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P. Yin et al. / Tetrahedron xxx (xxxx) xxx
2.4. Fluorescence imaging of Hg2þ in living cells
The HeLa cells were cultured in DMEM supplemented with 10%
FBS at 37 ꢀC (under 5% CO2), which were chosen for the cell imaging
experiments. Firstly, the living HeLa cells and sensor 3 TH (10 mM)
were cultured in cell culture media for 1 h at 37 ꢀC, washed with
PBS buffer (pH ¼ 7.4) for three times, and then imaging. Next, Hg2þ
(10 mM) was added to the pre-cultured cells of the 3 TH, cultured for
30 min at 37 ꢀC, and washed with PBS for three times, then imaging.
After 60 min, imaging the cells that loaded Hg2þ again were
detected under CLSM. The excitation wavelength was 488 nm, and
the emission filter was 500e550 nm.
Scheme 1. The synthetic route of sensor 3 TH.
2. Experiment procedure
3. Results and discussion
2.1. Reagents, materials and apparatus
3.1. Design and synthesis of 3 TH
All reagents were purchased from Sigma-Aldrich Chemical
Company and used without further purification. Deionized water
was used throughout all experiments. NMR spectra were recorded
on a BrukereAVANCE 400 NMR Spectrometer, with tetramethylsi-
lane (TMS) as an internal standard. FTIR spectra were performed on
Bruker ALPHA FT-IR spectrometer using KBr pellets. High resolu-
tion mass spectra (HRMS) were measured on an Agilent 6510
AccurateeMass QeTOF LC/MS system. Absorption and fluorescence
spectra were measured on Shimadzu UV-2600 and Hitachi Fe4600
spectrofluorometer, respectively. The pH measurements were per-
formed on a Model PHS-3C pH meter. The cell fluorescence images
were detected by a Leica TCS SP8 confocal-laser scanning micro-
scope (CL SM) with an objective oil lens of 63X magnification.
As described in Scheme 1, compound 3 TH was facilely synthe-
sized from 3T-CHO [53] and 1,3-malonylthiol. The structure of 3 TH
was well confirmed by 1H NMR, 13C NMR, FTIR and HRMS
(Figs. S1e4). Our design strategy was based on the well-known
Hg2þ-promoted desulfurization reaction of thioacetal to the cor-
responding aldehyde.
3.2. UVevis spectral response of 3 TH to Hg2þ
Initially, the selectivity study of sensor 3 TH towards Hg2þ was
evaluated. The UVevis response of 3 TH (10 mM) was carried out
after addition of a series of 2.0 equiv. metal ions (Kþ, Naþ, Agþ, Ca2þ
Mg2þ, Ba2þ, Al3þ, Co2þ, Cu2þ, Ni2þ, Pb2þ, Hg2þ, Zn2þ, Cd2þ, Cr3þ
,
,
Sr2þ, Fe2þ, and Fe3þ) in EtOH/H2O (1:1, v/v) solution. As shown in
Fig. 1, the 3 TH (10 mM) exhibited a maximal absorption band at
2.2. General procedure for the spectra measurement
360 nm, which should be ascribed to the absorption of oligothio-
phene moiety. Upon addition of various metal ions (2.0 equiv.), only
Hg2þ induced great changes: a typical absorption at 360 nm
reduced significantly while a new red-shift absorption band is
created at 400 nm along with the solution color change from
colorless to pale yellow. Whereas, other metal ions caused no sig-
nificant color and spectral changes. These results indicated that the
3 TH was dissolved in ethanol to prepare the 1.0 mM of stock
solution. The appropriate amount of each inorganic metallic salts
(Kþ, Naþ, Agþ, Ca2þ, Mg2þ, Ba2þ, Al3þ, Co2þ, Cu2þ, Ni2þ, Pb2þ, Hg2þ
,
Zn2þ, Cd2þ, Cr3þ, Sr2þ, Fe2þ, and Fe3þ) was dissolved in deionized
water at a concentration of 1.0 mM. Test solutions were prepared by
placing the stock solutions and diluted to the desired analytical
concentrations with a mixed solution of EtOH/H2O (1/1, v/v). All
measurements were carried out at room temperature, the excita-
tion wavelength was 360 nm with excitation and emission slit
widths of 5 and 5 nm, respectively.
colorimetric sensor 3 TH has high selectivity towards Hg2þ
.
2.3. Synthesis of sensor 3 TH
Compound 3T-CHO (50 mg, 0.18 mmol) and 1,3-malonylthiol
(39 mg, 0.36 mmol) were dissolved in dry dichloromethane
(10 mL), a catalytic amount of BF3$Et2O (0.02 mL, 0.15 mmol) as the
Lewis acid added into the above solution. The reaction mixture was
stirred at room temperature for 10 h under an atmosphere of argon.
Then, the reaction mixture was evaporated in vacuo, and the crude
product was purified by column chromatography to obtain com-
pound 3 TH, which is as a yellow solid (45 mg, 68% yield). Mp
136.8e137.5 ꢀC; FTIR (KBr, cmꢁ1) v ¼ 1059 (CeSeC), 1503 (C]C,
thiophene ring); 1H NMR (400 Hz, DMSO‑d6, ppm):
d
¼ 7.53 (d,
J ¼ 4.8 Hz, 1H), 7.34 (d, J ¼ 3.2 Hz, 1H), 7.26 (s, 2H), 7.18 (d, J ¼ 3.6 Hz,
1H), 7.09 (m, J ¼ 8.8 Hz, 2H), 5.73 (s, 1H), 3.05 (t, J ¼ 11.6 Hz, 2H),
2.89 (d, J ¼ 14 Hz, 2H), 2.08 (d, J ¼ 14 Hz, 1H), 1.76 (d, J ¼ 14 Hz, 1H);
13C NMR (100 Hz, DMSO‑d6, ppm):
134.8, 128.4, 127.2, 125.7, 125.1, 124.9, 124.3, 123.6, 79.6, 43.4, 30.1,
24.6; HRMS (ESI) m/z calcd for C16H14S5 [MþH]þ: 369.9777; Found
366.9782.
d
¼ 142.2, 135.9, 135.8, 135.6,
Fig. 1. The UVevis absorption spectra of 3 TH (10
various metal ions in EtOH/H2O (1/1, v/v) solution; Inset: Colorimetric responses of
3 TH (10 M) in an EtOH/H2O (1:1, v/v) solution upon the addition of 2.0 equiv. of
various metal ions.
mM) after addition of 2.0 equiv. of
m
Please cite this article as: P. Yin et al., A new “naked-eye” colorimetric and ratiometric fluorescent sensor for imaging Hg2þ in living cells,