D-PET coupled ESIPT phenomenon for fluorescent turn-on detection of hydrogen sulfide

Article abstract of DOI:10.1039/c3ra42499e

A d-PET coupled ESIPT based probe has been designed and synthesized for the selective detection of H₂S, among the other ions and various sulfur species such as cysteine and glutathione, in aqueous media via the hydrogen sulfide induced reduction of nitro functionalities, and has further been utilized for the imaging of H₂S in intracellular systems. The Royal Society of Chemistry.

Full text of DOI:10.1039/c3ra42499e

View Article Online
View Journal
RSC Advances
Accepted Manuscript
This article can be cited before page numbers have been issued, to do this please use: S. I. Reja, N. Kumar, R. Sachdeva, V.
Bhalla and M. Kumar, RSC Adv., 2013, DOI: 10.1039/C3RA42499E.
This is an Accepted Manuscript, which has been through the RSC Publishing peer
review process and has been accepted for publication.
Accepted Manuscripts are published online shortly after acceptance, which is prior
to technical editing, formatting and proof reading. This free service from RSC
Publishing allows authors to make their results available to the community, in
citable form, before publication of the edited article. This Accepted Manuscript will
be replaced by the edited and formatted Advance Article as soon as this is available.
To cite this manuscript please use its permanent Digital Object Identifier (DOI®),
which is identical for all formats of publication.
More information about Accepted Manuscripts can be found in the
Information for Authors.
Please note that technical editing may introduce minor changes to the text and/or
graphics contained in the manuscript submitted by the author(s) which may alter
content, and that the standard Terms & Conditions and the ethical guidelines
that apply to the journal are still applicable. In no event shall the RSC be held
responsible for any errors or omissions in these Accepted Manuscript manuscripts or
any consequences arising from the use of any information contained in them.
www.rsc.org/advances
Registered Charity Number 207890

Page 1 of 6
RSC Advances
Dynamic Article Links ►
Journal Name
View Article Online
DOI: 10.1039/C3RA42499E
Cite this: DOI: 10.1039/c0xx00000x
www.rsc.org/xxxxxx
ARTICLE TYPE
dꢀPET coupled ESIPT phenomenon for fluorescent turnꢀon detection of
hydrogen sulfide
Shahi Imam Reja, Naresh Kumar, Roopali Sachdeva, Vandana Bhalla and Manoj Kumar*
Received (in XXX, XXX) Xth XXXXXXXXX 20XX, Accepted Xth XXXXXXXXX 20XX
5
DOI: 10.1039/b000000x
A dꢀPET coupled ESIPT based probe has been designed and synthesized for the selective detection of
H S among the other ions and various sulfur species such as cysteine, glutathione in aqueous media via
2
the hydrogen sulfide induced reduction of nitro functionality and have further been utilized for the
imaging of H S in intracellular systems.
2
45
concentrations of biological sulfur species such as glutathione
and cysteine.
1
0
Introduction
Our research work involves the design and synthesis of
fluorescent chemosensors for the detection of soft metal ions and
evaluation of their switching behaviour. Recently, we reported a
Hydrogen sulfide (H S) is an endogenously produced gaseous
2
molecule along with nitric oxide (NO), carbon monoxide (CO)
10
1
which induces unique signaling response in cellular system. The
50
charge transfer assisted fluorescent probe for selective detection
production of H S is enzymatically regulated in mammals which
11
2
of hydrogen peroxide. In continuation of this work, we have
1
2
2
3
3
4
5
0
5
0
5
0
led to increased interest to evaluate the biological as well as
now synthesized compound 2 based on the 2ꢀhydroxynaphthalene
moiety appended with nitrophenylimino functionality as a
physiological role of H S. For example, it acts as a physiological
2
vasodilator, signal transmitter in the brain and in antitumor
fluorescence turnꢀon chemosensor for H S under physiological
55 conditions. Furthermore, biological application of probe 2 is
2
activity of the immune system. Further, H S is considered as
2
highly toxic and hazardous pollutant to the environment besides
its numerous physiological functions. Any imbalance in the H2S
has also been recognized to mediate a wide effects of
physiological processes, such as vasodilation, antiꢀoxidation,
evaluated for in vitro detection of H S in prostate cancer (PC3)
2
cell lines. We envisioned that the fluorescence properties of
compound 2 could be modulated via a photoꢀinduced electron
transfer (PET) process from the excited fluorophore to a strong
1
2
antiꢀapoptosis, antiꢀinflammation and the abnormal H S level was 60 electronꢀwithdrawing group (donorꢀexcited PET or dꢀPET).
2
connected to diseases such as Alzheimer’s and Downs syndrome
Therefore, nitro group, a strong electronꢀwithdrawing moiety was
2
selected as the signal modulator for the probe 2. The presence of
nitro group favours the dꢀPET from the excited fluorophore to the
nitro group responsible for the quenched fluorescence emission in
the aqueous media. The nitro group can be reduced to amino
including carcinogenesis as well as neurodegradation. Thus,
from both physiological and pathological point of view, it is
desirable to develop sensitive, specific and rapid techniques for
the detection of hydrogen sulfide. A number of techniques
including polarography, gas chromatography and colorimetry are
6
5
group under physiological conditions by H S which will then
2
3
trammel the dꢀPET mechanism and thus will result in the
enhancement of fluorescence emission owing to the excited state
available for the detection of gasotransmitters. However, these
techniques require complicated procedures which did not allow
the temporal monitoring and are destructive in nature. However,
the use of fluorescence approach provides unique advantages
13
intramolecular proton transfer (ESIPT) process.
70
Experimental
4
including high sensitivity and simplicity. Moreover fluorescence
imaging is the best technique for the determination and
measurement of intracellular molecules without the destruction of
tissues or cells which makes fluorescence approach superior to
General information
All reagents were purchased from Aldrich and were used without
further purification. Acetonitrile (AR grade) was used to perform
5
other analytical methods. In literature a few fluorescent
analytical studies. UVꢀvis spectra were recorded on
a
chemosensors for H S have been reported which involve H S
2
2
75 SHIMADZU UVꢀ2450 spectrophotometer, with a quartz cuvette
6
0
mediated reduction of azides to amines, nucleophilic addition of
H2S and copper sulfide precipitation approach but, H2S
promoted reduction of nitro group to amino group has not been
(
path length 1 cm). The cell holder was thermostatted at 25 C.
7
8
The fluorescence spectra were recorded with a SHIMADZU 5301
1
13
PC spectrofluorimeter. H and C spectra were recorded on a
9
explored much. Further, a key challenge for the selective
JEOLꢀFT NMRꢀAL 300 MHz spectrophotometer using CDCl as
3
detection of H S within the cellular system is comparatively high 80 a solvent and tetramethylsilane as the internal standard. Data are
2
This journal is © The Royal Society of Chemistry [year]
[journal], [year], [vol], 00–00 | 1

RSC Advances
Page 2 of 6
View Article Online
DOI: 10.1039/C3RA42499E
reported as follows: chemical shift in ppm (d), multiplicity (s = 50 for this compound (see ESI S5ꢀS8†).
singlet, d = doublet, t = triplet, q = quartet, m = multiplet, br =
broad singlet), coupling constants J (Hz), integration and
interpretation.
The photophysical behavior of 2 was studied toward different
analytes such as sulfur species, reactive oxygen species and
2
anions (H S, GSH, Cys, S O , H O , ClO, tꢀBuOOH, Cl, F,
2
2
3
2
2
2
Br, I, N , CN, AcO and CO ) by UVꢀvis and fluorescence
5
UVꢀvis and fluorescence titrations
3
3
5
6
6
7
7
8
8
9
9
5
0
5
0
5
0
5
0
5
spectroscopy. The absorption spectrum of compound 2 (5.0 ꢂM)
UVꢀvis and fluorescence titrations were performed on 5.0 ꢁM
solution of ligand in H O:CH CN (99.5:0.5, v/v) buffered with
in H O:CH CN (99.5:0.5, v/v) buffered with HEPES, pH = 7.4, is
2
3
2
3
characterized by an absorption band at 322 nm (see ESI,† S9)
HEPES, pH = 7.4; λex = 320 nm. Typically, aliquots of freshly
prepared anions (Cl, F, Br, I, CN and AcO as
14
4
ꢀ1
ꢀ1
with an extinction coefficient of∼6.4×10 M cm . However,
the addition of H S (Na S was used as a hydrogen sulfide source
2
2
2
10
tetrabutylammonium salt; CO  as K CO ; N  as NaN ), H S as
3
2
3
3
3
2
in all experiments) results in a slightly blue shifted absorption
ꢀ
1
ꢀ3
Na S; Cysteine and Glutathione standard solutions (10 M to 10
2
M) were added to record the UVꢀvis and fluorescence spectra.
3
0 ꢁM
a b
ꢀ
Hydrogen peroxide (H O ) and hypochlorite (OCl ) were
2
2
delivered from 30% and 5% aqueous solutions, respectively. In
titration experiments, each time a 3 ml solution of ligand was
filled in a quartz cuvette (path length, 1 cm) and spectra were
recorded after the addition of appropriate analyte.
2
H S
15
0 ꢁM
Fig. 1 Fluorescence spectra of 2 (5.0 ꢁM) with H
were recorded immediately after the additions of H
99.5:0.5, v/v) buffered with HEPES, pH = 7.4; λex = 320 nm at 25°C.
2
S (0ꢀ30 ꢁM); spectra
20
2
S in H O:CH CN
2
3
(
band at 314 nm along with the appearance of broad absorption
band at 278 nm. The UVꢀvis behavior of compound 2 in the
Scheme 1 Synthesis of compound 2.
presence of H S is ascribed to the selective reduction of acceptor
2
nitro to donor amino functionality, as a result it prevents the
charge transfer from donor naphthalene to acceptor nitro benzene
moiety leading to blue shift in the absorption band. The addition
of other analytes such as cysteine, ROS and anions did not alter
the absorption spectrum of compound 2 (see ESI,† S9).
Synthesis of compound 2
A mixture of compound 1 (0.1 g) and pꢀnitroaniline (0.08 g) were
dissolved in ethanol and refluxed for 24 h at 80ꢀ90˚C. After the
completion of the reaction, solvent was evaporated and the
residue was crystallized from CHCl /CH OH to give compound 2
25
30
35
3
3
In the fluorescence spectrum, receptor 2 exhibited no
fluorescence emission when excited at 320 nm in H O:CH CN
1
in 70% yield; mp 224 °C. H NMR (CDCl , 300 MHz): δ= 7.14
3
2
3
(
d, 1 H, J = 9 Hz, ArH), 7.42 (t, 1 H, J = 6 Hz, ArH), 7.47 (d, 2
H, J = 9 Hz, ArH), 7.59 (t, 1 H, J = 9 Hz, ArH), 7.78 (d, 1 H, J =
Hz, ArH), 7.89 (d, 1 H, J = 9 Hz, ArH), 8.15 (d, 1 H, J = 6 Hz,
ArH), 8.36 (d, 2 H, J = 9 Hz, ArH), 9.42 (d, 1 H, J = 3 Hz,
(99.5:0.5, v/v) buffered with HEPES, pH = 7.4 (Figure 1).
Although, the phenomenon of excited state intramolecular proton
transfer (ESIPT) could have resulted in the fluorescence emission
of 2 but owing to the photoꢀinduced electron transfer (PET) from
the excited fluorophore to the strong electronꢀwithdrawing group
6
1
3
CH=N), 14.92 (s, 1 H, OH) ppm. C NMR (CDCl , 75 MHz): δ
3
=
111.30, 117.69, 118.91, 120.92, 121.53, 122.39, 124.29,
125.43, 127.27, 128.70, 129.34, 137.66, 145.67, 151.96, 156.97
ppm. ESIꢀMS: m/z Calcd for C H N O Calcd: 293.0921 (M +
2
ꢀ
2
, Cys, GSH, S
, ClO, Cl, F,
Br, I, N3, CN,
2 3
O ,
1
7
12
2
3
H
2
S
2 2
H O
+
+
H) Found: 293.0921 (M+H) .
2
AcO and CO
SO .
3
3

,
2
ꢀ
Results and discussion
Condensation of βꢀhydroxynaphthaldehyde (1) with 4ꢀ
40
nitroaniline in ethanol gave the desired compound 2 in 70% yield
1
(Scheme 1). The H NMR spectrum of compound 2 showed five
doublets (1 H each) at 7.14, 7.78, 7.89, 8.15 and 9.42 ppm ( J = 9,
, 9, 6 and 3 Hz), two triplets (1 H each) at 7.42 and 7.59 (J = 6
6
100
and 9 Hz) ppm, two doublets (2 H each) at 7.47 and 8.36 ppm (J
= 9 Hz each) corresponding to the aromatic protons and imino
proton, one singlet (1 H) at 14.92 ppm corresponding to the –OH
45
Fig. 2 Fluorescence spectra of 2 (5 ꢁM) in the presence of H
2
S (30
+
proton. The molecular ion peak at m/z 293.0921 [M+H ]
ꢁM); spectra were recorded immediately after the additions of H S and
2
corresponds to the condensation product 2 observed in the ESIꢀ
MS spectrum. These spectroscopic data corroborate structure 2
other analyte (30 ꢁM each but in case of glutathione and cysteine
1
05 we used 5mM and 10mM) in H
2 3
O:CH CN (99.5:0.5, v/v) buffered
with HEPES, pH = 7.4; λex = 320 nm. .
2
| Journal Name, [year], [vol], 00–00
This journal is © The Royal Society of Chemistry [year]

Page 3 of 6
RSC Advances
View Article Online
DOI: 10.1039/C3RA42499E
aqueous media, we also carried out the fluorescence studies of
−
probe 2 toward other biologically relevant species (H O , ClO ,
2
2
−
2−
TBHP, N , S O , Cys and GSH). As shown in Figure 4, no
3
2
3
60
significant change was observed with other species in comparison
to the hydrogen sulfide. The time dependent fluorescence study
indicates that the 35 fold fluorescence enhancement was obtained
5
in 20 minutes after the addition of H S (Figure 4). Interestingly,
2
the emission enhancement was observed immediately after the
Fig. 3 Schematic diagram of dꢀPET mechanism
6
5
addition of H S which in turn indicates the highly reactive nature
2
1
5
1
1
2
2
3
3
4
4
5
5
0
5
0
5
0
5
0
5
0
5
(donorꢀexcited PET or dꢀPET) makes it nonꢀfluorescence. In
general, nitro substituted ring has lower LUMO energy level due
of probe 2 towards H S.
2
40
30
20
1
5b
to its electronꢀwithdrawing effect.
Thus, intramolecular
electron transfer occurs from excited fluorophore to to the
1
6
electron deficient benzene moiety. Upon addition of increasing
70
2
5
min
min
10 min
20 min
amounts of only H S to the aqueous solution of 2, a remarkable
2
enhancement in emission intensity was observed at 462 nm
(
Figure 1) along with the appearance of a blue coloured
fluorescence emission (inset of Figure 1) due to conversion of
nitro functionality to amino functionality which trammels the dꢀ
PET mechanism owing to higher LUMO energy level (Figure 3).
The fluorescence quantum yield (Φ ) of the compound 3 is found
to 0.194 as compared to that of receptor 2 (0.005) at 462 nm,
10
75
0
s
ꢀ
ꢀ
ꢀ
Na
Fig. 4 Fluorescence response of 2 (5 ꢁM) in H
buffered with HEPES, pH = 7.4 (λex= 320 nm) with various analytes (30
M each but in case of glutathione and cysteine we used 5mM
and 10mM). Bars represent selectivity (I/I ) (I = initial fluorescence
intensity at 462 nm; I = final fluorescence intensity at 462 nm after the
addition of analyte) of 2 upon addition of different analytes. Data were
given after incubation with the appropriate analyte at 25°C after 2, 5, 10
and 20 minutes.
Next, we investigated the effect of pH on the recognition
behavior of probe 2. At the acidic pH conditions (pH = 5.0 or 6.0)
2
S GSH Cys
H
2
O
2
TBHP N
3
S
2
O
3
OCl
2
O:CH
3
CN (99.5:0.5, v/v)
which shows good agreement with fluorescence spectra obtained
8
8
9
9
0
5
0
5
ꢁ
0
o
probe 2 exhibits low reactivity towards H S and hence we
2
observed small increase in the fluorescence emission (Figure 5).
However, as we increase the pH = 7.0, the enhancement in the
fluorescence emission was fast (Figure 5). On further increase of
the pH (pH = 8.0), more fast emission enhancement is observed.
This fast emission enhancement is due to the existance of H S as
HS under basic conditions which has more reducing power.
Further, we also investigated the effect of temperature on the
fluorescence behaviour and observed that at higher temperature
the reduction of nitro group to amino group is faster in
comparision to reduction at lower temperature conditions (see
ESI,† S12). We also tested the fluorescence response of
2
1
7
2
Scheme 2. (A) H Sꢀinduced reduction of 2; (B) Fast enol–imine to
keto–amine tautomerism in 3.
for receptor 2 in the presence of H S. Fluorescence enhancement
observed for receptor 2 upon addition of H S is attributed to the
reduction of nitro to amino group (Scheme 2A) which is 100
responsible for the inhibition of dꢀPET and hence fluorescence
turnꢀon behaviour due to the excited state intramolecular proton
transfer mechanism (Scheme 2B).
2
2
9
3
2
1
0 ꢁM
0 ꢁM
0 ꢁM
Thus, probe 2 acts as efficient platform for the fluorescence
turnꢀon detection of H S in aqueous environment. In the mass 105
2
spectrum peak at m/z 263.1173 (see ESI,† S10) confirmed the
formation of 3 which clearly indicates the reduction of nitro to
amino group in the presence of H S. As the reduction by H S
results in the formation of compound 3 containing an electronꢀ
donating group instead of an electronꢀwithdrawing group, the 110 Fig. 5 Fluorescence response of 2 (5 ꢁM) in H O:CH CN (99.5:0.5,
pH 8.0 pH 7.0 pH 6.0 pH 5.0
2
2
2
3
v/v) buffered with HEPES in different pH value (λex= 320 nm) to
addition of different concentration of H S. Data were given after
incubation with H S at 25 °C after 15 minutes.
fluorescence emission also underwent around 12 nm blue shift
4
e
2
(see ESI,† S11). Under the same conditions as used for H S in
2
2
This journal is © The Royal Society of Chemistry [year]
Journal Name, [year], [vol], 00–00 | 3

RSC Advances
Page 4 of 6
View Article Online
DOI: 10.1039/C3RA42499E
receptor 2 towards other analytes (Cl, F, Br, I, N , CN,
(5.0 ꢂM) show fluorescence in blue channel (Figure 7d). The
AcO, CO  and SO ꢃ). No significant change in fluorescence 60 appearance of blue fluorescence is attributed to the amino form of
3
2
2
3
3
occurred in the presence of any other analyte (Figure 2). Further,
we investigated the practical ability of receptor 2 as a H2S
selective fluorescent chemosensor by carrying out competitive
the receptor 2 produced by reduction of nitro functionality of
receptor 2 in presence of H S. These results suggest that receptor
2
2
is cell permeable and an effective intracellular H S imaging
2
5
0
5
agent with turnꢀon blue coloured fluorescence emissions.
experiments in the presence of H S mixed with other analytes
2
(
see ESI,† S13). No significant variation in fluorescence emission
6
5
Conclusions
was observed in comparison with or without the any other
analyte. Thus, 2 acts as efficient fluorescence turnꢀon probe for
the detection of H S in aqueous media. The detection limit of 10
2
×
In conclusion, we developed a reaction based fluorescent
chemosensor 2 which undergoes fluorescence enhancement in the
presence of only H S among the various sulfur, reactive oxygen
species and anions in aqueous media and living cells. The sensing
mechanism is based on the selective reduction of nitro group to
1
ꢀ
7
ꢀ1
10 mol L (see ESI,† S14) of probe 2 for H S suggests this
2
2
system as efficient sensing platform for H S.
2
70
amino moiety under physiological conditions by H S. The
2
a
b
1
H
2
S
addition of increasing amounts of H
receptor 2 resulted in a remarkable enhancement in emission
intensity along with the appearance of blue coloured
2
S to the aqueous solution of
a
75
fluorescence emission. Confocal microscopy studies indicate that
Fig. 6 Change in the fluorescence of 2 in the presence of H
2
S. (a) Testꢀ
our probe can detect the changes of H S level in living cells. We
2
−
4
strips of 2;(b) After adding 1 drop of H
2
S solution (10 M in H O) onto
2
envision that the fluorescent probe will be of great benefit for
detection of H S to investigate the effects of H S in biological
20
25
30
35
40
45
50
55
the testꢀstrips of 2. All images were taken under 365 nm UV
illumination.
2
2
systems.
In order to work as an efficient sensing system, a probe
molecule must have some practical applications. Therefore, we
introduced for the first time a fast and easiest method for the
8
0
Acknowledgements
MK and VB acknowledge the financial support from DST,
CSIR and UGC, New Delhi. S.I.R. & N.K. acknowledges UGC
and CSIR for JRF/SRF fellowships. We are also thankful to Dr.
Gurcharan Kaur and Shaffi Manchanda, Department of
Biotechnology, Guru Nanak Dev University, Amritsar for
biological studies.
detection of H S. We prepared testꢀstrips (see ESI,† S15) coated
2
with probe 2 which showed change in fluorescence after treating
with H S. The fluorescence enhancement was observed upon
2
ꢀ
4
dropping the saturated aqueous solution of Na S (10 M) onto the
2
85
test strips (Figure 6). These results show the practical
applicability of probe 2 toward the instant visualization of traces
of H S.
2
Notes and references
The potential biological application of the chemosensor 2 was
Department of Chemistry, UGC Sponsored Centre for Advanced Studies-
evaluated for in vitro detection of H S in prostate cancer (PC3)
2
1
, Guru Nanak Dev University, Amritsar, Punjab, India. Fax: +91
cell lines. The prostate cancer (PC3) cell lines were incubated
with receptor 2 (5.0 ꢂM) in an RPMIꢀ1640 medium for 20 min at
9
9
0
5
(0)183 2258820; Tel: +91 (0)183 2258802 9 ext. 3205;
E-mail: mksharmaa@yahoo.co.in
3
7°C and washed with phosphate buffered saline (PBS) buffer
† Electronic Supplementary Information (ESI) available: [Experimental
1
13
(pH 7.4) to remove excess receptor 2. Microscopic images
details, H NMR, CNMR, mass spectra, UVꢀvis and fluorescence
spectra]. See DOI: 10.1039/b000000x/
showed no intracellular fluorescence which indicated that
receptor 2 is nonꢀemissive in nature (Figure 7a). However, after
1
(a) K. R. Olson and J. A. Donald, Acta Histochem., 2009, 111, 244;
b) R. Schulz, M. Kelm and G. Heusch, Cardiovasc. Res., 2004, 61,
(
treatment with H S (5.0 ꢂM), the cells preꢀtreated with receptor 2
2
402; (c) R. Hosoki, N. Matsuki and H. Kimura, Biochem. Biophys.
Res., Commun., 1997, 237, 527; (d) N. Kumar, V. Bhalla and M.
Kumar, Coord. Chem. Rev., 2013, 257, 2335.
Blue Channel
Overlay
Brightfield
1
1
1
00
05
10
2
3
G. C. Brown and A. BalꢀPrice, Mol. Neurobiol., 2003, 27, 325.
(a) L. M. Siegel, Anal. Biochem., 1965, 11, 126; (b) J. E. Doeller, T.
S. Isbell, G. Benavides, J. Koenitzer, H. Patel, R. P. Patel, L. J. R. Jr,
V. M. DarleyꢀUsmar and D. W. Kraus, Anal. Biochem., 2005, 341,
a
b
c
4
0; (c) J. Furne, A. Saeed and M. D. Levitt, Am. J. Physiol. Regul.
Integr. Comp. Physiol., 2008, 295, 1479.
4
(a) A. R. Lippert, E. J. New and C. J. Chang, J. Am. Chem. Soc.,
2
011, 133, 10078; (b) Y. Qian, J. Karpus, O. Kabil, S. Y. Zhang, H.
L. Zhu, R. Banerjee, J. Zhao and C. He, Nat. Commun., 2011, 2,
495; (c) Peng, H.; Y. Cheng, C. Dai, A. L. King, B. L. Predmore, D.
J. Lefer and B. Wang, Angew Chem. Int. Ed., 2011, 50, 9672; (d) K.
Sasakura, K. Hanaoka, N. Shibuya, Y. Mikami, Y. Kimura, T.
Komatsu, T. Ueno, T. Terai, H. Kimura and T. Nagano, J. Am. Chem.
Soc., 2011, 133, 18003; (e) R. Wang, F. Yu, L. Chen, H. Chen, L.
Wang and W. Zhang, Chem. Commun., 2012, 48, 11757; (f) Z. Xu, L.
Xu, J. Zhou, Y. Xu, W. Zhu and X. Qian, Chem. Commun., 2012, 48,
10871; (g) C. Liu, B. Peng, S. Li, C.ꢀM. Park, A. R. Whorton and M.
Xian, Org. Lett., 2012, 14, 2184.
d
e
f
Fig. 7 Fluorescence and brightfield images of PC3 cell lines. (a)
Fluorescence image of cells in blue channel treated with probe 2 (5 ꢂM)
for 20 min at 37 °C. (b) Overlay image of (a) and (c). (c) Brightfield
image of (a). (d) Fluorescence images of cells in blue channel upon
treatment with probe 2 (5 ꢂM) and then Na
2
S (5 ꢂM) for 10 min at 37
115
°C. (e) Overlay image of (d) and (f). (f) Brightfield image of (d). λex =
4
05 nm; fluorescence images are recorded at blue channel.
4
| Journal Name, [year], [vol], 00–00
This journal is © The Royal Society of Chemistry [year]

Page 5 of 6
RSC Advances
View Article Online
DOI: 10.1039/C3RA42499E
5
(a) M. H. Lee, J. H. Han, P.ꢀS. Kwon, S. Bhuniya, J. Y. Kim, J. L.
Sessler, C. Kang and J. S. Kim, J. Am. Chem. Soc., 2012, 134, 1316;
b) S.ꢀK. Ko, X. Chen, Y. Yoon and I. Shin, Chem. Soc. Rev., 2011,
0, 2120.
(a) K. Zheng, W. Lin and L. Tan, Org. Biomol. Chem., 2012, 10,
683; (b) F. Yu, P. Li, P. Song, B. Wang, J. Zhao and K. Han, Chem.
(
4
5
6
7
9
Commun., 2012, 48, 2852.
(a) Y. Zhao, X. Zhu, H. Kan, W. Wang, B. Zhu, B. Du and X. Zhang,
Analyst, 2012, 137, 5576; (b) Y. Qian, L. Zhang, S. Ding, X. Deng,
C. He, X. E. Zheng, H. L. Zhu and J. Zhao, Chem. Sci., 2012, 3,
1
1
2
2
3
3
4
0
5
0
5
0
5
0
2
920; (c) C. Liu, J. Pan, S. Li, Y. Zhao, L. Y. Wu, C. E. Berkman, A.
R. Whorton and M. Xian, Angew. Chem. Int. Ed., 2011, 50, 10327;
d) Y. Chen, C. Zhu, Z. Yang, J. Chen, Y. He, Y. Jiao, W. He, L.
(
Qiu, J. Cen and Z. Guo, Angew. Chem. Int. Ed., 2013, 52, 1688; (e)
J. Liu, Y. Q. Sun, J. Zhang, T. Yang, J. Cao, L. Zhang and W. Guo,
Chem. Eur. J., 2013, 19, 4717; (f) T. Liu, Z. Xu, D. R. Spring and J.
Cui, Org. Lett., 2013, 15, 2310.
F. Hou, J. Cheng, P. Xi, F. Chen, L. Huang, G. Xie, Y. Shi, H. Liu,
D. Bai and Z. Zeng, Dalton Trans., 2012, 41, 5799; (b) J. Wang, L.
Long, D. Xie and Y. Zhan, J. Luminesc., 2013, 139, 40.
8
9
1
L. A. Montoya and M. D. Pluth, Chem. Commun., 2012, 48, 4767.
(a) M. Kumar, S. I. Reja and V. Bhalla, Org. Lett., 2012, 14, 6084;
0
(b) M. Kumar, N. Kumar and V. Bhalla, Chem. Commun., 2013, 49,
8
77.
11 M. Kumar, N. Kumar, V. Bhalla, P. R. Sharma and Y. Qurishi, Chem.
Commun., 2012, 48, 4719.
1
2
X. Zhang, L. Chi, S. Ji, Y. Wu, P. Song, K. Han, H. Guo, T. D. James
and J. Zhao, J. Am. Chem. Soc., 2009, 131, 17452.
(a) P. T. Chou, J. H. Liao, C. Y. Wei, C. Y. Yang, W. S. Yu and Y.
H. Chou, J. Am. Chem. Soc., 2000, 112, 986; (b) V. Bhalla, Roopa
and M. Kumar, Org. Lett., 2012, 14, 2802.
(a) C. C. Hsieh, Y. M. Cheng, C. J. Hsu, K. Y. Chen and P. T. Chou,
J. Phys. Chem. A, 2008, 112, 8323; (b) G. J. Zhao, J. Y. Liu, L. C.
Zhou and K. L. Han, J. Phys. Chem. B, 2007, 111, 8940.
1
3
1
4
15 (a) X. Zhang, Y. Wu, S. Ji, H. Guo, P. Song, K. Han, W. Wu, W.
Wu, T. D. James and J. Zhao, J. Org. Chem., 2010, 75, 2578; (b) T.
Ueno, Y. Urano, H. Kojima and T. Nagano, J. Am. Chem. Soc., 2006,
1
28, 10640; (c) X. Zhang, L. Chi, S. Ji, Y. Wu, P. Song, K. Han, H.
Guo, T. D. James and J. Zhao, J. Am. Chem. Soc., 2009, 131, 17452.
16 T. Ueno, Y. Urano, K. Setsukinai, H. Takakusa, H. Kojima, K.
Kikuchi, K. Ohkubo, S. Fukuzumi and T. Nagano, J. Am. Chem.
Soc., 2004, 126, 14079.
1
7 O. Kabil and R. Banerjee, J. Biol. Chem., 2010, 285, 21903.
This journal is © The Royal Society of Chemistry [year]
Journal Name, [year], [vol], 00–00 | 5

RSC Advances
Page 6 of 6
View Article Online
DOI: 10.1039/C3RA42499E
Graphical Abstract
A d-PET coupled ESIPT based probe has been designed and synthesized for the selective
detection of H S among the other ions and various sulfur species such as cysteine, glutathione in
2
aqueous media via the hydrogen sulfide induced reduction of nitro functionality and have further
been utilized for the imaging of H S in intracellular systems.
2