A 1,8-naphthalimide-based chemosensor with an off-on fluorescence and lifetime imaging response for intracellular Cr3+ and further for S2-

Article abstract of DOI:10.1016/j.dyepig.2015.12.001

A novel 1,8-naphthalimide-based chemosensor was designed and synthesized for rapid recognition of Cr³⁺. The desired sensor showed off-on fluorescent and lifetime-based response upon Cr³⁺ and S^2- in solution, on test paper and in cells. With the intensity-based method, the limit of quantification (LOQ) value was 0-5.5 × 10^-5 M and the detection limit could be as low as 0.60 ppm. The 'in situ' prepared Cr³⁺ complex can recognize S^2- among a series of common anions with high selectivity and sensitivity, the LOD can be as low as 307 nM. The lifetime of the sensor changes from 4.95 to 4.89 and further to 5.88 ns upon addition of Cr³⁺ and S^2- in turn.

Full text of DOI:10.1016/j.dyepig.2015.12.001

Accepted Manuscript
A 1,8-naphthalimide-based chemosensor with an off-on fluorescence and lifetime
3+
2−
imaging response for intracellular Cr and further for S
Mingming Yu, Weiwei Du, Wan Zhou, Haixia Li, Chunxia Liu, Liuhe Wei, Zhanxian Li,
Hongyan Zhang
PII:
S0143-7208(15)00488-X
10.1016/j.dyepig.2015.12.001
DYPI 5024
DOI:
Reference:
To appear in: Dyes and Pigments
Received Date: 18 August 2015
Revised Date: 28 November 2015
Accepted Date: 1 December 2015
Please cite this article as: Yu M, Du W, Zhou W, Li H, Liu C, Wei L, Li Z, Zhang H, A 1,8-naphthalimide-
3+
based chemosensor with an off-on fluorescence and lifetime imaging response for intracellular Cr and
2−
further for S , Dyes and Pigments (2016), doi: 10.1016/j.dyepig.2015.12.001.
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to
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ACCEPTED MANUSCRIPT
Graphical Abstract
The fluorescence and lifetime change of the chemosensor and the 'in situ' prepared
Cr3+ complex upon addition of Cr3+ and S2− on test paper and in living cells have been
successfully demonstrated.

ACCEPTED MANUSCRIPT
A 1,8-naphthalimide-based chemosensor with an off-on fluorescence and
lifetime imaging response for intracellular Cr3+ and further for S2−
Mingming Yua,*, Weiwei Dua, Wan Zhoua, Haixia Lia, Chunxia Liua, Liuhe Weia,
Zhanxian Lia,*, Hongyan Zhangb,*
* Corresponding author.
a. College of Chemistry and Molecular Engineering, Zhengzhou University, Zhengzhou,
450001, China.
E-mail address: yumm@zzu.edu.cn (M. Yu), lizx@zzu.edu.cn (Z. Li).
Tel: +(86)371-67781205
Fax: +(86)371-67781205
b. Key Laboratory of Photochemical Conversion and Optoelectronic Materials, Technical
Institute of Physics and Chemistry, Chinese Academy of Sciences, Beijing, 100190,
China
E-mail address: zhanghongyan@mail.ipc.ac.cn (H. Zhang).

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Abstract
A novel 1,8-naphthalimide-based chemosensor was designed and synthesized for rapid
recognition of Cr3+. The desired sensor showed off-on fluorescent and lifetime-based
response upon Cr3+ and S2− in solution, on test paper and in cells. With the intensity-based
method, the limit of quantification (LOQ) value was 0–5.5 × 10–5 M and the detection
limit could be as low as 0.60 ppm. The 'in situ' prepared Cr3+ complex can recognize S2−
among a series of common anions with high selectivity and sensitivity, the LOD can be as
low as 307 nM. The lifetime of the sensor changes from 4.95 to 4.89 and further to 5.88
ns upon addition of Cr3+ and S2− in turn.
Keywords: Sensor; Off-on fluorescence; Fluorescence lifetime imaging microscopy;
FLIM; Cr3+ ion; S2− anion

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1. Introduction
In recent years, great achievement has been achieved in the development of fluorescent
chemosensors toward environmentally and biologically important species such as metal
ions and anions due to several advantages over other methods such as high sensitivity,
specificity, and real-time monitoring with fast response [1−5]. Triple-charged metal cation
detection is of significance owing to its crucial influence in a wide range of
environmental and human health areas [6−8]. As a significant essential trace element in
biological systems, trivalent chromium (Cr3+) is an important component of glucose
tolerance factor, enhancing the biological function of insulin to promote the absorption of
glucose; Cr3+ can inhibit cholesterol biosynthesis which affects lipid metabolism process.
Cr3+ deficiency can cause diabetes, atherosclerosis, and growth retardation [9−10],
whereas excessive chromium can cause genotoxic effects [11]. Cr3+ has been proven to
nonspecifically bind to DNA and other cellular components resulting in inhibition of
transcription and possibly DNA replication [12]. In addition, as an environmental
contaminant and due to various industrial and agricultural activities, the build-up of Cr3+
especially in food and water is a matter of concern [13,14]. Besides, sulfide anions are
widely used in many fields, such as the production of sulfur and sulfuric acid, dyes and
cosmetic manufacturing [15]. It also exists in biosystems because of microbial reduction
of sulfate by anaerobic bacteria or sulfide generation from the sulfur-containing amino
acids in meat proteins [16]. Owing to its toxicity, exposure to a high level of sulfide
anions could cause loss of consciousness, irritation of mucous membranes and
suffocation [17]. In water, sulfide anions may be in the form of HS−, which becomes even
more toxic. High concentration of HS− can result in personal distress, permanent brain
damage, unconsciousness, or even asphyxiation [18]. Thus, it is urgent to develop
fluorescent chemosensors toward Cr3+ and S2−.

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It is generally believed that chemosensors with fluorescence enhancement are more
efficient than fluorescent "turn-off" based probes [19−25]. However, the paramagnetic
nature of Cr3+ leads fluorescence quenching of the fluorophore via the enhancement of
spin-orbit coupling [26−32], in the last few years, there are only few successful cases of
fluorescence enhancement type sensors for Cr3+ [33−38], and for S2− [16, 39−45].
Therefore, it is essential to explore new chemosesnors for Cr3+ and S2− in biological and
environmental samples.
In addition to fluorescence-based sensors, lifetime-based sensors have attracted great
attention [46−52]. These sensors provide the information on the analyte through obvious
changes in emission lifetimes upon interaction with the analyte. The rapid development of
fluorescence lifetime imaging microscopy (FLIM) makes the lifetime-based detection of
intracellular analytes possible. In FLIM imaging, luminescence lifetime is measured at
each spatially resolvable element of a microscope image [53], and the lifetime obtained is
independent of probe concentration and excitation laser intensity [54]. In addition, when
long-lived luminescent sensors are employed for FLIM imaging, the signals from the
sensors can be distinguished from the autofluorescence [55−57], which typically has a
fluorescence lifetime ranging from the picosecond to nanosecond level. As one of the
most promising photoresponsive materials, 1,8-naphthalimide-based derivatives have
been well recognized for excellent photostability, high fluorescence quantum yields and
large Stokes' shift. Moreover, fluorescent characteristics were anticipated to be tuned
through judicious structural modifications [58−64]. However, no 1,8-naphthalimide-based
fluorescent chemosensors for Cr3+ have been reported and the reported S2−-fluorescent
sensors are not many. Herein, as illustrated in Scheme 1, we synthesized a highly
sensitive and stable fluorescent 1,8-naphthalimide-based dye compound 1, which can
quantitatively detect the concentration of Cr3+ with dramatically enhanced fluorescence.

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The detection limit on fluorescence response of the sensor can be as low as 0.60 ppm. The
'in situ' prepared Cr3+ complex (1•Cr) showed high selective recognition of S2− with
distinct changes in its fluorescence. The LOD can be as low as 307 nM.
2. Experimental
2.1. Methods and materials
All commercial grade chemicals and solvents were purchased and were used without
further purification. Compound 1 was synthesized according to Scheme 1 [65,66].
(Insert: Scheme 1)
Mass spectra were obtained on high resolution mass spectrometer (IonSpec4.7 Tesla
1
FTMS-MALDI/DHB). H and 13C NMR spectra were recorded on a Bruker 400 NMR
spectrometer. Chemical shifts are reported in parts per million using tetramethylsilane
(TMS) as the internal standard.
2.2. Spectral characterizations
All spectral characterizations were carried out in HPLC-grade solvents at 20 °C within
a 10 mm quartz cell. Fluorescence spectroscopy was determined on a Hitachi F-4500
spectrometer. The fluorescence quantum yield was measured at 20 °C with quinine
bisulfate in 1 M H2SO4 (Фfr = 0.546) selected as the reference [67]. Time-resolved
fluorescence spectra were measured on a LifeSpec picosecond TRF spectrometer
(Edinburgh Instruments Ltd.).
2.3. Fluorescence and lifetime imaging experiments in living Hela cells
Hela cells were seeded in a 12-well plate in Dulbecco’s modified Eagle’s medium
(DMEM) supplemented with 10% fetal bovine serum and 1% penicillin. The cells were
incubated under an atmosphere of 5% CO2 and 95% air at 37 °C for 24 h before the cell

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imaging experiments. The cells were washed three times with PBS buffer before used.
The cells were firstly incubated with sensor 1 (10 ꢀM) for 10 min at 37 °C, then
incubated with Cr3+(40 ꢀM) for another 5 min and then incubated with S2− (80 ꢀM) for
another 2 min.
Fluorescence and lifetime imaging experiments in Living Hela cells were operated with
Nikon A1Rmp-PicoQuant FLIM. 405 nm semiconductor laser was used for fluorescence
imaging experiments, and LDH-P-C-405B laser was used for lifetime imaging
experiments.
2.4. Synthesis of compound 1
A mixture of compound 2 (0.2330 g, 0.7 mmol), K2CO3 (0.4070 g, 2.8 mmol) and
propargyl bromide (0.068 mL, 0.7 mmol) in 15 mL acetonitrile was refluxed for 8 h, then
cooled to room temperature, filtered, and the filter cake was washed with acetonitrile for
three times. The crude product was obtained from the concentration of the filtrate in
vacuum. The final product (0.1946 g) was purified by column chromatography over silica
gel column using ethyl acetate/petroleum ether (10:1) as eluent. The yield was 76.5%.
Characterization of compound 1: HRMS (EI) m/z: calcd for C21H21N3O3 [M + H],
364.1583; found, 364.1664. 1H NMR (400 MHz, CDCl3, TMS): δH 8.59 (d, 1H), 8.52 (d,
1H), 8.43 (d, 1H), 7.70 (m, 1H), 7.22 (d, 1H), 4.46 (t, 2H), 3.99 (t, 2H), 3.49 (s, 2H), 3.36
13
(t, 4H), 2.93 (t, 4H). C NMR (100 MHz, CDCl3): δC 165.39, 164.95, 132.96, 131.47,
130.66, 129.97, 126.08, 125.72, 122.93, 116.34, 115.03, 62.07, 52.87, 51.77, 46.87 and
42.75.
3. Results and Discussion
3.1. Fluorescence studies of chemosensor 1 towards Cr3+
To investigate the changes in fluorescence emission spectrum of 1 upon exposure to

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Cr(NO3)3 (Cr3+), fluorescence titrations were conducted with water solution of Cr3+ in
aqueous solution of 1 (1.0 × 10−5 M, water/ethanol = 6:4, v/v) (Fig. 1). Upon excitation at
385 nm, the fluorescence emission intensity of 1 gradually increased (Фfr changed from
0.018 to 0.335, I/I0 = 40) as the concentration of Cr3+ increased, indicating an efficient
Cr3+-selective fluorescent “off-on” behavior. The increased emission intensity is probably
due to the complex of Cr3+ and 1, in which, the coordination of Cr3+ and 1 inhibits the
photo-induced electron transfer (PET) from electron-donator piperazine moiety to
electron-receptor 1,8-naphthalimide moiety.66 Fig. S1 (in the Supporting Information)
indicates the relationship between the fluorescence peak intensity at 520 nm and the
concentration of Cr3+. A good linear relationship between the fluorescence peak at 520
nm and the concentration of Cr3+ was obtained at concentration range of 0 and 5.5 × 10−5
M (right of Fig. 1), implying that Cr3+ can be quantitatively detected at a wide
concentration range. According to this linear calibration graph, the detection limit of
probe 1 for Cr3+ is found to be about 1.68 × 10−6 M (0.60 ppm) based on signal-to-noise
ratio (S/N) = 3.68,69 This result proved that sensor 1 shows high sensitivity to Cr3+. Job's
plot indicated that sensor 1 chelates Cr3+ with 1:1 stiochiometry (Fig. S2 in the
Supporting Information).
(Insert: Fig. 1)
(Insert: Fig. 2)
(Insert: Fig. 3)
3.2. Counterion effect on the Cr3+-selective properties of sensor 1
Experiments to explore the counterion effect on the Cr3+-selective properties of sensor

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1 were also performed (Fig S3 in the Supporting Information). Sulfate and nitrate
counteranions had similar influence as chloride, demonstrating that the three kinds of
anions do not coordinate the metal ions while compound 1 was interacting with Cr3+. This
indicates that compound 1 as a fluorescent sensor has a much wide application range in
sensing Cr3+ ions.
3.3. The selectivity and anti-disturbance effect study of sensor 1 for Cr3+
To evaluate the selectivity of sensor 1 for Cr3+, various metal ions (K+, Ca2+, Na+, Mg2+,
Al3+, Co2+, Cr3+, Ni2+, Cu2+, Zn2+, Pb2+, Hg2+, Mn2+, Cd2+) were tested. As shown in Fig. 2
and black bars of Fig. 3, only the introduction of Cr3+to the sensor 1 solution induced a
significant enhancement in the fluorescent intensity at 520 nm. In the same condition,
other tested metal ions mentioned above did not induce any obvious fluorescence
enhancement to the sensor 1 solution.
To further assess its utility as a Cr3+-selective fluorescent sensor, its fluorescence
spectrum response to Cr3+ in the presence of other metal ions mentioned above (red bars
of Fig. 3) was also tested. The results demonstrated that all of the selected species have
no interference in the detection of Cr3+. This result strongly indicates that compound 1
could be an excellent fluorescent sensor towards Cr3+ with strong anti- interference
ability.
3.4. pH range of application of 1 toward Cr3+
For both environmental and biological applications of the fluorescent sensor, it would
be much better if the sensing works over a wide range of pH. The right panel of Fig. S4 in
the Supporting Information shows that in aqueous solution the suitable pH range for Cr3+
determination is 4-6, where the fluorescence off-on behavior can be operated by Cr3+
binding. The short pH application range may be ascribed to the strong protonation ability

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of sensor 1.
3.5. Reversibility study of the binding of 1 to Cr3+
To examine the reversibility of the binding of chemosensor 1 to Cr3+, a water solution
containing 4 equiv S2− was added to the 1•Cr3+ solution. When S2- was added to the
1•Cr3+ solution, fluorescence signals identical to those of 1 were restored (Fig S5 in the
Supporting Information and Fig. 4), demonstrating that Cr3+ was removed from the
1•Cr3+ complex by S2−. That is, the binding of 1 and Cr3+ is really chemically reversible.
(Insert: Fig. 4)
(Insert: Fig. 5)
3.6. Sensing mechanism of 1 to Cr3+
Reference compound 2 was applied to determine the coordination mode of 1 with Cr3+.
Upon addition of different metal ions to the solution of compound 2 respectively, no
fluorescence spectra change was observed (Fig S6 in the Supporting Information),
indicating that compound 2 cannot be a fluorescent sensor toward these metals. In
1
addition, the sensing mechanism was confirmed by the H NMR titration experiments of
compound 1 in 0.5 mL CD3OD with different amounts of the Cr3+ anion (0, 0.5, and 1
equiv). As shown in Fig S7 in the Supporting Information, upon addition of Cr3+, the four
peaks at 3.49, 3.33, 2.95 and 2.80 shifted to low field (3.69, 3.40, 3.16, and 2.98),
confirming that the two nitrogen atoms of the piperazine unit and the alkynyl group
coordinate the Cr3+ anion.
3.7. Fluorescence sensing properties of the 'in situ' prepared Cr3+ complex (1•Cr3+)
toward S2−
As we have mentioned above, only the addition of S2− can result in the fluorescence of

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the 'in situ' prepared 1•Cr3+ signals restoring. That is, the complex 1•Cr3+ can detect S2−
with fluorescence method.
To investigate the changes in fluorescence spectrum of the complex 1•Cr3+ upon
exposure to S2−, fluorescence titration experiments were conducted. As shown in left of
Fig. 4, upon addition of S2− water solution, fluorescence at 520 nm gradually decreased,
implying that Cr3+ is removed from the complex 1•Cr3+ by S2−, and the complex 1•Cr3+
can detect S2−. In concentration range of 0 and 530 ꢀM, the fluorescence intensity at 520
nm is in good linear relationship with S2− concentration (right of Fig. 4), implying that
S2− can be quantitatively detected in a wide concentration range. The LOD value from the
fluorescence titration experiment can be as low as 307 nM.
To examine the selectivity and anti-disturbance capacity toward S2−, further
experiments were carried out. Upon addition of different anions, only S2− can induce
fluorescence at 520 nm of the complex 1•Cr3+ decreasing (black bars of Fig 5), other
representative anions, such as F−, Cl−, Br−, I−, CO32−, CH3COO−, NO3 , PO43−, NO2 ,
−
−
2−
−
−
−
2−
−
2−
−
2−
SO3 , HSO3 , ClO3 , N3 , SiO3 , HCO3 , HPO4 , H2PO4 , SCN−, and SO4 showed
almost no effects on the fluorescence of the complex 1•Cr3+, implying high selectivity of
the complex 1•Cr3+ toward S2−. The competition experiments were conducted in the
presence of Cr3+ and S2− mixed with many anions (F−, Cl−, Br−, I−, CO32−, CH3COO−,
−
−
2−
−
−
−
2−
−
2−
−
NO3 , PO43−, NO2 , SO3 , HSO3 , ClO3 , N3 , SiO3 , HCO3 , HPO4 , H2PO4 , SCN−,
and SO42−) respectively (red bars). No significant fluorescence spectra change were found
by comparison with that without the other anions, which revealed strong anti-disturbance
capacity of the complex 1•Cr3+ toward S2−.
3.8. Application on Test Paper
As shown in the above, chemosensor 1 can detect Cr3+ in aqueous solution, it inspired

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us to further investigate the feasibility of solid materials for point-of-care detection
application. Test paper was selected and the fluorescence detection properties were
studied by a UV lamp with photography as well as solid fluorescence spectroscope. In the
detection process, the sensor spots were prepared by dropping 5 ꢀL 10 ꢀMsolutions of 1
Cr(NO3)3Cr3+portable ultraviolet lamp. As shown in Fig. 6, the sensor spots emitted
bright blue-green fluorescence upon the addition of Cr3+ under UV illumination,
meanwhile an enhancement of the fluorescence intensity at 495 nm could be recorded.
Additionally, the sensor spots emitted fluorescence only upon the addition of Cr3+ (Fig. 7),
demonstrating the high selectivity of sensor 1 as solid materials toward Cr3+.
(Insert: Fig. 6)
(Insert: Fig. 7)
3.9. Application of Sensor 1 in Cellular Imaging
The practical utility of using sensor 1 to detect Cr3+ and further S2− in an imagewise
manner within living Hela cells was explored (Fig. 8). When Hela cells were incubated
with 10 ꢀM probe 1 only for 10 min at 37 °C, no obvious fluorescence was observed (f of
Fig. 8). However, when Hela cells were preincubated with Cr3+ (40 ꢀM) and then
incubated with probe 1 (10 ꢀM) for 5 min, a significant green fluorescence inside the
cells was observed with the aid of an inverted fluorescence microscope (g of Fig.8),
implying that the stimulation of Cr3+ toward probe 1 only for 5 min can give green
emission. Furthermore, when Hela cells were preincubated with Cr3+ (40 ꢀM), probe 1
(10 ꢀM), and then incubated with 80 ꢀM S2− for 2 min, no obvious fluorescence was also
observed (h of Fig. 8), implying that addition of S2− lead to the fluorescence quenching of
the complex 1•Cr3+. Bright-field measurements indicated that the cells before and after
addition of Cr3+and S2− remained viable throughout the imaging experiments (a, b, c, and

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d of Fig. 8). Thus, probe 1 is capable of permeating into cells and sensing Cr3+ and S2− in
living cells.
Lifetime-based detection of Cr3+ and S2− was conducted in FLIM imaging. As shown in
Fig. 9, cells treated with 10 ꢀM sensor 1 for 2 h at 37 °C displayed a relatively short
lifetime of 4.89 ns. When 40 ꢀM Cr3+ was added in the growth media, the lifetime of
microregions hardly changed (4.95 ns). A lifetime of 5.88 ns was obtained for the above
system upon addition of 80 ꢀM S2−. FLIM imaging result demonstrates that S2− cannot
release Cr3+ from the complex, and the possible case is that S2− take the place of chloride
ion to coordinate Cr3+. This result implied that he 'in situ' prepared 1•Cr3+ complex was
able to serve as a Lifetime-based sensor for S2−.
(Insert: Fig. 8)
(Insert: Fig. 9)
4. Conclusion
In summary, a Cr3+ selective fluorescent "off-on" and lifetime-based chemosensor
1,8-naphthalimide derivative 1 was synthesized, which displays a high selectivity,
antidisturbance for Cr3+ among environmentally and biologically relevant metal ions, and
high sensitivity. The sensing mechanism may be ascribed to the inhibited PET process
from the coordination of Cr3+ and 1. The 'in situ' prepared Cr3+ complex (1•Cr) showed
high selectivity and sensitivity (the LOD can be as low as 307 nM) toward S2−, which can
be used to sense S2−. The lifetime of the sensor changes from 4.95 to 4.89 ns upon
addition of Cr3+, then the lifetime further increased to 5.88 ns upon addition of S2− to the
system above. In addition, the fluorescence and lifetime change of this chemosensor and
the 'in situ' prepared Cr3+ complex upon addition of Cr3+ and S2− on test paper and in
living cells have been successfully demonstrated.

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Acknowledgements
We are grateful for the financial supports from National Natural Science Foundation of
China (50903075 and J1210060), Key Laboratory of Photochemical Conversion and
Optoelectronic Materials and Zhengzhou University.
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Figure Captions
Scheme 1 Synthetic route to sensor 1.
Fig. 1. Emission spectra of 1 (1.0 × 10−5 M, Vwater:Vethanol = 6:4) upon titration of Cr3+
water solution (0–33 equiv to 1) with excitation at 385 nm. The linearity of peak intensity
with respect to Cr3+ concentrations (right).
Fig. 2. Fluorescence spectra (up) and photographs (down) of 1 (1.0 × 10−5 M,
Vwater:Vethanol = 6:4) upon addition of 2 equiv. various metal ions with excitation at 385 nm
(up) and under a 365 nm UV lamp (down). The metal ions used from left to right were
only 1, K+, Ca2+, Na+, Mg2+, Al3+, Zn2+, Co2+, Ni2+, Pb2+, Cu2+, Cr3+, Hg2+, Mn2+ and
Cd2+.
Fig. 3. Fluorescence responses of 1 (1.0 × 10−5 M, Vwater:Vethanol = 6:4) upon addition
of 10 equiv different metal ions (black bars), and fluorescence changes of the mixture of 1
and 3 equiv. Cr3+ after addition of 10 equiv metal ions (red bars). The excitation
wavelength was 385 nm. I0 represents the emission intensity at 520 nm in the
fluorescence spectroscopy of compound 1. I represents the emission intensity at 520 nm
in the fluorescence spectroscopy of compound 1 after addition of the species to the
solution of 1 (black bars) and of the mixture of 1 and Cr3+ after addition of an excess of
the species (red bars). The metal ions used were only 1, Cr3+, K+, Ca2+, Na+, Mg2+, Al3+,
Zn2+, Co2+, Ni2+, Pb2+, Cu2+, Hg2+, Mn2+, and Cd2+.
Fig. 4. Emission spectra of the 'in situ' prepared complex 1•Cr3+ (1.0 × 10–5 M,
water/ethanol = 6:4, v/v) in the presence of S2− water solution (left). Emission spectra
response of the 'in situ' prepared complex 1•Cr3+ at 520 nm as a linear dependence with
the concentration of S2− (right).
Fig. 5. Fluorescence emission spectra response of the 'in situ' prepared complex

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1•Cr3+ (1.0 × 10–5 M, water/ethanol = 6:4, v/v) at 520 nm in the presence of different
anions (4.0 × 10–5 M) (black bars) and fluorescence emission spectra response of the 'in
situ' prepared complex 1•Cr3+ (1.0 × 10–5 M, water/ethanol = 6:4, v/v) and S2− (4.0 × 10–5
M) after addition of other different anions (4.0 × 10–5 M) (red bars). The anions from 1
−
3−
−
2−
to 20 are only 1, S2−, F−, Cl−, Br−, I−, CO32−, CH3COO−, NO3 , PO4 , NO2 , SO3 ,
−
−
−
2−
−
2−
−
2−
HSO3 , ClO3 , N3 , SiO3 , HCO3 , HPO4 , H2PO4 , SCN−, and SO4
.
Fig. 6. (a) Fluorescence spectra and (b) fluorescence intensity at 495 nm of the sensor
spots of 1 (1.0 × 10−5 M, water/ethanol = 6:4, v/v) on test papers upon addition of various
concentrations of Cr3+ (0–10 mM).
Fig. 7. Images of test papers for the selectivity of Cr3+ upon addition of 10 mM
various metal ions (From left to right, the metal ions used were only 1, K+, Ca2+, Na+,
Mg2+, Al3+, Zn2+, Co2+, Ni2+, Pb2+, Cu2+, Cr3+, Hg2+, Mn2+ and Cd2+) under a UV lamp
(365 nm).
Fig. 8. Confocal fluorescence contrast images of living hela incubated with 10 ꢀM
sensor 1 at 37 °C for 10 min (a and d), incubated with 10 ꢀM sensor 1 and 40 ꢀM Cr3+ at
37 °C for 5 min (b and e), and incubated with 10 ꢀM sensor 1, 40 ꢀM Cr3+ and 80 ꢀM S2−
at 37 °C for 2 min (c and f).
Fig. 9. FLIM images in live hela cells constained by 10 ꢀM sensor 1 (a), cells
pretreated with 10 ꢀM sensor 1 followed by incubation with 40 ꢀM Cr3+ (b), and
incubated with 10 ꢀM sensor 1, 40 ꢀM Cr3+ and 80 ꢀM S2− (c).

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Scheme 1 Synthetic route to sensor 1.

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Fig. 1. Emission spectra of 1 (1.0 × 10−5 M, Vwater:Vethanol = 6:4) upon titration of Cr3+
water solution (0–33 equiv to 1) with excitation at 385 nm. The linearity of peak intensity
with respect to Cr3+ concentrations (right).

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ACCEPTED MANUSCRIPT
Fig. 2. Fluorescence spectra (up) and photographs (down) of 1 (1.0 × 10−5 M,
Vwater:Vethanol = 6:4) upon addition of 2 equiv. various metal ions with excitation at 385 nm
(up) and under a 365 nm UV lamp (down). The metal ions used from left to right were
only 1, K+, Ca2+, Na+, Mg2+, Al3+, Zn2+, Co2+, Ni2+, Pb2+, Cu2+, Cr3+, Hg2+, Mn2+ and
Cd2+.

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ACCEPTED MANUSCRIPT
Fig. 3. Fluorescence responses of 1 (1.0 × 10−5 M, Vwater:Vethanol = 6:4) upon addition of
10 equiv different metal ions (black bars), and fluorescence changes of the mixture of 1
and 3 equiv. Cr3+ after addition of 10 equiv metal ions (red bars). The excitation
wavelength was 385 nm. I0 represents the emission intensity at 520 nm in the
fluorescence spectroscopy of compound 1. I represents the emission intensity at 520 nm
in the fluorescence spectroscopy of compound 1 after addition of the species to the
solution of 1 (black bars) and of the mixture of 1 and Cr3+ after addition of an excess of
the species (red bars). The metal ions used were only 1, Cr3+, K+, Ca2+, Na+, Mg2+, Al3+,
Zn2+, Co2+, Ni2+, Pb2+, Cu2+, Hg2+, Mn2+, and Cd2+.

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ACCEPTED MANUSCRIPT
Fig. 4. Emission spectra of the 'in situ' prepared complex 1•Cr3+ (1.0 × 10–5 M,
water/ethanol = 6:4, v/v) in the presence of S2− water solution (left). Emission spectra
response of the 'in situ' prepared complex 1•Cr3+ at 520 nm as a linear dependence with
the concentration of S2− (right).