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. 2025 Jun 5;3(11):742–749. doi: 10.1021/cbmi.5c00046

Fast-Response Mitochondria-Targeted Fluorescent Probe for H2S Imaging in Tumors and Inflammation

Huixiang Wang †,‡,§, Erting Feng ‡,§,*, Huizhen Ma ‡,§, Fangfang Du ‡,§, Muyang Chen ∥, Zongjin Qu †,*, Fabiao Yu ‡,§,*
PMCID: PMC12648413  PMID: 41311901

Abstract

In order to explore the complex interaction between H2S and the development of related diseases, it is urgently necessary to develop effective visualization methods to monitor the dynamic changes of H2S in real time. Herein, we constructed the NIR fluorescent probe HCy-SSPy based on disulfide cleavage for the rapid imaging of H2S. The hemicyanine (HCy-NH2) unit was used as a NIR fluorophore, and asymmetric pyridyl disulfides (SSPy) acted as the specific recognition receptor for H2S. The synthesized HCy-SSPy showed a remarkable NIR turn-on signal at 765 nm activated by H2S. This probe also possessed excellent selectivity and high sensitivity, as well as rapid detection ability for H2S (∼5 s). Moreover, the low cytotoxicity, mitochondrial localization, and excellent cell imaging performance of HCy-SSPy were discussed. Further biological experiments revealed that the probe not only imaged H2S in tumor-bearing mice but also showed great potential for H2S detection in inflammatory processes.

Keywords: Near-infrared fluorescent probe, H2S imaging, Fast response, Mitochondria targeting, In vivo imaging

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1. Introduction

Hydrogen sulfide (H2S), as the third endogenous gaseous signaling molecule discovered after nitric oxide (NO) and carbon monoxide (CO), has attracted extensive attention in the field of life sciences in recent years. − Despite being a toxic gas with a pungent odor, research has gradually revealed its important physiological and pathological functions in the body. Physiological concentrations of H2S are involved in regulating key processes such as cardiovascular system relaxation, neural signal transmission, anti-inflammation and antioxidation, and mitochondrial energy metabolism. − However, abnormal levels of H2S are closely related to neurodegenerative diseases, cancer, diabetic complications, and cardiovascular diseases. − For instance, H2S inhibits myocardial ischemia-reperfusion injury by activating KATP channels, while its overexpression may promote tumor angiogenesis. This concentration-dependent dual role makes the precise detection of H2S dynamic changes crucial for understanding its biological mechanisms and developing related diagnostic and therapeutic approaches.

Although traditional H2S detection techniques such as electrochemical methods, gas chromatography, and colorimetry have certain reliability, they have significant limitations in practical applications. − For instance, electrochemical sensors are susceptible to interference from redox substances in complex biological samples and have difficulty in detection in situ within cells. Chromatography requires a cumbersome sample pretreatment and cannot meet the requirements of real-time monitoring. Colorimetry is limited by low sensitivity and poor selectivity. Alternatively, near-infrared (NIR) fluorescent imaging technology suitable for living systems has stood out for endogenous signaling molecule detection due to its noninvasive nature, high sensitivity, and real-time imaging capability. − The design principle mainly relies on the strong nucleophilicity and reducing property of H2S, triggering the “turn-on” fluorescence signal through specific reactions (such as azide reduction, nitro reduction, nucleophilic addition, etc.). , For instance, Xiao’s group reported the H2S-sensitive fluorescent probe QTZ-N3 to detect endogenous H2S in renal cancer cells and sensitively monitor H2S levels in the mouse tumor model. QTZ-N3 exhibited good sensitivity and selectivity to H2S based on azide reduction. Zhao et al. designed a resorufin-based fluorescent probe for H2S detection through nitro reduction. The probe exhibited turn-on fluorescence at 586 nm and was used to detect H2S in paper chips and in live zebrafish. Feng and co-workers developed the near-infrared fluorescent probe MI-H2S for imaging H2S in mitochondria and in vivo by nucleophilic addition. Of note, these fluorescent probes usually require long reaction times to respond to H2S (minute or even hour level, Table S1). It is difficult to monitor the rapid dynamic changes in H2S in vivo in real time.

It has been reported that asymmetric pyridyl disulfides (SSPy) can enhance the polarity and electrophilicity of the distal sulfur atom far from the pyridine, thereby effectively forming persulfides in the presence of H2S. These persulfides can then undergo intramolecular nucleophilic attack, expelling a fluorophore and a benzodithione as byproducts. Based on this disulfide cleavage, researchers have developed numerous H2S fluorescent probes with high selectivity based on the SSPy response group. − However, NIR fluorescent probes that respond quickly to H2S are still rarely reported. Herein, we report the NIR fluorescent probe HCy-SSPy for rapid H2S imaging in vitro and in vivo based on disulfide cleavage (Scheme ). HCy-SSPy consisted of two key segments. One was the hemicyanine (HCy-NH2) unit for the NIR fluorescence signal, and the other was the SSPy part, serving as the specific recognition receptor for H2S. HCy-SSPy exhibited weak fluorescence due to the quenching effect of SSPy. Noticeably, an intense emission peak at 765 nm occurred after the addition of nucleophilic H2S. The H2S-mediated reduction resulted in disulfide cleavage, and the NIR fluorophore (HCy-NH2) was released. Moreover, HCy-SSPy showed good sensitivity (LOD = 0.36 μM) and rapid response (∼5 s) to H2S. The probe also exhibited remarkable selectivity for H2S, excluding interference from sulfhydryl-containing biothiols. Especially, the positive characteristic of HCy-SSPy enabled it to effectively target mitochondria. Further biological applications suggested that HCy-SSPy can not only monitor exogenous and endogenous H2S in cells but also rapidly image H2S in inflammatory areas and tumor tissues.

1. Schematic Diagram of the Molecular Structure of Probe HCy-SSPy with Rapid Response to H2S.

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2. Experimental Section

2.1. Reagents and Instruments

The reagents and instruments employed in this study are detailed in the Supporting Information.

2.2. Synthesis of Fluorescent Probe HCy-SSPy

The synthetic route to the probe HCy-SSPy is described in Scheme S1. SSPy and HCy-NH2 were prepared according to the reported literature. , In detail, SSPy (52.6 mg, 0.2 mmol) was added into a single-neck bottle. Under ice bath conditions, dry CH2Cl2 (5 mL) containing 0.1 mL of oxaloyl chloride was dripped into the system. The mixture was stirred for 4 h at room temperature, and then distilled under reduced pressure to obtain a yellow solid, which was directly used in the next step. HCy-NH2 (50 mg, 0.1 mmol) dissolved in dry CH2Cl2 (10 mL) was dripped into a single-neck bottle. The reaction mixture was stirred at room temperature overnight. Then, the solution was concentrated under reduced pressure to give a crude blue solid, which was purified by column chromatography using CH2Cl2/MeOH (v/v = 10:1) as the eluent to obtain the desired product (33 mg, yield 42%). 1H NMR (400 MHz, DMSO-d 6): δ 12.05 (s, 1H), 8.90 (s, 1H), 8.27–8.27 (d, 1H), 7.92 (s, 1H), 7.87–7.81 (m, 3H), 7.68–7.53 (m, 6H), 7.36 (s, 1H), 7.29–7.24 (d, 3H), 7.11 (s, 1H), 6.91–6.87 (d, 1H), 4.55–4.54 (d, 2H), 2.77–2.72 (d, 4H), 1.86 (s, 2H), 1.76 (s, 9H). HRMS: calcd for C39H39N4OS3 + (M + NH4 + – H+ – I–)+, 675.2281; found, 675.6481.

2.3. Optical Measurements

Quartz cuvettes (1 cm) were used in all of the spectral measurements. All spectral measurements in solution were performed with 10 μM HCy-SSPy in a PBS-DMSO buffer mixture (4:1 ratio, 2 mL). The fluorescence was collected with an excitation wavelength at 660 nm.

2.4. Cell Imaging with HCy-SSPy

Prior to fluorescence imaging of live cells, a cytotoxicity assessment was performed to assess the biocompatibility of HCy-SSPy against the mouse breast cancer cell line 4T1 cells. The cells were grown in 96-well plates at a density of 1 × 104 cells per well and incubated at 37 °C with 5% CO2 for 24 h. Then MTT assays were performed to assess the cytotoxicity of HCy-SSPy (0–25 μM). For exogenous H2S imaging, 4T1 cells were pretreated at 37 °C for 30 min using 20, 40, 60, 80, and 100 μM NaHS. The cells were then stained with HCy-SSPy (10 μM) for 1 h and photographed using a confocal laser scanning microscope (CLSM). To perform endogenous H2S imaging with HCy-SSPy, 4T1 cells were pretreated with 50, 100, 150, and 200 μM Cys at 37 °C for 1 h, respectively, and then stained with HCy-SSPy (10 μM) for 1 h, and subsequently fluorescence images of the cells were collected. To observe the fluctuation of H2S in inflammatory cells, 4T1 cells were pretreated with 1.0 μg/mL lipopolysaccharide (LPS) at 37 °C for 1, 3, and 5 h, stained with HCy-SSPy (10 μM) for 1 h, and then imaged. In the colocalization experiment, 4T1 cells were preincubated with NaHS/HCy-SSPy (10 μM each) for 1 h, then cultured with Rhodamine 123 (Rh 123, 1 μM) for 20 min, and pictured with CLSM.

2.5. Mice Imaging with HCy-SSPy

Six-week-old female Balb/c mice were housed in an environmentally controlled animal facility for animal modeling. To detect H2S in tumor tissue, 4T1 tumor cells were injected subcutaneously into the right leg root of Balb/c mice to establish tumor models. The tumors were pretreated with Cys (1 mM, 100 μL) for 1 h to induce endogenous H2S. After injection of HCy-SSPy (0.1 mM, 100 μL) into tumors, imaging was performed at 0, 10, 20, 30, 40, and 50 min, respectively. After the mice were sacrificed, the main organs (such as heart, liver, spleen, lungs, kidneys) and tumors were dissected and subjected to imaging examinations. In order to detect the H2S level in the inflammation area, mice were intraperitoneally injected with LPS (1.0 mg/mL, 100 μL) for 8.0 h to induce local inflammation, and then HCy-SSPy (0.1 mM, 100 μL) was injected into the inflammation site; images were collected every 10 min for 50 min. The control group was injected with HCy-SSPy only. All the animal experiments were performed according to the relevant laws and guidelines and were approved by the Ethical Committee of Hainan Medical University (Haikou, China, HYLL-2023-006).

3. Results and Discussion

3.1. Sensing Property of HCy-SSPy for H2S

The UV–vis absorption and fluorescence spectra of the probe HCy-SSPy (10 μM) were first recorded to investigate the responses to H2S. NaHS was used as the equivalent of H2S. Figure A shows the absorption change of the probe before and after adding H2S (40 μM) in PBS buffer solution (containing 20% DMSO). HCy-SSPy exhibited an obvious absorption peak at 615 nm. Markedly, the maximal absorption spectrum of the probe showed a visible red-shift to 660 nm in the presence of H2S. The solution color also changed from blue to green immediately. Absorption titration experiments illustrated that with the addition of H2S, the peak at 615 nm gradually disappeared and a new absorption band around 660 nm appeared (Figure S5). These results proved that the probe HCy-SSPy had a rapid response characteristic of H2S that was visible to the naked eye. Next, we studied the fluorescence response of HCy-SSPy toward H2S as shown in Figure B. The individual probes had a weak fluorescence signal. With the increase in H2S concentration, the fluorescence emission at 765 nm was significantly enhanced. Furthermore, based on the linear relationship between the fluorescence intensity and H2S concentration (Figure C), the limit of detection (LOD) of the probe for H2S was calculated to be 0.36 μM using the 3σ/k method. This indicated that HCy-SSPy was capable of detecting H2S with “turn-on” fluorescence and high sensitivity. The stability of the probe after reaction with H2S was also discussed (Figure S6). The absorption and fluorescence spectra of HCy-SSPy+H2S remained stable within 30 min, indicating the prominent stability of the probe in response to H2S. Furthermore, the antiphotobleaching performance of the probe under continuous irradiation was investigated. As displayed in Figure S7, the absorption spectra of HCy-SSPy showed an ignorable change under continuous irradiation (610 nm, 20 mW/cm2), indicating that it had good photostability. Due to the enhanced NIR fluorescence, high sensitivity, and good photostability, the probe may be competent for detecting H2S in living organisms.

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(A) Absorption spectra of HCy-SSPy (10 μM) before and after the addition of H2S (40 μM). (B) Fluorescence spectra of HCy-SSPy (10 μM) with various concentrations of H2S (0–44 μM). (C) Linearity plots of the fluorescence intensity of HCy-SSPy against H2S concentrations. (D) Fluorescence spectra of HCy-SSPy (10 μM) in the presence of H2S and other analytes (including K+, Na+, Ca2+, Mg2+, Zn2+, Fe2+, Fe3+, Cu2+, SO4 2–, CO3 2–, HCO3 –, NO3 –, NO2 –, Ac–, Cl–, Br–, ClO–, H2O2, Cys, Hcy, and GSH; 200 μM each, except 40 μM for H2S, 1 mM for GSH). (E) Time-dependent fluorescence intensity of HCy-SSPy (10 μM) with/without the addition of H2S (40 μM). (F) Effect of the pH on the fluorescence intensity of HCy-SSPy (10 μM) with/without the addition of H2S (40 μM). λex = 660 nm.

Then, the selectivity of HCy-SSPy toward H2S was examined. Figure D displays the emission spectra of HCy-SSPy (10 μM) in the presence of H2S and other potential analytes, such as K+, Na+, Ca2+, Mg2+, Zn2+, Fe2+, Fe3+, Cu2+, SO4 2–, CO3 2–, HCO3 –, NO3 –, NO2 –, Ac–, Cl–, Br–, ClO–, H2O2, Cys, Hcy, and GSH (200 μM each, except 40 μM for H2S, 1 mM for GSH). The fluorescence signal of the solution increased significantly when H2S was added. Negligible fluorescence changes at 765 nm can be observed upon the addition of other analytes, including thiol-containing endogenous agents (GSH, Cys, and Hcy, Figure S8). Besides, the absorption spectra of the probe showed no appreciable change in the presence of other analytes (Figure S9). These results illustrated that HCy-SSPy possessed high selectivity to H2S. Then, the competition experiment was assessed by comparing the fluorescence intensity of HCy-SSPy with other substances in the presence of H2S as displayed in Figure S10. The enhanced fluorescence signals of HCy-SSPy were observed in all of the groups. The subsequently added analytes may combine with H2S (e.g., Zn2+). However, this did not affect the fluorescence reaction of HCy-SSPy with H2S, which benefited from the rapid response of the probe to H2S in advance. These results indicated that this probe had a high anti-interference ability for H2S detection. In addition, the time-dependent fluorescence response of HCy-SSPy (10 μM) to H2S (40 μM) was tested as demonstrated in Figure E. The fluorescence intensity markedly enhanced and rapidly reached a plateau within ∼5 s upon the addition of H2S. Compared with the reported H2S fluorescent probes (Table S1), the response time was significantly shortened, which was conducive to the rapid and real-time detection of the dynamic changes of H2S in biological systems. Over the following time, the fluorescence intensity remained stable, indicating that the probe had good stability after reacting with H2S. Subsequently, the fluorescence response of HCy-SSPy to H2S at different pH values was investigated (Figure F). Distinctly enhanced NIR fluorescence was observed under a wide range of pH (6.0–10.0), suggesting that the probe can be used to detect H2S at physiological pH. The aforementioned experiments indicated that HCy-SSPy can rapidly detect H2S through fluorescence activation in the physiological environment.

3.2. Response Mechanism

The proposed detection mechanism of HCy-SSPy to H2S was depicted as shown in Scheme . HCy-SSPy can react with H2S to form intermediate persulfides containing free SH. These persulfides can then undergo a spontaneous cyclization to form byproduct benzodithione and release the fluorophore HCy-NH2. The strong fluorescence emission ability of the probe was restored due to the strong electron-donating ability of the amino group. This detection mechanism has been further proved by HRMS. The peak of HCy-SSPy was located at m/z = 675.6481 (Figure S3). After reacting with H2S, the peak of HCy-SSPy disappeared and a new peak at m/z = 413.1849 emerged, which belonged to HCy-NH2 (calcd 413.2046, Figure S4). These results directly indicated that HCy-SSPy can be cleaved by H2S and converted into HCy-NH2.

2. Proposed Detection Mechanism of Probe HCy-SSPy to H2S.

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3.3. Fluorescence Imaging in Living Cells

In order to explore the bioimaging capability of the probe HCy-SSPy, we first evaluated their cytotoxicity on 4T1 cells by the MTT method. As shown in Figure S11, after incubating the cells with HCy-SSPy (0–25 μM) for 12 h, the cell survival rate was still greater than 80% even when the probe concentration reached 25 μM, indicating that this probe has relatively low cytotoxicity to cells. Then a colocalization experiment was performed to explore the organelle targeting of the probe (Figure S12). Due to the negative membrane potential of the inner mitochondrial membrane, the positively charged probe can be electrostatically attracted to cross the mitochondrial membrane and accumulate in the matrix. The commercial dye Rh 123 was used as a mitochondrial indicator. As can be seen, the green channel fluorescence of Rh 123 and the red channel fluorescence of HCy-SSPy showed good overlap. The colocalization Pearson’s coefficient reached 0.90, indicating that the probe had remarkable targeting specificity to mitochondria.

Encouraged by the prominent sensing property, low cytotoxicity, and mitochondrial targeting of HCy-SSPy, we then carried out the fluorescence imaging of H2S in 4T1 cells. Figure A exhibits the fluorescence images of 4T1 cells pretreated with NaHS (0–100 μM) for 30 min and then stained with HCy-SSPy (10 μM) for 1 h. The fluorescence signal was hardly observed without the addition of NaHS. When the cells were pretreated with NaHS (20–100 μM), remarkable fluorescence was seen. And the fluorescence signal gradually intensified with the increase of NaHS concentration (Figure B). These results manifested that HCy-SSPy can be utilized for the fluorescence detection of exogenous H2S in living cells.

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(A) Fluorescence images of exogenous H2S in 4T1 cells. The cells were pretreated with different concentrations of NaHS (0–100 μM) for 30 min and then stained with HCy-SSPy (10 μM). (B) Relative fluorescence intensity in (A). λex = 645 nm, λem = 700–800 nm; scale bar 20 μm.

Subsequently, the imaging ability of HCy-SSPy for endogenous H2S was further discussed. In general, cells use cysteine or homocysteine as substrates to catalyze the production of H2S through enzymatic reactions involving cystathionine β-synthase (CBS), cystathionine γ-lyase (CSE), and 3-mercaptopyruvate sulfurtransferase (3-MST). , Adding cysteine (Cys) is a simple and effective way to stimulate cells to produce more endogenous H2S. As depicted in Figure A,B, 4T1 cells were pretreated with Cys (0–200 μM) for 1 h and then stained with HCy-SSPy (10 μM). A significant Cys dose-dependent enhancement of the NIR fluorescence signal was observed. To verify that intracellular fluorescence was activated by endogenous H2S, we pretreated the cells with aminooxyacetic acid (AOAA, a CBS inhibitor) and then cultured them sequentially with Cys and HCy-SSPy. As expected, a negligible fluorescence signal was observed in the cells, which may be attributed to the potent inhibition of intracellular H2S synthesis by AOAA. The above results indicated that HCy-SSPy can be used for detecting endogenous H2S within living cells.

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(A) Fluorescence imaging of endogenous H2S in 4T1 cells. The cells were pretreated with Cys (0–200 μM) for 1 h and then stained with HCy-SSPy (10 μM). In addition, the cells pretreated with AOAA were cultured sequentially with Cys and HCy-SSPy as a control. (B) Relative fluorescence intensity in (A). λex = 645 nm, λem = 700–800 nm; scale bar 20 μm.

It has been reported that the H2S concentration in cells can be significantly upregulated by inflammatory stimulation. , Therefore, we used HCy-SSPy to further monitor H2S fluctuation in lipopolysaccharide (LPS)-induced inflammation in living cells as illustrated in Figure A,B. At the beginning, cells stained with the probe HCy-SSPy emitted a faint fluorescence signal. After LPS stimulation of the cells, a significant enhancement of fluorescence was seen over time (0–5 h). This indicated that the content of H2S in inflammatory cells was gradually increasing, and HCy-SSPy had the potential to monitor this process.

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(A) Fluorescence images of LPS-induced H2S in 4T1 cells stained with HCy-SSPy (10 μM). (B) Relative fluorescent intensity in (A). λex = 645 nm, λem = 700–800 nm; scale bar 20 μm.

3.4. Fluorescence Imaging in Mice

The NIR probe HCy-SSPy demonstrated remarkable H2S imaging capabilities in cells. Subsequently, we further investigated its potential for H2S fluorescence imaging in tumor-bearing mice. The tumors were pretreated with Cys (1 mM, 100 μL) for 1 h to induce endogenous H2S, followed by intratumoral injection of HCy-SSPy (0.1 mM, 100 μL). As shown in Figure A,B, obvious fluorescence was observed at the tumor site, and the fluorescence intensity gradually increased with time. To further evaluate the fluorescence detection performance of HCy-SSPy for H2S in a tumor, fluorescence imaging was performed on a tumor as well as major organs of tumor-bearing mice (Figure C). As expected, the tumor exhibited a significant fluorescence intensity. These results demonstrated that the probe could effectively detect endogenous H2S in tumor-bearing mice.

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H2S imaging in mouse tumor tissues. (A) Fluorescence imaging of tumor-bearing mice after intratumoral injection of HCy-SSPy (0.1 mM, 100 μL) with time (0–50 min). (B) Relative fluorescence intensity in (A). (C) Fluorescence imaging and relative fluorescent intensity of tumor and main organs: (1) heart, (2) kidney, (3) lung, (4) spleen, (5) liver, (6) tumor. λex = 675 nm, λem = 700–850 nm.

Finally, we used HCy-SSPy to conduct biological imaging of H2S in the inflammatory processes of mice. LPS was injected intraperitoneally into the abdomen of mice to induce local inflammation. After 8 h, the probe was injected into the inflammation area, and fluorescence imaging was conducted with time. As exhibited in Figure A,B, the mice treated with LPS and HCy-SSPy showed distinct fluorescence signals over time. However, in the control group (the mice were injected only with probe HCy-SSPy), no obvious fluorescence emerged. These results suggested that HCy-SSPy was a potential tool for H2S detection during inflammation in vivo.

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H2S imaging in mice inflammation areas. (A) Mice were intraperitoneally injected with LPS (1.0 mg/mL, 100 μL) for 8.0 h to cause inflammation. HCy-SSPy (0.1 mM, 100 μL) was then injected into the inflammatory areas and imaged every 10 min. The probe was also injected into the normal mice as a control. (B) Relative fluorescent intensity in (A). λex = 675 nm, λem = 700–850 nm.

4. Conclusion

In conclusion, we developed the NIR fluorescent probe HCy-SSPy based on disulfide cleavage for rapid H2S imaging in vitro and in vivo. HCy-SSPy had high selectivity, good sensitivity (LOD = 0.36 μM), and rapid response characteristic (∼5 s) for H2S detection. Moreover, this probe exhibited low cytotoxicity, mitochondrial targeting performance, and an outstanding imaging potential. The exogenous and endogenous H2S imaging in living cells and tumor-bearing mice was successfully performed. HCy-SSPy can also detect H2S in inflammation in vivo. Taken together, this study provides an effective NIR molecular probe for rapid H2S visualization, which may provide a new protocol for the early diagnosis of H2S-related diseases.

Supplementary Material

im5c00046_si_001.pdf (585.8KB, pdf)

Acknowledgments

This work was supported by the National Natural Science Foundation of China (No. 22264013), Hainan Province Science and Technology Special Fund (No. ZDKJ2021038), Key Research and Development Project of Hainan Province (No. ZDYF2025SHFZ043), and High-level Talents Project of Hainan Provincial Natural Science Foundation (No. 825RC763). We offer thanks for the support and assistance in terms of instruments and facilities provided by Public Research Center of Hainan Medical University.

The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/cbmi.5c00046.

  • Experimental section, synthetic route, NMR and MS spectra, and optical properties of HCy-SSPy, cytotoxicity, and mitochondrial colocalization assay (PDF)

H.W. and E.F. contributed equally to this work.

The authors declare no competing financial interest.

References

  1. Ranjana M., Kulkarni R. M., Sunil D.. Small Molecule Optical Probes for Detection of H2S in Water Samples: A Review. Acs Omega. 2024;9(13):14672–14691. doi: 10.1021/acsomega.3c08573. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Fosnacht K. G., Pluth M. D.. Activity-Based Fluorescent Probes for Hydrogen Sulfide and Related Reactive Sulfur Species. Chem. Rev. 2024;124(7):4124–4257. doi: 10.1021/acs.chemrev.3c00683. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Yu J., Xu J., Ma S., Wang C., Miao Q., Wang L., Chen G.. Visible Tracking of Small Molecules of Gases with Fluorescent Donors. Chem. Biomed. Imaging. 2024;2(6):401–412. doi: 10.1021/cbmi.4c00006. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Pang Q., Huo F., Yin C.. Research Progress in the Field of Hydrogen Sulfide Donors in the Last Five Years. ChemBioChem. 2025;26(5):e202400817. doi: 10.1002/cbic.202400817. [DOI] [PubMed] [Google Scholar]
  5. Li Y., Nouman N. M., Yin C., Huo F.. The Development of Near-infrared Hydrogen Sulfide Probes and Its Application in the Past Ten Years. Dyes Pigments. 2024;228:112246. doi: 10.1016/j.dyepig.2024.112246. [DOI] [Google Scholar]
  6. Hanc K., Janeková H., Štacko P.. Concurrent Subcellular Delivery of Hydrogen Sulfide and A Payload with Near-Infrared Light. JACS Au. 2024;4(7):2687–2694. doi: 10.1021/jacsau.4c00445. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chen Z., Ouyang C., Zhang H., Gu Y., Deng Y., Du C., Cui C., Li S., Wang W., Kong W., Chen J., Cai J., Geng B.. Vascular Smooth Muscle Cell-Derived Hydrogen Sulfide Promotes Atherosclerotic Plaque Stability via TFEB (transcription factor EB)-Mediated Autophagy. Autophagy. 2022;18(10):2270–2287. doi: 10.1080/15548627.2022.2026097. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Lv B., Chen S., Tang C., Jin H., Du J., Huang Y.. Hydrogen Sulfide and Vascular Regulation-An update. J. Adv. Res. 2021;27:85–97. doi: 10.1016/j.jare.2020.05.007. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Kolluru G. K., Shackelford R. E., Shen X., Dominic P., Kevil C. G.. Sulfide Regulation of Cardiovascular Function in Health and Disease. Nat. Rev. Cardiol. 2023;20(2):109–125. doi: 10.1038/s41569-022-00741-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Cirino G., Szabo C., Papapetropoulos A.. Physiological Roles of Hydrogen Sulfide in Mammalian Cells, Tissues, and Organs. Physiol. Rev. 2023;103(1):31–276. doi: 10.1152/physrev.00028.2021. [DOI] [PubMed] [Google Scholar]
  11. Zhang Y., Chen S., Zhu J., Guo S., Yue T., Xu H., Hu J., Huang Z., Chen Z., Wang P., Liu Y.. Overexpression of CBS/H2S Inhibits Proliferation and Metastasis of Colon Cancer Cells Through Downregulation of CD44. Cancer Cell Int. 2022;22(1):85. doi: 10.1186/s12935-022-02512-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Wei X., Wu S., Liu H., Huang M., Zhang J., He M., Wang F., Chen W.. Simple Strategy to Develop Multifunctional NIR Fluorescent Probes for Simultaneous Identification of H2S and SO2 . Anal. Chem. 2025;97(16):8852–8858. doi: 10.1021/acs.analchem.4c06842. [DOI] [PubMed] [Google Scholar]
  13. Li N., Li X., Li J., Li Y., Zhang T.. An AND-Gate Photoacoustic Probe for Cys and H2S Precise Photoacoustic Sensing in Localized Tumors. Anal. Chem. 2024;96(19):7342–7347. doi: 10.1021/acs.analchem.4c00341. [DOI] [PubMed] [Google Scholar]
  14. Hu C., Yang Z., Shi X., Xue Y., Huang L., Tang C., Wang F.. In Vivo Monitoring of Endogenous Hydrogen Sulfide and Evaluation of Natural Protectants in Liver Injury Mice using A Highly Selective Bioluminescent probe. Biosens. Bioelectron. 2025;278:117343. doi: 10.1016/j.bios.2025.117343. [DOI] [PubMed] [Google Scholar]
  15. Yang W., Yang G., Jia X., Wu L., Wang R.. Activation of KATP Channels by H2S in Rat Insulin-Secreting Cells and the Underlying Mechanisms. J. Physiol. 2005;569(2):519–531. doi: 10.1113/jphysiol.2005.097642. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Guo Z., Xiao Y., Wu W., Zhe M., Yu P., Shakya S., Li Z., Xing F.. Metal-Organic Framework-based Smart Stimuli-Responsive Drug Delivery Systems for Cancer Therapy: Advances, Challenges, and Future Perspectives. J. Nanobiotechnol. 2025;23(1):157. doi: 10.1186/s12951-025-03252-x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Liu Q., Liu Y., Wan Q., Lu Q., Liu J., Ren Y., Tang J., Su Q., Luo Y.. Label-Free, Reusable, Equipment-Free, and Visual Detection of Hydrogen Sulfide using A Colorimetric and Fluorescent Dual-Mode Sensing Platform. Anal. Chem. 2023;95(14):5920–5926. doi: 10.1021/acs.analchem.2c05364. [DOI] [PubMed] [Google Scholar]
  18. Chun H. W., Zheng J., Lee E. H., Oh B. M., Lee C. B., Min J. S., Kim E., Kim E., Lee W., Kim J. H.. Pure-Water-Soluble Colorimetric Chemosensors for Highly Sensitive and Rapid Detection of Hydrogen Sulfide: Applications to Evaluation of On-site Water Quality and Real-Time Gas Sensors. Sensor. Actuat. B-Chem. 2024;402:134989. doi: 10.1016/j.snb.2023.134989. [DOI] [Google Scholar]
  19. Mhatre S., Rai A., Ali H., Patil A., Singh N., Verma R., Auden J., Chandler C., Dash A., Opere C.. et al. Comparison of Colorimetric, Spectroscopic and Electrochemical Techniques for Quantification of Hydrogen Sulfide. BioTechniques. 2024;76(2):71–80. doi: 10.2144/btn-2023-0075. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Khan M. A. H., Rao M. V., Li Q.. Recent Advances in Electrochemical Sensors for Detecting Toxic Gases: NO2, SO2 and H2S. Sensors. 2019;19(4):905. doi: 10.3390/s19040905. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Chen Z., Yang Y., Tian Y., Yang J., Xiong H.. Diagnosis of Nonalcoholic Fatty Liver Disease via a H2S-Responsive Bioluminescent Probe Combined with Firefly Luciferase mRNA Delivery. Anal. Chem. 2024;96(22):9236–9243. doi: 10.1021/acs.analchem.4c01462. [DOI] [PubMed] [Google Scholar]
  22. Wu L., Tong Q., Cao X., Zhang D., Yang F., Lin H., Fan Q.. Dual-Locked Near-Infrared Fluorescent Probe for Real-Time Imaging of Hydrogen Sulfide/Matrix Metallopeptidase-2 In Vivo. ACS Nano. 2025;19(7):7294–7305. doi: 10.1021/acsnano.4c17799. [DOI] [PubMed] [Google Scholar]
  23. Gong S., Gu Y., Tian J., Zhou X., Chen Y., Chen J., Xu X., Wang Z., Wang S.. A Near-Infrared Fluorescent Probe with a Large Stokes Shift for Detecting Hydrogen Sulfide in Environmental Waters, Wine Samples, and Living Systems. J. Agric. Food Chem. 2025;73(8):4594–4604. doi: 10.1021/acs.jafc.4c11645. [DOI] [PubMed] [Google Scholar]
  24. He R., Tang D., Xu N., Liu H., Dou K., Zhou X., Yu F.. Evaluation of Erastin Synergized Cisplatin Anti-Nasopharyngeal Carcinoma Effect with A Glutathione-Activated Near-Infrared Fluorescent Probe. Chin. Chem. Lett. 2024;35(2):108658. doi: 10.1016/j.cclet.2023.108658. [DOI] [Google Scholar]
  25. Luo X., Yu F., Wang R., Su T., Luo P., Wen P., Yu F.. A Near-Infrared Two-Photon Fluorescent Probe for the Detection of HClO in Inflammatory and Tumor-bearing mice. Chin. Chem. Lett. 2025;36:110531. doi: 10.1016/j.cclet.2024.110531. [DOI] [Google Scholar]
  26. Su T., Shen R., Tu D., Han X., Luo X., Yu F.. A β-Galactosidase Activated Near-Infrared Fluorescent Probe for Tracking Cellular Senescence In Vitro and In Vivo. Smart Mol. 2025;3:e20240062. doi: 10.1002/smo.20240062. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Fang B., Yang J. Q., Wang L. M., Li H. Q., Guo J. Y., Zhang J. X., Guo Q. Y., Peng B., Liu K. D., Xi M. M., Bai H., Fu L., Li L.. A Mitochondria-Targeted H2S-Activatable Fluorogenic Probe for Tracking Hepatic Ischemia-Reperfusion Injury. Chin. Chem. Lett. 2024;35(6):108913. doi: 10.1016/j.cclet.2023.108913. [DOI] [Google Scholar]
  28. Lu K., Wang Y., Wang C., Liu R., Yang K., Zhang X., Xiao H.. A Bioluminescent Probe for H2S Detection in Tumor Microenvironment. Acs Bio Med. Chem. Au. 2025;5:175–183. doi: 10.1021/acsbiomedchemau.4c00102. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Sun Y., Tang X., Zhang K., Liu K., Li Z., Zhao L.. Hydrogen Sulfide Detection and Zebrafish Imaging by A Designed Sensitive and Selective Fluorescent Probe Based on Resorufin. Spectrochim. Acta, Part A. 2022;264:120265. doi: 10.1016/j.saa.2021.120265. [DOI] [PubMed] [Google Scholar]
  30. Gong S., Zheng Z., Guan X., Feng S., Feng G.. Near-Infrared Mitochondria-Targetable Fluorescent Probe for High-Contrast Bioimaging of H2S. Anal. Chem. 2021;93(14):5700–5708. doi: 10.1021/acs.analchem.0c04639. [DOI] [PubMed] [Google Scholar]
  31. Liu C., Pan J., Li S., Zhao Y., Wu L. Y., Berkman C. E., Whorton A. R., Xian M.. Capture and Visualization of Hydrogen Sulfide via A Fluorescent Probe. Angew. Chem., Int. Ed. 2011;50(44):10327–10329. doi: 10.1002/anie.201104305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Kang J., Huo F., Yin C.. A Novel Ratiometric Fluorescent H2S Probe Based on Tandem Nucleophilic Substitution/Cyclization Reaction and Its Bioimaging. Dyes Pigments. 2017;146:287–292. doi: 10.1016/j.dyepig.2017.07.016. [DOI] [Google Scholar]
  33. Wu Q., Huo F., Wang J., Yin C.. Fluorescent Probe for Detecting Hydrogen Sulfide Based on Disulfide Nucleophilic Substitution-Addition. Spectrochim. Acta, Part A. 2020;238:118437. doi: 10.1016/j.saa.2020.118437. [DOI] [PubMed] [Google Scholar]
  34. Zhang C., Zhang G., Feng L., Li J.. A Ratiometric Fluorescent Probe for Sensitive and Selective Detection of Hydrogen Sulfide and Its Application for Bioimaging. Sensor. Actuat. B-Chem. 2015;216:412–417. doi: 10.1016/j.snb.2015.04.073. [DOI] [Google Scholar]
  35. Zhang S., Chen H., Wang L., Qin X., Jiang B. P., Ji S. C., Shen X. C., Liang H.. A General Approach to Design Dual Ratiometric Fluorescent and Photoacoustic Probes for Quantitatively Visualizing Tumor Hypoxia Levels In Vivo. Angew. Chem., Int. Ed. 2022;61(7):e202107076. doi: 10.1002/anie.202107076. [DOI] [PubMed] [Google Scholar]
  36. Asghar R., Li Y., Huo F., Yin C.. Sensing Mechanism of Cysteine Specific Fluorescence Probes and Their Application of Cysteine Recognition. Chem. Biomed. Imaging. 2024;2:250–269. doi: 10.1021/cbmi.4c00001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Yan L., Gu Q. S., Jiang W. L., Tan M., Tan Z. K., Mao G. J., Xu F., Li C. Y.. Near-Infrared Fluorescent Probe with Large Stokes Shift for Imaging of Hydrogen Sulfide in Tumor-Bearing Mice. Anal. Chem. 2022;94(14):5514–5520. doi: 10.1021/acs.analchem.1c04169. [DOI] [PubMed] [Google Scholar]
  38. Cai W. J., Wang M. J., Ju L. H., Wang C., Zhu Y. C.. Hydrogen Sulfide Induces Human Colon Cancer Cell Proliferation: Role of Akt, ERK and p21. Cell Biol. Int. 2010;34(6):565–572. doi: 10.1042/CBI20090368. [DOI] [PubMed] [Google Scholar]
  39. Bhatia M., Gaddam R. R.. Hydrogen Sulfide in Inflammation: A Novel Mediator and Therapeutic Target. Antioxid. Redox Sign. 2021;34(17):1368–1377. doi: 10.1089/ars.2020.8211. [DOI] [PubMed] [Google Scholar]

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