A near-infrared and colorimetric fluorescent probe for palladium detection and bioimaging

Article abstract of DOI:10.1016/j.dyepig.2016.10.052

A near-infrared (NIR) and colorimetric fluorescent probe was developed for palladium species via a palladium catalyzed deallylation reaction. In this probe, 6-hydroxy-2,3-dihydro-xanthene-indolium (CyH) was acted as the signal unit and an allyl carbonate group was acted as the recognition unit. This non-fluorescent probe molecule can release the relevant fluorophore after interaction with palladium. The sensing mechanism was investigated by optical spectrum and NMR spectra. The probe can be used for “naked-eye” detection of palladium, and exhibited high selectivity to palladium over various other metal ions. Furthermore, the probe can be applied to imaging intracellular palladium ions in living HeLa cells, indicating its great potential for in vivo bioanalytical applications.

Full text of DOI:10.1016/j.dyepig.2016.10.052

Dyes and Pigments 137 (2017) 293e298
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Short communication
A near-infrared and colorimetric fluorescent probe for palladium
detection and bioimaging
Wei Su, Biao Gu, Xiaojun Hu, Xiaoli Duan, Youyu Zhang, Haitao Li*, Shouzhuo Yao
Key Laboratory of Chemical Biology and Traditional Chinese Medicine Research (Ministry of Education), College of Chemistry and Chemical Engineering,
Hunan Normal University, Changsha 410081, China
a r t i c l e i n f o
a b s t r a c t
Article history:
A near-infrared (NIR) and colorimetric fluorescent probe was developed for palladium species via a
palladium catalyzed deallylation reaction. In this probe, 6-hydroxy-2,3-dihydro-xanthene-indolium
(CyH) was acted as the signal unit and an allyl carbonate group was acted as the recognition unit. This
non-fluorescent probe molecule can release the relevant fluorophore after interaction with palladium.
The sensing mechanism was investigated by optical spectrum and NMR spectra. The probe can be used
for “naked-eye” detection of palladium, and exhibited high selectivity to palladium over various other
metal ions. Furthermore, the probe can be applied to imaging intracellular palladium ions in living HeLa
cells, indicating its great potential for in vivo bioanalytical applications.
Received 21 September 2016
Received in revised form
31 October 2016
Accepted 31 October 2016
Available online 3 November 2016
Keywords:
Near-infrared
Fluorescent probe
Palladium
© 2016 Elsevier Ltd. All rights reserved.
Bioanalytical applications
1. Introduction
In the past few years, several fluorescent probes have been re-
ported for the detection of palladium. For example, M. Kumar et al.
Palladium (Pd) as a precious metal was widely used in various
materials and pharmacy such as catalytic converters, fuel cells,
jewelry, dental crowns and so on [1e3], which led to high level of
residual palladium in the final product [4e6], thus resulting in
fearful environmental and health problems because it can bind to
biomolecules like proteins, DNA, thiol-containing amino acids and
disturb cellular processes [7e9]. Therefore, development of effi-
ciency methods for detective and imaging palladium species is
important for environment safe and human health.
Up to now, there are many traditional methods reported for the
detection of palladium species [10e13], such as inductively coupled
plasma atomic emission spectrometry (ICP-AES), solid-phase
microextraction high performance liquid chromatography (SPME-
HPLC), X-ray, atomic absorption spectroscopy (AAS), etc [14e17].
However, they suffer from complicated sample preparation pro-
cedures, complicated instrumentation and the requirement for
highly-trained individuals [18e20]. Compared with above tech-
nologies, fluoremetry is more attractive due to excellent sensitivity,
high selectivity, low detection limit and operational simplicity
[21e24].
design a fluorescent probe detection of palladium based on
hydroxyphenyl-benzothiazole [25]. And Liu et al. prepared a “turn-
on” fluorescent probe for palladium using rhodamine as the signal
group [26]. However, these probes need the ultraviolet or visible
light to excite, which severely limits in biological applications
because the fluorescence imaging in the visible region would be
easily disturbed by cell auto-fluorescence in living systems
[27e32]. Therefore, it is consequential to develop the near infrared
fluorescent probes for the detection of palladium [33]. Recently,
Wang's group has design a near-infrared fluorescent probe for
palladium, which exhibits high sensitivity and selectivity toward
both Pd(0) and Pd(II). But this probe has a main drawback of poor
water solubility, which is a disadvantage for bioimaging application
[34]. Zhang’ group reported a fluorescent probe for palladium.
Although this probe emit in the red light region, it cannot be used
for monitoring palladium in living cells due to the need high pro-
portion of organic solvents [35]. Until now, the fluorescent probe
based on near-infrared dyes for imaging intracellular palladium is
very rare. In addition, colorimetric method has also draw much
attention in terms of practical application due to its low-cost and
convenient operations [36]. As a consequence, it is still urgently
demanded to develop a novel NIR and colorimetric fluorescent
probe for sensing palladium species in biological sample.
* Corresponding author.
E-mail address: hatiao-li@hunnu.edu.cn (H. Li).
Based on above consideration, we choose heptamethine cyanine
http://dx.doi.org/10.1016/j.dyepig.2016.10.052
0143-7208/© 2016 Elsevier Ltd. All rights reserved.

294
W. Su et al. / Dyes and Pigments 137 (2017) 293e298
derivative (CyH) as the chromophore due to its NIR-emission,
excellent water solubility, easily synthetic and predictable colour
change in the solution after modification [37,38]. Herein, we
designed and synthesized a new NIR and colorimetric fluorescent
2.2.1. Synthesis of compound 1
Compound 1 was synthesized on the basis of the procedures
reported in the literature [39]. A mixture of dimethylformamide
(40 mL) and methylene chloride (40 mL) was chilled in an ice bath
for 30 min. Then phosphorus oxychloride (37 mL, 0.41 mol) and
cyclohexanone (10.0 g, 0.10 mol) was added dropwise to the above
mixture solution with stirring. The mixture solution was refluxed
for 3 h, cooled, poured onto 300 g of ice, and stand to overnight. The
yellow solid was collected with a yield of 7.8 g (44.5%).
probe,
6-((allylcarbonyl)oxy)-2,3-dihydro-xanthenes-indolium
(CyPd) detection of palladium. The probe is composed of CyH as the
fluorophore and the allyl carbonate group as the recognition unit
(in Scheme 1). In absence of any analyte, the free CyPd has almost
no fluorescence due to the intermolecular charge transfer (ICT)
possess from the fluorophore to the allyl carbonate group. Upon
addition of palladium, the depropargylation reaction occurs and
thus the protected hydroxyl group on the CyPd is liberated, which
lead to the significant enhancement of fluorescence. Indeed, the
synthesized probe showed many advantages as follow: high se-
lective and sensitive to palladium; emission in the NIR light region
2.2.2. Synthesis and characterization of Cy-7
2,3,3-trimethyl-3H-indole (3.66 g, 11.66 mmol), compound 1
(0.96 g, 5.44 mmol) and sodium acetate (0.47 g, 5.44 mmol) were
dissolved in 30 mL acetic anhydride under nitrogen atmosphere.
The mixture solvent was stirred for 2 h at room temperature. Then
the mixture solvents were removed under vacuum. The residual
were washed with ether to obtain 3.2 g of pure green solid (91%). 1H
(
lem ¼ 721 nm); excellent water solubility; simple synthesis;
“naked-eye” detection for palladium; the capability of monitoring
palladium in living cells. All of these performances make it
appropriate for potential application in biology.
NMR (500 MHz, CDCl3):
d
8.31 (d, J ¼ 14.1 Hz, 2H), 7.40e7.33 (m,
4H), 7.24e7.14 (m, 4H), 6.18e6.15 (s, 2H), 3.72 (s, 6H), 2.70 (t,
J ¼ 6.1 Hz, 4H), 1.97e1.88 (m, 2H), 1.69 (s, 12H); 13C NMR (126 MHz,
2. Experimental sections
CDCl3):
d 172.90, 150.67, 144.38, 142.75, 140.89, 128.84, 127.65,
125.36, 122.15, 110.88, 101.60, 77.40, 77.15, 76.90, 49.25, 32.69,
29.66, 28.08, 26.73, 20.68.
2.1. Reagents and apparatus
2,3,3-trimethyl-3H-indole, cyclohexanone, phosphoryl chloride,
sodium acetate, resorcinol, potassium carbonate, triethylamine,
allyl carbonochloridate, Pd(PPh3)4 (Pd(0)), PdCl2 (Pd(II)) and
(NH4)2PdCl6 (Pd(IV)) were purchased from Energy Chemical.
Acetonitrile (ACN), Chloroform, Dimethyl formamide (DMF),
Dichloromethane (DCM), Dimethyl sulfoxide (DMSO), and Acetic
anhydride were obtained from Sinopharm Chemical Reagent
Company. All chemicals used in this work were of analytical grade
and without further purification. Double distilled water was used in
this work.
Electrospray mass spectrometry (ESI-MS) spectra were acquired
on a ZQ2000 mass spectrometer (Manchester, UK). 1H and 13C
nuclear magnetic resonance (NMR) spectra were recorded on a
Bruker AVB-500 spectrometer using TMS as an internal standard.
UVevis spectra were recorded on a UV-2450 spectrophotometer
(Shimadzu). Time dependent fluorescence spectra were recorded at
37 ꢀC on a QM40 fluorescence spectrophotometer (PTI, Canada),
and other fluorescence spectra were recorded at room temperature
using an F-7000 fluorescence spectrophotometer (Hitachi Co.,
Japan) with the excitation and emission slit widths at 5 nm.
2.2.3. Synthesis and characterization of CyH
A stirred solution of resorcinol (220 mg, 2.0 mmol) and K2CO3
(276 mg, 2.0 mmol) in 15 mL ACN at room temperature under ni-
trogen atmosphere, stirred for 20 min, a solution of ACN (10 mL)
contain compound 2 (610 mg, 1.0 mmol) was added to the above
mixture solution via a syringe. The mixture solution was heated for
4 h at 50 ꢀC. The solvent was evaporated under reduced pressure,
the crude product was purified by silica gel column chromatog-
raphy (CH2Cl2/CH3OH ¼ 50:1), contain the desired CyH as a blue-
green solid (373 mg, yield 73%). 1H NMR (500 MHz, DMSO-d6):
d
8.15 (d, J ¼ 13.9,1H), 7.56 (s,1H), 7.53 (d, J ¼ 7.3, 1H), 7.37 (t, J ¼ 9.3,
2H), 7.28 (d, J ¼ 7.9, 1H), 7.17 (t, J ¼ 7.4, 1H), 6.59 (d, J ¼ 8.9, 1H), 6.44
(s, 1H), 6.01 (d, J ¼ 13.8, 1H), 3.55 (s, 3H), 2.64 (d, J ¼ 17.9, 2H), 2.63
(t, J ¼ 5.6 Hz, 2H), 1.79 (s, 2H), 1.66 (s, 6H). 13C NMR (126 MHz,
DMSO-d6):
d 169.76, 159.61, 143.83, 140.60, 138.46, 135.96, 130.38,
128.76, 123.96, 123.22, 122.60, 119.65, 115.88, 115.59, 110.42, 102.75,
98.32, 48.34, 40.50, 40.34e40.09, 40.00, 39.84, 39.67, 39.50, 31.25,
28.34, 28.01, 24.37, 21.10. MS (EI) m/z: 384.23 (Mþ).
2.2.4. Synthesis and characterization of CyPd
2.2. Synthesis of compounds
To a stirred solution of CyH (102.2 mg, 0.2 mmol, 1.0 equiv) in
CH2Cl2 (10 mL) was added triethylamine (Et3N, 56
equiv) and allyl chlorocarbonate (42.42 L, 0.4 mmol, 2.0 equiv) at
ꢀC under nitrogen atmosphere. Stirred for 30 min, the mixture
mL, 0.4 mmol, 2.0
The synthetic route of CyPd is shown in Scheme 2. The resulting
compounds were characterized by conventional ESI-MS, 1H NMR,
and 13C NMR spectroscopy (see Fig. S1eS8).
m
0
was heated to room temperature and stirred overnight. the reaction
mixture was concentrated under reduced pressure to give crude
solid, then purified by silica gel column chromatography (CH2Cl2/
CH3OH ¼ 50:1) to afford desired probe CyPd as a blue solid (51 mg,
yield 43%). 1H NMR (500 MHz, DMSO-d6):
d
8.56 (d, J ¼ 15.3 Hz, 1H),
Ha
O
Ha
C
7.78 (d, J ¼ 7.4 Hz, 1H), 7.74 (d, J ¼ 7.9 Hz, 1H), 7.58 (t, J ¼ 10.1 Hz,
3H), 7.51 (t, J ¼ 7.3 Hz, 1H), 7.38 (s, 1H), 7.22 (dd, J ¼ 8.4, 2.2 Hz, 1H),
6.66 (d, J ¼ 15.3 Hz, 1H), 6.03 (ddd, J ¼ 22.8, 10.7, 5.7 Hz, 1H), 5.44
(dd, J ¼ 17.2, 1.5 Hz, 1H), 5.34 (dd, J ¼ 10.5, 1.2 Hz, 1H), 4.78 (d,
J ¼ 5.6 Hz, 2H), 3.94 (s, 3H), 2.75e2.70 (m, 2H), 2.68 (t, J ¼ 5.9 Hz,
2H), 1.87e1.80 (m, 2H), 1.75 (s, 6H). 13C NMR (126 MHz, DMSO-d6):
Hc
Hc
C
OH
O
O
C
Hb
O
Pd(0),Pd(II),Pd(IV)
PBS, pH 7.45
O
N
N
I-
I-
d
179.40, 158.85, 152.93, 152.76, 152.65, 145.51, 142.85, 142.64,
CyPd
None emission
CyH
Red emission
132.07, 130.56, 130.33, 129.33, 128.76, 128.23, 123.13, 120.20, 119.65,
119.04, 114.61, 114.26, 109.97, 107.37, 69.60, 51.25, 33.62, 29.15,
27.42, 24.01, 20.27. MS (EI) m/z: 468.20 (Mþ).
Scheme 1. Recognition mechanism of CyPd toward palladium.

W. Su et al. / Dyes and Pigments 137 (2017) 293e298
295
O
OH Cl
H
POCl3,DMF
H
O
1
OH Cl
H
sodium acetate
Cl
H
O
acetic anhydride
N
N
-
N
I-
I
1
Cy-7
resorcinol
K2CO3 ,ACN
O
OH
O
O
allyl carbonochloridate
Et3N, CH2Cl
O
O
N
N
I-
I-
CyH
CyPd
Scheme 2. Synthetic route of CyPd.
2.3. Analytical procedure
Pd(0) (0.5 and 1.0
of CyPd displayed obvious changes after adding Pd(0) (0.5 and
1.0 M) (Fig. 1B). And the colour change from blue to cyan after
addition of Pd(0) (0.0, 0.05, 0.10 M) (see Fig. 1A inset).
Then, we investigate the kinetic profiles of the reaction, in order
to clarify the process of deallylation reaction. The responding of
CyPd to palladium was prove when added palladium species with
different valence states to the CyPd, as show in Fig. 2, The result
revealed that CyPd responds to palladium species in the following
rules: Pd(0) > Pd(II) > Pd(IV). Thus, Pd(0) was selected as the
representative palladium species in the following experiments.
mM) (Fig. 1A). Meanwhile, the fluorescence signal
Stock solutions of CyPd (1.0 mM) and CyH (1.0 mM) were pre-
pared in DMSO. Stock solutions of palladium species (1.0 mM) were
prepared from Pd(0) in DMSO, and Pd(II), Pd(IV) prepared in water.
Stock solutions of metal ions (20.0 mM) were prepared from
AgNO3, FeCl3$6H2O, FeCl2$4H2O, BaCl2$2H2O, AlCl3, Pb(NO3)2,
CaCl2$2H2O, MgCl2$6H2O, ZnCl2, NiCl2$6H2O, MnCl2$4H2O,
CdCl2$2.5H2O, NaCl, CoCl2$6H2O, CuCl2$2H2O, HgCl2, LiCl and
SrCl2$6H2O in water. Test solutions were prepared by dissolving
m
m
20 mL of CyPd stock solutions and a suitable amount of the analyte
stock solution into a phosphate buffer solution (PBS, 10 mM, pH
7.4). The mixture (final volume is 2.0 mL containing 5.0% v/v DMSO)
was incubated at 37 ꢀC for 30 min, Then, the fluorescence emission
spectra of the result solutions were recorded at an excitation
wavelength of 670 nm (unless otherwise noted, all spectral were
measured according to this method).
3.2. Mechanism study
New fluorescent probe CyPd contains a CyH as the signal unit
and an allyl carbonate group as the sensing unit. In CyPd, CyH was
weak absorption and fluorescence emission under the protection of
the eOH group with an allyl carbonate group. However, the allyl
carbonate group was left after incubated by Pd(0), the eOH group
in the probe will be released, and results in strong absorption and
fluorescent emission. The absorption and emission of CyPd in the
absence and presence of Pd(0) was investigated. As shown in
Fig. S9, the CyPd exhibited weak absorption and emission in the
absence of Pd(0). The absorbance and the emission were increased
notably after CyPd was treated with Pd(0), and the absorption and
emission spectrum was the same as CyH. Thus, these results indi-
cated that CyPd undergo the deallylation reaction followed by hy-
drolysis to generate CyH. The strong absorption and emission of
CyH at longer wavelength possible resulted from intermolecular
charge transfer (ICT). It is well known that the emission spectra of
molecules possessing the intermolecular charge transfer (ICT)
process show obvious red-shift with the increase of the polarity of
solvent. In order to better understand the mechanism in our work,
we examined the effect of polarity of solvents on the fluorophore
CyH. As shown in Fig. S10, the maximal emission of CyH displayed
red-shift from 685 nm to 737 nm (from DCM to MeOH), which
suggest the character of the ICT process in the CyH structure.
However, after the introduction allyl carbonate group to the beckon
of CyH, the ICT efficiency in the resulting probe molecule was
2.4. Cell cultures and fluorescence imaging
The HeLa cells were cultivated on a 96-well plate in the cultivate
medium and allowed to adhere for 24 h at 37 ꢀC. The cells were
washed with phosphate buffered saline (PBS) and incubated with
5 m
M CyPd for 30 min at 37 ꢀC, then washed with PBS for three
times, and imaged. After incubating with 5 mM Pd for another 1 h at
37 ꢀC, the HeLa cells were washed three times with PBS and imaged
again. Fluorescence imaging of intracellular Pd in HeLa cells was
recorded on an inverted fluorescence microscope (Nikon, Eclipse
TieS) with a 40 ꢁ objective lens. The excitation wavelength of the
laser was 540 nm.
3. Results and discussion
3.1. Spectral characteristics
The optical properties of CyPd were measured in 10 mM PBS
buffer (H2O/DMSO ¼ 19:1, v/v, pH ¼ 7.4) with or without Pd(0). As
shown in Fig. 1, CyPd showed weak absorption and weak fluores-
cence. However, the absorbance increased significantly after adding

296
W. Su et al. / Dyes and Pigments 137 (2017) 293e298
Fig. 1. UVe vis (A) and Fluorescence emission (B) spectra of CyPd (10
m
M), reacting with Pd(0) (0, 0.5, 1.0
m
M) in 10 mM PBS buffer (H2O/DMSO ¼ 19:1, v/v, pH ¼ 7.4, 37 ꢀC,
lex ¼ 670 nm). Inset image: (a) visible-light image of 10
mM probe; (b) visible-light image of 10 mM probe and 0.5 mM Pd(0); (c) visible-light image of 10 mM probe and 1.0 mM Pd(0).
However, the peak of the vinyl group in the product dis-
appeared, which indicated that CyPd undergoes the deallylation
reaction followed by hydrolysis to generate the CyH (Fig. S11).
1.2
1.0
Pd(0)
3.3. Effects of pH and incubation time
Pd(+4)
0.8
In order to obtain a excellent-sensitive response for Pd(0), the
effects of the experimental parameters, like pH and incubation time
were investigated. Fig. S12A illustrates the effect of pH on fluores-
cence properties of CyPd and it response to Pd(0). The pH value had
no obvious effect on the fluorescence intensity of CyPd itself,
however, it was notable that the fluorescence intensity increased in
the presence of Pd(0) in pH range from 5.0 to 7.45, and then reached
a constant value at alkaline condition. These results indicated that
the alkaline conditions in favour of the deallylation reaction.
Therefore, physiological pH (pH ¼ 7.4) could be act as an appro-
priate working pH in the following experiments. Fig. S12B shows
the time dependent fluorescence response of CyPd in the absence
and presence of Pd(0), suggesting that the reaction between CyPd
and Pd(0) was completed within 30 min. Hence, the incubation
time was controlled at 30 min.
0.6
0.4
0.2
Pd(+2)
0
20
40
60
80
100
Time (min)
Fig. 2. Time-dependent fluorescence changes of CyPd (10
palladium species (5.0
37 ꢀC, lex ¼ 670 nm).
m
M) in the presence of three
m
M) in 10 mM PBS buffer (H2O/DMSO ¼ 19:1, v/v, pH ¼ 7.4,
weaken, leading to the fluorescence quenching. Furthermore,
compared with a similar probe molecule in the reported literature
[37], it was found that the absorption change of referent probe is
similar as ours. Thus, the detection in our work is more likely to be
determined by the ICT process.
In order to further verify the above reaction mechanism, the
product of CyPd was isolated by a silica gel column after interaction
with Pd(0), and then was characterized by 1HNMR spectra. As
shown in Fig. 3, CyPd shows the characteristic alkenyl proton Hb
and Hc from 4.5 to 5.5 ppm and methylene proton Ha at 6.0 ppm,
respectively.
3.4. Pd detection performance
Under the optimal conditions, we studied the analysis capabil-
ities of CyPd by fluorescence response towards various concen-
trations of Pd(0). As shown in Fig. 4A, the fluorescence intensity at
721 nm increased with the increasing concentrations of Pd(0). The
normalized emission intensity (lex ¼ 721 nm) was linearly pro-
portional to the concentrations of Pd(0) in the range of 0e1.0 mM
(Fig.
4B).
A
linear
regression
equation
for
Pd(0),
F ¼ 0.0168 þ 0.9702x (R2 ¼ 0.9966), was obtained, where F refers to
the normalized intensities and x refers to the concentration of
Pd(0). The limit of detection (LOD) (S/N ¼ 3, the concentration
necessary to yield a net signal equal to three times the standard
deviation of the background) was calculated to be 2.24 ꢁ 10ꢂ8 M.
These results revealed that CyPd could monitor Pd(0) levels both
qualitatively and quantitatively. In addition, the increase of the
absorbance intensity at 695 nm corresponded to the addition of
Pd(0) to the CyPd (Fig. S13).
It is most important that a highly selective response to the target
species over other potentially competing species for a new fluo-
rescent probe with potential application in complex biological and
environmental samples. Therefore, the selectivity of CyPd to Pd was
investigated. As shown in Fig. 5, there was no notably fluorescence
change in the presence of the common cations, such as Agþ, Fe3þ
,
Fe2þ, Ba2þ, Al3þ, Pb2þ, Ca2þ, Mg2þ, Zn2þ, Ni2þ, Mn2þ, Cd2þ, Naþ,
Co2þ, Cu2þ, Hg2þ, Liþ, and Sr2þ (10
M for Pd(0), 100 M for other
Fig. 3. 1H NMR (500 MHz) spectra of CyPd (A) and the isolated product of
CyPd þ Pd(0) (B) in DMSO.
m
m

W. Su et al. / Dyes and Pigments 137 (2017) 293e298
297
1.2
1.0
0.8
0.6
0.4
0.2
0.0
1.2
(B)
(A)
1.0
10 μM
0 μM
0.8
0.6
0.4
0.2
0.0
Y=0.016+0.970X
R2=0.9966
700
720
740
760
780
800
0.0
0.2
0.4
0.6
0.8
1.0
Pd(0)/μM
Wavelength/nm
Fig. 4. Fluorescence spectrum evolution (A) of CyPd (10 mM) in the presence of different concentrations of Pd(0) (0e10 mM). Ratiometric calibration curve (B) of I670 nm as a function
of concentration of Pd(0), spectra were recorded in 10 mM PBS buffer (H2O/DMSO ¼ 19:1, v/v, pH ¼ 7.4, 37 ꢀC, lex ¼ 670 nm).
1.2
1.0
0.8
0.6
0.4
0.2
0.0
1
2
3
4
5
6
7
8
9 10 11 12 13 14 15 16 17 18 19 20
Fig. 5. Fluorescence responses of CyPd (10
cations (10 M for Pd(0), 100 M for other metal ions) (from left to right: blank, Pd(0),
Agþ, Fe2þ, Fe3þ, Ba2þ, Al3þ, Pb2þ, Ca2þ, Mg2þ, Zn2þ, Ni2þ, Mn2þ, Cd2þ, Naþ, Co2þ, Cu2þ
mM) in the presence of different various
m
m
,
Hg2þ
,
Liþ
,
and Sr2þ
,
respectively) in 10 mM PBS buffer (H2O/DMSO
¼
19:1, v/v,
pH ¼ 7.4, 37 ꢀC, lex ¼ 670 nm).
metal ions). However, a much larger fluorescence enhancement
were observed for CyPd upon addition of Pd(0). These results
clearly demonstrated that CyPd was a highly selective fluorescent
probe for Pd, and also verified that the hydrolysis of allyl carbonate
was specific toward Pd. In addition, the significant increase of the
emission intensity at 721 nm, along with the apparent colour
change only corresponding to the addition of Pd to the CyPd
(Fig. S14), the result indicated that CyPd can be detection of Pd by
naked-eye.
Fig. 6. Brightfield and fluorescence images of HeLa cells with CyPd: bright-field (A)
and fluorescence image (B) of the cells only incubated with CyPd (5 M) for 30 min;
bright-field (C) and fluorescence image (D) of the cells incubated with CyPd (5 M) for
30 min, and then addition of PdCl2 (5 M) for another 1 h.
m
m
m
indicating that PdCl2 was low cytotoxicity to the living cells within
a short time.
3.5. Detection of Pd in living cells
4. Conclusion
To further investigate the membrane permeability of CyPd, and
its ability to detect palladium in living cells, the cell imaging
experiment was carried out. Because PdCl2 is the most toxic species
among the palladium species, so we selected PdCl2 as the repre-
sentative palladium species in the cell fluorescent imaging exper-
iment. As shown in Fig. 6, the cell fluorescence images displayed
enhanced red fluorescence for the incubated HeLa cells with
CyPd þ PdCl2 at 540 nm excitation, which the relative emission
intensity was more intense compared to HeLa cells with CyPd
(Fig. 6B and D). These results indicated that CyPd was cell-
permeable and capable of imaging PdCl2 in living cells. The
bright-field images (Fig. 6A and C) confirmed that the cells were
living throughout the imaging experiments. Moreover, the cyto-
toxicity of PdCl2 in HeLa cells were investigated, the results from
MTT assays showed that the cells remained higher viability upon
In summary, a new near-infrared and colorimetric fluorescent
probe CyPd for palladium detection was successfully designed and
synthesized based on heptamethine cyanine dye derivative. The
probe exhibited high sensitivity and selectivity toward palladium
with excellent water solubility. In addition, the probe can be used
for “naked-eye” detection of palladium. Furthermore, CyPd has
been successfully applied to imaging of palladium in HeLa cells.
This research has demonstrated the potential of the probe for
environmental and biological applications.
Acknowledgments
This work was supported by the National Natural Science
Foundation of China (21375037, 21475043 and 21275051), Doctoral
Fund of Ministry of Education of China (NO: 20134306110006).
treatment with 2.0e10.0 mM PdCl2 for as long as 24 h (Fig. S15),

298
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