A small-molecule probe for ratiometric photoacoustic imaging of hydrogen sulfide in living mice

Article abstract of DOI:10.1039/c9cc02224d

A small-molecule photoacoustic probe based on cyanine dyes was developed by taking advantage of the nucleophilic substitution reaction of H₂S with chlorine. The probe demonstrated specific response to H₂S with ratiometric photoacoustic signals in the NIR region, which enabled real-time, accurate, high-resolution imaging of endogenous H₂S in vivo.

Full text of DOI:10.1039/c9cc02224d

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DOI: 10.1039/C9CC02224D.
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A small-molecule probe for ratiometric photoacoustic imaging of
hydrogen sulfide in living mice
Received 00th January 20xx,
Accepted 00th January 20xx
Xiang Li,a Yufu Tang,a Jie Li,a Xiaoming Hu,a Chao Yin,a Zhen Yang,a Qi Wang,a Zizi Wu,b Xiaomei
Lu,*b Wenjun Wang, Wei Huang ab and Quli Fan*a
DOI: 10.1039/x0xx00000x
www.rsc.org/
A small-molecule photoacustic probe based on cyanine dyes was short tissue penetration depth and strong photon scattering,
developed by taking advantage of the nucleophilic substitution which limit precise measurement of H2S in deep tissues. Thus,
reaction of H2S with chlorine. The probe demonstrated specific ratiometric probes with new imaging modality should be
response to H2S with ratiometric photoacustic signals in the NIR developed for accurate detection of H2S in the living body.
region, which enabled real-time, accurate, high-resolution imaging
of endogenous H2S in vivo.
An effective approach to address the aforementioned issues is
photoacoustic (PA) imaging, which converts optical excitation
into ultrasonic signal and avoids the influence of optical
scattering in principle.8 PA inherits the advantages of optical
imaging, such as high sensitivity and non-invasiveness, and
provides deep tissue penetration and excellent spatial
resolution.9 Given these extraordinary properties, ratiometric
PA probes with NIR excitation have been successfully
developed for accurate and high-resolution in vivo imaging of
biological targets, including reactive oxygen (ROS),10 metal
ions,11 Glutathione (GSH),12 and pH.13 However, the
development of ratiometric PA probes for H2S remains a huge
challenge because the ratio signal of two NIR absorption bands
is difficult to construct at the molecular level. At present, most
ratiometric PA probes are made of nanocomposites that
integrate two different contrast agents to set an internal
standard. Such multi-component systems experience leakage,
which significantly affects the stability of the ratio signal.
Moreover, most PA probes reported are highly hydrophobic
and require to be encapsulated with biocompatible polymers.
The increased size of the probes largely retains them in the
reticuloendothelial system. FDA-approved NIR contrast agents,
namely, methylene blue and indocyanine green (ICG), are small
molecules that can be rapidly excreted. Thence, these probes
are unsuitable for clinical application due to their potential
long-term toxicity. Therefore, the development of small-
molecule PA probes with ratiometric H2S response capability is
of great significance for accurate monitoring of H2S in vivo and
clinical translation of PA imaging.
In the last few decades, hydrogen sulfide (H2S) has been
recognized as a vital gaseous signaling molecule in living
organisms. Biologically relevant H2S is enzymatically
generated at micromolar levels from cysteine (Cys).1 H2S exerts
multifaceted regulatory functions in living systems, such as
maintaining redox homeostasis, modulating neurotransmission,
and stimulating vasodilation.2 Abnormal H2S production is
believed to be closely associated with diseases, such as
Parkinson’s disease, Down syndrome, and diabetes.3 Such
features make H2S a potential target for real-time imaging of
pathological and physiological conditions in the living body.1b
To date, various H2S responsive probes have been developed
based on different trigger mechanisms, including reduction of
azido or hydroxyamino,5 reaction with electrophilic moieties,5
and bonding to Cu(II) ion.6 Among them, ratiometric probes are
promising for reliable in vivo imaging because of the built-in
self-calibration system, which can eliminate interference from
the internal environment, such as heterogeneous probe
biodistribution and excitation intensity.7 However, these probes
are primarily based on fluorescence imaging modality and have
a. Key Laboratory for Organic Electronics and Information Displays & Jiangsu Key
Laboratory for Biosensors, Institute of Advanced Materials (IAM), Jiangsu
National Synergetic Innovation Center for Advanced Materials (SICAM), Nanjing
University of Posts & Telecommunications, Nanjing 210023, China. E-mail:
iamqlfan@njupt.edu.cn
b. Key Laboratory of Flexible Electronics & Institute of Advanced Materials,
Jiangsu National Synergetic Innovation Center for Advanced Materials, Nanjing
Tech University (NanjingTech), Nanjing 211816, China
c. Key Lab of Optical Communication Science and Technology of Shandong
Province & School of Physics Science and Information Engineering, Liaocheng
University, Liaocheng 252059, China
In this study, we developed an ICG-like cyanine dye as
ratiometric PA probe for real-time imaging of H2S in living
mice (Scheme 1a). A ratiometric response strategy was
constructed within a single small molecule based on the
† Electronic Supplementary Information (ESI) available: Experimental details,
NMR spectra etc. See DOI: 10.1039/x0xx00000x
Q.F. and X.L. supervised and conceived the research; X.L. performed experiments; nucleophilic substitution of H2S with chlorine. Nucleophilic
others analyzed the data and wrote the manuscript. All animal experiments were
aromatic substitution (SNAr) of electron-deficient aryl halides is
carried out in accordance with the guidelines of the Laboratory Animal Center of
Jiangsu KeyGEN Biotech Corp., Ltd.
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peak. Finally, the maximum absorption peak shifted to 720 nm
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(Figure S7). After 10 min of incubation,DtOheI: 1r0a.t1i0o39o/fCa9bCsCo0r2p2t2i4oDn
intensity at 720 nm to 800 nm (A720/A800) showed an
enhancement of 122.5-fold. Similar change in the absorption
band was observed for CyCl-2 with a small degree of shift from
775 nm to 710 nm. However, the absorption intensity ratio of
CyCl-2 (A710/A775) only enhanced by 10.2-fold (Figure 1a). The
evolution of the absorption spectra could be attributed to the
intramolecular charge transfer (ICT) of the probes before and
after the substitution of chlorine with electron-donating HS−,15
which caused a visible color change in the probe solutions from
green to blue (Figure S8). The response mechanism was
confirmed through mass spectrometry characterization. After
reaction with excess NaHS, new mass peaks at 776.909 and
791.015 were observed for CyCl-1 and CyCl-2, respectively,
which are ascribed to the [CySH−H+Na]+ species (Figure S9).
These results indicated that H2S can replace the active chlorine
of the probes through nucleophilic substitution under
physiological conditions and lead to ratio changes in the
absorption spectra in the NIR region.
Scheme 1 (a) The illustration of ratiomtetric PA detection of H2S in
vivo with CyCl-1; (b) Structures of CyCl-1 or CyCl-2 and the proposed
mechanism for H2S detection.
proven to be an efficient H2S detection strategy.7a, 14 Cyanine
dyes with active chlorine show strong electron deficiency due
to the indolium group, and it should react with H2S under
physiological conditions. In this regard, two chlorinated
cyanine dyes, namely, CyCl-1 and CyCl-2 (Scheme 1b), were
synthesized for H2S response studies. We envisioned that the
substitution of H2S versus chlorine of the probes would shift
their maximum absorption, which could provide a ratiometric
PA signal for accurate detection of H2S in vivo.
Then, we quantitatively investigated the response kinetics
and sensitivity of CyCl-1 and CyCl-2. The logarithmic value of
CyCl-1 and CyCl-2 were synthesized from cyclopentanone
and cyclohexanone to obtain different cycloolefin conjugated
centers. Two sulfonate-terminated butyl chains were introduced
to provide excellent aqueous solubility. The 1H NMR spectra of
the two probes were similar due to their structural similarity. In
CyCl-1, the peak of -CH2- adjacent to the olefinic bonds
presented as a singlet at 3.03 ppm (Figure S3) because the
protons of two -CH2- in the cyclopentenyl group are
magnetically equivalent. While, in CyCl-2, the coupling effect
from the additional -CH2- in the cyclohexenyl group led to a
triplet at 2.89 ppm (Figure S4). The mass spectrometry analysis
revealed the maximum mass peaks of CyCl-1 and CyCl-2 at
m/z 756.892 and 770. 936 (Figure S5), respectively, which are
ascribed to [M]+ species. These data indicated the successful
synthesis of the probes. The analysis of the photophysical
properties revealed a shift of 25 nm between the maximum
absorption peaks of the two probes. CyCl-1 exhibited a
maximum absorption peak at 800 nm and a shoulder peak at
725 nm, whereas CyCl-2 showed the maximum absorption
peak at 775 nm and shoulder peak at 715 nm. In addition,
CyCl-1 displayed higher absorbance than CyCl-2 (Figure S6).
These conditions were probably due to the better coplanarity of
the penta-cyclic than the hexa-cyclic structure.
Fig. 1 Normalized absorption of CyCl-1 and CyCl-2 (10 μM)
before and after incubation with NaHS (80 μM) for 10 min in
PBS buffer (pH 7.4); (b) The logarithmic value of the absorption
intensity ratios [ln(A720/A800) for CyCl-1 and ln(A710/A775) for
CyCl-2] of the probes as a function of incubation time;
Absorption spectra evolution of CyCl-1 (c) and CyCl-2 (b) upon
the addition of different concentrations of HS− from 0 to 80μM;
The logarithmic value of the absorption intensity ratios of CyCl-
1 (e) and CyCl-2 (f) as a function of HS− concentration.
The response of the probes to H2S was first investigated by
absorption spectroscopy in phosphate- buffered saline (PBS, pH
7.4). The two probes were treated with 80 μM NaHS. The
maximum absorption of CyCl-1 at 800 nm gradually decreased
accompanied by the continuous enhancement of the shoulder
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the absorption intensity ratio [ln(A720/A800) for CyCl-1 or
ln(A710/A775) for CyCl-2] was quantified as a function of
reaction time after treatment with 80 μM NaHS (Figure 1b).
The ratiometric signal of CyCl-1 [ln(A720/A800)] increased
rapidly after treatment with HS− and reached the equilibrium
within 5 min. In comparison, the ratiometric signal of CyCl-2
[ln(A710/A775)] showed slight increase even after 10 min of
incubation. The absorption spectra of the probes showed
gradual evolution over the HS− concentrations (Figures 1c and
d), which permitted the quantitative detection of H2S through
ratiometric absorption. The logarithmic value of the absorption
intensity ratios increased linearly with HS− concentrations for
the two probes (Figures 1e and f). The detection limits for H2S
with CyCl-1 and CyCl-2 were determined to be 0.16 and 0.37
μM, respectively (Figure S10). These results indicated that
CyCl-1 possessed faster response kinetics and higher sensitivity.
In view of the strong NIR absorption and excellent
responsiveness, CyCl-1 was selected as the PA probe for H2S
imaging.
The PA response of CyCl-1 to H2S was first evaluated in the
solution. The PA spectrum of CyCl-1 showed a maximum peak
at 800 nm and a shoulder peak at 730 nm in PBS (pH 7.4). In
the presence of HS−, the PA peak at 800 nm almost disappeared,
and the maximum peak shifted to 720 nm (Figure 2a). The
change in the PA spectrum was similar to the absorption data,
verifying the feasibility of CyCl-1 for ratiometric PA detection
of H2S. PA imaging of H2S in the solution with CyCl-1 was
conducted through excitation at 720 and 800 nm, which were
rendered with pseudo-colors of green and red, respectively
(Figure 2b). With increasing HS− concentration, the PA signal
at 720 nm continuously increased accompanied by a gradual
attenuation of the PA signal at 800 nm. The limit of detection
was caculated as 0.87 μM by using the logarithmic value of the
PA intensity ratio [ln(PA720/PA800)] as the detection signal
(Figures 2c and S11). Hence, the probe can be used for PA
detection of H2S under physiologically relevant concentrations.
We then evaluated the PA response selectivity of the probe
toward H2S. Compared with the significant enhancement of
ratiometric PA signal (PA720/PA800) triggered by H2S, other
analytes, such as metal cations, ROS, reactive nitrogen and
sulfur species, only induced negligible floating (Figure 2d).
Competition assays were conducted with biothiols such as GSH
and Cys. CyCl-1 exhibited potent response to NaHS in the
presence of excess Cys (1 mM) or GSH (5 mM) (Figure S12),
indicating the high response specificity to H2S over Cys and
GSH. This condition could be ascribed to the stronger
nucleophilicity of H2S than thiols.16 CyCl-1 exhibited stable
responsiveness to H2S within the biological pH range of 5.0 to
9.0 (Figure S13). Moreover, no obvious changes in the
maximum absorption of the probe were observed after
incubated in PBS containing 10% fetal bovine serum for 36 h
(Figure S14), reflecting the good stability of the probe in blood.
These data indicated the huge potential of the probe for H2S
detection with minimized interference in biological systems.
Furthermore, MTT assay revealed high cell viabilities under
test concentrations of CyCl-1. The low cytotoxicity of the probe
facilitated its application for PA imaging in vivo.
View Article Online
DOI: 10.1039/C9CC02224D
Fig. 2 (a) PA spectra of CyCl-1 (50 μM) in PBS (pH 7.4) before
(black line) and after (red line) incubation with 400 μM of HS−;
(b) PA images of CyCl-1 solution in the presence of different
concentrations of HS−, excitation at 800 nm (red) and 720 nm
(green); (c) The logarithmic value of PA720/PA800 as a function
of HS− concentration. (d) Ratiometric PA signals of CyCl-1 in the
presence of HS− and other biological analytes.
The in vivo PA imaging of H2S with CyCl-1 was investigated
in a mouse model. NaHS or PBS was subcutaneously injected
into the thigh of living mice. The mice were then injected with
CyCl-1 (50 μM, 10 μL) at the same locations, and the PA
signals of the probe were recorded under excitation at 720 nm
(green) and 800 nm (red). Figure 3 shows the PA images of the
probe-treated regions. The boundary of the injection areas
could be clearly demonstrated in these images, reflecting the
excellent spatial resolution of PA imaging. The PA signal
detected before injection of the probe was derived from the NIR
absorption of hemoglobin in the blood. In order to minimize the
background interference, the PA intensity increment (ΔPA),
defined as the PA intensity after injection of CyCl-1 minus the
PA intensity in the treated area before injection, was used to
analyze the images. The PA increment at 720 (ΔPA720) and 800
nm (ΔPA800) for PBS treated mice significantly increased after
injection of CyCl-1 and showed negligible variations over time
(Figure 3a). While ΔPA800 increased less than ΔPA720 for NaHS
treated mice after injection of CyCl-1 and showed opposite
changes over time (Figure 3b), which suggested the activation
of the probe by HS−. The probe was further used to detect
endogenous H2S in living mice. The mice were pretreated with
Cys (precursor of H2S) for 30 min, then CyCl-1 was injected.
ΔPA720 and ΔPA800 showed the same variation trend as the
mice treated with NaHS, after injection of CyCl-1 (Figure 3c).
The quantitative analysis revealed that ln(ΔPA720/ΔPA800
)
increased over time and reached 1.86 ± 0.12 and 0.93 ± 0.07 at
30 min for the mice treated with NaHS and Cys, respectively,
which are higher than those in the control group (− 0.119 ± 0.04,
Figure 3d). This result demonstrated the capability of CyCl-1
for real-time monitoring of endogenous H2S in vivo with
ratiometric PA imaging. Furthermore, the pharmacokinetics
studies revealed that CyCl-1 mainly accumulated in the liver
after systemic administration (Figure S16a). Fluorescence
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DOI: 10.1039/C9CC02224D
Fig. 3 In vivo PA images of saline (a), NaHS (b) and Cys (c) pretreated regions in the thigh of living mice before (0 min) and after
injection of CyCl-1 for 2, 10 and 30 min. The images were presented with pseudo-colors of green for excitation at 720 nm and
red for 800 nm. (d) The logarithmic value of ΔPA ratio [ln(ΔPA720/ΔPA800)] as a function of post-injection time.
J. Neurosci. Res., 2015, 93, 487; (b) E. Barone, E. Head, D. A.
Butterfield and M. Perluigi, Free Radical Biol. Med., 2016, 111,
S0891584916309972; (c) M. M. Safar and R. M. Abdelsalam,
Pharmacol. Rep., 2015, 67, 17.
intensity of the probe in liver reached the maximum at 3 min
and then rapidly decreased, with a decrease of 80% within 3 h
(Figure S16c). Bio-distribution of the probe was detected in
different organs. 30 min after systemic administration, strong
fluorescence signals were observed in the liver of mice, which
decreased to the same level as other organs at 3h (Figures S16b
and d). Moreover, strong fluorescence was detected from the
feces of mice after 2.5 h. These data indicated the rapid
clearance of CyCl-1 through the hepato-biliary excretion
4. (a) D. Sajal Kumar, L. C. Su, Y. S. Young, H. J. Hee and C. Bong Rae,
Cheminform, 2012, 48, 8395; (b) Q. Wan, Y. Song, Z. Li, X. Gao and H.
Ma, Chem. Commun., 2013, 49, 502.
5. (a) Y. Qian, J. Karpus, O. Kabil, S. Y. Zhang, H. L. Zhu, R. Banerjee, J.
Zhao and C. He, Nat. Commun., 2011, 2, 495; (b) W. Xu, J. Sun, W.
Zhang, X. Ma, J. Lv and T. Bo, Chem. Sci., 2013, 4, 2551.
6. 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.
pathway, which is quite similar to ICG.17
In conclusion, we developed a small-molecule PA probe
(CyCl-1) that exhibited a dual-peak ratiometric PA signal in the
NIR region when exposed to H2S. The probe was constructed
based on water soluble cyanine dyes through the nucleophilic
substitution of H2S to active chlorine, which led to significant
variations in the PA signals of 800 nm (attenuation) and 720
nm (enhancement). Such molecule design endowed the probe
with fast response and high selectivity to H2S. The probe could
be used for ratiometric PA monitoring of H2S in living mice,
and provided high imaging fidelity and rapid clearance. We
believe that the developed small-molecule ratiometric PA probe
can facilitate the application of advanced PA imaging
technology in H2S relevant biomedical and clinical studies.
This work was supported by the National Natural Science
7. (a) C. Zhao, X. Zhang, K. Li, S. Zhu, Z. Guo, L. Zhang, F. Wang, Q. Fei,
S. Luo and P. Shi, J. Am. Chem. Soc., 2015, 137, 8490; (b) X. Li, H.
Zhao, Y. Ji, C. Yin, J. Li, Z. Yang, Y. Tang, Q. Zhang, Q. Fan and W.
Huang, ACS Appl. Mater. Interfaces, 2018, 10, 39544.
8. (a) L. V. Wang and H. Song, Science, 2012, 335, 1458; (b) Y. Jiang and
K. Pu, Small, 2017, 13, 1700710; (c) H. J. Knox and J. Chan, Acc. Chem.
Res., 2018, 51, 2897.
9. (a) X. Hu, F. Lu, L. Chen, Y. Tang, W. Hu, X. Lu, Y. Ji, Z. Yang, W.
Zhang and C. Yin, ACS Appl. Mater. Interfaces, 2017, 9, 30458; (b) C.
Yin, G. Wen, C. Liu, B. Yang, S. Lin, J. Huang, P. Zhao, S. H. D. Wong,
K. Zhang, X. Chen, G. Li, X. Jiang, J. Huang, K. Pu, L. Wang and L.
Bian, Acs Nano, 2018, 12, 12201; (c) Q. Chen, C. Liang, X. Sun, J.
Chen, Z. Yang, H. Zhao, L. Feng and Z. Liu, Proc. Natl. Acad. Sci. U. S.
A., 2017, 114, 5343; (d) Q. Wang, B. Xia, J. Xu, X. Niu, J. Cai, Q. Shen,
W. Wang, W. Huang and Q. Fan, Mater. Chem. Front., 2019, DOI:
10.1039/C9QM00036D.
10. (a) J. Zhang, X. Zhen, P. K. Upputuri, M. Pramanik, P. Chen and K. Pu,
Adv. Mater., 2017, 29, 1604764; (b) C. Yin, X. Zhen, Q. Fan, W. Huang
and K. Pu, Acs Nano, 2017, 11, 4174; (c) Z. Yang, Y. Dai, C. Yin, Q.
Fan, W. Zhang, J. Song, G. Yu, W. Tang, W. Fan and B. C. Yung, Adv.
Mater., 2018, 30, 1707509.
Foundation of China (21602112, 21674048 and 21574064), 333
project of Jiangsu province (BRA2016379), Primary Research
& Development Plan of Jiangsu Province (BE2016770) and the
China Postdoctoral Science Foundation (No. 2017M621792).
11. Y. Liu, S. Wang, Y. Ma, J. Lin, H. Y. Wang, Y. Gu, X. Chen and P.
Huang, Adv. Mater., 2017, 29, 1606129.
12. C. Yin, Y. Tang, X. Li, Z. Yang, J. Li, X. Li, W. Huang and Q. Fan,
Small, 2018, 14, 1703400.
Conflicts of interest
There are no conflicts to declare.
13. (a) Q. Chen, X. Liu, J. Chen, J. Zeng, Z. Cheng and Z. Liu, Adv. Mater.,
2016, 27, 6820; (b) Z. Yang, W. Fan, W. Tang, Z. Shen, Y. Dai, J. Song,
Z. Wang, Y. Liu, L. Lin and L. Shan, Angew. Chem., 2018, 130, 14297;
(c) Z. Yang, J. Song, W. Tang, W. Fan, Y. Dai, Z. Shen, L. Lin, S.
Cheng, Y. Liu and G. Niu, Theranostics, 2019, 9, 526.
Notes and references
14. B. Shi, X. Gu, F. Qiang and C. Zhao, Chem. Sci., 2017, 8, 2150.
15. (a) F. Yu, P. Li, P. Song, B. Wang, J. Zhao and K. Han, Chem.
Commun., 2012, 48, 2852; (b) H. Xiao, C. Wu, P. Li, W. Gao, W. Zhang,
W. Zhang, L. Tong and B. Tang, Chem. Sci., 2017, 8, 7025.
16. Z. Yi, C. Wangqiao, Z. Jixin, P. Wenbo, W. Chengyuan, H. Ling, Y.
Cheng, Y. Qinyu, H. Wei and J. S. C. Loo, Small, 2015, 10, 4874.
17. H. Shinohara, A. Tanaka, T. Kitai, N. Yanabu, T. Inomoto, S. Satoh, E.
Hatano, Y. Yamaoka and K. Hirao, Hepatology, 2010, 23, 137.
1. (a) R. Wang, FASEB J., 2002, 16, 1792; (b) H. Peng, Y. Cheng, C. Dai,
A. L. King, B. L. Predmore, D. J. Lefer and B. Wang, Angew. Chem. Int.
Ed., 2011, 50, 9672; (c) H. Kimura, Amino Acids, 2011, 41, 113.
2. (a) T. V. Mishanina, M. Libiad and R. Banerjee, Nat. Chem. Biol., 2015,
11, 457; (b) P. Bindu D, S. Juan I, X. Risheng, V. M Scott, C. Jiyoung Y,
S. Adele M and S. Solomon H, Nature, 2016, 509, 96; (c) J. L. Greaney,
J. L. Kutz, S. W. Shank, S. Jandu, L. Santhanam and L. M. Alexander,
Hypertension, 2017, 69, 902.
3. (a) W. Min, Z. Jun, P. Yang, D. Jingde, Z. Lili, Z. Xiangrong and Z. Li,
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