- change is indicative of the aggregation of AuNPs. The sens-
ylation of the aniline moiety was expected to induce a de- ing experiments were carried out in buffered solutions to
crease in the charge-transfer character of dye 7, as we pre- prevent any acidic system slowing or even interfering in the
viously observed for a similar system.[6]
On the basis of these results we expected that the phos- cause they were hydrolysed in this medium.
sensing process. DFP and DCP could not be studied be-
phorylation of the pyridine moiety in nanoparticles
The ability of these functionalized AuNPs to act as
AuNP2–AuNP7 or the formation of an ammonium salt in probes for DCNP detection was studied by UV/Vis spec-
AuNP8 promoted by the presence of DCNP would change troscopy. The UV/Vis spectra are consistent with the ob-
the overall electrostatic charge on the surfaces of the nano- served color changes. Thus, the intensity of the surface plas-
particles, triggering an aggregation process coupled with a mon peak of the monodispersed AuNPs at 526 nm (A526
bathochromic shift of the SPR absorption band and a decreased and a new peak at around 660 nm (A660) ap-
)
change in the color of the solution.
peared as clusters of AuNP formed. Figure 5 shows the re-
In fact, when DCNP (5ϫ10–2 m, in acetonitrile) was sults of UV/Vis titration studies performed with AuNP3
added to a suspension of the prepared AuNP2–AuNP8 in and AuNP6, as representative examples, in the presence of
4774
www.eurjoc.org
© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Eur. J. Org. Chem. 2013, 4770–4779

Gold Nanoparticles for the Detection of Nerve Agent Simulants
increasing amounts of DCNP. Such changes in the spectra,
The variation in A670/A526 versus DCNP concentration
concomitant with the change in color of the solution, can for AuNP3 and AuNP6 is also presented in Figure 5. A
be well explained by the DCNP-induced aggregation of the very significant increase in the A660/A526 ratio was observed
nanoparticles through charge neutralization at the surface. as the concentration of DCNP was increased, above
The aggregation of the AuNPs was further confirmed by 600 ppm for AuNP3 and above 200 ppm for AuNP6, which
TEM analysis (see Figure 6 for AuNP3).
reflects the greater nucleophilic character of ligand 6. The
Figure 5. UV/Vis spectra of AuNP3 (top left) and AuNP6 (top right) on addition of increasing amounts of DCNP (5ϫ10–2 m in acetoni-
trile) expressed in ppm (mg/L). Plots of A660/A526 vs. DCNP concentration for AuNP3 (bottom left) and AuNP6 (bottom right).
Figure 6. TEM images of a stabilized AuNP3 dispersion (left) and its aggregates (right) upon addition of excess DCNP at a resolution
of 200 nm (inset: the same aggregates at a resolution of 1000 nm).
Eur. J. Org. Chem. 2013, 4770–4779
© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
4775

P. Gaviña, M. Parra et al.
FULL PAPER
visual limits of detection (visual LODs, defined as the mini- UV/Vis spectra, see the Supporting Information) observed
mum concentration of simulant necessary for an observable for DCNP are presented in Table 2.
colour change) and calculated LODs (from the changes in
As expected, AuNP4 and AuNP7 showed modest results,
probably due to strong steric hindance of the aliphatic ester
chain near the pyridine nucleophile, which prevents access
of the organophosphate agent to the nitrogen centre. The
greater nucleophilic character of the azo-substituted pyr-
idines in ligands 5 and 6 in comparison with ligand 3 would
explain the higher LOD observed for the latter.
The best results were obtained with the gold nanopar-
ticles functionalized with azo dye 6 (AuNP6) and the di-
methylaminophenylethanol ligand 8 (AuNP8; Figure 7),
which can detect 76 and 81 ppm of DCNP, respectively,
thus improving the limit previously obtained by us in solu-
tion by a similar approach (see Scheme 1, b).[6]
Table 2. Visual and calculated LODs of DCNP with AuNP1–
AuNP8.
AuNP2 AuNP3 AuNP4 AuNP5 AuNP6 AuNP7 AuNP8
Visual
LOD
[ppm]
Calcd.
LOD
[ppm]
456
597
1151
310
158
796
106
229
470 Ͼ1100 340
76
750
81
To demonstrate the selectivity of the system, studies were
undertaken that confirmed that AuNP3, AuNP4, AuNP5
and AuNP8 showed only modest responses in their UV/Vis
spectra with some of the interference agents that may be
present in military or civilian settings such as some pesti-
cides (malathion, dyfonate, 4,4Ј-DDD, 4,4Ј-DDE) and gas-
oline and diesel fuel at concentrations of 2.4 mm in buffered
aqueous solution (see Figure S15 in the Supporting Infor-
mation). No changes in the color of the solutions could be
observed with any of these interferents, the gold nanopar-
ticles remaining stable as a red monodispersion.
Conclusions
Functionalized AuNPs have proven to be an interesting
alternative for the direct colorimetric detection of nerve
agents in buffered aqueous media provided suitable ligands
are anchored to the surfaces of the AuNPs. We have
achieved detection limits of 76 and 81 ppm (mg/L) for li-
gands 6 and 8 and found that both steric and electronic
effects are crucial for obtaining good results.
Experimental Section
General Procedures: All reagents were commercially available and
used without purification. Silica gel 60 F254 (Merck) plates were
used for TLC. Milli-Q ultrapure water was used for the synthesis
of the AuNPs and in the sensing experiments. 1H and 13C NMR
spectra were recorded with a Bruker 300 MHz spectrometer. Chem-
ical shifts are reported in ppm with tetramethylsilane as an internal
standard. High-resolution mass spectra were recorded in the posi-
tive ion mode with a VG-AutoSpec mass spectrometer. UV/Vis ab-
sorption spectra were recorded in a 1 cm pathlength quartz cuvette
with a Shimadzu UV-2101PC spectrophotometer. All measure-
ments were carried out at 293 K (thermostatted). Zeta potentials
were measured with a Malvern Zetasizer ZS three times in 10–25
cycles. X-ray photoelectron spectroscopy (XPS) was employed to
verify the success of the molecular modification of the AuNP sur-
faces. The spectra were recorded with an Escalab 210 spectrometer
from Thermo VG Scientific. The base pressure in the analysis
chamber was 1.0ϫ10–10 mbar. Photoelectrons were extracted by
using the Mg-Kα excitation line (hν = 1253.6 eV). The binding en-
ergy of the spectra refers to the Fermi level. The electronic images
Figure 7. Representation of a AuNP functionalized with ligand 8
(top). Plot of A660/A526 vs. DCNP concentration (left).
4776
www.eurjoc.org
© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Eur. J. Org. Chem. 2013, 4770–4779

Gold Nanoparticles for the Detection of Nerve Agent Simulants
were obtained with a JEOL-1010 transmission electron microscope
operating at 100 kV.
= 9.1 Hz, 2 H), 3.68–3.54 (m, 1 H), 3.22–3.06 (m, 2 H), 2.67 (t, J
= 7.3 Hz, 2 H), 2.57–2.43 (m, 1 H), 1.95–1.86 (m, 1 H), 1.84–1.69
(m, 4 H), 1.65–1.55 (m, 2 H) ppm. 13C NMR (75 MHz, CD3OD):
δ = 172.37, 153.42, 144.38, 134.38, 134, 06, 129.61, 127.14, 123.05,
121.49, 56.81, 40.23, 38.56, 34.66, 33.57, 28.01, 24.64 ppm. HRMS:
calcd. for C19H22N3O2S2 [MH]+ 388.1153; found 388.1157.
Ligands 1 and 2 were commercially available and were used as re-
ceived.
Synthesis of Ligand 3: Dicyclohexylcarbodiimide (DCC; 5.19 g,
25.2 mmol) was added portionwise to a cold solution of pyridine-4-
methanol (2.72 g, 13.2 mmol) in dichloromethane (DCM; 20 mL).
Then thioctic acid (2.72 g, 13.2 mmol) and 4-(dimethylamino)pyr-
idine (DMAP; catalytic amounts) in DCM (20 mL) were added
and the mixture was stirred for 72 h at room temperature. Then aq.
NH4Cl was added and the resulting precipitate was removed and
the remaining solution was extracted with DCM (3ϫ 10 mL). The
combined organic layers were washed with water and then dried
with MgSO4. The solvent was evaporated and the crude product
was purified by column chromatography (silica gel, DCM/AcOEt,
1:1) to yield the corresponding ester 3 (2.04 g, 52%) as a yellow
Synthesis of Ligand 6: 4-aminopyridine (0.367 g, 4 mmol) was dis-
solved in a mixture of concentrated phosphoric acid (25 mL) and
concentrated nitric acid (12 mL). This solution was slowly added
to a solution containing sodium nitrite (0.334 mg, 4.8 mmol) and
water (8 mL) at –5 °C. The generated diazonium salt was immedi-
ately added to a solution containing 2-[methyl(phenyl)amino]eth-
anol (0.609 mg, 4.03 mmol) and 30% phosphoric acid (20 mL). The
reaction mixture was allowed to react for 30 min at –5 °C and then
for 60 min at room temperature. The final dark-red crude was neu-
tralized with a saturated sodium carbonate solution and the or-
ganic product extracted with DCM. The organic layer was dried
with MgSO4, filtered and the solvent eliminated in a rotary evapo-
rator. 4-Azidopyridine (0.234 g, 0.53 mmol, 45%) was isolated as a
dark-red solid by column chromatography using AcOET/hexane
(1:1) as eluent. 1H NMR (300 MHz, CDCl3): δ = 8.71 (dd, J = 4.6,
1.6 Hz, 2 H), 7.90 (d, J = 9.3 Hz, 2 H), 7.63 (dd, J = 4.6, 1.6 Hz,
2 H), 6.82 (d, J = 9.3 Hz, 2 H), 3.89 (m, 2 H), 3.66 (t, 2 H), 3.16
(s, 3 H) ppm. 13C NMR (75 MHz, CDCl3): δ = 149.72, 159.68,
148.34, 130.95, 120.83, 113.14, 11.52, 63.92, 58.47, 41.04 ppm.
HRMS: calcd. for C14H16N4O [M]+ 256.303; found 256.286.
1
oil. H NMR (300 MHz, CD3OD): δ = 8.52 (dd, J = 4.5, 1.7 Hz,
2 H), 7.45–7.38 (m, 2 H), 5.19 (s, 2 H) 3.56 (ddd, J = 12.2, 8.6,
6.3 Hz, 1 H), 3.22–3.03 (m, 2 H), 2.51–2.38 (m, 3 H), 1.94–1.80 (m,
1 H), 1.76–1.59 (m, 4 H), 1.55–1.41 (m,2 H) ppm. 13C NMR
(75 MHz, CD3OD): δ = 175.0, 150.7, 124.0, 65.5, 58.0, 41.7, 39.8,
36.1, 35.1, 30.2, 26.2 ppm. HRMS: calcd. for C14H19NO2S2 [M]+
297.0857; found 297.0832.
Synthesis of Ligand 4: Thioctic acid (2.00 g, 9.69 mmol), Ph3P
(3.05 g, 11.6 mmol) and 2,2Ј-dipyridyl disulfide (2.56 g, 11.6 mmol)
were dissolved in THF (30 mL). The solution was stirred at room
temperature for 24 h under argon and then concentrated in vacuo
to give the crude product. Purification by column chromatography
(hexane/EtOAc, 2:1) afforded 2.26 g (78%) of ligand 4[21] as a yel-
DCC (1.04 mmol) was added portionwise to a cold solution of (E)-
2-{methyl[4-(pyridin-4-yldiazenyl)phenyl]amino}ethanol (0.87 mmol)
in DCM (25 mL). Then a cold solution of thioctic acid (1.04 mmol)
and DMAP (catalytic amount) in DCM (25 mL) was added and
the mixture was stirred for 48 h at room temperature. After this
time a saturated solution of ammonium chloride was added and
the precipitated removed. The mixture was extracted with DCM
(3ϫ 10 mL) and the combined organic layers were washed with
water, dried with MgSO4, evaporated and further purified by col-
umn chromatography (DCM/ethyl acetate, 1:1) to yield the corre-
1
low oil. H NMR (400 MHz, CDCl3): δ = 8.58 (ddd, J = 4.8, 2.0,
0.8 Hz, 1 H), 7.59 (d, J = 7.6 Hz, 1 H), 7.71 (dt, J = 8.0, 2.0 Hz, 1
H), 7.26 (ddd, J = 7.6, 5.2, 1.2 Hz, 1 H), 3.53 (q, J = 6.4 Hz, 1 H),
3.17–3.04 (m, 2 H), 2.69 (t, J = 7.4 Hz, 2 H), 2.42 (sext, J = 6.8 Hz,
1 H), 1.87 (sext, J = 6.8 Hz, 1 H), 1.77–1.63 (m, 4 H), 1.43–1.53
(m, 2 H) ppm. 13C NMR (75 MHz, CDCl3): δ = 196.0, 151.2,
150.1, 137.2, 130.1, 123.5, 56.1, 43.8, 40.1, 38.4, 34.5, 28.4,
25.0 ppm.
1
sponding ester 6 (0.243 g, 45%) as a dark-orange solid. H NMR
(300 MHz, MeOD): δ = 8.70 (d, J = 6.2 Hz, 2 H), 7.96–7.85 (m, 2
H), 7.72–7.64 (m, 2 H), 6.84–6.73 (m, 2 H), 4.31 (t, J = 5.8 Hz, 2
H), 3.73 (t, J = 5.8 Hz, 2 H), 3.56–3.42 (m, 1 H), 3.14–3.09 (m, 2
H), 3.14–3.09 (s, 3 H), 2.48–2.33 (m, 1 H), 2.24 (t, J = 7.3 Hz, 2
H), 1.92–1.76 (m, 1 H), 1.69–1.51 (m, 4 H), 1.47–1.32 (m, 2
H) ppm. 13C NMR (75 MHz, CDCl3): δ = 173.70, 158.93, 150.09,
126.74, 116.82, 112.04, 60.81, 56.70, 51.22, 40.62, 39.29, 38.87,
34.95, 29.97, 24.88 ppm. 13C NMR (75 MHz, CDCl3): δ = 173.7,
159.2, 153.0, 144.4, 126.8, 116.9, 112.0, 61.6, 56.7, 51.2, 40.6, 38.9,
35.0, 34.3, 29.1, 24.9 ppm. HRMS: calcd. for C22H28N4O2S2 [M]+
444.165; found 444.169.
Synthesis of Ligand 5: Phenol (5.0 g, 53 mmol) and sodium nitrite
(4.0 g, 58 mmol) in aqueous NaOH (10% w/w, 20 mL) was added
dropwise to a solution of 4-aminopyridine (6.0 g, 64 mmol) in 7.3 m
HCl (45 mL) at 0 °C. The pH of the mixed solution was adjusted
to pH 6–7 by the addition of a 10% aqueous solution of NaOH.
The resulting mixture was stirred at 0 °C for 2 h. After neutraliza-
tion of the solution with sodium hydrogen carbonate, an orange
precipitate was obtained. Purification by crystallization with meth-
anol and DCM afforded 4-(4-hydroxyphenylazo)pyridine[22] (4.6 g,
1
44%) as an orange solid. H NMR (300 MHz, CDCl3): δ = 10.57
(s, 1 H), 8.77 (d, J = 6.0 Hz, 2 H), 7.90–7.85 (m, 2 H), 7.68 (dd, J
= 4.5, 1.6 Hz, 2 H), 7.02–6.94 (m, 2 H) ppm.
Synthesis of Ligand 7: 4-Aminopyridine (0.121 g, 1.29 mmol) was
DCC (0.988 g, 4.79 mmol) was added portionwise to a cold solu-
dissolved in water (10 mL). Then concentrated sulfuric acid
tion of 4-(4-hydroxyphenylazo)pyridine (0.454 g, 2.28 mmol) in (0.5 mL, 9.2 mmol) was added and the reaction mixture was heated
DCM (20 mL),. Then a cold solution of thioctic acid (0.518 g,
2.51 mmol) and a catalytic amount of DMAP in DCM (20 mL)
was added and the mixture was stirred for 72 h at room tempera-
ture. After this time, a saturated aqueous solution of ammonium
chloride was added and the resulting precipitate was removed. The
mixture was extracted with DCM (3ϫ 10 mL). The combined or-
ganic layers were washed with water, dried with MgSO4, evaporated
and further purified by column chromatography (DCM/AcOEt,
1:1) to yield the corresponding ester 5 (0, 353 g, 40%) as an orange
to give a solution. The reaction mixture was cooled to 0 °C and
then a solution of NaNO2 (0.089 g, 1.29 mmol) in water (5 mL)
was added dropwise. Then a solution of (dimethylamino)phenol
(0.206 g, 1.5 mmol) was added dropwise during 30 min. The re-
sulting orange solution was stirred for 30 min at 0 °C and then
for 30 min at room temperature. Then the reaction mixture was
neutralized with an aqueous saturated solution of potassium acet-
ate (30 mL). The precipitate was purified by column chromatog-
raphy (hexane/ethyl acetate, 9:1) to yield (E)-5-(dimethylamino)-2-
solid. 1H NMR (300 MHz, CD3OD): δ = 8.77 (d, J = 6.4 Hz, 2 (pyridin-4-yldiazenyl)phenol (0.122 g, 39%) as a brown solid. 1H
H), 8.06 (d, J = 9.1 Hz, 2 H), 7.84 (d, J = 6.4 Hz, 2 H), 7.36 (d, J NMR (300 MHz, CDCl3): δ = 15.62 (s, OH), 8.59 (s, 1 H), 7.45 (d,
Eur. J. Org. Chem. 2013, 4770–4779
© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
4777

P. Gaviña, M. Parra et al.
FULL PAPER
J = 6.1 Hz, 2 H), 7.32 (d, J = 9.3 Hz, 1 H), 6.52 (dd, J = 9.2,
2.6 Hz, 1 H), 5.91 (d, J = 2.6 Hz, 1 H), 5.30 (s, 1 H), 3.17 (s, 6
H) ppm. 13C NMR (75 MHz, CDCl3): δ = 159.72, 158.91, 151.87,
1
0.2106 g, 83%) as a pale-yellow oil. H NMR (300 MHz, CDCl3):
δ = 7.37–7.18 (m, 2 H), 6.94 (d, J = 8.5 Hz, 1 H), 3.81 (t, J =
7.25 Hz, 2 H), 2.87 (t, J = 7.0 Hz, 2 H), 2.62 (s, 6 H), 0.86 (s, 9 H),
148.37, 131.14, 114.01, 104.12, 99.18, 96.25, 41.29 ppm. HRMS: 0.00 (s, 6 H) ppm. 13C NMR (75 MHz, CDCl3): δ = 152, 137.03
calcd. for C13H14N4O [M]+ 242.1168; found 242.1194.
133.18, 131.2, 121.82, 118.1, 63.82, 45.60, 34.64, 26.33, 18.74,
–5.01 ppm. HRMS: calcd. for C16H28BrNOSi [M]+ 357.1124;
found 357.1192.
DCC (1.392 mmol) was added portionwise to a cold solution of
(E)-5-(dimethylamino)-2-(pyridin-4-yldiazenyl)phenol (0.87 mmol)
in DCM (25 mL). Then a cold solution of thioctic acid (1.04 mmol)
and DMAP (catalytic amount) in DCM (25 mL) was added and
the mixture was stirred for 48 h at room temperature. After this
time, a saturated solution of ammonium chloride was added and
the precipitate removed. The mixture was extracted with DCM (3ϫ
10 mL) and the combined organic layers were washed with water,
dried with MgSO4, evaporated and further purified by column
chromatography (DCM/ethyl acetate, 1:1) to yield the correspond-
ing ester 7 (0.150 g, 40%) as an orange solid. 1H NMR (300 MHz,
CDCl3): δ = 8.64 (dd, J = 4.6, 1.6 Hz, 1 H), 7.79 (d, J = 9.3 Hz, 1
H), 7.49 (dd, J = 4.6, 1.6 Hz, 2 H), 6.54 (dd, J = 9.3, 2.8 Hz, 1 H),
6.36 (d, J = 2.7 Hz, 1 H), 3.50 (dq, J = 13.2, 6.6 Hz, 1 H), 3.07 (s,
2 H), 3.08–3.04 (m, 3 H), 2.62 (t, J = 7.4 Hz, 1 H), 2.40 (dq, J =
6.6, 5.5 Hz, 1 H), 2.30 (t, J = 7.3 Hz, 2 H), 1.93–1.78 (m, 3 H) ppm.
A solution of bromide 12 (1.49 g, 4.15 mmol) dissolved in THF
(30 mL) was cooled to –78 °C under Ar. n-Butyllithium (6.5 mL,
5 mmol) was added dropwise with stirring and then warmed to
0 °C and kept at this temperature for an additional 1 h. Again the
solution was cooled to –78 °C and dry dimethylformamide
(0.96 mL, 12.48 mmol) was added. The reaction was stirred at
room temperature for 2 h and water (15 mL) was added. The sol-
vents were removed in vacuo and the residue redissolved in DCM.
The organic extract was dried with anhydrous MgSO4 and filtered.
3-{2-[(tert-Butyldimethylsilyl)oxy]ethyl}-4-dimethylaminobenzal-
dehyde (13) was obtained after silica gel column chromatography
(ethyl acetate/hexane, 1:5) in 98% yield (1.26 g). 1H NMR
(300 MHz, CDCl3): δ = 9.83 (s, 1 H), 7.76 (d, J = 2.1 Hz, 1 H),
7.68 (dd, J = 8.3 and 2.1 Hz, 1 H), 7.07 (d, J = 8.3 Hz, 1 H), 3.86
13C NMR (75 MHz, CDCl3): δ = 172.4, 159.7, 151.7, 148.2, 142.0, (t, J = 7.0 Hz, 2 H), 2.93 (t, J = 7.0 Hz, 2 H), 2.78 (s, 6 H), 0.84
130.1, 113.0, 108.3, 104.7, 96.4, 56.4, 41.2, 40.3, 38.7, 34.6, 33.5,
28.1, 24.6 ppm. HRMS: calcd. for C21H26N4O2S2 [M]+ 430.150;
found 430.146.
(s, 9 H), –0.02 (s, 6 H) ppm. 13C NMR (75 MHz, CDCl3): δ =
191.8, 159.2, 133.2, 132.8, 130.9, 129.7, 118.9, 67.9, 63.5, 50.3, 45.8,
44.7, 35.2, 31.4, 26.4, 26.3, 18.7, –5.0 ppm. HRMS: calcd. for
C17H29NO2Si [M]+ 307.1968; found 307.2036.
Synthesis of Ligand 8: 2-(2-Nitrophenyl)ethanol (2.3 g,
19.67 mmol), formaldehyde (4.83 mL, 32.3 mmol, 36%) and Pd/C NaBH4 (0.034 g, 9.0 mmol) was added to a solution of 13 (0.461 g,
(480 mg, 10%) were dissolved in absolute ethanol (200 mL) and
placed under H2 until uptake of hydrogen had ceased. After fil-
tration through Celite, the solvent was evaporated and the product
was purified by extraction with ethyl acetate and washed with
water. The organic layer was washed with brine and dried with
MgSO4 to give 2-(dimethylaminophenyl)ethanol (10; 3.22 g, 95%)
15 mmol) in MeOH (30 mL), and the mixture was heated at 60 °C
for 2.5 h. After addition of water (20 mL) at 0 °C, work-up was
performed with ethyl acetate and water, and the combined organic
layers were washed with brine and dried with MgSO4. After fil-
tration followed by evaporation, the residue was subjected to col-
umn chromatography through silica gel (ethyl acetate/hexane, 3:7)
to afford pure 3-{2-[(tert-butyldimethylsilyl)oxy]ethyl}-4-[(dimeth-
ylamino)phenyl]methanol (14; 0.424 g, 91%) as a pale-yellow solid.
1H NMR (300 MHz, CDCl3): δ = 7.23–7.13 (m, 2 H), 7.13–7.05
(m, 1 H), 4.59 (s, 2 H), 3.82 (t, J = 7.3 Hz, 2 H), 2.92 (t, J = 7.3 Hz,
2 H), 2.64 (s, 6 H), 1.61 (s, 2 H), 1.35–1.28 (m, 1 H), 0.87 (s, 9 H),
0.01 (s, 6 H) ppm. 13C NMR (75 MHz, CDCl3): δ = 153.3, 136.1,
134.7, 130.1, 126.3, 120.3, 65.7, 64.3, 45.7, 35.1, 26.4, 18.8 ppm.
HRMS: calcd. for C19H29NO2Si [M]+ 309.5190; found 309.1402.
1
as a yellow oil. H NMR (300 MHz, CDCl3): δ = 7.19–6.95 (m, 4
H), 3.76 (t, J = 5.9 Hz, 2 H), 2.91 (t, J = 5.7 Hz, 2 H), 2.62 (s, 6
H) ppm. 13C NMR (75 MHz, CDCl3): δ = 152.6, 136.5, 131.5,
128.0, 125.4, 120.4, 64.7, 45.4, 36.5 ppm.
NBS (0.187 g, 1.05 mmol) was added to a mixture of 2-(dimethyl-
aminophenyl)ethanol (10; 0.165 g, 1 mmol) and NH4OAc (10 mol-
%) in MeCN (5 mL) and the mixture was stirred at room tempera-
ture. After completion of the reaction as indicated by TLC, the
mixture was concentrated in vacuo and extracted with EtOAc/H2O
(1:1, 3ϫ 5 mL). The organic portion was separated from the ex-
tract, dried and concentrated. The residue was subjected to column
chromatography (silica gel, hexane/EtOAc, 10:1) to obtain pure 2-
(5-bromo-2-dimethylaminophenyl)ethanol (11) as a brown oil.
DCC (0.480 g, 2.32 mmol) was added portionwise to a cold of solu-
tion of 14 (0.343 g, 1.1 mmol) in DCM (25 mL). Then a solution of
thioctic acid (251 mg, 1.65 mmol) and 4-(dimethylamino)pyridine
(catalytic amount) in DCM (25 mL) was added and the mixture
was stirred for 24 h at room temperature. Then brine was added
and the mixture was extracted with diethyl ether (3ϫ 5 mL). The
combined organic layers were washed with water, dried with
1
(0.237, 97%). H NMR (300 MHz, CDCl3): δ = 7.23 (dt, J = 8.4,
2.4 Hz, 2 H), 6.95 (t, J = 7.1 Hz, 2 H), 4.86 (s, 1 H), 3.73 (t, J =
6.2 Hz, 2 H), 2.92–2.8 (m, 2 H), 2.57 (s, 6 H) ppm. 13C NMR MgSO4, evaporated and further purified by column chromatog-
(75 MHz, CDCl3): δ = 151.8, 138.9, 134.2, 130.9, 122.3, 118.2, 64.5,
45.4, 36.2 ppm.
raphy (dichloromethane/ethyl acetate, 1:1) to yield 15 (0.243 g,
44%) as a yellow oil. H NMR (300 MHz, CDCl3): δ = 7.16 (dd,
1
J = 10.2, 5.0 Hz, 2 H), 7.05 (dd, J = 8.0, 4.4 Hz, 1 H), 5.01 (s, 2
H), 3.81 (t, J = 7.1, 3.5 Hz, 2 H), 3.53 (dt, J = 13.0, 6.5 Hz, 1 H),
3.20–3.03 (m, 2 H), 2.90 (t, J = 7.3, 3.9 Hz, 2 H), 2.65 (s, 5 H),
2.48–2.38 (m, 1 H), 2.33 (t, J = 19.7, 7.0 Hz, 1 H), (dt, J = 19.7,
7.0 Hz, 1 H), 1.71–1.61 (m, 4 H), 1.46 (dt, J = 15.6, 6.8 Hz, 2 H),
0.87 (s, 9 H), 0.00 (s, 6 H) ppm. 13C NMR (75 MHz, CDCl3): δ =
173.8, 153.9, 134.6, 131.4, 130.9, 127.6, 120.1, 66.6, 64.2, 56.7, 45.6,
40.6, 38.9, 35.1, 35.0, 34.5, 29.1, 26.4, 25.1, 18.8, –4.94 ppm.
HRMS: calcd. for C25H43NO3S2Si [M]+ 497.893; found 497.8244.
2-(5-Bromo-2-dimethylaminophenyl)ethanol
(11;
1.75 g,
7.14 mmol) and imidazole (1.46 g, 21.44 mmol) was dissolved in
anhydrous DCM (5 mL). tert-Butyldimethylsilyl chloride (1.19 g,
7.86 mmol) was added and the reaction mixture was stirred at room
temperature until the complete disappearance of the starting mate-
rial (by TLC). The solvent was evaporated and the residue dis-
solved in ethyl acetate (20 mL) and washed with concentrated aque-
ous NaHCO3 (2ϫ 10 mL). The organic layer was dried with
MgSO4 and the solvents evaporated. The product was purified by
column chromatography (DCM/MeOH, 95:5) to give 4-bromo-2- TBAF·3H2O (0.73 mL, 0.7 mmol) was added to a solution of 15
{2-[(tert-butyldimethylsilyl)oxy]ethyl}-N,N-dimethylaniline (12; (0.243 mg, 0.5 mmol) in THF (4 mL) and the mixture was stirred
4778
www.eurjoc.org
© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Eur. J. Org. Chem. 2013, 4770–4779

Gold Nanoparticles for the Detection of Nerve Agent Simulants
at room temperature for 2 h. The reaction mixture was then diluted
with water (10 mL) and the aqueous phase extracted with diethyl
ether (5ϫ5 mL). The combined organic layers were dried with
MgSO4, filtered and the solvent was removed in vacuo. The crude
product was purified by column chromatography (hexane/ethyl
acetate, 1:1) to furnish the ligand 4-(dimethylamino)-3-(2-hydroxy-
ethyl)benzyl 5-(1,2-dithiolan-3-yl)pentanoate (8; 0.064 g, 35%) as a
SCSIE (Universidad de Valencia) is gratefully acknowledged for all
the equipment employed.
[1] a) H. H. Hill Jr., S. J. Martin, Pure Appl. Chem. 2002, 74, 2285–
2291; b) M. Wheelis, Pure Appl. Chem. 2002, 74, 2247–2251.
[2] S. M. Somani Chemical Warfare Agents, Academic Press, San
Diego, CA, 1992.
[3] a) J. J. Gooding, Anal. Chim. Acta 2006, 559, 137–151; b) L. M.
Eubanks, T. J. Dickerson, K. D. Janda, Chem. Soc. Rev. 2007,
36, 458–470.
[4] S. Royo, R. Martinez-Mañez, F. Sancenón, A. M. Costero, M.
Parra, S. Gil, Chem. Commun. 2007, 4839–4847.
[5] A. M. Costero, M. Parra, S. Gil, R. Gotor, P. M. E. Mancini,
R. Martinez-Mañez, F. Sancenón, R. Royo, Chem. Asian J.
2010, 5, 1573–1585.
[6] S. Royo, A. M. Costero, M. Parra, S. Gil, R. Martinez-Mañez,
F. Sancenón, Chem. Eur. J. 2011, 17, 6931–6934.
[7] a) R. Elghanian, J. J. Storhoff, R. C. Mucic, R. L. Letsinger,
C. A. Mirkin, Science 1997, 227, 1078–1081; b) L. M. Wang,
X. F. Liu, X. F. Hu, S. P. Song, C. H. Fan, Chem. Commun.
2006, 3780–3782; c) D. B. Liu, W. S. Qu, W. W. Chen, W.
Zhong, Z. Wang, X. X. Jiang, Anal. Chem. 2010, 82, 9606–
9610; d) K. Saha, S. S. Agasti, C. Kim, X. Li, V. M. Rotello,
Chem. Rev. 2012, 112, 2739–2779; e) V. K. K. Upadhyayula,
Anal. Chim. Acta 2013, 715, 1–18.
1
colourless liquid. H NMR (300 MHz, CDCl3): δ = 7.18 (dd, J =
10.2, 5.0 Hz, 2 H), 7.05 (dd, J = 8.0, 4.4 Hz, 1 H), 5.01 (s, 2 H),
3.84 (t, J = 7.1, 3.5 Hz, 2 H), 3.53 (dt, J = 13.0, 6.5 Hz, 1 H), 3.20–
3.03 (m, 2 H), 2.99 (t, J = 7.3, 3.9 Hz, 2 H), 2.65 (s, 5 H), 2.48–
2.38 (m, 1 H), 2.33 (t, J = 19.7, 7.0 Hz, 1 H), (dt, J = 19.7, 7.0 Hz,
1 H), 1.71–1.61 (m, 4 H), 1.46 (dt, J = 15.6, 6.8 Hz, 2 H) ppm. 13
C
NMR (75 MHz, CDCl3): δ = 173.7, 152.7, 136.8, 132.9, 131.7,
128.1, 120.6, 66.2, 64.7, 56.7, 45.4, 40.6, 38.9, 36.6, 35.0, 34.4, 29.1,
25.1 ppm. HRMS: calcd. for C19H29NO3S2 [M]+ 383.5683; found
383.1694.
Preparation of the Functionalized AuNPs: All glassware was thor-
oughly cleaned with freshly prepared aqua regia (HCl/HNO3, 3:1),
rinsed thoroughly with deionized water and dried in air. AuNPs
with a diameter of around 13 nm were synthesized as reported pre-
viously.[16] Briefly, an aqueous 38.8 mm trisodium citrate solution
(10 mL) was added to a boiling solution of 1 mm HAuCl4 (100 mL)
and the resulting solution boiled for 30 min until a red solution
was obtained. This solution was cooled to room temperature and
then stored in a refrigerator at 4 °C for use. The AuNPs were modi-
fied by ligand-exchange reaction at room temperature as follows:
A 0.01 m aqueous NaOH solution (20 μL) was added to the as-
[8] W. Haiss, N. T. K. Thanh, J. Aveyard, D. G. Fernig, Anal.
Chem. 2007, 79, 4215–4221.
[9] B. Kong, A. Zhu, Y. Luo, Y. Tian, Y. Yu, G. Shi, Angew. Chem.
2011, 123, 1877; Angew. Chem. Int. Ed. 2011, 50, 1837–1840.
[10] D. Liu, Z. Wang, X. Jiang, Nanoscale 2011, 3, 1421–1433.
[11] a) K. M. Mayer, J. H. Hafner, Chem. Rev. 2011, 111, 3828–
3857; b) E. E. Bedford, J. Spadovecchia, C. M. Pradier, F. X.
Gu, Macromol. Biosci. 2012, 12, 724–739.
prepared citrate-capped AuNPs (10 mL). Then TA (200 μL, 10–3
m
in ethanol) and the appropriate amount of ligand 2–8 (10–3 m in
ethanol) were added simultaneously and the solution was stirred
overnight. To purify the AuNPs, the mixture was centrifuged for
20 min at 14000 rpm and the supernatants were decanted. Then the
resulting AuNPs were resuspended in MOPS buffer solution (0.1 m
at pH 7). The size and shape of the modified AuNPs were charac-
terized by TEM. The UV/Vis spectra absorbance spectra of the
modified AuNPs were recorded.
[12] J. Sun, L. Guo, Y. Bao, J. Xie, Biosens. Bioelectron. 2011, 28,
152–157.
[13] H. Li, J. Guo, H. Ping, L. Liu, M. Zhang, F. Guan, C. Sun,
Q. Zhang, Talanta 2011, 87, 93–99.
[14] K. Bondyopadhyay, S. G. Lin, H. Lin, L. Echegoyen, Chem.
Eur. J. 2000, 6, 4385–4392.
[15] M. Kadalbajoo, J. Park, A. Opdahl, H. Suda, C. A. Kitchens,
J. C. Garnó, J. D. Batteas, M. J. Tarlov, P. Deshong, Langmuir
2007, 23, 700–707.
[16] P. Zhao, N. Li, D. Astruc, Coord. Chem. Rev. 2013, 257, 638–
665.
The limits of detection (LODs) were determined as described in
the Supporting Information
[17] N. R. Jana, L. Gearheart, C. J. Murphy, Langmuir 2001, 17,
Supporting Information (see footnote on the first page of this arti-
6782–6786.
1
cle): Copies of the H NMR and 13C NMR spectra (Figures S1 to
[18] a) K. C. Grabar, R. G. Freeman, M. B. Hommer, M. J. Natan,
Anal. Chem. 1995, 67, 735–743; b) K. R. Brow, A. P. Fox, M. J.
Natan, J. Am. Chem. Soc. 1996, 118, 1154–1157.
[19] S.-Y. Lin, Y.-T. Tsai, C.-C. Chen, C.-M. Lin, C.-h. Chen, J.
Phys. Chem. B 2004, 108, 2134–2139.
S11), UV/Vis spectra ligands 5–7 (Figure S12), UV/Vis spectra and
A660/A526 vs. DCNP concentration plots for AuNP2-AuNP8 and
method of determination of LODs.
[20] X. Liu, M. Atwater, J. Wang, Q. Huo, Colloids Surf. B 2007,
58, 3–7.
Acknowledgments
[21] M. Kadalbajoo, J. Park, A. Opdahl, H. Suda, C. A. Kitchens,
J. C. Garno, J. D. Batteas, M. J. Tarlov, P. DeShong, Langmuir
2007, 23, 700–707.
The authors thank the Spanish Government (grant numbers
MAT2009-14564-C04-03 and MAT2012-38429-C04-02) and the
Generalitat Valenciana (Project PROMETEO/2009/095) for sup-
port. A. M. is grateful to the Spanish Government for a fellowship.
[22] L. Cui, Y. Zhao, Chem. Mater. 2004, 16, 2076–2082.
Received: March 6, 2013
Published Online: June 17, 2013
Eur. J. Org. Chem. 2013, 4770–4779
© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
4779