A phthalimide-based fluorescent probe for thiophenol detection in water samples and living cells with a large Stokes shift

Article abstract of DOI:10.1016/j.tet.2015.08.074

A phthalimide-based fluorescent probe for the detection of thiophenol was developed based on the combination of photo-induced electron transfer (PET) and excited-state intramolecular proton transfer (ESIPT) mechanisms. This probe displays high sensitivity

Full text of DOI:10.1016/j.tet.2015.08.074

Accepted Manuscript
A phthalimide-based fluorescent probe for thiophenol detection in water samples and
living cells with a large Stokes shift
Xingjiang Liu, Liu Yang, Li Gao, Wenqiang Chen, Fengpei Qi, Xiangzhi Song
PII:
S0040-4020(15)30028-4
10.1016/j.tet.2015.08.074
TET 27096
DOI:
Reference:
To appear in: Tetrahedron
Received Date: 15 May 2015
Revised Date: 28 August 2015
Accepted Date: 31 August 2015
Please cite this article as: Liu X, Yang L, Gao L, Chen W, Qi F, Song X, A phthalimide-based fluorescent
probe for thiophenol detection in water samples and living cells with a large Stokes shift, Tetrahedron
(2015), doi: 10.1016/j.tet.2015.08.074.
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A phthalimide-based fluorescent probe for
thiophenol detection in water samples and
living cells with a large Stokes shift
Leave this area blank for abstract info.
Xingjiang Liu, Liu Yang, Li Gao,Wenqiang Chen, Fengpei Qi, and Xiangzhi Song
College of Chemistry & Chemical Engineering, Central South University, Changsha, Hunan Province, P. R.
China, 410083.

1
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Tetrahedron
journal homepage: www.elsevier.com
A phthalimide-based fluorescent probe for thiophenol detection in water samples and
living cells with a large Stokes shift
Xingjiang Liu a, Liu Yang a, Li Gao a, Wenqiang Chen a, Fengpei Qi a, and Xiangzhi Song abc,
a College of Chemistry & Chemical Engineering, Central South University, Changsha, Hunan Province, P. R. China, 410083.
b State Key Laboratory for Powder Metallurgy, Central South University, Changsha, Hunan province, P. R. China, 410083.
c State Key Laboratory of Fine Chemicals, Dalian University of Technology, Dalian 116024, Liaoning Province, P. R. China, 116024.
ARTICLE INFO
ABSTRACT
Article history:
Received
Received in revised form
Accepted
A phthalimide-based fluorescent probe for the detection of thiophenol was developed based on
the combination of photo-induced electron transfer (PET) and excited-state intramolecular
proton transfer (ESIPT) mechanisms. This probe displays high sensitivity and good selectivity
toward thiophenol with a large Stokes shift (161 nm) and a low detection limit (3.5 nM, based
on S/N = 3). Furthermore, the applications of this probe for quantitative detection of thiophenol
in real water samples and imaging intracellular thiophenol in living cells were successfully
demonstrated.
Available online
Keywords:
Fluorescent probe
Phthalimide
Photo-induced electron transfer (PET)
Excited-state intramolecular proton transfer (ESIPT)
Thiophenol
2009 Elsevier Ltd. All rights reserved.
1. Introduction
As a result, it would be an ideal candidate for the design of
fluorescent probes.28-31 We previously reported the first 3-
hydroxyphthalimide-based fluorescent probe for biothiols.32 It
has been demonstrated that 2,4-dinitrophenlate moiety is an
effective sensing group for the design of fluorescent probes to
discriminate thiophenols from aliphatic thiols by taking
Thiophenols, the environmental pollutants with high toxicity,
are widely used in production of pesticide, medicine and polymer
materials.1-3 Thiophenols can be readily autoxidized under
physiological conditions to form aromatic disulphides ， and
generate the harmful superoxide radical and hydrogen peroxide
in this process. On the other hand, aromatic diphenyl disulphides
have been found to undergo a cyclic reduction/autoxidation
reaction by virtue of glutathione (GSH) to generate ‘active
oxygen’ species which can induce oxidative damage to
erythrocytes cells.4-8 Thus, it has been shown that thiophenol
exposure not only induces grave central nervous system damage,
but also causes other related systemic injuries including increased
respiration, muscular weakness, hind limb paralysis, coma, and
advantage of their different pKa values (Ca. 6.5 and 8.5 for
33
thiophenols and aliphatic thiols, respectively).19,
Herein, we
reported
a
2,4-dinitrophenlate
derivative
of
3-
hydroxyphthalimide, Probe 1, that we designed as fluorescent
probe for the selective detection of thiophenols with a large
Stokes shift at a physiological condition. The synthetic route of
Probe 1 in four steps was shown in Scheme 1. Due to a photo-
9
even death.5, As a consequence, considerable attention have
been paid to techniques for the rapid, selective and sensitive
detection of thiophenols in environmental and biological
samples.10-19 In this regard, fluorescent sensing is well suited to
meet this need owning to its simplicity, high sensitivity, good
selectivity, rapid response and non-invasiveness.20-27
3-Hydroxyphthalimide and its derivatives exhibit a large
Stokes shift upon excited because of an excited-state
intramolecular proton transfer (ESIPT) process in the excited
state. Additionally, 3-hydroxyphthalimide possesses many
favorable optical properties such as good photo stability,
emission in green region, and relatively high fluorescent quantum.
———
Scheme 1. Synthetic route of Probe 1. (a) Butylamine, acetic acid, 120 °C,
2.5 h, yield 85.7%. (b) Palladium/C, H2, CH3OH, 65 °C, 12 h, 71.5%. (c) (1)
Corresponding author. Tel.: +86-731-88836954; fax: +86-731-88836954; e-mail: xzsong@csu.edu.cn

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NaNO2, 50% H2SO4, 0 °C, 0.5 h; (2) 90 °C, 1 h, yield 85.4%. (d) 1-Fluoro-
Fig. 1 Fluorescence spectra of Probe 1 (5.0 µM) in the presence of thiophenol
2,4-dinitro-benzene, K2CO3, DMF, 90 °C, 10 h, yield 74.5%.
(0.0 - 5.0 equiv.) in HEPES buffer (10.0 mM, pH = 7.4, containing 20%
CH3CN) with excitation wavelength at 358 nm. Inset: fluorescence images of
Probe 1 (5.0 µM) before and after addition of thiophenol (5.0 equiv.) in
HEPES buffer (10.0 mM, pH = 7.4 containing 20% CH3CN) .
induced electron transfer (PET) process and inhibition of proton
transfer (ESIPT) process induced by 2, 4-dinitrophenolate
moiety,34-35 we expected Probe 1 would be essentially non-
fluorescent. Thiophenol rather than aliphatic thiols would
selectively cleave the 2,4-dinitrophenolate moiety in Probe 1,
thus eliminating the possibility of PET-induced quenching and
concomitantly allowing the occurrence of ESIPT process (shown
in Scheme 2).
Fig. 2 Plot of the fluorescence intensity at 516 nm of Probe 1 (5.0 µM) as a
function of the thiophenol concentration in HEPES buffer (10.0 mM, pH =
7.4, containing 20% CH3CN). Inset: the linear relationship between the
fluorescence intensity at 516 nm of Probe 1 and the concentration of
thiophenol.
Scheme 2. The proposed sensing mechanism of Probe 1 with thiophenol
(PhSH).
2.2. Mechanism studies
2. Results and discussion
We reasoned that thiophenols could selectively cleave
dinitrobenzene moiety in Probe 1 generating dye 4 which is
highly fluorescent upon excited. To confirm the sensing
2.1. Sensing property of Probe 1 to thiophenol
Dye 4 exhibits strong green fluorescence (ΦF = 0.27) with a
maximum at 516 nm. Notably, it displays a large Stokes shift
(161 nm) which can reduce the interference from incident light.
As expected, Probe 1 is essentially non-fluorescent (ΦF < 0.001)
in HEPES buffer solution (10.0 mM, pH = 7.4, containing 20%
CH3CN), which is attributed to the PET process resulted from the
introduction of 2,4-dinitrobenzene moiety. The addition of
thiophenol to the solution of Probe 1 triggered a strong green
fluorescence (λemmax = 516 nm) with an concomitant appearance
of an absorption band at 350 nm, which are coincident with the
1
mechanism, we carried out H NMR spectral analysis on the
reaction product of Probe 1 with thiophenol, as shown in Fig. 3.
1
As expected, the H NMR spectrum of the reaction product is
very similar to that of dye 4. Furthermore, the mass spectra of the
reaction product of Probe 1 with thiophenol displayed a peak at
m/z = 218.0229 (Fig. S11) which is nearly identical to the molar
weight of dye 4 ([M-H] = 218.0817). Absorption and emission
1
spectra, along with H NMR and mass spectral analysis, confirm
that dye 4 is the product generated from Probe 1 with thiophenol.
spectral characteristic of dye
4 (Fig. 1 and Fig. S1).
Concentration-dependent studies found that the fluorescence
intensity at 516 nm increased with increasing thiophenol
concentration. The fluorescence enhancement could be up to 64-
fold when 5.0 equiv. of thiophenol was added to the solution of
Probe 1. A plot of fluorescence intensity vs the concentrations of
thiophenol in the range of 0.0 – 10.0 µM showed good linearity
(R = 0.99601) (Fig. 2). The detection limit was found to be 3.5
nM (based on S/N = 3), which was sufficiently low for the
detection of thiophenol in environmental and biological samples.
This result provided support for Probe 1 having high sensitivity
to thiophenol.
Fig. 3 Partial 1H NMR spectra of Probe 1 (a), the isolated product of Probe 1
with thiophenol (b) and reference dye 4 (c) in CDCl3.
2.3. Selectivity and competition studies
To evaluate the selectivity of Probe 1 (5.0 µM) towards
thiophenol, we observed its fluorescence response to various
analytes including aliphatic thiols (Cys, Hcy, mercaptoethanol,
thiohydracrylic acid, 0.5 mM for each, and 1.0 mM GSH), some
nucleophilic species (Ala, Gly, PhOH, PhNH2, NaN3, NaHS, 0.4
mM for each), and various anions and metal ions (NaF, NaBr, KI,
NaNO2, Na2S2O3, Na2SO4, Na3PO4, Na2SO3, ZnCl2, CaCl2,
MgCl2, 0.4 mM for each; 10.0 mM NaCl and KCl). As shown in

3
Fig. 4, only thiophenol resulted in a significant fluorescence
on Probe 1 under the same condition, which might be ascribed
to the different pKa values of thiophenol and biothiols.
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enhancement whereas other interfering analytes did cause
negligible fluoresce change. We were pleased that this probe
showed excellent selectivity to thiophenol over aliphatic thiols
(Cys, Hcy, and GSH) and NaHS. Moreover, the competition
studies were also carried out in the same concentration of
interfering analytes as mentioned above. The performance of
Probe 1 for the detection of thiophenol was not obviously
influenced by the co-existence of other interfering analytes
except that the fluorescence intensity was weakened in the
presence of Na3PO4. Therefore, it can be concluded that Probe 1
exhibits excellent selectivity for thiophenol.
Fig. 6 Time-dependent fluorescence intensity at 516 nm of Probe
1 (5.0
µ
M) in response to thiophenol, Cys, Hcy, and GSH in HEPES buffer
(10.0 mM, pH = 7.4, containing 20% CH3CN).
2.5. pH effect studies
The potential pH effects on the fluorescence signal of Probe 1
with thiophenol was also investigated. As shown in Fig. 7, the
fluorescence spectra of Probe 1 remained essentially unchanged
within a wide pH range from 2 to 13, indicating that Probe 1 was
very stable. In contrast, remarkable fluorescence enhancement
was observed when this probe was treated with thiophenol within
a pH range from 6.0 to 10.0. The data implied that Probe 1 was
able to detect thiophenol in a relatively wide pH range.
Fig. 4 Fluorescence spectra of Probe 1 (5.0 µM) with various tested analytes
in HEPES buffer (10.0 mM, pH = 7.4, containing 20% CH3CN). The analytes
include thiophenol, Cys, Hcy, GSH, mercaptoethanol, thiohydracrylic acid,
Ala, Gly, PhOH, PhNH2, NaN3, NaHS, NaF, NaCl, NaBr, KI, NaNO2,
Na2S2O3, Na2SO4, Na3PO4, Na2SO3, ZnCl2, CaCl2, MgCl2, KCl.
Fig. 5 The fluorescent responses of Probe 1 (5.0 µM) to thiophenol in the
presence of various relevant species in HEPES buffer (10.0 mM, pH = 7.4,
containing 20% CH3CN). 1, thiophenol; 2, Cys; 3, Hcy; 4, GSH; 5,
mercaptoethanol; 6, thiohydracrylic acid; 7, Ala; 8, Gly; 9, PhOH; 10,
PhNH2; 11, NaN3; 12, NaHS; 13, NaF; 14, NaCl; 15, NaBr; 16, KI; 17,
NaNO2; 18, Na2S2O3; 19, Na2SO4; 20, Na3PO4; 21, Na2SO3; 22, ZnCl2; 23,
CaCl2; 24, MgCl2; 25, KCl.
Fig. 7 Fluorescence intensity at 516 nm of Probe 1 (5.0 µM) in the absence
(black line) and presence (red line) of thiophenol (25.0 µM) at different pH
values.
2.6. Preliminary application of Probe 1 in water samples and
living cells
Firstly, we applied Probe 1 to detect toxic thiophenol in real
water samples to demonstrate its practical utility in
environmental science. The water samples were collected from
the Yangtze River and Tap water in Suzhou City, respectively.
All the samples were filtrated before used. The fluorescence
changes of Probe 1 (5.0 µM) were examined in all these samples
which were spiked with thiophenol at different levels (0.0, 1.0,
2.0, 3.0, 4.0, 5.0, 6.0, 7.0, 8.0, 9.0, 10.0 µM). From the results
shown in Fig. 8a, we could see that fluorescence enhancements in
real samples and in distilled water were coincident. As shown in
Fig. 8b-c, a good linear relationship between the fluorescence
2.4. Kinetic studies
We next studied time-dependent fluorescence behavior of
Probe 1 with thiophenol by monitoring changes in fluorescence
intensity at 516 nm as a function of time (Fig. 6). Upon the
addition of 5.0 equiv. of thiophenol to Probe 1, a strong
fluorescence signal was initiated within
3 min (7-fold
enhancement) and reached a stable intensity plateau within 60
min. However, in the absence of thiophenol, no fluorescence
signal was observed in the solution of Probe 1. It’s noted that
GSH, Cys and Hcy didn’t produce fluorescence enhancement
when time-dependent fluorescence experiments were performed

4
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Fig. 8 (a) Fluorescent response of Probe 1 (5.0 µM) to different concentrations of thiophenol in different water samples (10.0 mM HEPES buffer, pH = 7.4,
containing 20% CH3CN). (b) and (c) Linear plots of fluorescent intensity of Probe 1 at 516 nm against the spiked concentrations of thiophenol from 0.0 to 10.0
µM in Yangtze River and tap water samples.
intensity at 516 nm and the spiked thiophenol concentrations was
obtained for each real water sample. Moreover, the added
thiophenol in these samples could be accurately measured with a
good recovery (see Table S1). These results suggest that Probe 1
has potential application for quantitative detection of thiophenol
in water samples.
1H NMR and 13C NMR spectra were obtained on a Bruker 500
NMR spectrometer using tetramethylsilane (TMS) as the internal
standard for chemical shifts. Mass spectra were recorded on
MICROTOF-Q II mass spectrometer (Bruker Daltonics,
Germany). All spectral characterizations were carried out in
HPLC-grade solvents at 20 °C within a 10 mm quartz cell. UV-
vis absorption spectroscopy was measured with a UV-2450
spectrophotometer and fluorescence spectra was recorded on a
Hitachi F-7000 spectrometer. The fluorescence quantum yields
were measured at 20 °C with coumarin 151 as the reference (Фfl
= 0.53 in ethanol).36 Fluorescence imaging was performed with
an Olympus IX83 inverted microscope. The pH measurement
was carried out on a Leici PHS-3C meter. TLC silica gel plates
and silica gel (mesh 200-300) for column chromatography were
purchased from Qingdao Ocean Chemicals, China. All chemicals
and solvents were obtained from commercial suppliers and used
as received. HNE-2 cells were provided by Xiangya Third
People’s Hospital of Central South University (China).
Next, we evaluated the ability of Probe 1 to perform within
living cells. Living HNE-2 cells, nasopharyngeal carcinoma cell
lines, only incubated with Probe 1 showed little fluorescence, as
depicted in Fig. 9a. However, cells pretreated with thiophenol
and then stained with Probe 1 resulted in a strong green
fluorescence (shown in Fig. 9b). These results indicate that Probe
1 is cell membrane permeable and can response to thiophenol in
living cells.
4.2. Synthesis
4.2.1. Synthesis of compound 2
To a solution of 3-nitrophthalic anhydride (1.9361 g, 10.0
mmol) in acetic acid (20.0 mL) was slowly added butylamine
(1.0951 g, 15.0 mmol) within 5 min. After stirring at room
temperature for 10 min, the resulting mixture was refluxed at 120
°C for 2.5 h. Next, cool the reaction mixture to room temperature
and pour into 50.0 mL cold water. The precipitated solid was
collected and washed with water (3.0 mL × 3) to give compound
1
2 as a white solid (2.1252 g, 85.7% yield). H NMR (500 MHz,
DMSO-d6, TMS) δH 8.27 (d, J = 8.1 Hz, 1H), 8.16 (d, J = 7.5 Hz,
1H), 8.05 (t, J = 7.8 Hz, 1H), 3.57 (t, 2H), 1.63-1.51 (m, 2H),
1.38-1.24 (m, 2H), 0.90 (t, 3H). 13C NMR (125 MHz, DMSO-d6)
δC 166.5, 163.8, 144.6, 136.5, 134.1, 128.6, 127.2, 123.5, 38.1,
30.2, 19.9, 13.9.
Fig. 9 Living-cell fluorescence images of thiophenol in living HNE-2 cells. (a)
Fluorescence and (c) bright field images of cells incubated with Probe 1 (5.0
µM) for 30 min at 37 °C. (b) Fluorescence and (d) bright-field images of cells
pretreated with thiophenol (25.0 µM) and then incubated with Probe 1 (5.0
µM) for 30 min at 37 °C. The scale bars mean 100 µm. The cells were
magnified 20 times.
4.2.2. Synthesis of compound 3
Compound 2 (0.4964 g, 2.0 mmol) was hydrogenated in
methanol (15.0 mL) under reflux at 65 °C for 12 h with 10%
Pd/C (0.0496 g) as a catalyst. Then, the reaction mixture was
filtered through celite to remove the catalyst. Next, the filtrate
was concentrated in vacuum to give a residue. Pure compound 3
(0.6240 g, 71.5% yield) was obtained by silica gel column
chromatography using petroleum ether/dichloromethane (v/v,
1:1) as eluent. 1H NMR (500 MHz, CDCl3, TMS) δH 7.42 (dd, J =
8.3, 7.1 Hz, 1H), 7.16 (d, J = 7.6 Hz, 1H), 6.86 (d, J = 8.8 Hz,
1H), 5.13 (s, 1H), 3.65 (t, 2H), 1.63-1.70 (m, 2H), 1.35-1.42 (m ,
2H). 0.95 (t, 3H). 13C NMR (125 MHz, CDCl3) δC 170.4, 168.8,
145.2, 135.02, 132.9, 121, 112.6, 111.4, 37.4, 30.8, 20.1, 13.7.
3. Conclusions
In conclusion, we presented the design, synthesis and
preliminary evaluation of the 2,4-dinitrophenyl ether of 3-
hydroxyphthalimide derivative, Probe 1, as a turn-on fluorescent
probe for the detection of thiophenol in both the real water
sample and living cells. This probe exhibited a good selectivity
toward thiophenol against aliphatic thiols and other biologically
relevant anions and metal ions and high sensitivity with a 3.5 nM
detection limit. This probe featured a 64-fold fluorescence
enhancement with a 161 nm large Stokes shift in response to
thiophenol.
4. Experimental section
4.2.3. Synthesis of compound 4
4.1. Instruments and Materials

5
To a solution of compound 3 (0.2180 g, 1.0 mmol) in 50%
Acknowledgements
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sulfuric acid (10.0 mL) at 0 °C was added a solution of NaNO2
(0.0692 g，1.0 mmol) in 2.0 mL water dropwise. After stirring
for 30 min at 0 °C, the mixture was heated to 90 °C and stirred
for 1 h. The reaction mixture was diluted with 15.0 mL water,
then extracted with ethyl acetate (20.0 mL × 3). The organic
layer was combined, washed with brine and dried over anhydrous
Na2SO4. After removal of the solvent, the solid was purified by
The research was supported by the State Key Laboratory of
Fine Chemicals (KF1202) and Hunan Nonferrous Metals
Holding Group Co. Ltd.
References and notes
†
Electronic supplementary information (ESI) available. See DOI:
10.1016/j.tet.2015.xx.xxx.
silica
gel
flash
chromatography
using
petroleum
ether/dichloromethane (v/v, 1:1) as eluent to afford compound 4
(0.1862 g, 85.4% yield). H NMR (500 MHz, CDCl3, TMS) δH
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7.70 (s, 1H), 7.56 (t, J = 7.6 Hz, 1H), 7.36 (d, J = 7.2 Hz, 1H),
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32. Liu, X.; Gao, L.; Yang, L.; Zou, L.; Chen, W.; Song, X. RSC Adv.,
2015, 5, 18177.
33. Lin, W.; Long, L.; Tan, W. Chem. Commun., 2010, 46, 1503.
34. Chen, S.; Hou, P.; Zhou, B.; Song, X.; Wu, J.; Zhang, H.; Foley, J. W.
RSC Adv., 2013, 3, 11543.
35. Lan, M.; Wu, J.; Liu, W.; Zhang, H.; Zhang, W.; Zhuang, X.; Wang, P.
Sens. Actuators B, 2011, 156, 332.
4.2.5. Synthesis of the reaction product of Probe 1
with thiophenol
To a solution of Probe 1 (0.0385 g, 0.1 mmol) in a mixture of
acetonitrile (2.00 mL) and distilled water (10.0 mL) was added
thiophenol (0.0552 g, 0.5 mmol) at room temperature. Stir the
reaction mixture for 4 h and extract with dichloromethane (20
mL × 3). The organic layer was combined and dried over
anhydrous Na2SO4. After removal of the solvent, the solid was
purified by silica gel flash chromatography using petroleum
ether/dichloromethane (v/v, 2:1) as eluent to afford the target
1
product (0.0142 g, 52.3% yield). H NMR (500 MHz, CDCl3,
TMS) δ 7.66 (s, 1H), 7.59 (t, J = 7.6, 7.3 Hz, 1H), 7.39 (d, J = 7.2
Hz, 1H), 7.17 (d, J = 8.4 Hz, 1H), 3.67 (t, J = 7.3 Hz, 1H), 1.70-
1.64 (m, 7.6 Hz, 1H), 1.35-1.41 (m, 2H), 0.97 (t, 3H).
4.3. Imaging of HNE-2 cells
HNE-2 cells were seeded in a 6-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.
The cells were washed three times with PBS buffer before used.
The cells were firstly incubated with thiophenol (25.0 µM) for 30
min at 37 °C, washed with PBS buffer and then incubated with
Probe 1 (5.0 µM) for another 30 min. After washing with PBS
buffer, cell imaging were performed. For a control experiment,
HNE-2 cells were only incubated with Probe 1 (5.0 µM) for 30
min at 37 °C.
36. Reynolds, G. A.; Drexhage, K. H. Opt. Commun., 1975, 13, 222.

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Tetrahedron
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Supporting Information
A phthalimide-based fluorescent probe for thiophenol
detection in water samples and living cells with a large
Stokes shift
a
a
a
a
a
Xingjiang Liu , Liu Yang , Li Gao , Wenqiang Chen , Fengpei Qi and Xiangzhi
Song abc,
*
a College of Chemistry & Chemical Engineering, Central South University, Changsha,
Hunan Province, P. R. China, 410083.
b State Key Laboratory for Powder Metallurgy, Central South University, Changsha,
Hunan province, P. R. China, 410083.
c State Key Laboratory of Fine Chemicals, Dalian University of Technology, Dalian,
Liaoning Province, P. R. China, 116024.
* Corresponding author. Fax: +86-731-88836954; Tel: +86-731-88836954; Email:
xzsong@csu.edu.cn (XZ Song).
Contents
Fig. S1............................................................................................................................2
Table S1.........................................................................................................................2
Fig. S2............................................................................................................................3
Fig. S3............................................................................................................................3
Fig. S4............................................................................................................................4
Fig. S5............................................................................................................................4
Fig. S6............................................................................................................................5
Fig. S7............................................................................................................................5
Fig. S8............................................................................................................................6
Fig. S9............................................................................................................................6
Fig. S10..........................................................................................................................7
Fig. S11..........................................................................................................................8

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Fig. S1 Absorption spectra of Probe 1 (5.0 µM) upon the addition of thiophenol
(0.0-5.0 equiv.) in HEPES buffer (10.0 mM, pH = 7.4, containing 20% CH3CN).
Table S1 Determination of thiophenol concentration in water samples.
Thiophenol spiked
Thiophenol recovered
Sample
recovery (%)
(µM)
(µM)
0
1
not detected
0.99 ± 0.02
99
2
3
103
108
103
106
103
103
102
101
99
2.06 ± 0.04
3.24 ± 0.08
4.10 ± 0.09
5.29 ± 0.11
6.23 ± 0.15
7.18 ± 0.14
8.12 ± 0.21
9.13 ± 0.16
9.91 ± 0.11
not detected
0.92 ± 0.02
1.83 ± 0.03
3.04 ± 0.02
4.01 ± 0.05
4.94 ± 0.09
5.84 ± 0.11
6.76 ± 0.13
7.63 ± 0.11
8.29 ± 0.17
9.07 ± 0.15
4
5
Yangtze River water
6
7
8
9
10
0
1
92
92
2
3
101
100
99
4
5
Tap water
6
97
7
97
8
95
92
9
91
10

3
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Fig. S2 1H NMR spectrum of compound 2.
Fig. S3 13C NMR spectrum of compound 2.

4
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Fig. S4 1H NMR spectrum of compound 3.
Fig. S5 13C NMR spectrum of compound 3.

5
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Fig. S6 1H NMR spectrum of compound 4.
Fig. S7 13C NMR spectrum of compound 4.

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Fig. S8 1H NMR spectrum of Probe 1.
Fig. S9 13C NMR spectrum of Probe 1.

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Fig. S10 HRMS spectrum of Probe 1.

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Fig. S11 HRMS spectrum of the reaction product of Probe 1 with thiophenol, dye 4.