C. Li, P. Yin, T. Li et al.
Journal of Molecular Liquids 320 (2020) 114396
Scheme 1. The design and synthesis of sensor 2TD.
2. Experimental section
(DMSO/H2O, 1:99, v/v). Stock solutions (10 mM) of various anions
(ClO−, Br−, F−, Cl−, ClO4−, HCO3−, CO32−, PO34−, SO42−, NO3−, NO−2
,
2.1. Instruments and chemicals
H2PO4−, HPO24−, S2O32−, AcO−), metal cations (Al3+, Cr3+, Fe3+, Ni2+
,
Pb2+, Cu2+, Hg2+) and various reactive ROS or RNS species (H2O2,
ONOO−, •OH, •NO, ROO•) were dissolved in deionized water and used
freshly. All spectroscopic measurements were measured at room tem-
perature in ~100% aqueous solution (DMSO/H2O, 1/99, v/v) with pH 7.
The spectra of absorption, fluorescence, NMR, FTIR, and HRMS were
completed by the spectrometer of Shimadzu UV-2600, Hitachi F-4600,
Bruker AV-400, Bruker ALPHA FT-IR, and Q-TOF LC/MS Agilent 6510, re-
spectively. The absolute fluorescence quantum yields (ΦF) were ob-
tained by an Edinburgh FLS1000 spectrophotometer. The pH value
was determined by PHS-25 pH-meter. MTT assay was tested by micro-
plate reader (Thermo Scientific, USA), and fluorescence images were
obtained by confocal laser scanning microscope (CLSM) Leica TCS SP8
with a 63× magnification target oil lens. The DFT studies at the B3LYP/
6-311G(d) level were performed via Gaussian 09 package. All the re-
agents involved in the experiments are analytically pure, which are pur-
chased from commercial suppliers (Adamas, China) and were used
directly. 2,2′-Bithiophene (2T) and [2,2′-bithiophene]-5-carbaldehyde
(2T-CHO) were synthesized through the previous method [52,53]. The
ONOO−, ROO•, NO• and •OH were prepared according to the previous
method [5,54]. Deionized water was used throughout the test process.
For fluorescence measurements, the slits for excitation and emission
were set at 5/5 nm, respectively, the voltage was set at 500 V, the scan
rate was set at 2400 nm/min, and the excitation wavelength was set
at 350 nm.
2.4. Cell incubation and fluorescence imaging
The HeLa cells were firstly seeded at a 96-well plate at 37 °C for 24 h,
and then were treated with 10 μM sensor 2TD for 60 min. After the cells
being washed for 3 times with PBS buffer, and further incubated with
30 μM ClO− for 30 min. Finally, the HeLa cell imaging was achieved by
using CLSM under the 430–470 nm blue channel with an excitation at
405 nm.
3. Results and discussion
3.1. Selectivity and anti-interference studies
The sensor 2TD itself is water-insoluble, the effect of solvent (differ-
ent ratios between DMSO and H2O) on the fluorescence spectra was
firstly explored (Fig. S5), and the result indicated that the DMSO/H2O
(1/99, v/v) solution is the best detection media for all the sensing exper-
iments. To evaluate the selectivity of sensor 2TD to ClO−, the absorption
and fluorescence spectra of 10 μM 2TD were performed upon treating
with 20 μM various biological analytes in ~100% aqueous solution
(DMSO/H2O, 1/99, v/v). The anions including ClO−, Br−, F−, Cl−, ClO4−,
2.2. Synthesis of 2-(-([2,2′-bithiophen]-5-ylmethylene)amino)-3-
aminomale-onitrile (2TD)
A mixture of compound 2T-CHO (388 mg, 2.0 mmol) and
diaminomaleonitrile (DAMN) (250 mg, 2.3 mmol) in dry EtOH
(15 mL) was added catalytic amount of acetic acid glacial (2 drops).
The above reaction mixture was stirred for 4 h under reflux. Upon com-
pletion, the orange precipitate was filtered-off, washed with ethanol
and dried in vacuum to produce the final compound 2TD (511 mg,
yield: 90.1%). 1H NMR (400 MHz, DMSO‑d6, ppm): δ = 8.40 (s, 1H),
7.77 (s, 2H), 7.72 (d, J = 4.0 Hz, 1H), 7.64 (d, J = 4.0 Hz, 1H), 7.45 (d,
J = 4.0 Hz, 1H), 7.42 (d, J = 4.0 Hz, 1H), 7.16 (t, J = 4.0 Hz, 1H)
(Fig. S1); 13C NMR (100 MHz, DMSO‑d6, ppm): δ = 149.1, 142.4,
140.5, 135.6, 129.3, 127.7, 126.4, 126.0, 125.5, 115.0, 114.3, 103.4
(Fig. S2); FTIR (KBr, cm−1): v = 3455, 3306 (NH2), 2207 (C`N), 1600
(C=N) (Fig. S3); HRMS (ESI) m/z calcd for C13H8N4S2: 284.0190;
Found: 285.0247, [M+H]+ (Fig. S4).
HCO−3 , CO32−, PO43−, SO24−, NO−3 , NO−2 , H2PO−4 , HPO42−, S2O23−, AcO−
,
metal ions including (Al3+, Cr3+, Fe3+, Ni2+, Pb2+, Cu2+, Hg2+), and re-
active ROS or RNS species including H2O2, ONOO−, •OH, ROO•, and •NO,
were selected for the selectivity and competition tests, because some
oxidizing and reducing species as well as some metal ions may have
some effects on the sensor 2TD through a redox or coordination reac-
tion. As depicted in Fig. 1a, upon adding ClO−, the main absorption
peak at 378 nm disappeared, which was accompanied by a new absorp-
tion peak at 350 nm, with obvious yellow color change to colorless,
while other tested analytes showed negligible response, suggesting
that 2TD had a highly specific selectivity for ClO− over other tested spe-
cies. Under the same conditions, the selectivity of the 2TD towards ClO−
is further evaluated by fluorescence response. As displayed in Fig. 1b,
the free 2TD exhibited almost no fluorescence (ФF = 12.47%) after exci-
tation at 350 nm, only the introduction of ClO− resulted in a notable
fluorescence enhancement at 423 nm (ФF = 20.09%) along with obvi-
ous fluorescence color change, indicating that 2TD can effectively and
selectively detect ClO−.
2.3. Spectral measurements
A stock solution of sensor 2TD (1 mM) was prepared in DMSO, and
then the sensor 2TD was diluted to 10 μM with ~100% aqueous solution
2
Chen, Jianbin
