A rapid and highly sensitive fluorescent imaging materials for thiophenols

Article abstract of DOI:10.1016/j.dyepig.2016.06.009

Thiophenols, a class of toxic and polluting compounds, are widely used in industrial production. Meanwhile, some aliphatic thiols play important roles in living organisms. Therefore, the development of probes for specific thiophenol detection is of great importance. Herein, a novel highly sensitive and selective 'off-on' fluorescent probe for detecting thiophenols has been developed by an ICT mechanism through a rational design. The probe displays a large Stokes shift (140 nm) and 60-fold fluorescence intensity enhancement. More importantly, the probe features a rapid signal response time (within 100 s), a good linearity range and the detection limit is as low as 13 nM. In addition, the ability of the probe to detect thiophenols in living cells (HepG2 cells) via an enhancement of the fluorescence has also been demonstrated.

Full text of DOI:10.1016/j.dyepig.2016.06.009

Accepted Manuscript
A rapid and highly sensitive fluorescent imaging materials for thiophenols
Weijie Zhang, Fangjun Huo, Tao Liu, Yin Wen, Caixia Yin
PII:
S0143-7208(16)30260-1
10.1016/j.dyepig.2016.06.009
DYPI 5293
DOI:
Reference:
To appear in: Dyes and Pigments
Received Date: 24 April 2016
Revised Date: 1 June 2016
Accepted Date: 4 June 2016
Please cite this article as: Zhang W, Huo F, Liu T, Wen Y, Yin C, A rapid and highly sensitive fluorescent
imaging materials for thiophenols, Dyes and Pigments (2016), doi: 10.1016/j.dyepig.2016.06.009.
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Abstract Graphic
The Title:
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A rapid and highly sensitive fluorescent imaging materials for thiophenols
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List of Authors:
Weijie Zhang,a,1 Fangjun Huo, b,* Tao Liu,a,1 Yin Wen,a Caixia Yin a,**
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The statement:
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A novel highly sensitive and selective ‘off-on’ fluorescent probe for detecting
thiophenols has been developed by an ICT mechanism through a rational design. It
displays a large Stokes shift (140 nm), a good linearity range, a rapid signal response
time (within 100 s) and the detection limit is as low as 13 nM. In addition, the ability
of the probe to detect thiophenols in living cells (HepG2 cells) via an enhancement of
the fluorescence has also been proved.
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A rapid and highly sensitive fluorescent imaging materials for thiophenols
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Weijie Zhang,a,1 Fangjun Huo, b,* Tao Liu,a,1 Yin Wen,a Caixia Yin a,**
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Institute of Molecular Science, Key Laboratory of Materials for Energy Conversion
and Storage of Shanxi Province, Shanxi University, Taiyuan, 030006, China.
b Research Institute of Applied Chemistry, Shanxi University, Taiyuan, 030006, China.
*Corresponding author:
C.X. Yin, E-mail: yincx@sxu.edu.cn,Tel/Fax: +86-351-7011022;
F.J. Huo, E-mail: huofj@sxu.edu.cn.
1Weijie Zhang, Tao Liu contributed equally to this work.
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Abstract: Thiophenols, a class of toxic and polluting compounds, are widely used in
industrial production. Meanwhile, some aliphatic thiols play important roles in living
organisms. Therefore, the development of probes for specific thiophenol detection is
of great importance. Herein, a novel highly sensitive and selective ‘off-on’ fluorescent
probe for detecting thiophenols has been developed by an ICT mechanism through a
rational design. The probe displays a large Stokes shift (140 nm) and 60-fold
fluorescence intensity enhancement. More importantly, the probe features a rapid
signal response time (within 100 s), a good linearity range and the detection limit is as
low as 13 nM. In addition, the ability of the probe to detect thiophenols in living cells
(HepG2 cells) via an enhancement of the fluorescence has also been demonstrated.
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Keywords: Thiophenols; Detection; Fluorescent probe; Bioimaging.
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1. Introduction
Thiophenols, also named benzenethiols, are extensively used in organic synthesis for
preparing various products in the agrochemical industry and pharmaceutical industry.
However, thiophenols are poisonous to aquatic bio-organisms and animals, possessing a
median lethal concentration (LC 50 ) of 0.01−0.4 mM in fish and a median lethal dose
(LD 50 ) of 46.2 mg/kg in mouse. It was also reported that thiophenols can enter into the
human body easily by inhalation and skin absorption to induce systemic injuries
including shortness of breath, muscular weakness, nausea, vomiting, and even death [1-3].
Despite high toxicity, thiophenols are essential and widely used chemicals for the
preparation of agrochemicals, pharmaceuticals, and various industrial products [4-6].
Considering their toxicity and the continuing environmental concerns, simple, rapid,
sensitive, and selective detection of thiophenols is therefore of considerable interest in
both environmental and biological science.
Fluorescence detection using fluorescent probes has been recognized as one of the
most attractive methods to monitor and visualize molecules due to its simplicity,
convenience, and great potential for use in a wide range of chemical, biological, and
environmental applications. Since Wang et al. [7] reported the first reaction based
fluorescent turn-on probe for selective detection of thiophenols, a number of fluorescent
probes have been developed for these compounds [8-20]. Because of the similarity of
chemical properties among aliphatic thiols and thiophenols, most of these probes are
designed mainly for discrimination of aliphatic thiols such as cysteine and glutathione
from other amino acids, and in general, they cannot clearly discriminate thiophenols
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over aliphatic thiols [21-24]. 2,4-Dinitrobenzenesulfonyl (DNBS) was first used by
Maeda et al. as a sulfonate ester of a fluorophore for the detection of thiols [25]. The
high level of electron deficiency enables the DNBS moiety to act as an electron sink
and incur intramolecular charge transfer (ICT), resulting in the quenching of the
fluorescence. The DNBS ester can easily undergo desulfonylation in the presence of
thiophenols through a nucleophilic aromatic substitution (SNAr) mechanism,
liberating SO2 and the attached fluorophore, thus resulting in an increase in
fluorescence.
Herein, we report a novel type of highly sensitive and selective fluorescent probe
toward thiophenols which operates through an ICT pathway by selecting
N-butyl-4-amino-1,8-naphthalimide as a fluorophore, the 2,4-dinitrobenzene-
sulfonamide group as a recognition unit, and a piperazine moiety as a linker to extend
the fluorophore and reactive sulfonamide bond. The masked sulfonamide moiety can
be easily removed by a highly nucleophilic thiolate anion through the SNAr process
(see Scheme 1)[26]. More importantly, we found that this probe not only shows high
selectivity and sensitivity (DL = 13 nM) but also offers a rapid fluorescence turn-on
sensing process (100s) for detecting thiophenols. Furthermore, this probe was
successfully applied in fluorescent imaging in living cells.
2. Materials and methods
2.1. Materials
4-(2-Hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) was purchased
from Sigma-Aldrich (St. Louis, MO). Sodium hydroxide solution (0.1 mol/L) was
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added to aqueous HEPES (10 mmol/L) to adjust the pH to 7.4. Amino acids were
purchased from Shanghai Experiment Reagent Co., Ltd (Shanghai, China). All other
chemicals used were of analytical grade.
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2.2. Instruments
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A pH meter (Mettler Toledo, Switzerland) was used to determine the pH.
Ultraviolet–visible (UV–vis) spectra were recorded on an Agilent 8453 UV-Visible
spectrophotometer. Fluorescence spectra were measured on F-7000 FL
Spectrophotometer. A PO-120 quartz cuvette (10 mm) was purchased from Shanghai
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Huamei Experiment Instrument Plants, China. H NMR, 13C NMR spectra were
recorded on a Bruker AVANCE-600 MHz and 150 MHz NMR spectrometer,
respectively (Bruker, Billerica, MA). ESI determinations were carried out on AB
Triple TOF 5600plus System (AB SCIEX, Framingham, USA). The ability of the
probe to react with thiophenols in living cells was also evaluated by laser confocal
fluorescence imaging using an Olympus FV1000 laser scanning microscope.
2.3. Preparation and characterization of the probe
2.3.1.Synthesis of N-butyl-4-Br-1,8-naphthalimide
The route employed to synthesise of the probe is summarized in scheme 2.
4-Bromo-1,8-naphthalic anhydride (2.76 g, 10 mmol) and N-butylamine (0.8 g, 11
mmol) was taken in EtOH (20 mL) and stirred at 55◦C monitoring with TLC. After the
reaction was complete, the reaction mixture was evaporated in vacuum and the
residue was washed with water and dried to give a solid product. The solid obtained
was purified by column chromatography (silica gel dichloromethane) to give a white
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product in 88% yield. mp, 104℃-106℃. IR (KBr): 3389.40, 3340.32, 3308.66,
3069.14, 2954 45, 2930.41, 2871.51, 1700.09, 1658.69, 1616.26, 1590.03, 1569.12,
1502.94, 1460.23, 1435.22, 1402.59, 1380.89, 1359.43, 1342.57, 1259.15, 1229.36,
1191.63, 1151.33, 1116.74, 1098.79, 1075.34, 1042.95, 996.06, 940.45, 899.29,
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873.13, 858.94, 848.36, 780.97, 748.66, 730.67, 711.70, 659.48, 567.65, 424.73. H
NMR (DMSO-d6, 600 MHz): δ (ppm): 8.54 (d, J = 7.2 Hz, 1H), 8.51 (d, J = 8.7 Hz,
1H), 8.30 (d, J = 7.6 Hz, 1H), 8.19 (d, J = 7.7 Hz, 1H), 7.98 (t, J = 7.8 Hz, 1H), 4.02
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(t, J = 7.3 Hz, 1H), 1.61 (m, 1H), 1.35 (m, 1H), 0.93 (t, J = 7.4 Hz, 2H); C NMR
(DMSO-d6, 150 MHz): δ 163.22, 163.17, 132.95, 131.94, 131.74, 131.32, 130.14,
129.51, 129.17, 128.61, 123.10, 122.32, 30.02, 20.27, 14.17. ESI–MS m/z: [B + H]+
Calcd. for C16H15BrNO2 332.0286, Found 332.0286 (Fig. S1).
2.3.2. Synthesis of N-butyl-4-(piperazin-1-yl)-1,8-naphthalimide
N-butyl-4-Br-1,8-naphthalimide (1.65 g, 5 mmol) and Piperazine(0.52 g, 6 mmol)
was taken in 20 mL of 2-methoxyethanol. The mixture was stirred at 130℃ with
monitoring by TLC. After the reaction was complete, the clear solution obtained was
concentrated and left to cool. The brown solid was collected by filtration, washed
with water and dried to give a solid product. After crystallization from MeCN, a solid
was obtained to give a brown product in 82% yield. mp, 102℃-104℃. IR (KBr):
3374.44, 3324.79, 3291.76, 3066.96, 2956.74, 2927.13, 2862.04, 2837.13, 2748.11,
1691.30, 1652.99, 1613.22, 1588.15, 1512.96, 1456.36, 1427.33, 1384.64, 1355.30,
1287.47, 1243.90, 1181.73, 1164.87, 1137.39, 1118.77, 1089.48, 1070.96, 1024.50,
1001.53, 949.56, 929.37, 893.57, 873.89, 848.70, 835.18, 783.44, 758.58, 746.45,
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671.98, 656.48, 412. 20. H NMR (600 MHz, DMSO) δ 8.41 (dd, J = 11.9, 8.0 Hz,
2H), 8.35 (d, J = 7.9 Hz, 1H), 7.76 (t, J = 7.6 Hz, 1H), 7.27 (d, J = 7.9 Hz, 1H), 4.00 (t,
J = 6.7 Hz, 2H), 3.13 (s, 4H), 3.00 (s, 4H), 1.58 (d, J = 6.4 Hz, 2H), 1.39 – 1.25 (m,
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2H), 0.91 (t, J = 7.0 Hz, 3H). C NMR (151 MHz, DMSO) δ 163.98 (s), 163.45 (s),
156.74 (s), 132.69 (s), 131.16 – 130.91 (m), 129.59 (s), 126.30 (s), 125.74 (s), 122.98
(s), 115.70 (s), 115.32 (s), 54.57 (s), 46.14 (s), 40.48 (d, J = 16.2 Hz), 30.18 (s), 20.28
(s), 14.19 (s). ESI–MS m/z: [C + H]+ Calcd. for C20H23N3O2 337.179, Found
338.1839 (Fig. S2).
2.3.3.Synthesis of the probe
2,4-Dinitrobenzen-esulfonylchloride (0.662 g, 2.5 mmol) and N-butyl-4-
(piperazin-1-yl)-1,8-naphthalimide (0.674 g, 2 mmol) in CH2Cl2 (15 mL) with 2 mmol
(0.202 g) Et3N. The mixture was stirred overnight at 0-5 in ice-water with
monitoring by TLC. After the reaction was completed, removing the solvent under
reduced pressure gave a solid which was purified by column chromatography (silica
gel dichloromethane) to give probe 1 in 77% yields. mp 218℃-220℃. IR (KBr):
3426.36, 3105.53, 2959.04, 2924.03, 2852.91, 2170.33, 1697.12, 1650.66, 1590.96,
1556.47, 1536.51, 1513.89, 1447.13, 1425.28, 1390.92, 1351.29, 1267.81, 1234.48,
1162.78, 1125.57, 1073.18, 956.88, 909.84, 845.95, 832.77, 782.04, 755.27, 703.44,
662.57, 620.28, 598.58, 571.76, 545.76, 515.76, 464.38. 1H NMR (600 MHz, DMSO)
δ 9.08 (s, 1H), 8.65 (d, J = 6.6 Hz, 1H), 8.48 (d, J = 7.2 Hz, 1H), 8.47 – 8.42 (m, 1H),
8.40 (t, J = 9.9 Hz, 1H), 8.37 (d, J = 8.7 Hz, 1H), 7.81 (t, J = 7.7 Hz, 1H), 7.39 (d, J =
8.0 Hz, 1H), 4.03 (s, 2H), 3.60 (s, 4H), 3.34 (s, 4H), 1.70 – 1.54 (m, 2H), 1.34 (dd, J =
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14.8, 7.3 Hz, 2H), 0.90 (dd, J = 25.9, 18.6 Hz, 3H). 13C NMR (151 MHz, DMSO) δ
163.97 (s), 163.46 (s), 155.16 (s), 150.78 (s), 148.40 (s), 134.50 (s), 132.88 (s),
132.48 (s), 131.23 (s), 130.84 (s), 129.45 (s), 127.51 (s), 126.86 (s), 125.96 (s),
123.10 (s), 120.59 (s), 117.01 (s), 116.41 (s), 52.56 (s), 46.37 (s), 40.52 (s), 30.16 (s),
20.26 (s), 14.20 (s). ESI–MS m/z: [Probe + H]+ Calcd. for C26H25N5O8S 567.14238,
Found 568.1474 (Fig. S3).
<Inserted Scheme 2>
2.4. Optical studies of probe 1
Probe stock solutions were prepared in DMSO. Other analyte solutions were
prepared by using deionized water. Fluorescence measurements were carried out with
a slit width of 5 nm (λex = 383 nm). UV–Vis and fluorescence spectra were obtained
in DMSO–HEPES buffer (10 mmol/L, pH 7.4, 1 : 1, v/v).
2.5. General UV–vis and fluorescence spectra measurements
Thiophenol stock and Probe stock were prepared in DMSO. Other analytes were
prepared by using deionized water. Fluorescence measurements were carried out with
a slit width of 5 nm (λex = 383 nm). UV–Vis and fluorescence spectra were obtained
in HEPES buffer (10 mmol/L, pH 7.4)/DMSO (v/v, 1:1).
2.6. Detection wavelength range
Fluorescence spectra were measured from 450 to 650 nm with excitation at 383
nm (slit width: 5 nm/5 nm) in HEPES buffer (10 mmol/L, pH 7.4)/DMSO (v/v, 1:1).
The detection threshold for thiophenol was 13 nM, and at this level the fluorescence
color change was very obvious.
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3. Results and discussion
3.1 The Selective response of probe to thiophenols
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An important feature of the probe is that it has special selectivity for for one kind
of analyte over other substances. In order to study its special recognition ability, we
carried out the experiment by UV-Vis and fluorescence spectrometer. Figure 1 shows
the fluorescence spectral changes upon addition of various analytes (10 equiv.) in
HEPES buffer (10 mmol/L, pH 7.4)/DMSO (v/v, 1:1) solution, including PhOH,
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2-
PhNH2, PhCH3, KCN, H2O2, Na2S, Cl-, Br-, I-, N3 , NO3 , HSO3 , SO3 , ME, Cys, Hcy,
GSH, p-Nitrothiophenol, thiosalicyclic acid they all triggered very minor changes.
However, the probe displayed obvious fluorescence enhancement (Fig.1) (λex = 383
nm, λem = 523 nm) in the presence of PhSH, p-NH2-C6H4SH, p-CH3-C6H4SH in
HEPES buffer (10 mmol/L, pH 7.4)/DMSO (v/v, 1:1) solution, Similarly, the probe
displayed obvious UV-Vis spectral (Fig. S4a) changes in the presence of PhSH, and
p-NH2-C6H4SH, p-CH3-C6H4SH. These signal changes indicate that the probe can
serve as a selective chemosensor for thiophenols .
<Inserted Figure 1>
3.2 The UV–vis and Fluorescence spectra titration for probe 1
A detailed spectral titration for thiophenol was carried out. Fig. 2 showed the
fluorescence spectra change of probe 1 (2 µmol/L) upon addition of thiophenol (0-6
µmol/L) in HEPES (10 mmol/L, pH 7.4)/DMSO (v/v, 1:1). Initially, the probe (2
ꢀmol/L) has no fluorescence (λex = 383 nm). The addition of thiophenol caused a
remarkable enhancement (60-fold) of emission intensity at 523 nm (Fig. 2b). The
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sensitivity of probe 1 was then examined by the fluorescence response of the probe
toward different concentrations of thiophenol under the same reaction condition as
described above (Figure 3). The increase of fluorescence intensity was displayed in a
concentration dependent manner. However, when more than 3 equiv of thiophenol
were used, the enhancement of fluorescence intensity reached a maximum without
further alteration.
The UV-Visible spectrum response of probe 1 (10 µmol/L) toward thiophenol
(0-10 µmol/L) was studied in HEPES (10 mmol/L, pH 7.4)/DMSO (v/v, 1:1) solution.
Fig. S4b shows the change of probe 1 (0.5 µmol/L) in the UV-Visible spectrum upon
addition of thiophenol (0-10 µmol/L), the absorption peak of the probe at 375 nm is
gradually increased.
<Inserted Figure 2>
To investigate the detection limit of probe 1 for thiophenol, probe 1 (2 µmol/L)
was treated with various concentrations of thiophenol (0-6 µmol/L) and the relative
emission intensity at 523 nm in HEPES (10 mmol/L, pH 7.4)/DMSO (v/v, 1:1) (λex =
383 nm, slit: 5 nm/5 nm) was plotted as a function of the thiophenol concentration
(Fig. 3). Subsequent data analysis revealed an excellent linear relationship (R2 =
0.9999) between the relative fluorescence intensity at 523 nm and the thiophenol
concentration (0-6 µmol/L). The detection limit for thiophenol, based on the definition
by IUPAC (CDL=3 Sb/m) [27-31], was found to be 13 nM, which was much lower
than that of the ICT probe reported by Wang et al. A summary of the comparison of
recently developed fluorescent probes for thiophenol is given in Table 1. The probe 1
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showed its excellent analytical performance.
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<Inserted Figure 3>
<Inserted Table 1>
3.3. pH dependence
As the pH value of system is often considered as a significant influencing factor on
interactions. The effect of pH on the fluorescence intensity of the probe in the absence
and presence of thiophenol were also investigated (Figure S5). Experimental results
indicated that the probe was pH insensitive in the pH range from pH 2 to 13. As
expected, due to the enhanced ionization of thiophenol to result in a faster SNAr
process, the fluorescence intensity increase with the pH ranging from pH 5 to 11, .
Notably, the fluorescence changes of probe 1 to thiophenol reached a maximum and
became almost constant in the pH range of 6−8, which is consistent with the pKa
value of thiophenol (pK a ≈ 6.5). The weaker response at pH >8.0 was presumably
attributed to diminished concentration of thiophenol, because of the disulfide
formation under this conditions. Therefore, physiological pH (pH=7.4) could be
selected as an appropriate working pH for the sensing of thiophenols with the
designed probe.
3.4. Time-dependence in the detection process of thiophenol
Time-dependent modulations in the fluorescence spectra of probe 1 were
monitored in the presence of 10 eq. of thiophenol. The kinetic study (Fig. 4) showed
that the reaction was complete within 100 s for thiophenol, indicating that the sensor
can achieve real-time detection of thiophenols.
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<Inserted Figure 4>
3.5. Proposed mechanism
Similar to previously reported DNBS-based thiophenol probes [33, 34], the
fluorescence enhancement response of probe 1 is attributed to a thiolate anion induced
SNAr process which resulted in the cleavage of the DNBS group from the probe
molecule (Scheme 1). Zhao etc has reported a new fluorescent probe for thiophenols
based on 2,4-Dinitrosulfonyl functional group with 1-amino BODIPY. Under their
experimental conditions, the probe displayed a selective response to thiophenols over
aliphatic thiols and the detection limit could be improved to 34 nM. In our work, upon
addition of thiophenol, the probe caused a remarkable enhancement (60-fold) of
emission intensity at 523 nm and the detecting limit is as low as 13 nM, which was
much lower compared with the literature reported. But, there are still room to improve
in terms of detection limit and application in vivo. As mentioned before, after reacting
with thiophenol, probe produced an obviously fluorescence enhancement due to a
nucleophilic aromatic substitution (SNAr) mechanism, releasing SO2 gas and
N-butyl-4-(piperazin-1-yl)-1,8-naphthalimide, thus resulting in an increase in
fluorescence. Furthermore, The identification of the adduct in the ESI-MS analysis
made it possible to propose the signaling mechanism: a peak at m/z 338.1852
corresponding to [probe-PhSH + H]+ was clearly observed (Fig. S6). Further NMR
spectroscopic analysis also provided the evidence for thiophenol nucleophilic
substitution reactions with probe 1. With addition of thiophenol to probe in DMSO-d6,
the resonance of the original proton at 9.08, 8.66, 8.64 ppm disappeared and new
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peaks at 8.90, 7.58, 7.56 ppm appeared (Fig. S7). Thus, the sensing mechanism of
probe 1 towards thiophenol, is based on the nucleophilic aromatic substitution (SNAr)
reactions as shown in Scheme 1.
3.6. Cellular Imaging
Ions play important roles in many industrial and biochemical processes[35, 36].
In order to evaluate the cell permeability and capability of probe 1 to selectively
detect intracellular thiophenol, live-cell imaging studies were carried out (Fig. 5). As
shown in Fig. 5a, HepG2 cells incubated with 10 µmol/L of probe for 30 min at 37
showed no fluorescence. In a further experiment it was found that HepG2 cells
displayed yellow-green fluorescence when the cells were first incubated with 10
ꢀmol/L of probe for 30 min at 37 and then incubated with 20 ꢀmol/L thiolphenol
(Fig. 5b). These data demonstrated the good cell-membrane permeability of probe and
it can thus be used to mark thiolphenol within living cells.
4. Conclusions
In summary, we have developed a “turn-on” fluorescent probe for thiolphenols based
on a 2,4-dinitrosulfonyl functional group with high selectivity and sensitivity in
aqueous-DMSO media. Besides, owing the merits of a high selectivity and sensitivity,
this probe shows a rapid detection process (100 s), high fluorescence enhancement
(up to 60-fold), large Stokes shift and the detection limit is as low as 13 nM.
Moreover, fluorescence scanning microscopic experiments demonstrated that the
probe can readily be used to detect the intracellular thiophenol in living HepG2 cells.
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Acknowledgments
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The work was supported by the National Natural Science Foundation of China (No.
21472118), the Program for the Top Young and Middle-aged Innovative Talents of
Higher Learning Institutions of Shanxi (TYMIT, No. 2013802), talents Support
Program of Shanxi Province (No. 2014401), Shanxi Province Outstanding Youth
Fund (No. 2014021002), and Shanxi University funds for study aboard 2014.
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Figure captions
Scheme 1 Proposed detection mechanism of the probe to thiophenol.
Scheme 2 The synthesis of the probe 1.
Fig. 1 (a) Fluorescence spectra spectra of the probe 1 (2 µmol/L) in DMSO–HEPES
buffer (10 mmol/L, pH=7.4, 1 : 1, v/v) in the presence of 60 µmol/L PhOH, PhNH2,
PhCH3, KCN, H2O2, Na2S, ME, Cys, Hcy, GSH, and 6 µmol/L PhSH, p-NH2-C6H4SH,
p-CH3-C6H4SH (b) Optical density two-dimensional graph of the probe at 523 nm
upon the addition of various analytes.
Fig. 2 Fluorescence spectra of the probe 1 (2 µmol/L) in the presence of various
concentrations of thiophenol (0-6 µmol/L) in DMSO–HEPES buffer (10 mmol/L, pH
7.4, 1 : 1, v/v) (λex = 383 nm, slit: 5 nm/5 nm).
Fig. 3 The linearity of the relative fluorescence intensity versus thiophenol
concentration.
Table. 1 Summary of fluorescent probes for thiophenols.
Fig. 4 Reaction time profiles of probe (2 µmol/L) with thiophenol (60 µmol/L).
Fig. 5 Confocal fluorescence images in HepG2 cells. (a) Fluorescence image of
HepG2 cells with adding probe (10 ꢀM) and its bright field image (c); (b)
Fluorescence image of HepG2 cells incubated with 10 ꢀM probe for 30 min at 37 °C,
then incubated with 20 ꢀM thiophenol for 30 min at 37 °C and its bright field image
(d).
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Scheme 1
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Scheme 2
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Fig. 1 (a)
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Table 1
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Fig.4.
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Fig. 5.
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Support Information
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A rapid and highly sensitive fluorescent imaging materials for thiophenols
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Figure S1: Structure characterization of compound B
Figure S2: Structure characterization of compound C
Figure S3: Structure characterization of the probe
Figure S4: UV-Vis titration
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Figure S5: PH interference to the detection
Figure S6: The ESI-MS of product obtained by reaction of probe and thiophenol
Figure S7: 1D 1H NMR the probe and probe-thiophenol
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Figure S1: 1D 1H NMR, 13C NMR, ESI-MS and IR of compound B.
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The 1H NMR (600MHz) spectra of compound B in DMSO-d6.
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The 13C NMR (150 MHz) spectra of compound B in DMSO-d6.
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The ESI-MS of compound B: m/z: [B + H] Calcd. for C16H15BrNO2 332.0286,
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Found 332.0286.
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The IR spectra of compound B
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