Benzo[d]imidazo[2,1-b]thiazole-based fluorescent sensor for Zn2+ ion detection

Article abstract of DOI:10.1016/j.jphotochem.2019.112184

In this study, we have synthesized and evaluated three benzo[d]imidazo[2,1-b]thiazole-based sensors (BIT-1, BIT-2 and BIT-3). Among them, BIT-3 namely 2-(benzo[d]imidazo[2,1-b]thiazol-2-yl)-5-methoxyphenol was introduced as a selective fluorescence chemosensor for Zn²⁺ ion detection. The presence of zinc ions enhanced fluorescence property of BIT-3 at 404 nm due to formation of a 1:1 complex between the chemosensor and the Zn²⁺ ion. Control experiments showed that the analogous ions do not have fluorescence enhancement effect and some of them even quench the fluorescence dramatically. The data obtained from UV–vis absorption analysis, fluorescence measurements and ¹H NMR spectroscopy approved the interaction between BIT-3 and Zn²⁺ ion. The sensor probe BIT-3 exhibits a selective fluorescence enhancing property via a chelation-enhanced fluorescence (CHEF) upon addition of Zn²⁺. Thus, a selective fluorescent chemosensor BIT-3 establishes an important sensing platform for real-time monitoring of Zn²⁺ ion in aqueous environment.

Full text of DOI:10.1016/j.jphotochem.2019.112184

JournalofPhotochemistry&PhotobiologyA:Chemistryxxx(xxxx)xxxx
Contents lists available at ScienceDirect
Journal of Photochemistry & Photobiology A: Chemistry
journal homepage: www.elsevier.com/locate/jphotochem
Benzo[d]imidazo[2,1-b]thiazole-based fluorescent sensor for Zn2+ ion
detection
Seyed Ershad Moradia, Sajjad Molavipordanjania, Seyed Jalal Hosseinimehrb, Saeed Emamic,*
a Pharmaceutical Sciences Research Center, Student Research Committee, Faculty of Pharmacy, Mazandaran University of Medical Sciences, Sari, Iran
b Department of Radiopharmacy, Faculty of Pharmacy, Mazandaran University of Medical Sciences, Sari, Iran
c Department of Medicinal Chemistry and Pharmaceutical Sciences Research Center, Faculty of Pharmacy, Mazandaran University of Medical Sciences, Sari, Iran
A R T I C L E I N F O
A B S T R A C T
Keywords:
Benzo[d]imidazo[2,1-b]thiazole
Zinc ions
Fluorescent probe
Chemosensor
CHEF
In this study, we have synthesized and evaluated three benzo[d]imidazo[2,1-b]thiazole-based sensors (BIT-1,
BIT-2 and BIT-3). Among them, BIT-3 namely 2-(benzo[d]imidazo[2,1-b]thiazol-2-yl)-5-methoxyphenol was
introduced as a selective fluorescence chemosensor for Zn2+ ion detection. The presence of zinc ions enhanced
fluorescence property of BIT-3 at 404 nm due to formation of a 1:1 complex between the chemosensor and the
Zn2+ ion. Control experiments showed that the analogous ions do not have fluorescence enhancement effect and
some of them even quench the fluorescence dramatically. The data obtained from UV–vis absorption analysis,
fluorescence measurements and 1H NMR spectroscopy approved the interaction between BIT-3 and Zn2+ ion.
The sensor probe BIT-3 exhibits a selective fluorescence enhancing property via a chelation-enhanced fluores-
cence (CHEF) upon addition of Zn2+. Thus, a selective fluorescent chemosensor BIT-3 establishes an important
sensing platform for real-time monitoring of Zn2+ ion in aqueous environment.
1. Introduction
investigations have focused on the design, development and discovery
of fluorescent sensors for Zn2+ ions [13,21–24]. However, only a few
Zinc has been recently recognized as an essential element in or-
ganisms [1]. It has attracted a considerable attention due to its ex-
istential role in many biological processes [2]. Appropriate levels of
zinc ions prevent teeth caries and osteoporosis [3]. The lack of zinc in
human body have deleterious effect on health including growth re-
tardation, diarrhea, malfunctioning of wound healing and dermatitis
[4], while high concentration of zinc can be toxic. To that end, the
elevated levels of zinc compromises cell survival and is a risk factor in
stroke [5] and Alzheimer's disease [6]. Blood and urine Zn2+ ion con-
centration depends on a wide range of parameters such as biological,
habitual, and environmental conditions [7]. Thus, quantification of zinc
ions concentration in real water and biological samples could provide a
unique diagnostic tool in the clinical, environmental and industrial
areas [8].
Currently, many methods are available for determination of zinc
ion, including atomic absorption spectrometry [9], inductively coupled
plasma [10], electrochemical techniques [11], UV–Vis methods based
on surface plasmon resonance [12] and fluorescence assays [13]. In
particular, fluorescent chemosensors are at the center of attentions due
to their high response speed, good selectivity, high sensitivity and easy
operation character [13–20]. In the recent years, intensive
sensors surmount the drawbacks including weak selectivity, low sen-
sitivity and detection in aqueous medium. Also, the prominent obstacle
for most of the sensors is the requirement of toxic organic solvent for
their operation and single ion detection. Therefore, developing che-
mosensors capable to overcome these limitations is troublesome
[25,26].
In this work, we designed and synthesized a fluorescence probe BIT-
3 derived from benzo[d]imidazo[2,1-b]thiazole (BIT) for Zn2+ ion
detection (Fig. 1). Fluorescent sensor BIT-3 was very selective and
sensitive for Zn2+ in comparison with other cations. We also copiously
studied the optical properties and recognition behaviors of BIT-3 and
related compounds in detail using fluorescence and UV–vis spectro-
scopy. Furthermore, we have also investigated the usefulness of de-
signed sensor for the detection of the Zn2+ ions in real water and
biological samples.
2. Experimental
2.1. Reagents and materials
All the starting materials including 2-aminobenzothiazole, 2-
⁎ Corresponding author at: Department of Medicinal Chemistry, Faculty of Pharmacy, Mazandaran University of Medical Sciences, Sari, Iran.
E-mail addresses: semami@mazums.ac.ir, sdemami12@gmail.com (S. Emami).
https://doi.org/10.1016/j.jphotochem.2019.112184
Received 21 June 2019; Received in revised form 15 September 2019; Accepted 19 October 2019
1010-6030/©2019ElsevierB.V.Allrightsreserved.
Pleasecitethisarticleas:SeyedErshadMoradi,etal.,JournalofPhotochemistry&PhotobiologyA:Chemistry,
https://doi.org/10.1016/j.jphotochem.2019.112184

S.E. Moradi, et al.
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Fig. 1. Chemical structures of designed fluorescent sensors.
bromoacetophenone, 1-(2-hydroxyphenyl)ethanone and 1-(2-hydroxy-
4-methoxyphenyl)ethanone were purchased from Sigma-Aldrich and
used for synthesis of sensors without further purification unless other-
wise mentioned. All solvents used in spectroscopic tests were spectro-
78%; mp 107–108 °C; 1H NMR (400 MHz, DMSO-d6) δ: 8.89 (s, 1 H),
8.40 (d, J =8.0 Hz, 1 H), 8.04 (d, J =8.0 Hz, 1 H), 7.97 (d, J =7.2 Hz,
2 H), 7.58 (t, J =8.0 Hz, 1 H), 7.31–7.51 (m, 4 H). 13C NMR (100 MHz,
CDCl3) δ: 147.94 (1C), 147.41 (1C), 133.63 (1C), 131.95 (1C), 130.06
(1C), 128.64 (2C), 127.43 (1C), 126.06 (1C), 125.06 (2C), 124.77 (1C),
124.19 (1C), 112.53 (1C), 106.80 (1C). MS (m/z, %): 250 (M+, 100),
223 (5), 125 (9), 116 (8), 103 (5), 52 (7), 63 (4). Anal. Calcd for
scopic grade without fluorescent impurity. The solutions of Na+, K+
,
,
Cs+, Ag+, Cd2+, Ba2+, Pb2+, Zn2+, Mg2+, Ca2+, Fe2+, Cu2+, Ni2+
Hg2+, Co2+, Pd2+, Fe3+ and Al3+ ions were prepared from their sul-
fate salts (Sigma-Aldrich).
C
15H10N2S: C, 71.97; H, 4.03; N, 11.19. Found: C, 72.09; H, 3.96; N,
11.31.
2.2. Apparatus
2.3.2. Preparation of 2-(benzo[d]imidazo[2,1-b]thiazol-2-yl)phenol (BIT-
2)
General information about instruments used in this study can be
found in Supplementary Material.
A solution of 2-bromo-1-(2-hydroxyphenyl)ethanone (1b, 215 mg,
1.0 mmol) in ethanol (2 mL) was added to a solution of 2-amino-
benzothiazole (2, 150 mg, 1.0 mmol) in ethanol (3 mL). After stirring at
room temperature (2 h), the reaction mixture was refluxed for 24 h. The
precipitated solid was filtered, washed with cold ethanol and dried to
give BIT-2. Yield 56%; mp 190–193 °C 1H NMR (400 MHz, DMSO-d6) δ:
8.99 (s, 1 H), 8.27 (d, J = 7.6 Hz, 1 H), 8.18 (d, J = 8.0 Hz, 1 H), 7.83
(d, J = 7.6 Hz, 1 H), 7.67 (t, J = 8.0 Hz, 1 H), 7.56 (t, J = 7.6 Hz, 1 H),
7.25 (t, J = 8.0 Hz, 1 H), 7.04 (d, J = 8.0 Hz, 1 H), 6.97 (t, J = 7.6 Hz,
1 H). 13C NMR (100 MHz, DMSO-d6) δ: 154.84 (1C), 146.53 (1C),
138.08 (1C), 131.98 (1C), 130.45 (1C), 130.03 (1C), 127.86 (1C),
127.05 (1C), 125.87 (1C), 120.13 (1C), 117.00 (1C), 115.61 (1C),
115.10 (1C), 111.79 (1C). MS (m/z, %): 266 (M+, 100), 237 (24), 211
(6), 135 (14), 108 (9), 82 (10). Anal. Calcd for C15H10N2OS: C, 67.65;
H, 3.78; N, 10.52. Found: C, 67.82; H, 3.86; N, 10.44.
2.3. Synthesis of chemosensors
Scheme 1 presents the synthetic routes to BIT-1, BIT-2 and BIT-3.
The bromo- intermediates 1b and 1c were prepared according to the
reported methods [27,28]. The synthetic procedures for preparation of
other compounds are described here in details.
2.3.1. Preparation of 2-phenylbenzo[d]imidazo[2,1-b]thiazole (BIT-1)
A solution of 2-bromoacetophenone (1a, 199 mg, 1.0 mmol) in
ethanol (2 mL) was added to a solution of 2-aminobenzothiazole (2,
150 mg, 1.0 mmol) in ethanol (3 mL). After stirring at room tempera-
ture (2 h), the reaction mixture was refluxed for 24 h. A white pre-
cipitated solid was formed after cooling. The solid was separated by
filtration, then washed with cold ethanol and dried to give BIT-1. Yield
Scheme 1. Synthesis of sensors BIT-1, BIT-2 and BIT-3.
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Fig. 2. Absorption spectral changes of BIT-3 (100 μM) in combination with 1.0 equiv of ions in DMSO/H2O (95:5, v/v) solution.
2.3.3. Preparation
of
2-(benzo[d]imidazo[2,1-b]thiazol-2-yl)-5-
2.5. Determination of figures of merit
methoxyphenol (BIT-3)
A
solution of 2-bromo-1-(2-hydroxy-4-methoxyphenyl)ethanone
To obtain the detection limit of BIT-3, fluorescence titration curve
was obtained by addition of Zn2+ (0.1–20 μM). Fluorescence intensity
of BIT-3 was measured five times and the standard deviation of blank
control was calculated. The following equation was used for de-
termining detection limit:
(1c, 245 mg, 1.0 mmol) in ethanol (2 mL) was added to a solution of 2-
aminobenzothiazole (2, 150 mg, 1.0 mmol) in ethanol (3 mL). After
stirring at room temperature (2 h), the reaction mixture was refluxed
for 24 h. The precipitated product (brown solid) was filtered and wa-
shed with cold ethanol. Further purification was carried out by column
chromatography (silica gel, eluting with CHCl3:MeOH = 98:2, v/v) to
afford compound BIT-3. Yield 48%; mp 173–175 °C 1H NMR (400 MHz,
DMSO-d6) δ: 10.85 (s, 1 H), 8.59 (s, 1 H), 8.06 (d, J = 7.6 Hz, 1 H), 8.03
(d, J = 8.4 Hz, 1 H), 7.80 (d, J = 6.4 Hz, 1 H), 7.55 (t, J = 7.6 Hz, 1 H),
7.42 (t, J = 7.6 Hz, 1 H), 6.45–6.55 (m, 2 H), 3.75 (s, 3 H). 13C NMR
(100 MHz, DMSO-d6) δ: 159.99 (1C), 156.31 (1C), 145.84 (1C), 144.47
(1C), 132.27 (1C), 129.56 (1C), 127.62 (1C), 127.11 (1C), 125.49 (1C),
125.38 (1C), 113.93 (1C), 112.41 (1C), 109.42 (1C), 105.99 (1C),
101.94 (1C), 55.47 (1C). MS (m/z, %): 296 (M+, 100), 253 (44), 225
(11), 199 (5), 134 (10), 108 (4), 63 (4). Anal. Calcd for C16H12N2O2S: C,
64.85; H, 4.08; N, 9.45. Found: C, 64.98; H, 3.97; N, 9.44.
3σ
k
i
Detection limit =
(1)
Where σi stands for the standard deviation of the blank, and k is the
slope between the fluorescence intensity versus Zn2+ concentration.
2.6. Quantum yield measurements
The fluorescence quantum yield was measured by using anthracene
as a standard fluorophore (Фs = 0.27, in ethanol) and the following
equation:
n2·F ·A
x
x
Φx =
s ·Φs
n2·F·Ax
(2)
s
s
where, subscripts s and x refer to the reference and the sample, re-
spectively. Letter A stands for absorbance at the excitation wavelength,
n is the solution refractive index, and F represents emission integrated
area. BIT-3 and Zn2+ were dissolved in DMSO/H2O to make a 10 μM
solution. Fluorescence spectra of anthracene (10 μM) were taken in
ethanol.
2.4. UV–vis and fluorescence spectroscopic studies
The titrations for recording absorption and fluorescence spectra
were performed in DMSO/H2O (95:5, v/v) solution. Inorganic salts
solutions were prepared by using distilled water to acquire 2.0 mM
aqueous solution. Due to the high lipophilicity of ligands, the stock
solutions of ligands (2.0 mM) were prepared in DMSO. Aliquot of stock
solution of each ligand was diluted to 2 mL DMSO/H2O (95:5, v/v)
solution in order to provide the final concentration of 20 μM. The re-
sulting solutions contained 20 μM of ligand and cation. The UV–vis
spectra were provided at 200–600 nm using a 1 cm quartz cell. The
fluorescence spectra of ligand (20 μM) in the presence of various
competitive species under the same conditions were recorded at 404 nm
(excitation was provided at 343 nm) in DMSO/H2O (95:5). Both the
excitation and emission slits were set at 5 nm. The selectivity of the
ligand toward Zn2+ was confirmed by adding excess amount (1 equiv.)
of competitive metal ions in the mentioned experimental medium and
the fluorescence emissions were recorded in the absence and presence
of ions.
2.7. Binding constant
Emission titration studies of BIT-3 with a Zn2+ solution (effective
concentration between 0–60 μM) were carried out at its effective con-
centration (20 μM). The spectrophotometric titration data were applied
to calculate the apparent binding constants by Benesi–Hildebrand
(B–H) plot.
1
1
1
=
+
F − F0
K (Fmax − F0)[Zn+2
]
Fmax − F0
(3)
F0 and F are symbols for the BIT-3 fluorescence intensity at maximum
and particular wavelength in the presence of a certain concentration of
the Zn2+, respectively. Fmax is the maximum emission intensity ob-
served in the presence of Zn2+ at 404 nm during titration with varying
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Fig. 3. Fluorescence emission spectra of (a) BIT-1, (b) BIT-2 and (c) BIT-3 (20 μM) in absence and presence of different metal ions in DMSO/H2O (95:5, v/v) solution
(1.0 equiv., λex = 343 nm).
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Fig. 4. (a) Fluorescence emission intensity changes and (b) F/F0 values of BITs (20 μM) in the presence of Zn2+ ions in a DMSO/H2O (95:5, v/v) mixture (1.0 equiv.,
ex = 343 nm).
λ
Fig. 5. Fluorescence enhancement mechanism and probable binding mode of BIT-3-Zn2+ complex.
analytes concentration, and K (M−1) is the slope of the linear plot,
which is equal to apparent binding constant.
2.9. Preparation of real water and bio-fluid samples for Zn2+ determination
The potential applicability of BIT-3 to detect Zn2+ ion in real water
and bio-fluid samples was studied using spikes and recovery method.
Tap water sample was attained at our laboratory water pipe and the
river water samples were acquired from Tajan River (Sari, Mazandaran,
Iran). Human blood serum samples were collected from the Tooba
clinical laboratory (Sari, Mazandaran, Iran). The stock solution of blood
serum sample was prepared by diluting 1 mL of each blood serum
2.8. 1H NMR titration of BIT-3 with Zn2+
The BIT-3 stock solution (1.0 mM) was prepared in DMSO-d6 while
Zn2+ ions solution (1.0 mM) was separately prepared in D2O. After
addition of Zn2+ (1 equiv.) to BIT-3 solution, the 1H NMR spectrums
were recorded immediately.
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Fig. 6. (a) Fluorescence emission intensity change of BIT-3 (20 μM) alone and (b) BIT-3 (20 μM) in the presence of Zn2+ ions in different solvent systems (1.0 equiv.,
λ
ex = 343 nm).
Fig. 7. Effect of pH on the fluorescence intensity of BIT-3 (20 μM) in DMSO/H2O mixture (95:5, v/v) in the absence and presence of Zn2+ ions (20 μM).
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Fig. 8. Fluorescence emission intensity change of BIT-3 (20 μM) in the presence of Zn2+ ion salts (1.0 equiv., λex = 343 nm).
Fig. 9. Competitive selectivity of BIT-3 (20 μM) toward Zn2+ (20 μM) in the presence of other metal ions (1.0 equiv.) with the excitation of 343 nm in a DMSO/H2O
(95:5, v/v) mixture.
Table 1
The obtained tolerance limits of different interfering ions in the determination of Zn2+ (20 μM).
Interfering ions
Tolerance limit (x/Zn2+
)
Li+, Na+, K+, Ca2+, Mg2+, Ba2+, Ni2+,Cu2+, Ag+, Pb2+, Hg2+, and Al3+
Co2+, Cd2+, Mn2+ and Fe2+
500
300
samples into 50 mL of double distillated water. The human urine
samples were collected from 2 male volunteers. Prior to experiment
ultrapure concentrated HNO3 (about 2.0 mL) was added to the urine
samples and the acidified samples were stored at −4 °C. In order to
assure of homogeneity of subsamples, all samples were shaken vigor-
ously for 1 min before each experiment. The quantification was con-
ducted by standard spiked sample addition and recovery.
Fluorescence spectra measurement of real water samples containing
Zn2+ were carried by adding 100 μL of 400 μM stock solution of BIT-3
(final concentration 20 μM) to 1.90 mL sample solutions. After well
mixing, the solutions were allowed to stand at 25 °C for 1 min before
test.
3. Results and discussion
3.1. Spectroscopic studies of BIT-3
3.1.1. UV–vis absorption studies
The UV–vis sensing properties of BIT-3 (100 μM) in DMSO/H2O
(95:5, v/v) were investigated with the addition of metal ions Na+, K+
Cs+, Ag+, Cd2+, Ba2+, Pb2+, Zn2+, Mg2+, Ca2+, Fe2+, Cu2+, Ni2+
,
,
Hg2+, Co2+, Pd2+, Fe3+ and Al3+. As shown in Fig. 2, UV–vis spectra
of free BIT-3 exhibited two maximal absorptions at 272 and 343 nm
(broad band) before titrations. When Zn2+ ions (1.0 equiv) were added,
the absorbance band at 272 nm gradually decreased in intensity, while
the intensity of absorbance band at 343 nm was gradually increased.
The isosbestic point at 286 nm might be attributable to the coordination
between BIT-3 and Zn2+. Whereas, the other tested metal ions dis-
played an obvious decrease in absorption bands at 272 nm with a week
bond at 343 nm. So, a good distinction between Zn2+ and other ions
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Fig. 10. UV–vis spectra for gradual addition of Zn2+ (0 equivalence to 2.0 equivalence) to BIT-3 (100 μM in DMSO/H2O (95:5, v/v)).
Fig. 11. Job’s plot of BIT-3-Zn2+ complex resulted from the measurement of fluorescence emission at 404 nm (λex = 343 nm).
was observed at 343 nm. Based on these results, 343 nm was selected as
role in the fluorescence properties of BIT-3.
excitation wavelength in fluorescence emission studies.
As a parent ligand, BIT-1 didn’t show any selectivity toward the
aforementioned cations with lower fluorescence signal enhancement
(Fig. 4). The 2-hydroxyphenyl derivative BIT-2 showed better se-
lectivity and fluorescence enhancement in comparison with BIT-1. The
introduction of an electron donating group such as the methoxy on the
phenyl moiety of BIT-2 resulted in BIT-3 with fluorescence enhancing
property and better selectivity towards Zn2+ ions. Accordingly, chela-
tion enhanced fluorescence (CHEF) could be a main mechanism for BIT-
3 (Fig. 5). Chemosensors with CHEF mechanism generally have sepa-
rated conjugated units in their semi-rigid structure and would be in-
tegrated into a rigid coplanar conjugation system via metal coordina-
tion. When the Zn(II) ion is coordinated with nitrogen and oxygen
atoms of BIT-3, the free rotation around single bond between aryl ring
and tricyclic benzo[d]imidazo[2,1-b]thiazole system is restricted and
eventually π-conjugation between them is increased, resulting in CHEF
phenomena. The electron donating characteristic of the methoxy group
on the phenyl moiety of BIT-3 facilitates the 2-phenoxy group for more
strongly coordination with Zn ion, and preferring CHEF to be as main
mechanism.
3.1.2. Fluorescence emission studies
3.1.2.1. Selectivity. The selectivity of BITs ligands as chemosensors was
investigated by fluorescence titration (Fig. 3) using a wide range of
metal ions (Na+, K+, Cs+, Ag+, Cd2+, Ba2+, Pb2+, Zn2+, Mg2+, Ca2+
,
Fe2+, Cu2+, Ni2+, Hg2+, Co2+, Pd2+, Fe3+ and Al3+). Addition of 1.0
equiv of all ions except Zn2+ caused
a complete quenching or
decreasing of fluorescence at 404 nm. Certainly, 1.0 equiv Zn2+
resulted in a significant fluorescence increment at 404 nm, which was
completely distinguishable from other metal ions spectrum.
3.1.2.2. Effect of chemical structure. The presence of hydroxy group in
ortho position and methoxy group in para position of phenyl moiety in
BITs chemosensors affect their fluorescence sensitivity and intensity for
Zn2+ ions (Fig. 4a). To that end, BITs relative fluorescence intensity
(Fig. 4b) was calculated and compared. As seen in Fig. 4b, the 2-
hydroxy-4-methoxy derivative BIT-3 showed a higher F/F0 value in
comparison to 2-hydroxy analog BIT-2 and unsubstituted compound
BIT-1. These findings demonstrate that the 2-hydroxy and 4-methoxy
substituents as strong electron-donating fluorophores have important
3.1.2.3. The effect of solvents. The fluorescence emission of a solution
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Table 3
Determination of Zn(II) in real water and biological samples (n = 2).
Table 2
Comparison of organic fluorescence chemosensors for the detection of Zn2+
.
Sensors
Detection
limit (μM)
Binding
Reference
Sample
Added
Found
Recovery
rate (%)
Relative
constant (M−1
)
(μmol
(μmol L−1
)
standard
deviation (RSD)/
%
L−1
)
2.5
1.5 × 104
2.82 × 108
3.4 × 103
[31]
Rain water
0
2
10
0
2
10
0
2
10
0
2
10
0
2
10
0
2
10
0
2
10
0
2
N.D.a
2.082
10.051
N.D.a
2.018
10.149
N.D.a
2.020
9.950
N.D.a
1.989
9.918
N.D.a
1.989
9.885
N.D.a
1.998
10.007
N.D.a
2.018
9.918
N.D.a
2.003
9.971
–
–
104.10
100.51
–
100.90
101.49
–
101.00
99.50
–
99.45
99.18
–
99.45
98.85
–
99.90
100.07
–
100.90
99.18
–
100.15
99.71
1.29
0.62
–
1.76
0.39
–
1.77
0.28
–
1.35
0.90
–
1.86
0.47
–
0.14
2.16
–
1.75
1.01
–
2.23
0.61
River water
(Tajan)
Tap water
0.86
[32]
Urine sample 1
Urine sample 2
Blood serum
sample 1
0.52
[33]
Blood serum
sample 2
Blood serum
sample 3
10
0.06
No data
[34]
a
Not detected.
3.1.2.4. The effect of pH. A series of buffered solutions of BIT-3 [20 μM,
with various pH values (2–12)] with or without Zn2+ (20 μM) was
prepared in HEPES buffer solution in DMSO/H2O (95:5, v/v) mixture.
The fluorescence intensity of the sensor BIT-3 and BIT-3–Zn2+ over a
wide range of pH (2.0 to 12.0) was studied as shown in Fig. 7. Almost
no obvious change in the fluorescence intensity of BIT-3 at 404 nm was
observed in the pH range from 2.0 to 12.0, revealing that the free BIT-3
is stable over the wide pH range.
0.03
0.03
5 × 104
[25]
[35]
1.1 × 107
As seen in Fig. 7, BIT-3 exhibited a sensitive fluorescence en-
hancement response to Zn2+ in the pH range 6–12, but at pH below 5,
BIT-3 exhibited very weak fluorescence intensity. Thus, results of the
pH study confirmed that BIT-3 exhibited a sensitive response to Zn2+ in
the 6–12 pH range, making it suitable for detecting Zn2+ at different pH
levels.
0.07
2.7 × 106
[26]
3.1.2.5. The effect of common anions on BIT-3-Zn2+ complex. The
impact of Zn2+ companion anion on fluorescence emission response
of BIT-3 was evaluated as well. The fluorescence emission of the
solution containing BIT-3 was investigated in the presence of common
salts of zinc (1.0 equiv. of Zn2+) including ZnCl2, Zn(NO3)2, Zn(SO4)2
and Zn(Ac)2 (Fig. 8). Fluorescence spectroscopic results showed that the
type of anion negligibly affects fluorescence intensity of the BIT-3-Zn2+
complex.
0.018
0.03
1.25 × 104
6.8 × 109
[36]
[This work]
3.1.2.6. Competition experiments. In order to evaluate the impact of
other cations on fluorescence emission intensity, the fluorescence was
obtained in the absence and presence of interfering ions in BIT-3 or
BIT-3-Zn2+ and the data were compared. Addition of 20 μM of aqueous
composed of BIT-3 alone or BIT-3 with 1.0 equiv. of Zn2+ was studied
in different solvents including THF, DMF, ethanol, methanol,
acetonitrile, ethyl acetate, DMSO/H2O (10:90, v/v) and DMSO/H2O
(95:5, v:v), and the obtained data are presented in Fig. 6 (a and b,
respectively). BIT-3 in different solvents showed weak and same
fluorescence peak in whole studied domain (350–500 nm). As
solution to the solutions of Li+, Na+, K+, Ag+, Cd2+, Ba2+, Pb2+
,
Mg2+, Ca2+, Fe2+, Cu2+, Ni2+, Hg2+, Co2+, Mn2+, Al3+ and Bi2+
ions, did not produce significant changes in fluorescence emission
intensity of BIT-3-Zn2+ complex (Fig. 9).
Moreover, in order to find the maximum tolerable amounts of var-
ious interfering ions for determination of Zn2+ ion, 20 μM of Zn2+ ion
alone and various concentrations of some diverse ions were added into
the BIT-3–Zn2+ complex solution (20 μM of Zn2+ ion and BIT-3). Then,
the inhibition effects of Zn2+ ion and the mixtures of Zn2+ + Mn+ on
presented in Fig.
6
b, the BIT-3-Zn2+ complex showed good
fluorescence emission intensity in the DMSO/H2O (95:5, v/v) solution
compared with other solvents; therefore this solvent system was used
for further studies.
9

S.E. Moradi, et al.
JournalofPhotochemistry&PhotobiologyA:Chemistryxxx(xxxx)xxxx
Fig. 12. Reversibility experiment of BIT-3 (20 μM) in DMSO/H2O (95:5, v/v) with EDTA and Zn2+ (10 equivalents amount of Zn2+ and EDTA was used).
Fig. 13. The cycle index of BIT-3–Zn2+ (20.0 μM) reacting with Na2EDTA (20.0 μM).
Fig. 14. Time-dependent fluorescence spectroscopy of BIT-3−Zn2+ complex (20 μM, 1 to 1 equiv.).
10

S.E. Moradi, et al.
JournalofPhotochemistry&PhotobiologyA:Chemistryxxx(xxxx)xxxx
and RSD values (lower than 2.23) for all samples.
the BIT-3–Zn2+ complex were examined. As seen in Table 1, most of
the tested ions do not have significant effect on the determination of
Zn2+ (> 5.0% is considered tolerated).
3.4. Reversibility and response time for detection
3.1.2.7. Determination of quantum yield. The fluorescence quantum
yield of BIT-3 was calculated in comparison with anthracene (as the
standard) in the absence and presence of Zn2+ ions in DMSO/H2O
(95:5, v/v). In the absence of Zn2+ ions, the quantum yield of BIT-3
was 6.2%; which increased to 16.5% when 1 equiv. of Zn2+ ions was
added. Therefore the presence of Zn2+ has a significant impact on
quantum yield of BIT-3.
The reversibility of the reaction between BIT-3 and Zn2+ to form a
complex during recognition process was checked by adding ethylene-
diamine tetraacetic acid disodium salt (EDTA) as the competing che-
lating agent, to the BIT-3 and Zn2+ solution (10 equiv of BIT-3). As
shown in Fig. 12, addition of EDTA to the Zn–BIT-3 system immediately
quenches the fluorescence peak at 404 nm. After addition of EDTA,
introduction of Zn2+ to the solution recovers the main of primary
fluorescence, which indicates that BIT-3 and Zn2+ reaction in the
sensing process is reversible to a certain degree. The probe reversibility
can grantee its reusability in the presence of most interfering metal
ions. This result indicates the high reversibility of BIT-3 toward Zn2+
ion sensing and its potential for application in real medium monitoring.
To study the reversible cycle times of the BIT-3 and Zn2+ ion
complex formation and deformation, Zn2+ and EDTA periodically
added to BIT-3 (20 μM) in DMSO/H2O mixture (95:5, v/v), and the
fluorescent spectra (at 343 nm) after 10 cycles was scanned (Fig. 13).
Obviously, the fluorescence intensity of recovered BIT-3 appeared
again by further adding Zn2+ ions, and restored steadily upon repeated
addition of EDTA. As can be seen from Fig. 13, the BIT-3 shows ex-
cellent stability with no significant fluorescence intensity decay for
several successive cycles, which is important for the practical applica-
tions of the chemosensors.
3.1.2.8. Determination of binding constant (Ka). By applying the
obtained fluorescence emission spectroscopy data to Benesi-
Hildebrand equation, the association constant (Ka) was calculated to
be 6.8 × 109 M−1 for the BIT-3–Zn2+ complex [29].
3.1.3. Binding studies
In UV–vis titration study, different concentrations of Zn2+ ions
(0.1–2 equiv.) were added to the BIT-3 solution (100 μM) and the re-
sulted data led to the calculation of BIT-3 to Zn2+ ions ratio (Fig. 10).
As seen, by increasing concentration of Zn2+ in solution, the absorption
of BIT-3 around 343 nm increased gradually. Conversely, the increasing
amounts of Zn2+resuled in decrease of absorption bond at 272 nm. The
presence of isosbestic points at 282 nm revealed that a stable complex
with a certain stoichiometric ratio is formed after addition of Zn2+
.
In addition to UV–vis titration experiment, to verify the ratio of BIT-
3 and Zn2+ ions, Job’s plot experiment also was performed by using
different concentration of ligand and the Zn2+ ion with a total con-
centration of 20 μM. The maximum point in Job’s plot appears at the
mole fraction of 0.5, indicating the 1:1 ratio of BIT-3 and Zn2+ ion
(Fig. 11).
Subsequently, the time-dependent changes of fluorescence intensity
of BIT-3 to Zn2+ were recorded and presented in Fig. 14. The fluores-
cence intensity of BIT-3 at 404 nm was increased rapidly within 25 s
upon addition of 20 μM Zn2+ ions to 20 μM BIT-3 solutions, and even if
the reaction time was extended to 180 s, the fluorescence intensity were
substantially unchanged. It can conclude that BIT-3 exhibits fast re-
cognition to Zn2+ ions in aqueous solutions.
Moreover, we examined the complexation of BIT-3 with Zn2+ by 1H
NMR titration experiments. After addition of Zn2+ (1.0 equiv), the
protons of −OH at 13.72 ppm disappeared completely. It is mainly due
to the addition of D2O and deuterium exchanges with hydrogen.
Moreover, the protons of 6.52, 7.44, 7.55, 7.81 and 8.59 ppm shifted
slightly downfield, which indicates Zn2+ ions can interact with BIT-3 to
form Zn2+–BIT-3 complex.
4. Conclusion
We have introduced a benzo[d]imidazo[2,1-b]thiazole-based fluor-
escent probe BIT-3 as a novel chemosensor for the rapid detection of
Zn2+ ions. The chemosensor BIT-3 was synthesized from 2-amino-
benzothiazole and an appropriate 2-bromoacetophenone. Job’s plot
disclosed a 1:1 stoichiometry between BIT-3 and Zn2+. The association
3.2. Analytical features
constant was evaluated as 6.8 × 109
M
−1. BIT-3 displayed high se-
The validity of procedure was evaluated by determination of linear
dynamic range, coefficient of determination (R2) and limit of detection
as quality parameters. Under optimum conditions, the calibration graph
was linear over the range of 0.10–20.00 μM of Zn2+ with the de-
termination coefficient of 0.9993 (n = 5), which means that BIT-3 is
suitable chemosensor for the detection of Zn2+ ions quantitatively. At
the 0.2 μM concentration of Zn2+ (n = 5), the relative standard de-
viation was found to be 1.82%. According to the definition by IUPAC
(CDL = 3Sb/m) the calculated detection limit was 0.03 μM, which is
thousand times smaller than that of the value of WHO guideline for the
detection of Zn2+ ions in the drinking water samples [30]. The per-
formance of the developed chemosensor was compared with some of
the figures of merit of the recently reported chemosensors for the de-
termination of Zn2+ (Table 2). Obviously, the developed procedure
with the newly designed compound BIT-3 has a better or comparable
detection limit and lower relative standard deviation in comparison
with other reported compounds.
lectivity toward Zn2+ ions over other interfering ions with over 12-fold
fluorescence enhancement, as well as high sensitivity with the detection
limit of 0.03 μM. The recovery studies of the water and biological
samples demonstrated the value of BIT-3 in the practical application as
a sensitive and selective probe for Zn2+ ion.
Declaration of Competing Interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influ-
ence the work reported in this paper.
Acknowledgement
The authors are grateful to Mazandaran University of Medical
Sciences for financial support of this project (grant No. 10387). The
post-doctoral researcher SEM was supported by the Deputy of Research
and Technology, Ministry of Health and Medical Education, Tehran,
Iran.
3.3. Real water and biological samples analysis
The BIT-3 ligand as a chemosensor was applied to detect the content
of Zn(II) in real samples such as different type of water (drinking, rain
and tap water), urine and blood. The results are presented in Table 3.
The data showed satisfactory recovery (in the range of 98.85–104.10%)
Appendix A. Supplementary data
Supplementary material related to this article can be found, in the
online version, at doi:https://doi.org/10.1016/j.jphotochem.2019.
11

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JournalofPhotochemistry&PhotobiologyA:Chemistryxxx(xxxx)xxxx
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