A turn-on chemosensor based on naphthol-triazole for Al(III) and its application in bioimaging

Article abstract of DOI:10.1016/j.tetlet.2013.04.115

A new Schiff base 1-[(1H-1,2,4-triazole-3-ylimino)-methyl]-naphthalene-2-ol (H₂L) exhibiting high selectivity for Al³⁺ ion over other metal ions, such as Li⁺, Na⁺, K⁺, Ca ²⁺, Mg²⁺, Cu²⁺, Co²⁺, Mn ²⁺, Ni²⁺, Zn²⁺, Cd²⁺, Pb ²⁺, Fe³⁺, and Cr³⁺ was prepared, and it is sensitive for Al³⁺ with the detection limit reaching 0.69 μM in DMF. Upon addition of Al³⁺, the significant enhancement (32-fold) of fluorescence intensity for H₂L at 466 nm is ascribed to the formation of a 1:1 complex between Al³⁺ and H₂L, which is denoted as the chelation-enhanced fluorescence (CHEF) effect. The confocal fluorescence microscopy experiments demonstrate that H₂L could be used as a fluorescent probe for Al³⁺ in living cells.

Full text of DOI:10.1016/j.tetlet.2013.04.115

Tetrahedron Letters 54 (2013) 3471–3474
Contents lists available at SciVerse ScienceDirect
Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
A turn-on chemosensor based on naphthol–triazole for Al(III)
and its application in bioimaging
⇑
Tian-Jing Jia, Wei Cao, Xiang-Jun Zheng , Lin-Pei Jin
Beijing Key Laboratory of Energy Conversion and Storage Materials, College of Chemistry, Beijing Normal University, Beijing 100875, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
A new Schiff base 1-[(1H-1,2,4-triazole-3-ylimino)-methyl]-naphthalene-2-ol (H2L) exhibiting high
selectivity for Al3+ ion over other metal ions, such as Li+, Na+, K+, Ca2+, Mg2+, Cu2+, Co2+, Mn2+, Ni2+
,
Received 13 February 2013
Revised 17 April 2013
Accepted 26 April 2013
Available online 3 May 2013
Zn2+, Cd2+, Pb2+, Fe3+, and Cr3+ was prepared, and it is sensitive for Al3+ with the detection limit reaching
0.69 l
M in DMF. Upon addition of Al3+, the significant enhancement (32-fold) of fluorescence intensity
for H2L at 466 nm is ascribed to the formation of a 1:1 complex between Al3+ and H2L, which is denoted
as the chelation-enhanced fluorescence (CHEF) effect. The confocal fluorescence microscopy experiments
demonstrate that H2L could be used as a fluorescent probe for Al3+ in living cells.
Ó 2013 Elsevier Ltd. All rights reserved.
Keywords:
Al(III) sensor
Schiff base
Fluorescence
Turn-on
Cell imaging
Aluminum is the third most prevalent element and the most
abundant metal in the biosphere, accounting for approximately
8% of the Earth’s mass, and extensively used in modern life.1
Aluminum in its ionic form (Al3+) is found in most animal and plant
tissues, and in natural waters. High concentration of Al3+ hampers
plant performance,2 killing fish, algae, bacteria, and other species
in aquatic ecosystems.3 Also, Al3+ causes neurofibrillary, enzy-
matic, and neurotransmitter changes in the central nervous sys-
tem. Human illnesses such as dementia and encephalopathy,
Parkinson and Alzheimer diseases are believed to be attributed to
the nitrogen and oxygen donor sites can contribute to the high
stability of the resultant complexes.8 Thus, we selected 2-hydro-
xy-1-naphthaldehyde as a fluorophore and binding moiety, and
3-amino-1H-1,2,4-triazole as a synergic ligand to provide more
coordination sites for metal ions to synthesize a new Schiff base,
named as 1-[(1H-1,2,4-triazole-3-ylimino)-methyl]-naphthalene-
2-ol (H2L).
H2L was synthesized as shown in Scheme 1 and characterized
by elemental analysis, IR, and single-crystal X-ray diffraction (see
Supplementary data). The crystallographic data and structure
refinement, and key bond distances and bond angles of H2L are
listed in Tables S1 and S2. The crystal structure of H2L is shown
in Figure 1. The bond distance of C3–N4 is 1.292 Å, indicating the
feature of a double bond. The double bond, –C@N– links the tria-
zole and naphthalene moieties to a coplanar molecule. Single crys-
tal X-ray diffraction reveals that intramolecular and intermolecular
hydrogen bonds exist in H2L. The O–HÁÁÁN (1.817 Å) intramolecular
the toxicity of Al3+ 4,5 Therefore, the facile detection of Al3+ is
.
crucial in environmental monitoring and biological assays, and
remains a major requirement. However, compared to other transi-
tion metal ions, limited examples of fluorescence sensors based on
small molecules for Al3+ have been reported.6,7
Nitrogen and oxygen-rich coordination environments can pro-
vide a hard-base environment for the hard-acid Al3+. Moreover,
CHO
HO
N
N
H
C
OH
80 ഒ
HN
N
HN
NH
+
2
isopropanol
N
N
Scheme 1. The synthesis of H2L (2-hydroxy-1-naphthaldehyde, 3-amino-1H-1,2,4-triazole, isopropanol, 80 °C, yield: 57%).
⇑
Corresponding author. Tel.: +86 1058805522.
E-mail address: xjzheng@bnu.edu.cn (X.-J. Zheng).
0040-4039/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tetlet.2013.04.115

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T.-J. Jia et al. / Tetrahedron Letters 54 (2013) 3471–3474
200
(a)
Al3+
150
100
50
free H L, and
2
other metal ions
Zn2+
0
450
500
550
600
650
Figure 1. The crystal structure of chemosensor H2L.
Wavelength (nm)
hydrogen bonds are formed between the oxygen atom (O1) of
naphthol hydroxyl and the nitrogen atom (N4), and the N–HÁÁÁN
(2.191 Å) intermolecular hydrogen bonds are formed between
the nitrogen atom (N2) of triazole moiety and nitrogen atom
(N3) from another molecule.
180
150
120
90
(b)
(c)
The excitation and emission spectra of H2L are shown in Figure
S1. When H2L is excited at 288 nm, a broad emission band can be
observed at 348 nm, while H2L shows weak emission upon excita-
tion at 442 nm. However, upon addition of Al(NO3)3, the fluores-
cence intensity of H2L increased by a factor of 32 at 466 nm
when excited at 442 nm owing to the formation of a complex
between H2L and Al3+. In contrast, addition of other relevant metal
60
30
ions, such as Li+, Na+, K+, Ca2+, Mg2+, Cu2+, Co2+, Mn2+, Ni2+, Zn3+
,
0
Cd2+, Pb2+, Fe3+, and Cr3+ has almost no fluorescence enhancement.
The fluorescence response behavior of H2L upon addition of various
metal ions in DMF is shown in Figure 2a. The selectivity of H2L to
Al3+ was plotted as a bar graph in Figure 2b; only Al3+ resulted in
a pronounced fluorescence enhancement relative to the control
ions. To evaluate the selectivity of H2L to Al3+ in practice, the sys-
tems of Al3+ coexisting with other metal ions were examined in
DMF. As shown in Figure 2c, relatively low fluorescence intensities
were observed in the presence of other metal ions except Pb2+. The
response of H2L for Al3+ in the presence of Fe3+ was obviously low
but can be detectable. There was almost no fluorescence of H2L
upon addition of Pb2+ alone, however, the remarkable enhance-
ment in fluorescence intensity of H2L with Al3+ in the presence of
Pb2+ ion was observed and should be attributed to a cooperative
interaction which was triggered by Pb2+ ion.9 Pb2+ ion can combine
with nitrogen atom of the triazole group, as well as the oxygen
atom from naphthol hydroxyl, which may lead to fluorescence
enhancement. When H2L was titrated with Al3+, the fluorescence
intensity increased steeply up to 7 equiv and then remained almost
the same at the concentration over 13 equiv (Fig. 3). This could be
attributed to the triazole group in H2L to provide nitrogen atoms
and more Al3+ ions are coordinated with them. Therefore, H2L pro-
Al3+Ca2+Cd2+Co2+Cr2+Cu2+Fe2+K+Li+Mg2+Mn2+Na+Ni2+Pb2+Zn2+no
240
200
160
120
80
40
0
Al3+ Al3+ Al3+ Al3+ Al3+ Al3+ Al3+ Al3+Al3+ Al3+ Al3+ Al3+Al3+Al3+Al3+
Ca2+Cd2+Co2+Cr3+Cu2+Fe3+ K+ Li + Mg2+Mn2+Na+Ni2+Pb2+Zn2+
Figure 2. (a) Fluorescence spectra of H2L (50.0
(1.0 equiv) of Li+, Na+, K+, Ca2+, Mg2+, Cu2+, Co2+, Mn2+, Ni2+, Zn2+, Cd2+, Pb2+, Fe3+
Cr3+, and Al3+ in DMF. kex = 442 nm. (b) A bar graph for fluorescence responses of
H2L (50.0 M) to various cations (1.0 equiv) in DMF at 466 nm. (c) Fluorescence
responses of H2L (50.0 M) to various metal ions (1.0 equiv) in DMF at 466 nm.
lM) upon the addition of metal salts
,
l
l
vided a wide sensing range for Al3+
.
kex = 442 nm. The black bars represent the emission intensity of H2L in the presence
of other metal ions. The light gray bars represent emission intensity of a mixture of
H2L with other metal ions followed by addition of Al3+ to the solution, respectively.
1H NMR for H2L is shown in Figure 4a. The assignment for H2L
was also confirmed by 1H MBC, 1H SQC, and H-H COSY (Figs. S2–
S4). The binding mode of Al3+ with H2L was examined by 1H
NMR spectra of H2L in DMF-d7 recorded upon addition of various
amounts of Al3+ (Fig. 4). The proton signal of the naphthol hydroxyl
at 14.37 ppm disappeared when Al3+ and H2L were mixed in a 1:1
ratio (Fig. 4b), and at the same time, the proton signal of C@N was
downfield shifted by 0.71 ppm. Furthermore, compared to 1H NMR
spectrum of H2L upon addition of 1.0 equiv Al3+, 1H NMR remained
nearly unchanged upon addition of 1.5 equiv Al3+ (Fig. 4c). These
results indicated that the nitrogen atom of imine and oxygen atom
exhibited three absorption bands at 267, 333, and 380 nm. We fur-
ther observed changes in the absorbance of H2L upon addition of
Al(NO3)3 (Fig. 5a). As there was a gradual addition of Al3+ in
increasing concentration (0–1.4 equiv), the regular decrease and
increase occurred at the bands 380 nm and 435 nm, respectively.
A well-defined isosbestic point at ca. 402 nm was observed, indic-
ative of a clean conversion of H2L into the Al3+–H2L complex. Job’s
plot (Fig. 5b) which was based on the changes of absorbance at
435 nm, confirmed a 1:1 stoichiometry between Al3+ ion and H2L.
of naphthol hydroxyl were the binding sites for Al3+
.
We then studied the absorbance changes of H2L upon addition
of metal ions (Fig. S5). The electronic absorption spectrum of H2L
Moreover,
which we could observe
a
positive-ion ESI mass spectrum (Fig. S6), from
peak at m/z 380.0 assigned to
a

T.-J. Jia et al. / Tetrahedron Letters 54 (2013) 3471–3474
3473
160
140
120
100
80
0.8
(a)
140
120
100
80
16 eq
0.4
0.3
0.2
60
0.6
40
20
0
1.4eq
0.0
0.4
0.8
1.2
0
2
4
6
8
10 12 14 16
3+
Amount of Al (eq)
3+
Amount of Al (eq)
0.4
0.2
0.0
60
40
20
0
450
500
550
Wavelength (nm)
600
650
250
300
350
400
450
500
550
600
Wavelength (nm)
Figure 3. Fluorescence spectra of H2L (50.0 lM) in DMF upon the addition of
Al(NO3)3 (0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, and 16 equiv) with an
(b)
excitation of 442 nm. Inset: Fluorescence intensity at 466 nm as a function of the
amount of Al3+
.
0.4
0.3
0.2
0.1
0.0
0.0
0.2
0.4
0.6
0.8
1.0
3+
3+
[Al
]
/
[
Al +H L]
2
Figure 5. (a) UV–vis spectral changes of H2L (50.0 lM) in DMF upon the addition of
Al(NO3)3 (0, 0.2, 0.4, 0.6, 0.8, 1.0, 1.2, 1.4 equiv). Inset: Absorbance at 435 nm was
plotted as a function of the amount of Al3+. (b) Job’s plot for the binding of Al3+ with
H2L. Absorbance at 435 nm was plotted as a function of the molar ratio of [Al3+]/
[Al3++H2L].
Figure 4. 1H NMR spectra of H2L without and with Al(NO3)3 in DMF-d7. (a) H2L
only, (b) H2L and 1.0 equiv of Al3+, (c) H2L and 1.5 equiv of Al3+
.
[Al(HL)(CH3CN)2(H2O)2]2+ (CH3CN was solvent), provided addi-
tional evidence for the formation of a 1:1 complex of Al3+ and
H2L. Furthermore, perfect linear relationships were obtained from
the absorption titration profile, from which the association con-
stant (logKa) of H2L for Al3+ was calculated to be 5.14 and the
detection limit was 0.69 lM based on the 3a/slope (Fig. S7). The
logKa of H2L was within the range of 2.9–14.5 of the reported logKa
for Al3+-binding sensors.7 The detection limit is lower than the US
EPA and FDA guideline of 7.41
l
M Al3+ for bottled drinking
water.10
Having studied the interesting photophysical properties of H2L
such as high sensitivity and selectivity toward Al3+ ions, we further
extended our study to evaluating their potential applications in
imaging Al3+ in cancer cell lines. The confocal fluorescence micros-
copy measurement was carried out. The human cervical HeLa can-
cer cells were grown in DMEM (Dulbecco’s modified Eagle’s
medium) supplemented with 10% FBS (fetal bovine serum) at
37 °C and saturated humidity in an incubator in the atmosphere
of 5% CO2 and 99% air. Cells (5 Â 108/L) were plated on 14 mm glass
cover slips and allowed to adhere for 24 h. After being supple-
Figure 6. Confocal fluorescence images of Al3+ in HeLa cells. Bright-field (a),
fluorescent (b) and the overlay (c) images of HeLa cells incubated with Al(NO3)3 for
8 h. Bright-field (d), fluorescent (e) and the overlay (f) images of HeLa cells
incubated with Al(NO3)3 for 8 h and then exposed to 20 lM H2L for 30 min.
kex = 405 nm.
mented with 100 lM of Al(NO3)3 in the growth media for 8 h at
37 °C, there was no intracellular fluorescence (Fig. 6a–c). After
washing with PBS three times to remove any excess Al(NO3)3,

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rescence from the intracellular area was observed (Fig. 6d–f). These
results demonstrated that H2L was membrane permeable and
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.
In summary, we have successfully developed a new Schiff-base
fluorescent turn-on chemosensor H2L which shows high selectivity
and sensitivity to Al3+ ion in DMF, and displays the 1:1 binding
stoichiometry with Al3+. In our assay, Al3+ could participate in com-
plex formation with the receptor, which resulted in fluorescence
enhancement by CHEF effect. Moreover, the result provides a use-
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Acknowledgments
This work is supported by the National Natural Science Founda-
tion of China (20971015) and the Fundamental Research Funds for
the Central Universities.
Supplementary data
Crystallographic data (excluding structure factors) for the
structures in this paper have been deposited with the Cambridge
Crystallographic Data Centre as supplementary publication no.
CCDC 923333. Copies of the data can be obtained free of charge
on application to CCDC, 12 Union Road, Cambridge CB2 1EZ, UK,
(fax: +44-(0)1223-336033 or e-mail: deposit@ccdc.cam.ac. uk).
Supplementary data (including synthesis and characterization
by elemental analysis, IR and single-crystal X-ray diffraction))
associated with this article can be found, in the online version, at
http://dx.doi.org/10.1016/j.tetlet.2013.04.115.
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