Combinatorial synthesis of a triphenylmethine library and their application in the development of Surface Enhanced Raman Scattering (SERS) probes

Article abstract of DOI:10.1039/b921550f

The first synthesis of a triphenylmethine (TM) library of compounds and screening of their Surface Enhanced Raman Scattering (SERS) capability was carried out to identify novel Raman reporters with high sensitivity. We identified three novel SERS reporters (B2, B7, and C7) with higher signal intensity than that of commonly used crystal violet (CV). These reporters may find potential applications in developing sensitive SERS based biosensors.

Full text of DOI:10.1039/b921550f

View Article Online / Journal Homepage / Table of Contents for this issue
COMMUNICATION
www.rsc.org/chemcomm | ChemComm
Combinatorial synthesis of a triphenylmethine library and their
application in the development of Surface Enhanced Raman
Scattering (SERS) probesw
b
Sung Ju Cho,a Young-Hoon Ahn,z Kaustabh Kumar Maiti,a U. S. Dinish,a Chit Yaw Fu,a
Praveen Thoniyot,a Malini Olivo*acd and Young-Tae Chang*ae
Received (in Cambridge, UK) 14th October 2009, Accepted 26th November 2009
First published as an Advance Article on the web 17th December 2009
DOI: 10.1039/b921550f
The first synthesis of a triphenylmethine (TM) library of
compounds and screening of their Surface Enhanced Raman
Scattering (SERS) capability was carried out to identify novel
Raman reporters with high sensitivity. We identified three novel
SERS reporters (B2, B7, and C7) with higher signal intensity
than that of commonly used crystal violet (CV). These reporters
may find potential applications in developing sensitive SERS
based biosensors.
(TM) compounds exhibit absorption at visible ranges that
enable the compounds to be a useful Raman reporter in
visible-NIR excitation.8a–c Although a few TM compounds,
i.e. malachite green isothiocyanate (MGITC) and crystal violet
(CV), have been used as reporters,4–9 there is a growing need
for reporter molecules that are easily identifiable within a
multiplexed analysis platform which can also generate higher
SERS intensity. However, a systematic study for diverse SERS
reporter generation and screening has not been carried out. In
this context, we report the first combinatorial synthesis of a
TM dye library and the screening of their SERS properties
for development of highly sensitive SERS nanotags. The
conventional syntheses of the TM dyes were mostly based
on electrophilic aromatic substitutions and the common
electrophilic reagents are phosgene, formaldehyde, chloro-
form, and carbon tetrachloride.10 Our diversity oriented
library approach incorporates11,12 solid-phase chemistry to
generate the final product TM, avoiding the use of toxic
reagents and time-consuming purification steps. A general
approach to the synthesis of the TM library is outlined in
In surface enhanced Raman scattering (SERS), the intensity of
the vibrational spectra of a molecule is enhanced by several
orders of magnitude when the molecule is in close proximity to
metallic nanoparticles such as gold or silver. SERS has been
successfully applied for labeling biological systems even in cells
and tissues to sense multiplexed biomarkers.1 Nanoparticle
tags that use SERS to generate detectable Raman signals have
been shown to be a successful alternative to fluorescence
labeling, which has the drawbacks of photobleaching, peaks
overlapping in multiplexed experiments, and inability to
function in some extreme environments in biological systems.2
SERS nanotags have been used for molecular, cellular and in
several in vivo bioimaging techniques by bioconjugating with
multiple diagnostic and therapeutic agents.1–3 Recently Qian
et al.4 and Keren et al.5 reported the use of SERS nanotags for
imaging on a rodent model. These nanotags were developed by
immobilizing a Raman active dye (Raman reporter) on a metal
colloid followed by bioconjugation to target specific locations.
Such a nanoparticle–Raman reporter assembly is called a
Raman tag in analogy with quantum dots and can provide a
platform for multiplexing, targeting and tracking in bio-
imaging and sensing applications.6,7
Scheme 1. Building block
A is commercially available
(Aldrich) and B–D were synthesized as described in ESI.w
Each intermediate A–D was then loaded on 2-chlorotrityl
chloride resin, and reacted with 29 different Grignard reagents
(R2 building block) for diversity. An acidic cleavage from the
resin resulted in the dehydration of the corresponding tertiary
alcohols, giving the fully conjugated TM derivatives. The
choice of 4-aminophenyl groups as the R1 building block
(S1) has the advantages of providing a linker group to the
resin and also of providing a reactive group to the metal
surface in the final TM library compound. All the library
compounds were characterized by LCMS and 52 relatively
pure compounds were selected for further studies with the
purity and full characterization in ESI.w
The reporter molecule is a major factor to enhance the
sensitivity of a Raman tag, as well as metal nanoparticles.
Among the different reporter molecules, triphenylmethine
The primary screening was carried out by incubating 60 nm
citrate stabilized gold colloidal solution with each dye compound
to measure SERS spectra under a confocal Raman microscope.
The highest SERS intensity from each spectrum was identified
and compared with that of CV as a reference in Fig. 1. The result
clearly shows that the SERS signal varies significantly across the
different TM library compounds, and at least 13 compounds
exhibited a stronger SERS signal than CV. In general,
compounds with building blocks B and C show higher SERS
intensity than those with building blocks A and D.
a Singapore Bioimaging Consortium, Agency for Science,
Technology and Research (A*STAR), 11 Biopolis Way, Singapore
138667. E-mail: chmcyt@nus.edu.sg; Fax: +65 6478 9957;
Tel: +65 6478 8761
b Department of Chemistry, New York University, New York,
NY 10003, USA
c National Cancer Center, 11 Hospital Drive, Singapore 169610
d School of Physics, National University Ireland, Galway, Ireland
e Department of Chemistry, National University of Singapore,
Singapore 11754
w Electronic supplementary information (ESI) available: Synthesis,
LCMS, UV-Vis, SERS and NMR data. See DOI: 10.1039/b921550f
z Current address: Johns Hopkins University School of Medicine.
We resynthesized the best five compounds from the initial
screening (B2, B7, C3, C7, and C9) and carried out the SERS
ꢀc
This journal is The Royal Society of Chemistry 2010
722 | Chem. Commun., 2010, 46, 722–724

View Article Online
Fig. 2 SERS intensities of PEG encapsulated nanotags. The average
intensities of five individual measurements with error bars denoting
their standard deviation. SERS spectra were obtained from excitation
at 633 nm with laser power of 3 mW.
Scheme 1 Synthesis of triphenylmethine library. Reagents and conditions:
(a) 2-chlorotrityl chloride resin (1.37 mmol gꢁ1), pyridine, CH2Cl2–DMF;
(b) Grignard reagents, THF, 62 1C; (c) 1% TFA, CH2Cl2.
Fig. 3 SERS spectra of PEG encapsulated nanotags and structure of
TM compounds: (a) CV, (b) B2, (c) B7, and (d) C7.
In summary, we have synthesized a structurally diverse
triphenylmethine library by a combinatorial solid phase
approach and have systematically screened their SERS
intensity. By modifying the hit compounds with surface
stabilization, we finally identified three highly Raman active
compounds (B2, B7, and C7) with high potential as SERS
reporters for biological applications. Further studies towards
in vivo bioimaging and the combinatorial synthesis and
screening of a wider spectrum of libraries are in progress.
The authors thank the A*STAR Cross Council Office (CCO),
Singapore for financial support (Grant CCOGA02_005_2008)
and gratefully acknowledge the National University of
Singapore (NUS) for financial support (Young Investigator
Award: R-143-000-353-101). The authors thank Dr Loyola
D’silva for assistance with NMR instrument at Singapore
Bioimaging Consortium (SBIC).
Fig. 1 Comparative SERS intensities of TM compounds from A to D
building blocks with CV. SERS spectra were obtained from excitation
at 633 nm with laser power of 3 mW.
study in various conditions. In many cases, we observed that
the SERS signal was fluctuating over the measurement and
storage time, perhaps due to the aggregation of Au nano-
particles. To stabilize the SERS signal, we modified the
nanotags by encapsulating the gold surface with thiolated
PEG which protected the nanotags from aggregation in
aqueous media. We observed that PEG encapsulated nanotags
did not show significant aggregation under ambient conditions
and furnished stable SERS intensity for all five compounds.
Moreover, the SERS intensities of the PEG encapsulated
nanotags could be optimized by changing the dye concentration
to that which gave the most stable enhanced signals as shown
in Fig. 2. The novel reporters B2, B7, and C7 showed 2.3-, 2.0-,
and 1.8-fold increased signal respectively compared with CV
(Fig. 2) under optimum conditions (10 mM of each dye). The
SERS spectra of our best three reporters, B2, B7 and C7, and
CV are shown in Fig. 3. The highest intensity peak has been
chosen for comparison.
Notes and references
1 C. L. Zavaleta, B. R. Smith, I. Walton, W. Doering, G. Davis,
G. B. Shojaei, M. J. Natan and S. S. Gambhir, Proc. Natl. Acad.
Sci. U. S. A., 2009, 106, 13511.
2 D. Graham, D. G. Thimpson, W. E. Smith and K. Faults, Nat.
Nanotechnol., 2008, 3, 548.
3 X. M. Qian, X. Zhou and S. Nie, J. Am. Chem. Soc., 2008, 130, 14934.
4 X. Qian, X.-H. Peng, D. O. Ansari, Q. Yin-Geon, G. Z. Chen,
D. M. Shin, L. Yang, A. N. Young, M. D. Wang and S. Nie, Nat.
Biotechnol., 2008, 26, 83.
ꢀc
This journal is The Royal Society of Chemistry 2010
Chem. Commun., 2010, 46, 722–724 | 723

View Article Online
5 S. Keren, C. Zavaleta, Z. Cheng, A. de la Zerda, O. Gheysens and
S. S. Gambhir, Proc. Natl. Acad. Sci. U. S. A., 2008, 105, 5844.
6 M. Sanles-Sobrido, W. Exner, L. Rodriguez-Lorenzo, B. Gonzalez
Rodriguez, M.-A. Correa-Duarte, R. Puebla-Alvarez and
L. Marzan-Liz, J. Am. Chem. Soc., 2009, 131, 2699.
7 P. Zhang and Y. Guo, J. Am. Chem. Soc., 2009, 131, 3808.
8 (a) A. Kudelski, Chem. Phys. Lett., 2005, 414, 271; (b) N.-P. W.
Pieczonka and R.-F. Aroca, Chem. Soc. Rev., 2008, 37, 946;
(c) K. Kneipp, H. Kneipp, G. Deinum, I. Itzkan, R.-R. Dasari
and M.-S. Feld, Appl. Spectrosc., 1998, 52(2), 175.
9 S. Lee, H. Chon, M. Lee, J. Choo, S. Y. Shin, Y. H. Lee,
I. J. Rhyu, S. W. Son and C. H. Oh, Biosens. Bioelectron., 2009,
24, 2260.
10 H. Zollinger, in Color Chemistry, Synthesis, Properties, and
Applications of Organic Dyes and Pigments, Wiley-VCH,
Switzerland, 2004, ch. 4, pp. 101–122.
11 Y. H. Ahn, J. S. Lee and Y. T. Chang, J. Am. Chem. Soc., 2007,
129, 4510.
12 J. S. Lee, Y. K. Kim, M. Vendrell and Y. T. Chang, Mol. BioSyst.,
2009, 5, 411.
ꢀc
This journal is The Royal Society of Chemistry 2010
724 | Chem. Commun., 2010, 46, 722–724