Thiazole-based chemosensor: synthesis and ratiometric fluorescence sensing of zinc

Article abstract of DOI:10.1016/j.tetlet.2009.07.078

A new ratiometric and selective fluorescent chemosensor (1), based on thiazole for quantification of zinc ions in aqueous ethanol, was synthesized and investigated. The mechanism of fluorescence was based on the cation-induced inhibition of excited-state

Full text of DOI:10.1016/j.tetlet.2009.07.078

Tetrahedron Letters 50 (2009) 5510–5515
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Tetrahedron Letters
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Thiazole-based chemosensor: synthesis and ratiometric fluorescence sensing
of zinc
*
Aasif Helal, Hong-Seok Kim
Department of Applied Chemistry, Kyungpook National University, Daegu 702-701, Republic of Korea
a r t i c l e i n f o
a b s t r a c t
Article history:
A new ratiometric and selective fluorescent chemosensor (1), based on thiazole for quantification of zinc
ions in aqueous ethanol, was synthesized and investigated. The mechanism of fluorescence was based on
the cation-induced inhibition of excited-state intramolecular proton transfer (ESIPT).
Ó 2009 Elsevier Ltd. All rights reserved.
Received 31 May 2009
Revised 14 July 2009
Accepted 16 July 2009
Available online 18 July 2009
Keywords:
Thiazole
Ratiometric sensing of Zn2+
ESIPT
Fluorescence
A fluorescent chemosensor consists of a molecular system for
which the physicochemical properties change upon interaction
with a chemical species, in such a way as to produce a detect-
able fluorescent signal.1 Zinc(II) is the second most abundant
transition-metal form of cation in the biological system.2
Approximately 300 enzymes contain zinc(II) as an essential com-
ponent, either for a structural purpose or as a part of a catalytic
site. For example, zinc is essential for the regulation of DNA syn-
thesis during the proliferation and differentiation of cells.3 Zinc
is also known to have a role in neurological disorders, such as
Parkinson’s disease, Alzheimer’s disease, amyotrophic lateral
sclerosis, and epileptic seizures.4 Therefore, the design and
development of a fluorescent chemosensor selective to zinc are
of considerable interest.
A variety of zinc ion selective fluorescent probes that are based
on quinoline,5 fluorescein,6 coumarin,7 indole,8 and other fluoro-
phores9 or proteins10 have been developed. But a majority of these
zinc sensors function as cation-responsive optical switches that
translate the binding event into either an increase or decrease of
the emission intensity.11 However, the emission intensity is also
dependent on other factors, such as the sensor concentration,
bleaching, optical path length, and illumination intensity. It is
therefore desirable to eliminate the effects of these factors by using
a ratiometric sensor that exhibits a spectral shift upon reaction or
binding to the analyte of interest. The ratio between the two emis-
sion intensities can be used to evaluate the analyte concentra-
tion.12 Excited-state intramolecular proton transfer (ESIPT) is one
of the most common photophysical processes that occurs in ben-
zazoles and used to develop ratiometric probes. Inhibition of the
ESIPT process by cation binding yields a significant hypsochromic
shift of the fluorescence emission maximum.13,14
Crown ethers that contain thiazole moieties15 have been re-
ported to exhibit large ammonium ion selectivity.14b Benzene-
based tripodal receptors16,17 and steroidal tweezers18,19 have been
used for selective detection of silver(I) by introduction of soft het-
eroatoms N and S, as electron donors to metal cations. However
application of heteroaromatic ring system such as thiazole, which
contains soft heteroatoms N and S and has substitutes at positions
2 and 4 as a zinc chemosensor has not yet been reported.
In this Letter, design and development of a novel thiazole-based
ratiometric fluorescent chemosensor with substitutes at positions
2 and 4 regarding zinc ion in aqueous media are reported. It exhib-
its a ratiometric fluorescent response upon addition of zinc ion into
10% water in ethanol, buffered at pH 7.4. The results obtained from
investigation on effect of solvents and competitive processes of
zinc cation with other cations are reported.
ESIPT is one of the most common photophysical properties that
occurs in benzazoles used to develop ratiometric probes. In order
to understand the crucial role of both phenol and pyridine rings
providing a suitable binding site for zinc ion, 1 and 2 were pre-
pared in good yields by the reaction of 2-hydroxythiobenzamide
with 2-(2-bromoacetyl)pyridine and 2-bromoacetophenone in
refluxing ethanol, as shown in Scheme 1. The structures of 1 and
2 were confirmed by 1H NMR, 13C NMR, and elemental analyses
data.
Initial studies on the UV–vis absorption and fluorescent emis-
sion processes revealed that 1 showed selectivity toward zinc
* Corresponding author. Tel.: +82 53 950 5588; fax: +82 53 950 6594.
E-mail address: kimhs@knu.ac.kr (H.-S. Kim).
0040-4039/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2009.07.078

A. Helal, H.-S. Kim / Tetrahedron Letters 50 (2009) 5510–5515
5511
S
at 481 nm, whereas in the presence of zinc 1 revealed absorption
and emission bands at 390 and 461 nm, respectively. Coordination
of zinc cation to 1 causes a red shift in UV–vis absorption and a
blue shift in fluorescent spectrum along with enhancement in
the degree of intensity.
In contrast to 4-pyridylthiazole (1), 4-phenylthiazole (2) which
has no pyridine moiety has not revealed any significant changes in
absorption and fluorescence emissions upon addition of up to
10 equiv of zinc ion as shown in Figure 1.
O
O
Br
Br
EtOH
EtOH
+
NH2
N
N
N
OH
S
OH
1
S
N
OH
2
Compound 1, bearing the 2-hydroxyphenyl-thiazole signaling
unit and pyridine as the binding site, is anticipated to act as an
ESIPT-based chemosensor. The changes of absorption spectra of 1
upon addition of zinc perchlorate showed three different stages
Scheme 1. Synthesis of 4-pyridylthiazole (1) and 4-phenythiazole (2).
cation in CH3CN. As shown in Figure 1, in the absence of zinc 1
showed an absorption band at 331 nm and a fluorescent emission
0.5
0.5
(a)
(b)
0.4
0.4
333 nm
333 nm
0.3
0.3
331 nm
390 nm
0.2
0.2
0.1
0.0
0.1
0.0
300
350
400
450
300
461 nm
350
400
450
120
90
60
30
0
120
90
60
30
0
(c)
(d)
501 nm
501 nm
481 nm
400
450
500
550
600
650
400
450
500
550
600
650
Wavelength (nm)
Figure 1. UV–vis spectra of 1 (__) and 2 (–) (a) in the absence, and (b) in the presence of 10 equiv of Zn(ClO4)2 in dry CH3CN. Fluorescence spectra of 1 (__) and 2 (–) (c) in the
absence and (d) in the presence of 10 equiv of Zn(ClO4)2 in dry CH3CN.
0.5
0.4
0-10 eq
0.3
0.2
0.1
0.0
289 nm
0-10 eq
0-10 eq
331 nm
390 nm
366 nm
350 nm
401 nm
300
350
400
450
500
Wavelength (nm)
Figure 2. Changes in UV–vis spectra of 1 (20
lM) upon addition of Zn(ClO4)2 in dry CH3CN.

5512
A. Helal, H.-S. Kim / Tetrahedron Letters 50 (2009) 5510–5515
as shown in Figure 2. At the initial stage of addition (0–0.3 equiv)
peak develops at 401 nm which is due to the binding of zinc with
the nitrogen of pyridine (Form I). Upon the further addition of zinc
(0.3–0.6 equiv) this peak gradually disappears and a new peak at
366 nm appears in the second stage which is attributed to the con-
comitant deprotonation of the phenolic proton and protonation of
the pyridine nitrogen (Form II). In the final step (0.6–1 equiv) zinc
coordinates with the nitrogen of the pyridine ring to give peak at
390 nm (Form III). Thus the absorption band at 390 nm becomes
saturated up to 1 equiv of zinc. These possible forms of 1 with dif-
ferent amounts of zinc in ground and excited states are depicted in
Scheme 2.
nolic proton. On further addition of zinc (0.6–1.0 equiv) this peak
undergoes quenching due to binding with the nitrogen of the
pyridine and saturates within the addition of 1 equiv of zinc.
From the fluorescence titration, the binding constant for zinc
ion is observed to be 5.0 Â 105 MÀ1 (Error estimated to be
610%).
In order to achieve a more physiologically acceptable condition
the photophysical properties of 1 were examined in an ethanol–
water system. UV–vis study was carried out in 10% (v/v) water/eth-
anol buffered by 10 mM HEPES at pH 7.4 at a concentration level of
20
l
M. Sensor 1 displayed an obvious absorption band at 329 nm.
This can be attributed to a
p
–
p* transition; this is favored by the
The fluorescence titration spectra of 1 with zinc cation show
an emission maximum peak at 481 nm (Fig. 3). Upon addition of
zinc up to 0.6 equiv emission band at 481 nm undergoes a hyp-
sochromic shift and enhancement to 461 nm gradually. This is
attributed to the inhibition of ESIPT by deprotonation of the phe-
planar orientation enforced by the intramolecular hydrogen bond-
ing.20 However, upon addition of zinc ion to the solution of 1, the
absorption band at 384 nm becomes enhanced gradually while
the absorption band at 329 nm decreased synchronously, as shown
in Figure 4. The absorption bands at 329 and 384 nm linearly
S
S
-HL
+HL
O
N
O
L
N
H
Zn
N
N
L Zn
H
L
L
-HL
I
S
N
II
λ
max = 366 nm
λmax= 401 nm
-
O
N
L = ClO4
ZnL2
-HL
H
1
-HL
S
N
S
λmax= 331 nm
O
N
O
L
N
Zn
III
H
N
Tautomer
λ
max= 390 nm
λem = 461 nm
λem = 481 nm
Scheme 2. Possible forms of sensor 1 with Zn(ClO4)2 in the ground and excited states in dry CH3CN.
250
200
0.6-10 eq
0-0.6 eq
150
461 nm
100
50
481 nm
481 nm
0
400
450
500
550
600
650 400
450
500
550
600
650
Wavelength (nm)
Figure 3. Changes in fluorescence spectra of 1 (20 lM) upon addition of Zn(ClO4)2 in dry CH3CN. (kex at 350 nm.)

A. Helal, H.-S. Kim / Tetrahedron Letters 50 (2009) 5510–5515
5513
0.5
0.4
0.3
0.2
0.1
0.0
384 nm
0.9
0.6
0.3
0.0
-0.3
-0.6
-0.9
329 nm
0.0
0.5
1.0
1.5
2.0
Concentration of Zn+2
0-10 eq
0-10 eq
329 nm
384 nm
345 nm
300
350
400
450
Wavelength (nm)
Figure 4. Changes in UV–vis spectra of 1 (20 lM) upon addition of Zn(ClO4)2 in EtOH/H2O (9:1) containing HEPES buffer (10 mM, pH 7.4). Inset: mol ratio plots of absorbance
at 329 and 384 nm.
decreased and increased, respectively, up to 1 equiv of zinc (Fig. 4
inset), indicating the formation of a 1:1 complex with a strong
binding affinity.
The Job’s plot of 1 with zinc also indicates the formation of a 1:1
complex (Fig. S-1). The binding constant calculated in an ethanol–
water system at pH 7.4 was 3.5 Â 104 MÀ1 (Error estimated to be
610%).
Similarly, fluorescence titration of 1 with zinc was carried out in
10% (v/v) water/ethanol buffered by 10 mM HEPES at pH 7.4 at a
concentration level of 20 lM. Addition of zinc ion to the solution
of 1 causes a simultaneous blue shift of fluorescent emission from
502 to 461 nm when excited at 345 nm, with an isoemission point
at 485 nm (Fig. 5). Like the benzothiazole derivatives13 compounds
1 and 2 contain an intramolecular hydrogen bond that undergoes
ESIPT and yields a highly Stokes’-shifted emission from the pro-
ton-transfer tautomer.21–23 Coordination of zinc removes the phe-
nolic proton and disrupts the ESIPT, thus causing emission with a
normal Stokes’ shift. The binding mode of 1 with zinc from the
250
461 nm
0.9
0.6
0.3
0.0
200
-0.3
-0.6
502 nm
-0.9
0-10 eq
-1.2
0.0
0.5
1.0
1.5
2.0
Concentration of Zn+2
0-10 eq
150
100
50
461 nm
502 nm
485 nm
0
400
450
500
550
600
650
Wavelength (nm)
Figure 5. Changes in fluorescence spectra of 1 (20 lM) in EtOH/H2O (9:1) containing HEPES buffer (10 mM, pH 7.4) upon addition of Zn(ClO4)2. (kex = 345 nm) Inset: mol ratio
plots of emission at 461 and 502 nm.

5514
A. Helal, H.-S. Kim / Tetrahedron Letters 50 (2009) 5510–5515
results of fluorescence titration spectra (Fig. 5 inset) and Job’s
2.5
2.0
1.5
1.0
0.5
0.0
plot (Fig. S-2) showed to be 1:1 with
a binding constant
3.2 Â 104 MÀ1. This value is smaller than that of the one obtained
in an aprotic solvent CH3CN. The photophysical properties of 1
and 2 are summarized in Table 1.
The selectivity and tolerance of 1 for zinc ion over other biolog-
ically relevant metal cations such as Na+, K+, Mg2+, Ca2+, and non-
biologically relevant metal cations were investigated by adding
10 equiv of metal cations to 20 lM solution of 1. There was no
obvious blue shift to 461 nm in any other metal ions except zinc.
The paramagnetic transition-metal cations Co2+, Fe2+, Ni2+, and
Cu2+ coordinate to 1, but partially or completely quench the fluo-
rescence emission (Fig. 6). The quenching results obtained with
addition of these cations suggest that Co2+, Fe2+, Ni2+, and Cu2+
which occupy open shell d-orbitals provide a very fast and efficient
non-radiative decay of the excited states due to the electron or en-
ergy transfer between the metal cations and 1. While Zn2+ cation,
which has close shell d-orbitals, does not introduce low-energy
metal-centered or charge-separated excited states energy and elec-
tron transfer processes cannot take place.24 Fluorescence intensity
ratio curve also suggests that only zinc undergoes a blue shift on
coordination with 1 (Fig. 7). Competition binding experiment car-
ried out with different metal ions (1.0 equiv) and zinc (Fig. S-3),
showed that they did not interfere with the ratiometric sensing
of zinc by 1, except in the case of Co2+, Ni2+, Fe2+, and Cu2+ the
enhancement was less when compared to others. The detection
a
b
c
d
e
f
g
h
i
j
k
l
m
n
o
p
Figure 7. Enhancement of fluorescence intensity of 1 (20
containing HEPES buffer (10 mM, pH 7.4) by addition of 10 equiv of respective
metal cation. (a) 1 only, (b) 1 + Ag+, (c) 1 + Hg2+, (d) 1 + Pb2+, (e) 1 + Ca2+, (f) 1 + Cu2+
(g) 1 + Ni2+, (h) 1 + Co2+, (i) 1 + K+, (j) 1 + Zn2+, (k) 1 + Fe2+, (l) 1 + Na+, (m) 1 + Cs+, (n)
1 + Rb+, (o) 1 + Cd2+, (p) 1 + Mg2+
lM) in EtOH/H2O (9:1)
,
.
limit of 1 for zinc ion was found to be 10 lM (Fig. S-5) which
was found to be sufficient for sensing the zinc ion in the biological
system such as brain tissue (0.1–0.5 mM).25
Table 1
pKa value and wavelength of the absorption and emission maxima of sensors 1 and 2 in EtOH/H2O (9:1)
b
Sensor
Absorption max. nm (log
e
)
D
k (nm)
Emission max. (nm)
Complex with Zn2+
D
k (nm)
pKa
Free ligand
Complex with Zn2+
Free ligand
1
329 (3.74)
333 (3.91)
331 (3.74)
384 (3.79)
333 (3.91)
390 (3.74)
55
0
59
502
499
481
461
499
461
41
0
20
12.02
11.55
2
1a
a
In dry CH3CN.
Calculated with 20
b
l
M of 1 and 2 in EtOH/H2O (9:1) containing 0.1 M KCl within a pH range 8–13 (Fig. S-4).14b
250
200
150
Only 1, Ag+1, Pb+2, Ca+2, K+, Na+,
Cs+, Rb+, Hg+2, Mg+2
Zn+2
.
100
50
0
Fe+2
Cd+2
Co+2, Cu+2, Ni+2
400
450
500
550
600
650
Wavelength (nm)
Figure 6. Fluorescence spectra of 1 (20
lM) in EtOH/H2O (9:1) containing HEPES buffer (10 mM, pH 7.4) upon addition of various metal cations (each concentration was
200 M) with an excitation wavelength of 345 nm.
l

A. Helal, H.-S. Kim / Tetrahedron Letters 50 (2009) 5510–5515
5515
Figure 8. Partial 1H NMR spectra of 1 with Zn(ClO4)2 in CD3CN.
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shown in Figure 8. Notably, when 1 equiv of zinc cation was added,
the signals of Ha and Hb shifted downfield due to the deshielding
effect of the metal ion. But in the case of protons Hc, Hd, He, and
Hf experienced a clear upfield shift. It possibly resulted from phe-
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–d orbital interaction through space.26 The –OH proton
peak disappears due to deprotonation.
In summary a new ratiometric chemosensor 1 has been devel-
oped for zinc ion utilizing strong coordination of zinc cation on
the phenolic oxygen and thiazole and pyridine nitrogen atoms,
and thus disruption of ESIPT mechanism. Upon complexation with
zinc ion, this sensor exhibits a blue shift (41 nm) in the emission
spectrum. The strong binding affinity with zinc ion is successfully
achieved by incorporating a pyridine group as the binding ligand
into the fluorophore.
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Acknowledgment
This work was supported by the Kyungpook National University
Research Fund, 2009.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.tetlet.2009.07.078.
19. Shim, J. H.; Jeong, I. S.; Lee, M. H.; Hong, H. P.; On, J. H.; Kim, K. S.; Kim, H.-S.;
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