A benzothiazole-based chemosensor for significant fluorescent turn-on and ratiometric detection of Al3+ and its application in cell imaging

Article abstract of DOI:10.1016/j.ica.2018.11.042

Chemosensor Z including benzothiazole fluorophore was synthesized and characterized with NMR and HRMS spectroscopic techniques. Z exhibited fluorescence turn-on and ratiometric absorbance detection of Al³⁺ ions based on the combination mechanis

Full text of DOI:10.1016/j.ica.2018.11.042

Accepted Manuscript
A benzothiazole-based chemosensor for significant fluorescent turn-on and ra-
tiometric detection of Al3+ and its application in cell imaging
Shuang Zeng, Shi-Jie Li, Xue-Jiao Sun, Ming-Qiang Li, Zhi-Yong Xing, Jin-
Long Li
PII:
S0020-1693(18)31376-8
https://doi.org/10.1016/j.ica.2018.11.042
ICA 18657
DOI:
Reference:
Inorganica Chimica Acta
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Revised Date:
Accepted Date:
1 September 2018
23 November 2018
26 November 2018
Please cite this article as: S. Zeng, S-J. Li, X-J. Sun, M-Q. Li, Z-Y. Xing, J-L. Li, A benzothiazole-based chemosensor
for significant fluorescent turn-on and ratiometric detection of Al3+ and its application in cell imaging, Inorganica
Chimica Acta (2018), doi: https://doi.org/10.1016/j.ica.2018.11.042
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A benzothiazole-based chemosensor for significant
fluorescent turn-on and ratiometric detection of Al3+ and its
application in cell imaging
Shuang Zeng a,1, Shi-Jie Li b,1, Xue-Jiao Sun a, Ming-Qiang Li a, Zhi-Yong Xing a,*, Jin-Long Li c
a Department of Applied Chemistry, College of Science, Northeast Agricultural University, Harbin
150030, PR China
b College of Life Science, Northeast Agricultural University, Harbin 150030, PR China
c School of Chemistry and Chemical Engineering, Qiqihar University, Qiqihar 161006, PR China
∗
Corresponding author at: Department of Applied Chemistry, College of Science, Northeast
Agricultural University, Harbin 150030, PR China
E-mail addresses: zyxing@neau.edu.cn, Tel: +86 451 5519 1810, Fax: +86 451 5519 0317
1 These authors contributed equally to this work and should be considered co-first authors.
Abstract: Chemosensor Z including benzothiazole fluorophore was synthesized and characterized
with NMR and HRMS spectroscopic techniques. Z exhibited fluorescence turn-on and
ratiometric absorbance detection of Al3+ ions based on the combination mechanism of CHEF and
ESIPT processes. Moreover, the stiochiometry between Z and Al3+ were determined 2:1 through
job’s plot and HRMS, and the limit of detection was determined as 8.47 nM which was qualified
in the detection of Al3+ in drinking water. Especially, Z was achieved in fast identification the
existence of Al3+ by filter paper, and cell imaging in human stromal cell (HSC).
Key Words: benzothiazole; Al3+; ratiometric detection; filter paper; cells HSC
1

1. Introduction
Aluminum, one of the most abundant metal elements in the earth's crust, is closely associated
with our daily life [1-4]. However, abnormal concentrations of Al3+ in the human body is
detrimental to the health of human owing to it can cause a number of diseases, such as
Alzheimer’s disease, Parkinson’s disease, liver damages, amyotrophic lateral sclerosis, anemia,
and hemochromatosis [5-8]. Moreover, high concentration of Al3+ has obvious reverse effects on
the growth of seeds and roots [9, 10] and can kill fish in acidified water [11]. So, it is a challenge
to find an efficient method for qualitative and quantitative analysis of Al3+ in complex
environments and vivo as well.
Recently, many researchers have paid more attention on the development of fluorescent
probe, but the detection and quantification of Al3+ is always a challenging work owing to the
strong hydration of Al3+ in aqueous media, thus leading to its weak coordination ability compared
with transition metals. Although a lot of fluorescent probe were successfully developed in the
detection of Al3+, there are still some deficiencies, such as weak selectivity, poor solubility in
water and muti-step synthetic methods, which limit their further application. Therefore,
advancement in the synthesis of sensitive, selective and water solubility of Al3+ probes is still in
high demands. 2-(2’-Hydroxyphenyl) benzothiazole (HBT) as a fluorophore have been widely
used in construction chemosensors in the recent years because of its distinguished properties in
photophysical and photochemical fields mainly embodied in its facile structural modification,
dual emission via the excited state intramolecular proton transfer (ESIPT), and excellent photo
stability [12-14]. Furthermore, Al3+, known as a hard acid, prefers a hard-base coordination sphere
containing N and O as the binding sites [15-17], in order to improve the binding ability with Al3+,
2

as show in Scheme 1, a novel HBT-based fluorescent chemosensor Z, affording N and O as
binding sites, had been synthesized by condensation reaction of compound 2 with
4-Hydroxybenzohydrazide. The chemosensor Z itself had weak light yellow fluorescence, due to
the C=N isomerization and possible two-way ESIPT process (Scheme 2). After the chemosensor Z
coordinate with Al3+, the C=N isomerization and ESIPT processes were both inhibited, hence
leading to the chelation induced enhanced fluorescence (CHEF). The exciting results showed that
chemosensor Z exhibited highly selectivity to Al3+ with a significant fluorescence enhancement
and ratiometric absorption detection in DMF-H2O (1/9, v/v, pH = 7.4, 0.01 M HEPES). Moreover,
chemosensor Z was achieved in the detection of Al3+ in real water samples, test paper, CIE
diagram and cell imaging in human stromal cell (HSC).
2. Experimental
2.1. Materials and instruments
Unless otherwise specified, all the solvents and reagents (analytical or spectroscopic grade)
were obtained commercially and used without further purification. 1H NMR spectra and 13C NMR
spectra were recorded on a Bruck AV-600 spectrometer. Chemical shifts (δ) are reported in ppm
using DMSO-d6 as the solvent. Absorption spectra were recorded on a Pgeneral TU-1901 UV–vis
spectrometer at 25 ℃. Fluorescence measurements were measured on a Perkin Elmer LS55
fluorescence spectrometer. Mass spectra were measured on a Waters Xevo UPLC/G2-SQ Tof MS
spectrometer. The melting point was measured on a Beijing XT4-100X microscopic melting point
apparatus. The pH measurements were recorded using a model PHS-3C meter (Shanghai, China).
Cell images were collected using a laser confocal microscope (Leica, TCS SP2 AOBS).
3

2.2. Synthesis
2.2.1 Synthesis of compound 1-2
According to reported methods, compound 2-(benzo[d]thiazol-2-yl) phenol (1), which was
synthesized
through
condensation
[18],
reaction
was
of
used
2-aminobenzenethiol
for the synthesis
with
of
2-hydroxy-5-methylbenzaldehyde
3-(benzo[d]thiazol-2-yl)-2-hydroxybenzaldehyde (2) through Duff reaction [19]. Sensor Z was
prepared through condensation reaction between compound 2 and 4-hydroxybenzohydrazide
(Scheme 1).
2.2.2 Synthesis of Sensor Z
The compound 2 (99.5 mg, 0.37 mmol) and 4-Hydroxybenzohydrazide (58 mg, 0.38 mmol)
were dissolved in ethanol (20 mL), and then the reaction mixture was refluxed 4 h (monitored by
TLC). After completion of reaction, the mixture was cooled to room temperature. The solid was
filtered, washed 5 times with ethanol and dried under vacuum to get the light yellow product Z
1
(122 mg, yield 82%). H NMR (600 MHz, DMSO-d6) (Fig. S1) δ (ppm) 13.42 (s, 1H), 12.18 (s,
1H), 10.23 (s, 1H), 8.66 (s, 1H), 8.22 (s, 1H), 8.17 (d, J = 8.0 Hz, 1H), 8.09 (d, J = 8.0 Hz, 1H),
7.87 (d, J = 8.5 Hz, 2H), 7.60 – 7.52 (m, 2H), 7.47 (td, J = 7.7, 1.0 Hz, 1H), 6.91 (d, J = 8.7 Hz,
2H), 2.40 (s, 3H). 13C NMR (151 MHz, DMSO-d6) (Fig. S2) δ (ppm) 163.03, 162.45, 161.00,
153.88, 151.26, 147.08, 134.84, 133.01, 130.05, 129.74, 128.47, 126.34, 124.98, 122.80, 122.18,
121.91, 119.42, 119.39, 115.09, 19.83. HRMS (m/z) (TOF MS ES-) (Fig. S3): calcd for
C22H16N3O3S: 402.0912 [M-H]-, found: 402.0903.
4

Scheme 1. Synthesis of sensor Z.
2.3. General information
The stock solution of Z (0.1 mM) was prepared in DMF, then diluted with HEPES buffer
(10 mM, pH 7.4)/DMF (1/9, v/v) to 10 μM for the measurement of UV-vis absorption and
fluorescence spectra, respectively. The stock solutions (10 mM) of the cationic salts (Na+, K+,
Mg2+, Ca2+, Ba2+, Cr3+, Mn2+, Fe2+, Fe3+, Co2+, Ni2+, Cu2+, Ag+, Zn2+, Cd2+, Hg2+, Al3+ and Pb2+)
were prepared with ultrapure water, respectively. The excitation was set at 350 nm for the
measurement of fluorescence, and slit widths of the excitation and emission were 10 nm and 10
nm, respectively.
2.4. Preparation of [Z-Al3+]
The compound Z (10 mg, 0.025 mmol) and Al(NO3)3·9H2O (9.4 mg, 0.025 mmol) were
dissolved in ethanol (10 mL). Then the mixture was refluxed for 2 h under stirring. The solution
turned to deep yellow and cooled to the temperature. The Z-Al3+ complexes were obtained by
removing the solvent under reduced pressure. Yield: 86%, color: yellow solid.
2.5. Cell culture and staining
The human stromal cell (HSC) purchased from ATCC (CRL-4003) was routinely cultured in
1:1 mixture of DMEM medium and F-12 medium supplemented with 10% heat-inactivated FBS,
100 U/mL penicillin, 100 μg/mL streptomycin, and 1mM sodium pyruvate at 37 ℃, 5% CO2 for
5

maintained. After plant HSC into 35 mm plates at concentration of 5×104 cells/cm2, the media
without FBS or antibiotic, was used for culture cells and chemical treatment. These cells were
incubated 1 hour with different amounts of Al3+ (0, 5 and 50 µM). Then fibroblast cells of every
group were washed with PBS 3 times and were fixed by using a standard paraformaldehyde
fixation protocol. After fixation, fibroblast cells were rinsed with 4:6 mixture solution of DMF and
water and then stained by incubating for 2 hours with Z (1×10-4 M). Lastly, the cells were
mounted in standard mounting media and imaged by laser confocal microscope. The excitation
wavelength and an emission wavelength were set at 405 nm and 490 nm, respectively.
3. Results and Discussion
The chemosensor Z was synthesized and characterized by 1H NMR, 13C NMR and HRMS
spectrum analysis depicted in scheme 1.
3.1. UV-vis and Fluorescence spectra characteristics for ions
The fluorescence spectral responses of Z toward different metal ions (Mn2+, Na+, Ni2+, Ca2+,
K+,Mg2+, Ba2+, Cr3+, Fe2+, Fe3+, Co2+, Cu2+, Zn2+, Ag+, Hg2+, Cd2+, Al3+ and Pb2+) were
investigated to ascertain the selectivity of Z in DMF/H2O(1/9, v/v, pH = 7.4, 0.01 M HEPES). As
shown in Fig. 1a, probe Z displayed a weak fluorescence-emission (λem = 506 nm) which may be
due to the two aspects as followed. Firstly, the isomerization of C=N moiety, reported by some
researchers [20-22], was the dominant decay process. Secondly, the possible two-way ESIPT
process of Z could lead to fluorescence quenching caused by the nonradiative decay of the excited
state (Scheme 2) [23-25]. Interestingly only Al3+ caused a drastic enhancement in the emission
intensity of Z with a prominent peak at 496 nm, which induced by the inhibition of C=N
6

isomerization and ESIPT processes upon the coordination of Al3+ with Z, hence causing chelation
induced enhanced fluorescence (CHEF) [23, 24]. Moreover, the solution of Z displayed a visible
color change from light yellow to brilliant cyan, which easily detected by naked-eye under UV
light of 365 nm. As for UV-visible absorbance spectrum, the free Z showed two absorption bands
at 300 and 375 nm in DMF/H2O (1/9, v/v, pH = 7.4, 0.01 M HEPES) solution. However, upon
addition of Al3+, the absorption band at 375 nm disappeared and new bands at 325 nm, 340 nm,
425 nm were observed, indicated the formation of complexation between Z and Al3+ (Fig. 1b).
These results indicated the possibility of probe Z for the identification of Al3+ with good
selectivity.
Fig. 1. (a) Fluorescence emission spectra (λex=350 nm) of Z (10 μM) in the presence of 5 equiv. of various metal
ions in DMF-H2O(1/9, v/v, pH = 7.4, 0.01 M HEPES). Inset: The color changes of Z (10 μM) in the presence of
Al3+ ions (5 equiv.) in DMF-H2O (1/9, v/v, containing 0.01 M HEPES, pH = 7.4) under UV light of 365 nm. (b)
Absorption spectra of Z (10 μM) recorded without and with 5.0 equiv. Al3+ in DMF-H2O(1/9, v/v, pH = 7.4, 0.01
M HEPES).
Scheme 2. Probable two-way ESIPT of sensor Z.
7

Subsequently, the experiments including fluorescence and UV–vis titration were measured in
order to evaluate the sensing ability of Z to Al3+, respectively (Fig. 2). Upon the addition of Al3+
(0–1.0 equiv.) to the solution of Z, the emission intensity at 496 nm increased gradually and then
almost reached a platform when the addition of Al3+ was 5μM (0.5 equiv.) (Fig. 2a), indicating the
2:1 binding ratio of Z with Al3+. Plotting of the fluorescence intensity at 496 nm versus the
concentration of Al3+ (0-3 μM) showed a good linear relationship (R2 = 0.99636) (Fig. S4).
Moreover, a significant red-shift with two isobestic points at 356 and 388 nm was observed upon
the gradual addition of Al3+ (Fig. 2b), and the absorbance intensity ratios of Z at 427 nm and 367
nm (A427 /A367) increased linearly with the amount of Al3+ ranging from 1.2 μM to 3.2 μM (Fig.
S5), showing a clear ratiometric absorbance response of the probe Z towards Al3+. The detection
limits of probe Z for Al3+ were found as low as 4.08×10-8 M and 8.47×10-9 M for fluorescence and
absorbance-ratiometric detection, respectively, both of which were quite lower than the
requirement (maximum contaminant is 7.4 μM) of the U.S. Environmental Protection Agency
(EPA). This result showed that probe Z had high sensitivity towards Al3+ ion, and was qualified in
the detection of Al3+ in drinking water.
Fig. 2. (a) Fluorescence spectra of Z (10 μM) on addition of different amount of Al3+ in DMF/H2O (1/9, v/v, pH =
7.4, 0.01 M HEPES). Inset: fluorescence intensity at 496 nm versus the number of equiv. of Al3+ added. (b)
UV–vis absorption spectra of Z (10 μM) in DMF/H2O (1/9, v/v, pH = 7.4, 0.01 M HEPES) upon the addition of
8

Al3+ (1.2-3.2 μM). Inset: absorbance intensity at 427 nm/367 nm as a function of Al3+ (0.12–0.32 equiv.) in
DMF/H2O(1/9, v/v, pH = 7.4, 0.01 M HEPES).
To further confirm the 2:1 stoichiometry between Z and Al3+, Job’s plot analysis was
performed in DMF/H2O (1/9, v/v, pH = 7.4, 0.01 M HEPES). As show in Fig. 3, the maximum
emission intensity of the tested solution was seen when the mole ratio of Al3+ appeared at 0.3,
indicating the 2:1 binding mode between Z and Al3+. This result was further confirmed by a peak
at m/z 831.1619, which was assignable to [2Z - 2H+ + Al3+]+ (calcd. m/z 831.1640) in the ESI
mass spectrum (Fig.4). According to the nonlinear curve fitting of the fluorescence and UV–vis
titration data, the binding constants of probe Z for Al3+ were determined as 2.139×105 M-1 and
1.556×105 M-1 (Fig. S6-S7), respectively, based on the Benesi-Hildebrand plot. This result showed
the excellent stability of Z-Al3+ complex.
Fig. 3. Job plot of Al3+ complex formation. {[Al3+]/([Al3+] + [Z])} is the molar fraction of Al3+ ion.
9

Fig. 4. ESI–MS spectrum of Z (50 μM) upon addition of 5 equiv. of Al3+ in DMF.
Moreover, the high stability of Z-Al3+ complex was evaluated through reversibility analysis
of Z for Al3+ in DMF/H2O (1/9, v/v, pH = 7.4, 0.01 M HEPES). According to recent reports, F-
and EDTA2- had been used to study the reversibility of Z for aluminum ions [26, 27], so we
conducted the reversibility experiment on the probe Z (10 μM) for Al3+ (10 μM) mixed with F-
and EDTA2- (10 μM) (Fig. 5), respectively. The absorption and fluorescence spectra of Z-Al3+ +
EDTA2- and Z-Al3++ F- were almost same as that of Z-Al3+, respectively, which verified the higher
binding constant of Z-Al3+ complex. The comparative analysis between Z and some previously
reported sensors were summarized in Table 1.
Fig. 5. (a) The absorption properties of the probe Z in response to Al3+ (1.0 equiv.) in the presence of F- and
EDTA2- (1.0 equiv.) in DMF/H2O(1/9, v/v, pH = 7.4, 0.01 M HEPES). (b) Fluorescence spectra of Z in response to
Al3+ (1.0 equiv.) in the presence of F- and EDTA2- (1.0 equiv.) in DMF/H2O (1/9, v/v, pH = 7.4, 0.01 M HEPES).
10

Table 1. Comparison of different properties of Z with recently reported probes used as fluorescent sensor for Al3+.
Methods of
detection
Binding
pH
Ref.
Detection Medium
LOD
Recovery
Constants
range
[28]
[29]
Ratiometric fluorescent
Fluorescent
CH 3CN/H2O (1:3)
Methanol
8×10-7
M
4.9×104 M-1
9.1×104 M-1
NR
F-
3-10
NR
9.24×10-7
3.48×10-8
2.72×10-9
9.2×10-8
M
M
M
Colorimetric
[30]
[31]
[32]
Ethanol/H2O (3:1)
Ethanol
1.3×104 M-1
5.06×104 M-1
NR
EDTA2-
EDTA2-
F-
5-11
NR
and fluorescent
Fluorescent
Fluorescent
Tris-HCl aqueous buffer
(0.2% methanol, pH=7.0)
DMSO/H2O (1:5)
M
NR
[33]
Fluorescent
1.05×10-8
M
8.5×105 M-1
F-
4-10
HEPES pH = 7.2
[34]
[35]
[36]
Fluorescent
Fluorescent
Fluorescent
CH 3CN
Methanol
3.1×10-7
M
5.44×104 M-1
6.37×104 M-1
3.21×106 M-1
NR
NR
NR
1.59×10-7
M
EDTA2-
EDTA2-
DMF/H2O (9:1)
6.7×10-6 M
5-13
This
Ratiometric absorbance
fluorescent
DMF/H2O (1:9)
HEPES pH = 7.4
8.47×10-9 Ma
4.08×10-8 Mb
1.556×105 M-1
2.139×105 M-1
None of
4-10
work
EDTA2- and F-
LOD: The limit of detection; NR: Not reported in the corresponding paper.
In order to determine the selectivity of the probe Z to aluminum ions in the presence of other
cations, we also conducted the competitive experiment on the probe Z (10 μM) for Al3+ (50 μM)
mixed with other tested metal ions (Na+, K+, Mg2+, Ca2+, Ba2+, Cr3+, Mn2+, Co2+, Ni2+, Cu2+, Zn2+,
Cd2+, Hg2+ and Pb2+) (50 μM) (Fig. 6).The fluorescence intensity of Z-Al3+ complexation was
almost unaffected except for Co2+, Ni+, and Cu2+, which could inhibit the emission intensity (496
nm) to different extent. Although Co2+, Ni2+ had interference on the detection of Al3+, the
fluorescence intensity was still strong enough for the Al3+detection. The existence of Cu2+ almost
quenched the fluorescence intensity due to its paramagnetic effect. Hence, Z has its limitation in
that it can detect Al3+ in the presence of most metal ion except Cu2+.
11

Fig. 6. Competition experiment of Z toward Al3+ in the presence of 5 equiv. of other cations. [Z] = 10 μM, [Al3+]
= 50 μM, and [Xn+] = 50 μM in DMF/H2O (1/9, v/v, pH = 7.4, 0.01 M HEPES). λex = 350 nm.
3.2. Binding mode and sensing mechanism
The FT-IR spectra of Z and Z-Al3+ complexes were measured (Fig. 7), respectively.
Compared with the FT-IR spectra of Z, the OH absorption at 3409 cm-1 vanished and the C-O of
phenolic hydroxyl shifted from 1278 cm-1 to 1373 cm-1 in the FT-IR spectra of Z-Al3+ complexes,
indicated the deprotonation of phenolic hydroxyl and the coordination of oxygen atom with the
Al3+ ion. Moreover, the typical imide absorption of -C=N- shifted from 1595 cm-1 to 1542 cm-1,
which could be caused by the coordination of the imide N atom to the Al3+ ions according to
previous reports [37].
Fig. 7. FT-IR spectra of Z and Z-Al3+ complex.
12

The 1H NMR experiments in the absence and presence of Al3+ were carried out in DMSO-d6
to understand the exact binding mode of Z-Al3+. As show in Fig. 8, the proton signal of phenolic
hydroxyl (Ha) at 13.42 ppm disappeared, indicating that the oxygen atom of phenolic hydroxyl (a)
of Z was involved in the coordination to Al3+. The proton signal of amide (Hc) disappeared
gradually and a new signal of amide (Hc’) appeared at 9.25 ppm, indicating the isomerization from
(E)-configuration to (Z)-configuration upon the coordination of nitrogen atom of amide with Al3+.
Fig. 8. 1H NMR spectra of Z with Al3+ in DMSO-d6.
1
According to the analysis of Job's plot, FT–IR spectra, ESI–MS and H NMR, a rational
bonding mode of Z for Al3+ was proposed (Scheme 3).
Scheme 3. Probable binding mode of sensor Z with Al3+ ions.
13

3.3. Effect of pH on the fluorescence intensity
In order to get a good view in the practical applicability of the probe Z, the effect of pH on
the fluorescence spectrum of Z in the absence and presence of Al3+ were measured, respectively.
As show in Fig. 9a, the probe Z showed a weak fluorescence at 496 nm at pH 7.0, when the pH
was changed from 8 to 10, the fluorescence spectrum was red-shift accompanied with the
increased intensity centered at 514 nm. The drastic enhancement in emission intensity of Z in
basic environment was attributed to the deprotonation of phenolic hydroxyl, resulting in the
inhibition of ESIPT processes which had been explained in scheme 2. However, when the pH was
changed from 6 to 2, the fluorescence maximum emission peak was shifted to 551 nm, and the
fluorescence intensity increased gradually. The red-shift in the acidic environment might be
attributed to the enhancement of the electron-withdrawing ability of the benzothiazol group upon
binding the H+, thereby enhancing the ICT process upon photoexcitation [38, 39] (Scheme S1).
Furthermore, as show in Fig. 9b, the fluorescence intensity of Z almost did not change at
551 nm in the range of pH 2-3 upon the addition of Al3+. However, significant changes in the
fluorescence spectrum and fluorescent intensity were observed whatever in acidic or alkaline
condition (ranging from the pH 4 to pH 10) compared with Z itself in correspondence pH
condition. These results showed that probe Z could be used as a distinguished probe for Al3+
detection in a wide range from acidic (pH 4) to alkaline (pH 10) medium. Taking above results
into consideration, the most suitable of pH range for the detection of Al3+ was from 4 to 10, based
on the result of fluorescence intensity at 496 nm as recorded signal at different pH range from 2
to12 (Fig.S8).
14

Fig. 9. (a) Plot showing the influence of Z (10 μM) in DMF/H 2 O (1:9, v/v) with pH; (b) plot showing the
influence of solution Z (10 μM)-Al3+ (50 μM) in DMF/H 2 O (1:9, v/v) at different pH medium.
4. Analytical application
4.1. Application of Z for Al3+ analysis in water samples
In order to further explore the practical application of Z, experimental verification was
employed to detect aluminum ions in tap water and pure water. The accuracy was investigated by
adding a known amount of standard Al3+ to the samples (Table 2). The results showed that probe Z
had good recoverability and high accuracy for the practical application of aluminum ions in water,
which was of great practical value for the detection of Al3+ in environmental analysis.
Table 2. Determination of Al3+ in water samples from different water sources by standard-addition method (n = 3).
Amount of
Water samples
studied
Total Al3+ found
Recovery of Al3+
(n=3) added (%)
RSD
(%)
Relative
standard Al3+
(n=3) (μmol/L)
error (%)
added (μmol/L)
0.200
0.400
0.200
0.400
0.203
0.406
0.193
0.405
101.50
101.50
96.50
1.06
1.16
1.18
1.44
1.50
1.50
-3.50
1.25
ultrapure water
Tap water
(Department of
Chemistry)
101.25
15

4.2. Application in Test Paper
Considering Z has good selectivity to Al3+, the colormetric experiments of probe Z for Al3+
on test paper were examined (Fig. 10). Test papers were emerged in the solution of Z mixed with
different concentrations of Al3+ (0-3 μM) and then dried in air. With increased concentration of
Al3+, the test paper displayed a visible color change from light yellow to brilliant cyan, facilitating
in the “naked-eye” detection of Al3+ under UV light of 365 nm. Hence, this interesting result
showed that such test papers had potential application in the fast detection the existence of Al3+ in
a certain system.
Fig. 10. The photographs of probe Z on test strips with different concentrations of Al3+ at room temperature under
365 nm UV light.
4.3. Application in CIE diagram
For better understand the trace of the fluorescence changes of the probe Z with addition of
aluminum ions, the CIE chromaticity coordinates were also calculated from the emission spectrum
[40]. The CIE system is a two-dimensional space (XY plane), each point on the chromaticity
diagram represents a certain color (Fig. S9). The CIE chromaticity coordinates of Z was found to
be x=0.2579, y=0.5294 at room temperature. Upon the addition of Al3+ (0-3 μM) to the solution of
Z, the CIE chromaticity coordinates of Z-Al3+ complex was finally found to be x=0.1441,
y=0.4183 when the aluminum ions concentration was 3 μM. This result indicates that the color
coordinates shifts gradually from chartreuse to cyan color region upon progressive addition of Al3+
16

into the solution of Z, which is also consistent with the result of the test paper experiment.
4.4. Cells imaging
For evaluation of the potential of detecting Al3+ in cells with Z, the human stromal cell
(HSC), this endometrium fibroblast cell line was derived from the human stromal cells and
immortalized with hTERT, has been used as an in vetro model. As determined by laser confocal
microscope (λex= 405 nm, λem= 490 nm) (Fig. 11 A), the cells incubated with 0 µM of Al3+ and 0.1
mM of Z for 2 hours did not give any intracellular fluorescence. When cells were incubated with 5
µM of Al3+ and 0.1 mM of Z for 2 hours, weak fluorescence was observed (Fig. 11 B). After cells
were incubated with 50 µM of Al3+ and 0.1 mM of Z for 2 hours, strong fluorescence appeared
(Fig. 11 C). These results suggest that the probe Z have potential bio-medical applications.
Fig. 11. (A) The cells was incubated 2 hours with Z (0.1 mM); (B) cells was incubated 5 μM of Al3+ and 0.1 mM
of Z; (C) cells was incubated 50 μM of Al3+ and 0.1 mM of Z.
17

5. Conclusions
In conclution, chemosensor Z with benzothiazole fluorophore was synthesized and employed
in the detection of Al3+ ions based on the combination mechanism of CHEF and ESIPT processes.
Z exhibited fluorescence turn-on and ratiometric absorbance detection of Al3+ ions. A 2:1 binding
mode of Z and Al3+ was confirmed by job’s plot and HRMS, and the detection limit (8.47 nM)
was qualified in the detection of Al3+ in drinking water. Moreover, Z was achieved in fast
determination the existence of Al3+ by filter paper, and cell imaging in human stromal cell (HSC).
Acknowledgements
This work was supported by the Postdoctoral Scientific Research Developmental Fund of
Heilongjiang Province (No. LBH-Q14023).
Appendix A. Supporting information
Supplementary data associated with this article can be found in the online version at
References
[1] Q. P. Diao, P. Y. Ma, L. L. Lv, T. C. Li, Y. Sun, X. H. Wang, D. Q. Song, A water-
soluble and reversible fluorescent probe for Al3+ and F- in living cells, Sens. Actuators B:
Chem. 229 (2016) 138-144.
[2] B. Sen, S. K. Sheet, R. Thounaojam, R. Jamatia, A. K. Pal, K. Aguan, S. Khatua, A coumarin
based Schiff base probe for selective fluorescence detection of Al3+ and its application in live
cell imaging. Spectrochim. Acta Part A 173 (2017) 537–543.
18

[3] U. Panda, S. Roy, D. Mallick, A. Layek, P. P. Ray, C. Sinha, Aggregation induced emission
enhancement (AIEE) of fluorenyl appended Schiff base : A turn on fluorescent probe for Al3+,
and its photovoltaic effect, J. Lumin. 181 (2017) 56–62.
[4] Y. S. Dong, T. Q. Liu, X. J. Wan, H. Pei, L. S. Wu, Y. W. Yao, Facile one-pot synthesis of
bipyridine-based dual-channel chemosensor for the highly selective and sensitive detection of
aluminum ion, Sens. Actuators B: Chem. 241 (2017) 1139–1144.
[5] P. F. Good, C. W. Olanow, D. P. Perl, Neuromelanin-containing neurons of the substantia nigra
accumulate iron and aluminum in Parkinson's disease: a LAMMA study, Brain Res. 593
(1992) 343–346.
[6] G. D. Fasman, Aluminum and Alzheimer's disease: model studies, Coord. Chem. Rev. 149
(1996) 125-165.
[7] M. Kawahara, K. Muramoto, K. Kobayashi, H. Mori, Y. Kuroda, Aluminum Promotes the
Aggregation of Alzheimer's Amyloid β-Protein in Vitro, Biochem. Biophys. Res.
Commun. 198 (1994) 531–535.
[8] Y. Wang, Z. Y. Ma, D. L. Zhang, J. L. Deng, X. Chen, C. Z. Xie, X. Qiao, Q. Z. Li, J. Y. Xu,
Highly selective and sensitive turn-on fluorescent sensor for detection of Al3+ based on
quinoline-base Schiff base, Spectrochim. Acta, Part A 195 (2018) 157–164.
[9] D. L. Godbold, E. Fritz, A. Hüttermann, Aluminum toxicity and forest decline, Proc. Natl.
Acad. Sci. U. S. A. 85 (1988) 3888–3892.
[10] H. Sade, B. Meriga, V. Surapu, J. Gadi, M. S. L. Sunita, P. Suravajhala, P. B. K. Kishor,
Toxicity and tolerance of aluminum in plants: tailoring plants to suit to acid soils, Biometals,
29 (2016) 187–210.
19

[11] J. Barceló, C. Poschenrieder, Fast root growth responses, root exudates, and internal
detoxification as clues to the mechanisms of aluminium toxicity and resistance: a review.
Environ. Exp. Bot. 48 (2002) 75–92.
[12] Z. Huang, S. Ding, D. Yu, F. Huang. G. Feng, Aldehyde group assisted thiolysis of
dinitrophenyl ether: a new promising approach for efficient hydrogen sulfide probes, Chem.
Commun. 50 (2014) 9185–9187.
[13] C. Y. Yang, Y. Chen, K. Wu, T. Wei, J. L. Wang, S. S. Zhang, Y. F. Han, A reactive probe for
Cu2+ based on the ESIPT mechanism and its application in live-cell imaging, Anal. Methods,
7 (2015) 3327–3330.
[14] B. Gu, Y. Chen, L. Y. Huang, W. Su, X. L. Duan, H. T. Li, S. Z. Yao,
A benzothiazole-based fluorescent probe for distinguishing and bioimaging of Hg2+ and Cu2+,
Anal. Chim. Acta 954 (2017) 97-104.
[15] D. Maity, T. Govindaraju, Conformationally Constrained (Coumarin-Triazolyl-Bipyridyl)
Click Fluoroionophore as a Selective Al3+ Sensor, Inorg. Chem. 49 (2010) 7229-7231.
[16] C. S. Liang, W. H. Bu, C. L. Li, G. W. Men, M. Y. Deng, Y. K. Jiangyao, H. C. Sun, S. M.
Jiang, A highly selective fluorescent sensor for Al3+ and the use of the resulting complex as a
secondary sensor for PPi in aqueous media: its applicability in live cell imaging, Dalton
Trans. 44 (2015) 11352-11359.
[17] W. H. Wang, Z. F. Mao, M. D. Wang, L. J. Liu, D. W. J. Kwong, C. H. Leung, D. L. Ma, A
long lifetime luminescent iridium(III) complex chemosensor for selective switch-on detection
of Al3+ ions, Chem. Commun. 52 (2016) 3611–3614.
[18] C. C. Chang, F. Wang, J. Qiang, Z. J. Zhang, Y. H. Chen, W. Zhang, Y. Wang, X. Q. Chen,
20

Benzothiazole-based fluorescent sensor for hypochlorite detection and its application for
biological imaging, Sens. Actuators B: Chem. 243 (2016) 22–28.
[19] L. J. Tang, X. Dai, X. Wen, D. Wu, Q. Zhang, A rhodamine–benzothiazole conjugated sensor
for colorimetric, ratiometric and sequential recognition of copper (II) and sulfide in aqueous
media, Spectrochim. Acta, Part A 139 (2015) 329–334.
[20] E. T. Feng, R. M. Lu, C. B. Fan, C. H. Zheng, S. Z. Pu, A fluorescent sensor for Al3+ based on
a photochromic diarylethene with a hydrazinobenzothiazole Schiff base unit, Tetrahedron
Lett. 58 (2017) 1390–1394.
[21] E. T. Feng, C. B. Fan, N. S. Wang, G. Liu, S. Z. Pu, A highly selective diarylethene
chemosensor for colorimetric detection of CN− and fluorescent relay-detection of Al3+ /Cr3+,
Dyes Pigm. 151 (2018) 22–27.
[22] S. Y. Li, D. B. Zhang, J. Y. Wang, R. M. Lu, C. H. Zheng, S. Z. Pu, A novel
diarylethene-hydrazinopyridine-based probe for fluorescent detection of aluminum ion and
naked-eye detection of hydroxide ion, Sens. Actuators B: Chem. 245 (2017) 263–272.
[23] L. J. Tang, S. L. Ding, K. L. Zhong, S. H. Hou, Y. J. Bian, X. M. Yan, A new
2-(2′-hydroxyphenyl)quinazolin-4(3H)-one derived acylhydrazone for fluorescence
recognition of Al3+, Spectrochim. Acta, Part A 174 (2017) 70–74.
[24] Z. Liao, Y. Liu, S. F. Han, D. Wang, J. Q. Zheng, X. J. Zheng, L. P. Jin, A novel
acylhydrazone-based derivative as dual-mode chemosensor for Al3+, Zn2+and Fe3+ and its
applications in cell imaging, Sens. Actuators B: Chem. 244 (2017) 914–921.
[25] S. B. Roy, K. K. Rajak, A quinoline appended naphthalene derivative based AIE
active“turn–on” fluorescent probe for the selective recognition of Al3+ and colourimetric
21

sensor for Cu2+ : Experimental and computational studies, J. Photochem. Photobiol. A: Chem.
332 (2017) 505–514.
[26] A. Roy, S. Dey, P. Roy, A ratiometric chemosensor for Al3+ based on naphthalene-quinoline
Conjugate with the resultant complex as secondary sensor for F−: Interpretation of molecular
logic gates, Sens. Actuators B: Chem. 237 (2016) 628–642.
[27] F. Yu, L. J. Hou, L. Y. Qin, J. B. Chao, Y. Wang, W. J. Jin, A new colorimetric and turn-on
fluorescent chemosensor for Al3+ in aqueous medium and its application in live-cell imaging,
J. Photochem. Photobiol. A: Chem. 315 (2016) 8–13.
[28] D. J. Wang, X. P. Fan, S. G. Sun, S. Z. Du, H. J. Li, J. L. Zhu, Y. F. Tang, M. X. Chang, Y. Q.
Xu, Substituent effect: A new strategy to construct a ratiometric fluorescent probe for
detection of Al3+ and imaging in vivo, Sens. Actuators B: Chem. 264 (2018) 304–311.
[29] C. Lim, H. Seo, J. H. Choi, K. S. Kim, A. Helal, H. S. Kim, Highly selective fluorescent
probe for switch-on Al3+ detection and switch-off F- detection, J. Photochem. Photobiol. A:
Chem. 356 (2018) 312–320.
[30] B. J. Pang, C. R. Li, Z. Y. Yang, Design of a colorimetric and turn-on fluorescent probe for
the detection of Al(III), J. Photochem. Photobiol. A: Chem. 356 (2018) 159–165.
[31] X. L. Yue, Z. Q. Wang, C. R. Li, Z. Y. Yang, A highly selective and sensitive fluorescent
chemosensor and its application for rapid on-site detection of Al3+, Spectrochim. Acta, Part A
193 (2018) 415–421.
[32] Q. P. Diao, P. Y. Ma, L. L. Lv, T. C. Li, Y. Sun, X. H. Wang, D. Q. Song, A water-soluble and
reversible fluorescent probe for Al3+ and F− in living cells, Sens. Actuators B: Chem. 229
(2016) 138–144.
22

[33] D. Sarkar, P. Ghosh, S. Gharami, T. K. Mondal, N. Murmu, A novel coumarin based
molecular switch for the sequential detection of Al3+ and F−: Application in lung cancer live
cell imaging and construction of logic gate, Sens. Actuators B: Chem. 242 (2017) 338–346.
[34] M. Tajbakhsh, G. B. Chalmardi, A. Bekhradnia, R. Hosseinzadeh, N. Hasani, M. A. Amiri, A
new fluorene-based Schiff-base as fluorescent chemosensor for selective detection of Cr3+
and Al3+, Spectrochim. Acta, Part A 189 (2018) 22–31.
[35] N. N. Li, S. Zeng, M. Q. Li, Y. Q. Ma, X. J. Sun, Z. Y. Xing, J. L. Li, A Highly Selective
Naphthalimide-Based Chemosensor: “Naked-Eye” Colorimetric and Fluorescent Turn-On
Recognition of Al3+ and Its Application in Practical Samples, Test Paper and Logic Gate, J.
Fluoresc. 28 (2018) 347–357.
[36] S. M. Hossain, K. Singh, A. Lakma, R. N. Pradhan, A. K. Singh, A schiff base ligand of
coumarin derivative as an ICT-Based fluorescence chemosensor for Al3+, Sens. Actuators B:
Chem. 239 (2017) 1109–1117.
[37] H. Y. Liu, B. B. Zhang, C. Y. Tan, F. Liu, J. K. Cao, Y. Tan, Y. Y. Jiang, Simultaneous
bioimaging recognition of Al3+ and Cu2+ in living-cell, and further detection of F-and S2- by a
simple fluorogenic benzimidazole-based chemosensor, Talanta 161 (2016) 309–319.
[38] W. F. Niu, Z. W. Wei, J. Jia, S. M. Shuang, C. Dong, K. M. Yun, A ratiometric emission
NIR-fluorescent probe for sensing and imaging pH changes in live cells, Dyes Pigm. 152
(2018) 155–160.
[39] M. T. Sun, L. B. Du, H. Yu, K. Zhang, Y. Liu, S. H. Wang, An intramolecular charge transfer
process based fluorescent probe for monitoring subtle pH fluctuation in living cells, Talanta
162 (2017) 180–186.
23

[40] M. Kumar, A. Kumar, M. S. H. Faizi, S. Kumar, M. K. Singh, S. K. Sahu, S. Kishor, R. P.
John, A selective ‘turn-on’ fluorescent chemosensor for detection of Al3+ in aqueous medium:
Experimental and theoretical studies, Sens. Actuators B: Chem. 260 (2018) 888–899.
24

25

Highlights
●Chemosensor Z with benzothiazole fluorophore was synthesized and
characterized.
●Chemosensor Z indeed exhibits highly selective fluorescence
responses to Al3+ with a 9-fold fluorescence enhancement and
ratiometric absorption detection
●The limit of detection (LOD) for Al3+ was 4.08×10-8 M and 8.47×10-9 M
for fluorescence and absorbance-ratiometric detection, respectively.
● Z was successfully applied to form paper strip sensor for Al3+ and cell
imaging in HSC.
26