Novel thieno-imidazole based probe for colorimetric detection of Hg 2+ and fluorescence turn-on response of Zn2+

Article abstract of DOI:10.1021/ol300867e

Novel thieno-imidazole based polymer P showed both colorimetric and ratiometric detections of Hg²⁺ as well as fluorometric detection of Zn²⁺ via fluorescence turn-on response with augmented lifetime. Its model polymer M did not show any such sensing capability under similar conditions, which further confirmed the unique sensitivity of P toward Hg ²⁺ and Zn²⁺ via the chelation of metal ions to both "S" and "N" heteroatoms.

Full text of DOI:10.1021/ol300867e

ORGANIC
LETTERS
2012
Vol. 14, No. 10
2564–2567
Novel Thieno-imidazole Based Probe for
Colorimetric Detection of Hg2þ and
Fluorescence Turn-on Response of Zn2þ
Rudrakanta Satapathy, Yen-Hsing Wu, and Hong-Cheu Lin*
Department of Materials Science and Engineering, National Chiao Tung University,
Hsinchu 30049, Taiwan (ROC)
linhc@mail.nctu.edu.tw
Received April 5, 2012
ABSTRACT
Novel thieno-imidazole based polymer P showed both colorimetric and ratiometric detections of Hg2þ as well as fluorometric detection of Zn2þ via
fluorescence turn-on response with augmented lifetime. Its model polymer M did not show any such sensing capability under similar conditions,
which further confirmed the unique sensitivity of P toward Hg2þ and Zn2þ via the chelation of metal ions to both “S” and “N” heteroatoms.
Owing to the ghastly immunotoxic, genotoxic, neuro-
toxic, malnutrition, and digestive disorder effects, mercury
is considered to be a highly dangerous element in the
environment, which can be easily bioaccumulated.1 Zinc,
a pervasive second most essential transition element in the
human body after iron, is involved in numerous biological
processes, such as cellular metabolism, neurotransmission,
signal transduction, gene expression, apoptosis, as well as
pathological processes in many diseases (including Alzhei-
mer’s disease, epilepsy, ischemic stroke, and so forth).2 It
is thus imperative to develop analytic and detective meth-
ods for sensitive sensing of Zn2þ and Hg2þ. Unlike other
biological transition metal ions, such as Fe2þ, Mn2þ, and
Cu2þ ions, both Zn2þ and Hg2þ ions are spectroscopically
and magnetically neutral and thus do not give any signals
due to their 3d104s0 (Zn2þ) and 5d106s0 (Hg2þ) electronic
configurations. Thus common analytic techniques, such as
Mossbauer, nuclear magnetic resonance (NMR), and elec-
tron paramagnetic resonance (EPR), are unable to detect
this archetypal metal ion in biological systems. Further-
more, fluorescence offers significant advantages over other
methods for detection of these metal ions because of
(3) (a) Amendola, V.; Fabbrizzi, L.; Forti, F.; Licchelli, M.; Mangano,
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Y.; Fan, J. Org. Biomol. Chem. 2007, 5, 226. (h) Quang, D. T.; Kim, J. S.
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Y.; Duan, L.; He, Y.; Feng, Z. Chem. Res. Toxicol. 2005, 18, 1814.
(b) Harada, M. Crit. Rev. Toxicol. 1995, 25, 1. (c) Renzoni, A.; Zino, F.;
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r
10.1021/ol300867e
Published on Web 05/09/2012
2012 American Chemical Society

simplicity, nondestructive character, high sensitivity, and
instantaneous response.3 Consequently, noninvasive fluor-
escence technique is a mode of choice for metal ion
imagining.4
model polymerM (where only a single nitrogen atom in the
imidazole ring is available for binding) was synthesized.
Synthesis of model polymer M is depicted in Supplemen-
tary Scheme S1.
The furthermost advantage of fluorescence “turn-on”
sensors related to “turn-off” sensors is the ease of detecting
low-concentration contrast relative to a “dark” back-
ground, which reduces the possibility of false positive
signals and enhances the sensitivity, as demonstrated by
numerous studies.5 The intramolecular charge transfer
(ICT)-based fluorescence mechanism has been widely used
to illustrate fluorescence turn-on and turn-off behaviors
for various sensory materials.6 Furthermore, ratiometric
fluorescent probes have immense significance because they
permit signal ratio corresponding to more than one wave-
length and provide built-in correction for environmental
effects.7 The naked eye perceived color change would be
beneficial for instantaneous visual sensing.
Scheme 1. Synthesis of Polymer P
Numerous probes have been reported for sensing of
Hg2þ, for example, bis(N-methylindolyl)methane,8a amino-
acid,8b,c indole,8d sulfonamide,8e triazole-based dansyl,8f
methionine,8g and rhodamine.8h Likewise several probes
have been designed for Zn2þ sensing, such as dipicoly-
lamine,9a,b DPA,9c TPEN,9d and Schiff base derivatives.9e
Since the sulfur atom is considered to be a ‘‘soft’’ donor atom,
it can act as a chelating agent and also increase the sensor’s
affinity and selectivity for binding transition metal cations.
Herein, we synthesized thieno-imidazole based polymer
P (Scheme 1), with neighboring nitrogen and sulfur het-
eroatoms as chelating sites for metal ions. This was found
to be a suitable design for colorimetric detection of Hg2þ
and fluorometric detection of Zn2þ. 3,30-Bithiopene was
prepared from 3-bromothiophene by treating with n-BuLi
and CuCl2, which was further acylated without any Lewis
acid to produce compound 3. Then, it was coupled with
4-((2-ethylhexyl)oxy)benzaldehyde by refluxing in the pre-
sence of acetic acid and NH4OAc to obtain compound 4.
N-Alkylation of 4 by hexyliodide and K2CO3 yielded
compound 5, which was brominated by NBS to obtain
monomer 6. Finally, Grignard polymerization of mono-
mer 6 by CH3MgBr and NiDPPCl2 produced polymer P.
To compare the selectivity of polymerP toward metal ions,
To verify the sensing abilities of metal ions, polymer P
was titrated over a wide range of metal ions, such as Naþ,
Kþ, Ba2þ, Ca2þ, Ni2þ, Cuþ, Co2þ, Agþ, Zn2þ, and Hg2þ
(Supplementary Figure S1). Originally, two absorption
maxima at 248 and 303 nm appeared in polymer P. Upon
the sequential addition of Hg2þ, a significant change in
absorption pattern was observed. As shown in Figure 1a,
the final intensities of the peak at 248 nm increased ca. 4.9-
fold, and the peak at 303 nm decreased ca. 50%. Further-
more, a new peakappearedat395 nm and increasedalmost
9.7-fold. Two clear isosbestic points at 281 and 367 nm
were obtained upon titration with Hg2þ. The intensities at
305 and 248 nm (Figure 1b) as well as 395 and 305 nm
(Figure 1c) were compared, which showed the intensity
changed almost linearly with the concentration of Hg2þ
.
The color of the polymer solution changed from colorless
to yellow, which could be easily detected by the naked eye.
However, other metal ions did not show such alteration in
absorption pattern. A Job plot was plotted to find the
stoichiometry during the binding of Hg2þ (Supplementary
Figure S2). The variation of absorption intensities at
385 and 305 nm (A385 ꢀ A305) as a function of molar ratio
XM = [Hg2þ]/{[Hg2þ] þ [P]} showed 1:1 stoichiometry.
1H NMR titration was further conducted to elucidate the
binding mode, and the NMR titration spectra of monomer
6 (in d-THF) upon the addition of 0ꢀ1.1 equiv of Hg2þ (in
D2O) are depicted in Figure 2. Notable downfield shifts of
peaks corresponding to the aromatic protons of monomer
(5) (a) Rurack, K.; Kollmannsberger, M.; Resch-Genger, U.; Daub,
J. J. Am. Chem. Soc. 2000, 122, 968. (b) McQuade, D. T.; Hegedus,
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(8) (a) Kaur, P.; Kaur, S.; Singh, K Org. Biomol. Chem. 2012, 10,
1497. (b) Yang, M. H.; Thirupathi, P.; Lee, K. H. Org. Lett. 2011, 13,
5028. (c) Lohani, C. R.; Kim, J. M.; Lee, K. H. Tetrahedron 2011, 67,
4130. (d) Kaur, P.; Kaur, S.; Singh, K.; Sharma, P. R.; Kaur, T. Dalton
Trans. 2011, 40, 10818. (e) Joshi, B. P.; Lohani, C. R.; Lee, K. H. Org.
Biomol. Chem. 2010, 8, 3220. (f) Neupane, L. N.; Kim, J. M.; Lohani,
C. R.; Lee., K. H. J. Mater. Chem. 2012, 22, 4003. (g) Yang, M. H.;
Lohani, C. R.; Cho, H.; Lee, K. H. Org. Biomol. Chem. 2011, 9, 2350.
(h) Kumar, M.; Kumar, N.; Bhalla, V.; Singh, H.; Sharma, P. R.; Kaur.,
T. Org. Lett. 2011, 13, 1422.
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Q. Y.; Lee, S. J.; Kang, C.; Kim, J. S. Org. Lett. 2011, 13, 5294. (b) Xu, Y.;
Pang., Y. Dalton Trans. 2011, 40, 1503. (c) Xu, Y.; Pang., Y. Chem.
Commun. 2010, 46, 4070. (d) Hanaoka, K.; Muramatsu, Y.; Urano, Y.;
Terai, T.; Nagano., T. Chem.;Eur. J. 2010, 16, 568. (e) Wang, L.; Qin, W.;
Tang, X; Dou, W.; Liu., W. J. Phys. Chem. A 2011, 115, 1609.
6 were observed upon the gradual addition of Hg2þ
.
1
However, no further changes in H NMR signals were
observed at higher equivalents of Hg2þ. This result is
consistent with 1:1 binding stoichiometry of Hg2þ with
Org. Lett., Vol. 14, No. 10, 2012
2565

Figure 3. Fluorescence spectral response of P (1.1 ꢁ 10ꢀ5 M) in
THF/H2O (1/1) acquired by the sequential addition of 0ꢀ10 equiv
of Zn2þ. Inset: Intensity of P as a function of equiv of Zn2þ added.
Similarly, the fluorescence patterns of polymer P upon
the addition of various metal ions under similar conditions
are depicted in Supplementary Figure S5, and their histo-
grams are shown in Supplementary Figure S6. As observed
in Figure 3, upon the addition of Zn2þ, the fluorescence
intensity increased ca. 10-fold, which was the most sig-
nificant change in fluorescence signals compared with the
other metal ions. The lone-pair electrons in the thieno-
imidazole receptor undergo rapid electron transfer, which
causes the week fluorescence of P. In the presence of
suitable cationic analytes (here Zn2þ), the nonbonding
electrons get coordinated and thus the ICT effect is
inhibited, which resulted in the strong fluorescence of P.
Thus, in the presence of Zn2þ, P showed a remarkable
increase in the intensity of the emission band with a chela-
tion enhanced fluorescence factor (CHEF)10 = 10.5. Again,
it resulted in a 14.3-fold enhancement of the quantum yield
for polymer P (from Φ = 0.03 to Φ = 0.43). From the
fluorescence binding isotherm, the association constant
was calculated to be Ka = 5.14 ꢁ 106 Mꢀ1. To find the
interference of other back ground cations, we conducted
the titrations by using a dual metal system (Supplementary
Figure S7). The enhanced fluorescence of polymer P in the
presence of Zn2þ was not substantially perturbed by the
background cations. To gain insight into the binding of
Zn2þ with the repeating unit of P, 1H NMR titration was
carried out by the gradual addition of Zn2þ (in D2O) to
monomer 6 (in d-THF). As shown in Figure 4, a momen-
tous upfield shift of peaks corresponding to the aromatic
protons of monomer 6 was observed upon the sequential
addition of Hg2þ (0ꢀ1.1 equiv). Fluorometric titration of
Zn2þ was conducted for model polymer M to investigate
the effect of Zn2þ. Model polymer M did not show any
significant fluorescence enhancement in the presence of
Zn2þ under similar conditions (see Supplementary Figure S8).
As shown in Table 1, the quantum yield of M revealed
only a 2.25-fold increase (from Φ = 0.04 to Φ = 0.09), and
Figure 1. (a) Absorption spectral changes of P (1.1 ꢁ 10ꢀ5 M) in
THF/H2O (1:1) upon titration with (0ꢀ1.1 ꢁ 10ꢀ4 M) of Hg2þ
.
Upper inset: absorption pattern from 275 to 600 nm. Lower inset:
color change upon the addition of Hg2þ. Absorption spectral ratios (b)
A
305/A248 and (c) A395/A205 as a function of equivalents of Hg2þ added.
polymer P. This result is consistent with 1:1 binding
stoichiometry of Hg2þ with polymer P. The binding con-
stant for binding of Hg2þ was calculated to be 0.74 ꢁ 105
from Supplementary Figure S3.
Model polymer M (where the binding site is only nitro-
gen heteroatom in the imidazole ring) showed two absorp-
tion maxima at 267 and 290 nm. Polymer M was titrated
with Hg2þ under the similar condition as that of polymer
P. However, unlike polymer P, no additional peak ap-
peared and the absorption pattern was not changed sub-
stantially (Supplementary Figure S4). This may be
attributed to the susceptibility of Hg2þ to be bound
between “S” and “N” of thio-imidazole unit in polymer
P. Thus, it is concluded that the thio-imidazole based probe
is a suitable design for the colorimetric detection of Hg2þ
.
Figure 2. 1H NMR spectra (aromatic region) of monomer 6
upon the addition of 0ꢀ1.1 equiv of Hg2þ wrt [6].
(10) Satapathy, R.; Wu, Y. H.; Lin, H. C. Chem. Commun. 2012, DOI:
10.1039/C2CC31131C.
2566
Org. Lett., Vol. 14, No. 10, 2012

Table 1. Quantum Yields, Fluorescence Lifetimes, Association
Constants, and CHEF Values of Polymers P and M upon the
Addition of Zn2þ
Φa
lifetimeb
CHEFd
c
Ka
P
0.03
0.43
0.04
0.09
0.49
2.96
0.34
0.37
NA
NA
P þ Zn2þ
5.14 ꢁ 106
10.5
NA
M
NA
1.35 ꢁ 103
M þ Zn2þ
2.26
Figure 4. 1H NMR spectra (aromatic region) of monomer 6
upon the addition of 0ꢀ1.1 equiv of Zn2þ wrt [6].
a Quantum yields calculated using tryptophan (in H2O) as a standard.
b Obtained from time-resolved fluorescence measurements. c Association
constants calculated from the slopes of binding isotherms. d CHEF is defined
as I/I0, where I is the maximum emission intensity of the receptorꢀmetal
complex and I0 is the maximum emission intensity of the free receptor.
correspondingly the CHEF was increased just up to
2.2 times. Furthermore, the association constant of poly-
mer P with Zn2þ was calculated as 5.14 ꢁ 106, which is
almost 3.8 ꢁ 103 times higher than that of polymer M. To
elucidate the effect of polymerization, we further carried
oxidation process for polymer P due to the presence of
close proximity of a positively charged Zn cation center.
Furthermore, the HOMO level of polymer P was lowered
(from ꢀ5.36 to ꢀ5.56), and its LUMO level was increased
out the fluorescence titration of compound 5 with Zn2þ
.
Upon the addition of equivalent amounts of Zn2þ, the
fluorescence intensities of compound 5 were sequentially
increased merely up to 4 times (Supplementary Figure S9).
Thus, in contrast to the small molecular analogue 5
(CHEF = 3.8), the increased sensitivity of polymer P
(CHEF = 10.5) is due to the molecular ion effect with a
larger number of binding sites in polymer P.
(from ꢀ3.92 to ꢀ3.42) after complexation with Zn2þ
.
Thus, the obstruction of the reductive as well as oxidative
ICT caused the enhancement of the fluorescent intensity.
To check the affinities of different anions toward a P-Zn
complex, we added various anions, such as Fꢀ, Clꢀ, Brꢀ,
Iꢀ, S2ꢀ, NO3ꢀ, Acꢀ, PO43ꢀ, and SCNꢀ, to the solution of
P-Zn complex. Surprisingly, unlike other anions, upon the
addition of Iꢀ, the color of the solution changed from
colorless to yellow (Supplementary Figure S12a). The pro-
gressive appearance of a new peak at 375 nm was observed
upon the sequential addition of Iꢀ (Supplementary Figure
S12b). Thus, P-Zn complex is susceptible for the colorimetric
detection of Iꢀ ions. Reversible association of Zn2þ with
polymer P was achieved by the addition of Na2-EDTA as a
suitable counter ligand (Supplementary Figure S13). There-
fore, the onꢀoffꢀon fluorescence etiquette was successfully
achieved via the alternate addition of Zn2þ and Na2-EDTA
for 5 successive cycles (Supplementary Figure S14).
Time-resolved fluorescence measurements10 were per-
formed for both polymer P and its Zn2þ-complex probed at
390 nm (excited at 375 nm), as shown in Figure 5. In the
absence of Zn2þ, a single exponential fitting for the fluores-
cence lifetime of polymer P was estimated as 0.49 ns, which
corresponded to the lifetime of the S1 state. Upon the addi-
tion of Zn2þ, the fluorescence lifetime corresponding to the S1
state was elevated to 2.96 ns. No remarkable change in fluo-
rescence lifetime of model polymer M was observed under
similar conditions (see Table 1 and Supplementary Figure S10).
In summary, the naked-eye detection of Hg2þ was
achieved by polymer P with a thieno-imidazole backbone.
Furthermore, polymer P showed a fluorescence turn-on
response with an enhanced fluorescence lifetime in the
presence of Zn2þ. Its model polymer M could not show
any such remarkable sensitivity toward Hg2þ and Zn2þ
,
which confirmed the unique sensitivity of P via the chela-
tion of metal ions to both “S” and “N” heteroatoms.
Moreover, the colorimetric detection of Iꢀ was selectively
accomplished by P-Zn complex over other anions.
Acknowledgment. Financial support of this project was
provided by the National Science Council of Taiwan
(ROC) through NSC 99-2113-M-009-006-MY2.
Figure 5. Time-resolved fluorescence spectral responses for
polymer P before and after the addition of Zn2þ
.
Upon the progressive addition of Zn2þ, two oxidation
peaks Ox1 and Ox2 of polymer P were anodically shifted
and disappeared, respectively (see Supplementary Figure
S11), and the half-wave oxidation potential (E1/2) of Ox1
(appeared at 1520 mV originally) was shifted to 1200 mV
with ΔE1/2 = 320 mV. This indicated a less favorable
Supporting Information Available. Synthetic proce-
dures, Figures S1ꢀS12, and NMR spectral data. This
material is available free of charge via the Internet at
http://pubs.acs.org.
The authors declare no competing financial interest.
Org. Lett., Vol. 14, No. 10, 2012
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