I-type photosensitizer with NIR-I absorption and NIR-II emission characteristics as well as preparation method and application of I-type photosensitizer

Abstract

The invention belongs to the technical field of biochemical materials, and particularly discloses an I-type photosensitizer with NIR-I (Near Infrared Region I) absorption and NIR-II (Near Infrared Region II) emission characteristics as well as a preparation method and application of the I-type photosensitizer. The photosensitizer molecule provided by the invention has a D-pi-A molecular structure as shown in a formula I, a dithienopyrrole derivative is used as an electron donor (Donor), benzo [cd] indole salt is used as an electron acceptor (Acceptor), and the photosensitizer molecule is prepared by single / double thienyl pi bridge connection and Knoevenagel condensation reaction. The I-type photosensitizer disclosed by the invention can emit NIR-II fluorescence under the excitation of an NIR-I light source, has excellent I-type active oxygen generation capability and photo-thermal generation capability, and can realize biological tissue fluorescence imaging detection with high signal-to-noise ratio and efficient photodynamic-photo-thermal combined tumor treatment under a hypoxic condition; the method has a huge potential application value in the field of biological medicines, especially in the aspect of tumor diagnosis and treatment.

Description

Technical Field

[0001] The present invention belongs to the technical field of biochemical materials, and particularly relates to a type I photosensitizer with NIR-I absorption and NIR-II emission characteristics, and a preparation method and application thereof. Background Art

[0002] Cancer (malignant tumor) seriously endangers human life and health and has become one of the major public health problems worldwide. In recent decades, research on cancer diagnosis and treatment has attracted much attention. Among them, as a non-invasive treatment technology, photodynamic therapy (PDT) has shown great application potential in the field of cancer treatment due to its significant advantages such as low toxic side effects, high spatiotemporal precision controllability, and low drug resistance.

[0003] PDT achieves precise treatment through the ternary synergistic action of a photosensitizer, an excitation light source, and tissue oxygen. When the photosensitizer is excited by light of a specific wavelength, it will generate free radicals and radical anion species (type I PDT) through electron transfer with nearby biomolecules in the cell microenvironment, or directly transfer energy to nearby triplet oxygen to generate singlet oxygen (type II PDT). These reactive oxygen species have extremely strong cytotoxicity and can cause oxidative damage to cell membranes, DNA, proteins, etc., ultimately inducing apoptosis or necrosis of cells. However, type II PDT highly depends on the oxygen concentration, and the interior of tumors is in a hypoxic state, which makes its effect in tumor treatment not ideal. In contrast, hypoxia-dependent type I PDT can efficiently generate reactive oxygen in the tumor hypoxic microenvironment, thereby effectively killing tumor cells. Thus, photosensitizers capable of generating type I reactive oxygen are excellent candidates for tumor PDT therapeutic agents.

[0004] As a non-invasive imaging technology, fluorescence imaging has the advantages of high resolution, high sensitivity, easy access to probes, and low cost of imaging equipment, and has been widely used in biological imaging research. However, due to the strong absorption and scattering of short-wavelength light by biological tissues, compared with visible light (400 - 700 nm) and NIR-I (700 - 900 nm) fluorescence imaging, NIR-II (1000 - 1700 nm) fluorescence imaging has higher imaging resolution, deeper tissue detection depth, and lower background interference signals, and thus shows more significant application advantages in cancer diagnosis and surgical navigation.

[0005] Although researchers have developed a variety of photosensitizers for cancer diagnosis and treatment, the absorption wavelengths of most photosensitizers still lie in the visible light region (<700 nm), and the emission wavelengths are in the NIR-I region, which greatly weakens their diagnostic and therapeutic effects on deep lesion tissues. Therefore, developing type-I photosensitizers with long absorption in NIR-I and long emission in NIR-II is of great significance for improving the photodynamic therapy effect of deep tumors. Summary of the Invention

[0006] The purpose of the present invention is to provide a type-I photosensitizer with NIR-I absorption and NIR-II emission characteristics, as well as its preparation method and application. The photosensitizer provided by the present invention can emit NIR-II fluorescence under the excitation of NIR-I light source, and has excellent type-I reactive oxygen species generation ability and photothermal generation ability, which can realize high signal-to-noise ratio fluorescence imaging detection of biological tissues and efficient photodynamic-photothermal combined tumor treatment under hypoxic conditions, and has great potential application value in the biomedical field, especially in tumor diagnosis and treatment.

[0007] To achieve the above purpose, the present invention provides the following solutions:

[0008] The present invention provides a type-I photosensitizer with NIR-I absorption and NIR-II emission characteristics, having the structure shown in Formula I:

[0009] Wherein, R1 is a straight-chain or branched-chain alkyl with 1-10 carbon atoms (preferably 1-6 carbon atoms) or a straight-chain or branched-chain alkynyl with 3-6 carbon atoms (preferably 5 carbon atoms), R2 is a straight-chain or branched-chain alkyl with 1-20 carbon atoms or R3 is a straight-chain or branched-chain alkyl with 1-10 carbon atoms (preferably 1-6 carbon atoms), n is 1 or 2, X - represents an anion, such as Cl - 、Br - 、I - 、NO3 - or PF4 - .

[0010] Preferably, R1 is methyl, ethyl, isopropyl or

[0011] Preferably, R2 is

[0012] Preferably, n is 1.

[0013] Preferably, X - is I - and PF4 - .

[0014] The present invention provides a method for preparing the above-mentioned photosensitizer, comprising the following steps:

[0015] (1) React compound 1 with IR1 under the action of sodium hydride to obtain compound 2; under nitrogen protection, add the Grignard reagent CH3ClMg to the solution of compound 2, heat the reaction, cool to room temperature, and react with hydrochloric acid solution and saturated KX solution to obtain compound 3;

[0016]

[0017] (2) Under nitrogen protection, carry out Buchwald-Harwig coupling reaction on compound 4 and R2NH2 under the action of a palladium catalyst to obtain compound 5; under nitrogen protection, add n-butyllithium to compound 5 and react with 2-isopropoxy-4,4,5,5-tetramethyl-1,3,2-dioxaborolane to generate compound 6;

[0018]

[0019] (3) Under nitrogen protection, carry out Suzuki coupling reaction on compound 6 and 5-bromothiophene-2-carbaldehyde or 5-bromo-2,2'-bithiophene-5'-carbaldehyde under the action of a palladium catalyst to obtain compound 7;

[0020] n is 1 or 2,

[0021] (4) Under nitrogen protection, carry out Knoevenagel condensation reaction on compound 3 and compound 7 to obtain the photosensitizer.

[0022] n is 1 or 2.

[0023] Further, in the step (1), the molar ratio of compound 1 to IR1 is 1:1 - 2, the molar ratio of compound 1 to NaH is 1:1.5 - 2, the reaction temperature is 0 - 40 °C, and the reaction time is 6 - 24 hours; the molar ratio of compound 2 to CH3ClMg is 1:1 - 2, the reaction temperature is 40 - 100 °C, and the reaction time is 1 - 6 hours.

[0024] Further, in the step (2), the molar ratio of compound 4 to R2NH2 is 1:1 - 2, and the palladium catalyst is one or more of tris(dibenzylideneacetone)dipalladium, palladium acetate, tetrakis(triphenylphosphine)palladium, and (diphenylphosphino)ferrocene dichloropalladium. The reaction temperature is 90 - 150 °C, and the reaction time is 12 - 48 hours. The molar ratio of compound 5 to n-BuLi is 1:1 - 1.5, and the molar ratio of compound 5 to 2-isopropoxy-4,4,5,5-tetramethyl-1,3,2-dioxaborolane is 1:1 - 2. The reaction temperature is -78 °C, and the reaction time is 1 - 6 hours.

[0025] Further, in the step (3), the molar ratio of compound 6 to 5-bromothiophene-2-carbaldehyde or 5-bromo-2,2'-bithiophene-5'-carbaldehyde is 1:1 - 3, and the palladium catalyst is one or more of tris(dibenzylideneacetone)dipalladium, palladium acetate, tetrakis(triphenylphosphine)palladium, and (diphenylphosphino)ferrocene dichloropalladium. The reaction temperature is 60 - 150 °C, and the reaction time is 12 - 48 hours.

[0026] Further, in the step (4), the molar ratio of compound 3 to compound 7 is 1:0.5 - 2, and the solvent used is one or more of acetonitrile, methanol, ethanol, isopropanol, and n-butanol. The reaction temperature is 50 - 100 °C, and the reaction time is 1 - 24 hours.

[0027] Another object of the present invention is to provide a nanoparticle prepared from the type I photosensitizer of the present invention.

[0028] Preferably, the type I photosensitizer of the present invention is mixed with a coating agent (such as DSPE-PEG, PS-PEG, F127) and an organic solvent (such as THF, DMSO, and DMF), then dropped into deionized water and ultrasonicated, and then ultrafiltered and concentrated to remove the organic solvent to obtain a water-dispersed nanoparticle.

[0029] Another object of the present invention is to provide the application of the type I photosensitizer or nanoparticle of the present invention in the preparation of a cell or tissue fluorescence imaging detection reagent.

[0030] Another object of the present invention is to provide the application of the type I photosensitizer or nanoparticle of the present invention in the preparation of a photodynamic and photothermal therapy drug.

[0031] The beneficial effects of the present invention are as follows:

[0032] The photosensitizer molecule provided by the present invention uses a dithienopyrrole derivative with strong electron-donating ability as the electron donor, a benzo[cd]indolium salt with strong electron-withdrawing ability as the electron acceptor, and introduces mono / dithienyl as the π-bridge, significantly enhancing the intramolecular charge transfer (ICT) effect, effectively reducing its HOMO-LUMO energy gap, shifting its absorption wavelength and emission wavelength to NIR-I and NIR-II respectively, thereby improving the penetration ability of biological tissues, achieving high-resolution and high signal-to-noise ratio NIR-II fluorescence imaging detection in living mice, and providing strong support for precision biomedical imaging. In addition, due to its strong ICT effect and long conjugated structure, the energy difference between the lowest excited singlet state (S1) and the excited triplet state (T) of the photosensitizer molecule provided by the present invention is significantly reduced, effectively promoting the intersystem crossing (ISC) process, and further improving its reactive oxygen species generation ability. More importantly, the photosensitizer provided by the present invention can generate a large amount of reactive oxygen species through a hypoxia-dependent type I reaction pathway, thus overcoming the limitation of the tumor hypoxic microenvironment on PDT treatment and making it more advantageous in tumor treatment. At the same time, the photosensitizer provided by the present invention also has excellent photothermal generation ability and photothermal stability, and can cooperate with PDT to achieve efficient combined treatment of tumors. In summary, the photosensitizer provided by the present invention can not only achieve high signal-to-noise ratio fluorescence imaging detection of biological tissues, but also perform efficient photodynamic-photothermal combined tumor treatment under hypoxic conditions, and has great potential application value in the biomedical field, especially in tumor diagnosis and treatment.

[0033] The present invention also provides a preparation method of the type I photosensitizer with NIR-I absorption and NIR-II emission characteristics described in the above technical solution. The reaction conditions are simple and easy to implement, with strong controllability and high repeatability, and have universal applicability. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 is the nuclear magnetic resonance spectrum of DTP-T-Indol of the present invention ( 1 H NMR).

[0035] Figure 2 is the nuclear magnetic resonance spectrum of DTP-T-Indol of the present invention ( 13 C NMR).

[0036] Figure 3 is the ESI-MS mass spectrum of DTP-T-Indol of the present invention.

[0037] Figure 4Normalized absorption spectra and fluorescence spectra of DTP-T-Indol (a), DTP-TT-Indol (b), DTP-OMe-TT-Indol (c) of the present invention, and comparative examples DTP-vinyl-Indol (d) and DTP-OMe-divinyl-Indol (e) in dichloromethane solution.

[0038] Figure 5 Curves showing the change of fluorescence intensity at 525 nm with irradiation time under 808 nm laser irradiation for the mixed solution of DTP-T-Indol (a) and DTP-TT-Indol (b) of the present invention and the total ROS scavenger DCFH.

[0039] Figure 6 For DTP-T-Indol (a) and DTP-TT-Indol (b) of the present invention and 1 Curves showing the change of absorbance at 418 nm with irradiation time under 808 nm laser irradiation for the mixed solution of DTP-T-Indol (a) and DTP-TT-Indol (b) of the present invention and the O2 scavenger DPBF.

[0040] Figure 7 Curves showing the change of fluorescence intensity at 515 nm with irradiation time under 808 nm laser irradiation for the mixed solution of DTP-T-Indol (a) and DTP-TT-Indol (b) of the present invention and the ·OH scavenger HPF.

[0041] Figure 8 For DTP-T-Indol (a) and DTP-TT-Indol (b) of the present invention and O2· - Curves showing the change of fluorescence intensity at 527 nm with irradiation time under 808 nm laser irradiation for the mixed solution of DTP-T-Indol (a) and DTP-TT-Indol (b) of the present invention and the O2· scavenger DHR123.

[0042] Figure 9 Hydrodynamic particle size and transmission electron microscope images of DTP-T-Indol NPs (a) and DTP-TT-Indol NPs (b) of the present invention.

[0043] Figure 10 Normalized absorption spectra and fluorescence spectra of DTP-T-Indol NPs (a) and DTP-TT-Indo NPs (b) of the present invention in aqueous solution.

[0044] Figure 11 Curves showing the change of fluorescence intensity at 525 nm with irradiation time under 808 nm laser irradiation for the mixed solution of DTP-T-Indol NPs of the present invention and the total ROS scavenger DCFH.

[0045] Figure 12 For DTP-T-Indol NPs of the present invention and1 The change curve of the absorbance at 418 nm of the mixed solution of O2 scavenger DPBF with the irradiation time under 808 nm laser irradiation.

[0046] Figure 13 This is the change curve of the fluorescence intensity at 515 nm of the mixed solution of DTP-T-Indol NPs and ·OH scavenger HPF of the present invention with the irradiation time under 808 nm laser irradiation.

[0047] Figure 14 This is for DTP-T-Indol NPs and O2· of the present invention - The change curve of the fluorescence intensity at 527 nm of the mixed solution of scavenger DHR123 with the irradiation time under 808 nm laser irradiation.

[0048] Figure 15 This is the infrared thermal image (a) and the temperature rise curve (b) of the 50 μg / mL aqueous solution of DTP-T-Indol NPs of the present invention continuously irradiated with 808 nm (0.8 W / cm 2 ) laser for 6 minutes.

[0049] Figure 16 This is the temperature rise curve of the aqueous solutions of DTP-T-Indol NPs with different concentrations of the present invention continuously irradiated with 808 nm (0.8 W / cm 2 ) laser for 6 minutes and the temperature rise curve of the 50 μg / mL aqueous solution of DTP-T-Indol NPs continuously irradiated with 808 nm laser with different powers for 6 minutes.

[0050] Figure 17 This is the cooling cycle photothermal stability diagram of the 50 μg / mL aqueous solution of DTP-T-Indol NPs of the present invention under 808 nm laser (0.8 W / cm 2 ) irradiation.

[0051] Figure 18 This is the cell survival rate after the co-incubation of DTP-T-Indol NPs of the present invention with NIH 3T3 mouse embryonic fibroblasts, MCF-7 human breast cancer cells and 4T1 mouse breast cancer cells for 24 hours.

[0052] Figure 19 This is the cell survival rate of 4T1 mouse breast cancer cells co-incubated with the present invention and irradiated with 808 nm laser (0.8 W / cm 2 ) for 5 minutes.

[0053] Figure 20Calcein-AM (green, live cells) / PI (red, dead cells) live / dead staining fluorescence images of 4T1 murine breast cancer cells co-incubated with the present invention and DTP-T-Indol NPs after different experimental treatments. Scale bar: 100 μm.

[0054] Figure 21 Analysis chart of the generation of total ROS, 2 O2, ·OH, and O2· in 4T1 murine breast cancer cells co-incubated with the present invention and DTP-T-Indol NPs after irradiation with 808 nm laser (0.8 W / cm 1 ) for 5 minutes. Scale bar: 20 μm. -

[0055] Figure 22 NIR-II fluorescence imaging maps of the abdominal blood vessels of mice injected with DTP-T-Indol NPs via the tail vein of the present invention, irradiated with 808 nm laser, and collected under 1000 nm and 1300 nm long-pass filters, as well as the relative fluorescence intensity distribution maps and Gaussian fitting curves along the red dotted line in the corresponding images.

[0056] Figure 23 NIR-II fluorescence imaging maps of the lymphatic vessels of mice injected with DTP-T-Indol NPs via the tail vein of the present invention, irradiated with 808 nm laser, and collected under 1000 nm and 1300 nm long-pass filters, as well as the relative fluorescence intensity distribution maps and Gaussian fitting curves along the red dotted line in the corresponding images.

[0057] Figure 24 NIR-II fluorescence imaging maps of tumor-bearing mice injected with DTP-T-Indol NPs via the tail vein of the present invention at different time points, irradiated with 808 nm laser, under 1000 nm long-pass filter, and the fluorescence signal intensity ratio (T / N) of tumor to normal tissue at different time points.

[0058] Figure 25 Thermal imaging maps of tumor-bearing mice injected with PBS and DTP-T-Indol NPs via the tail vein of the present invention respectively during continuous irradiation with 808 nm laser for 5 minutes and the change curve of tumor temperature with irradiation time.

[0059] Figure 26 Change curves of relative tumor volume and body weight of tumor-bearing mice of the present invention after different treatments (PBS, PBS+Laser, DTP-T-Indol NPs, and DTP-T-Indol NPs+Laser) within fourteen days (n = 5).

[0060] Figure 27Ex vivo tumor masses and photographs of tumor-bearing mice in different experimental groups (PBS, PBS+Laser, DTP-T-Indol NPs, and DTP-T-Indol NPs+Laser) on the 14th day after treatment with the present invention.

[0061] Figure 28 H&E staining pictures of important tissues and organs of tumor-bearing mice in different experimental groups (PBS, PBS+Laser, DTP-T-Indol NPs, and DTP-T-Indol NPs+Laser) on the 14th day after treatment with the present invention (n = 5), scale bar: 100 μm. Detailed implementation manners

[0062] The present invention provides a type I photosensitizer with NIR-I absorption and NIR-II emission characteristics, its preparation method and application. To make the objectives, technical solutions and effects of the present invention clearer and more definite, the present invention is further described in detail below. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention.

[0063] Example 1 Preparation of type I photosensitizer DTP-T-Indol

[0064]

[0065] The preparation method of the above-mentioned photosensitizer is as follows:

[0066]

[0067] Synthesis of compound 2: Sodium hydride (NaH, 60% mineral oil mixture, 2.40 g, 60 mmol) was added to a 250 mL round-bottom flask. Compound 1 (6.65 g, 39 mmol) was dissolved in 100 mL of anhydrous N,N-dimethylformamide (DMF), and under the condition of an ice-water bath, it was slowly dropped into the round-bottom flask. After the dropping was completed, the reaction was stirred for 20 minutes. Subsequently, iodoethane (7.33 g, 47 mmol) was dissolved in 20 mL of DMF and slowly dropped into the reaction flask, and the mixture was continuously stirred in an ice-water bath for 30 minutes. The reaction was transferred to room temperature and continued for 24 hours. After the reaction was completed, saturated ammonium chloride solution was slowly added to the flask, extracted with dichloromethane (DCM), washed three times with water, the organic phases were combined, and dried over anhydrous sodium sulfate. The solid was filtered off, the solvent was removed by reduced pressure concentration, and the crude product was separated by silica gel column chromatography. The eluent was a mixed solvent of petroleum ether and ethyl acetate (v / v = 10:1), and yellow solid compound 2 (7.37 g, yield: 95.0%) was obtained. 1 H NMR (400 MHz, DMSO-d6) δ8.21–8.15 (m, 1 H), 8.04 (d, J = 6.9 Hz,1 H), 7.80 (dd, J = 8.1, 7.0 Hz, 1H), 7.64 (d, J = 8.4 Hz, 1H), 7.55 (dd, J = 8.4, 7.0 Hz, 1H), 7.20 (d, J = 7.0 Hz, 1H), 3.93 (q, J = 7.1 Hz, 2H), 1.27 (t, J = 7.2 Hz, 3H). MS(ESI+): m / z calc. for C 13 H 11 NO: 197.08; found: 198.03 [M + H] + 。

[0068]

[0069] Synthesis of Compound 3: Under nitrogen protection, dissolve Compound 2 (1.97 g, 10.0 mmol) in 30 mL of anhydrous tetrahydrofuran (THF) and add it to a 100 mL three-necked flask, and stir under an ice-water bath. Then, add CH3ClMg (4 mL, 3.0 mmol / mL in THF) dropwise to the reaction system, and react in an oil bath at 60 °C for two hours. Cool to room temperature, slowly add 1 M hydrochloric acid solution (20 mL), then add 10 mL of saturated potassium iodide (KI) solution, and stir at room temperature for 30 minutes. Finally, filter under reduced pressure to obtain red solid Compound 3 (2.42 g, yield: 75%), and no further treatment is required. 1 H NMR (400 MHz, DMSO-d6) δ 8.99 (d, J = 7.2 Hz, 1H), 8.81 (d, J = 8.1 Hz, 1H), 8.55 (d, J = 7.3 Hz, 1H), 8.46 (d, J = 8.3 Hz, 1H), 8.18 (t, J = 7.6 Hz, 1H), 8.02 (t, J = 7.8 Hz, 1H), 4.72 (q, J = 7.3 Hz, 2H), 3.25 (s, 3H), 1.55 (t, J = 7.3 Hz, 3H). MS(ESI-): m / z calc. for C 14 H 14 N + : 196.11; found: 196.12 [M - I] + 。

[0070]

[0071] Synthesis of Compound 5a: Compound 4 (8.0 g, 24.7 mmol), 1-octylamine (3.99 g, 30.9 mmol), 1,1'-binaphthalene-2,2'-bis(diphenylphosphine) (BINAP, 1.54 g, 2.5 mmol), sodium tert-butoxide (5.93 g, 61.7 mmol) and 200 mL of anhydrous toluene were added to a 500 mL Schlenk flask. After two cycles of freezing with liquid nitrogen - pumping - thawing to remove oxygen and nitrogen supplementation, tris(dibenzylideneacetone)dipalladium(0) (Pd2(dba)3, 0.57 g, 0.62 mmol) was added. After three more cycles of freezing with liquid nitrogen - pumping - thawing to remove oxygen and backfilling with nitrogen, the reaction mixture was stirred at 110 °C for 24 hours under nitrogen protection. After cooling to room temperature, the reaction solution was extracted with DCM, washed twice with saturated NaCl solution and once with distilled water. The organic phases were combined, dried over anhydrous sodium sulfate, filtered to remove solids, and the solvent was removed by concentration under reduced pressure. The crude product was separated by silica gel column chromatography, and the eluent was a mixed solvent of petroleum ether and dichloromethane (v / v = 15:1), to obtain a pale yellow transparent liquid compound 5a (6.69 g, yield: 93%). 1 H NMR(400MHz,Chloroform - d)δ7.12(d,J=5.3Hz,2H),7.00(d,J=5.3Hz,2H),4.19(t,J=7.1Hz,2H),1.91–1.81(m,2H),1.34–1.21(m,10H),0.86(t,J=6.8Hz,3H).MS(ESI+):m / z calc.for C 16 H 21 NS2:291.11;found:314.12[M+Na] + 。

[0072]

[0073] Synthesis of Compound 6a: Compound 5a (4.0 g, 13.7 mmol) and anhydrous THF (40 mL) were added to a 100 mL Schlenk reaction flask. After two cycles of freezing with liquid nitrogen, pumping, and thawing to remove oxygen and then replenishing with nitrogen, it was placed in a -78 °C low-temperature reaction bath and stirred for 0.5 h. n-Butyllithium (n-BuLi, 6.6 mL, 2.5 mmol / L in hexane) was slowly added dropwise to the reaction flask with a syringe and stirring continued for 1 h. Subsequently, isopropoxyboronic acid pinacol ester (2.81 g, 15.1 mmol) was slowly added to the reaction flask with a syringe. After 4 h, it was transferred to room temperature and stirred for 4 h. The reaction was quenched by slowly adding saturated ammonium chloride solution, extracted with DCM, washed twice with saturated NaCl solution and twice with distilled water respectively. The organic phases were combined, dried over anhydrous sodium sulfate, and the solids were filtered off. The solvent was removed by concentration under reduced pressure to obtain a black oily liquid, Compound 6a. Due to its instability, it could be used directly in the next step without further purification and characterization.

[0074]

[0075] Synthesis of Compound 7a-I: Compound 6a (0.50 mmol), 5-bromo-2-thiophenecarboxaldehyde (124.2 mg, 0.65 mmol), potassium carbonate (207.3 mg, 1.50 mmol), toluene (16 mL), ethanol (4 mL), and distilled water (4 mL) were added to a 50 mL Schlenk reaction flask. After two cycles of freezing with liquid nitrogen, pumping, and thawing to remove oxygen and then replenishing with nitrogen, tetrakis(triphenylphosphine)palladium (Pd(PPh3)4, 57.8 mg, 0.05 mmol) was quickly added. After three more cycles of freezing with liquid nitrogen, pumping, and thawing to remove oxygen and then backfilling with nitrogen, the reaction was stirred at 90 °C under nitrogen protection for 24 h. After cooling to room temperature, the reaction solution was extracted with DCM, washed twice with saturated NaCl solution and once with distilled water. The organic phases were combined, dried over anhydrous sodium sulfate, the solids were filtered off, and the solvent was removed by concentration under reduced pressure. The crude product was separated by silica gel column chromatography, and the eluent was a mixed solvent of petroleum ether and dichloromethane (v / v = 1:1) to obtain an orange-yellow solid, Compound 7a-I (144.0 mg, two-step yield: 72%). 1 1H NMR (400 MHz, Chloroform-d) δ 7.12 (d, J = 5.3 Hz, 2H), 7.00 (d, J = 5.3 Hz, 2H), 4.19 (t, J = 7.1 Hz, 2H), 1.91–1.81 (m, 2H), 1.34–1.21 (m, 10H), 0.86 (t, J = 6.8 Hz, 3H). MS (ESI+): m / z calc. for C 21 1H 23 1NOS3: 401.09, found: 401.28.

[0076]

[0077] Synthesis of DTP-T-Indol: Compound 3 (97.0 mg, 0.3 mmol), compound 7a-I (132.5 mg, 0.33 mmol), and anhydrous ethanol (10 mL) were added to a 25 mL Schlenk flask. After three cycles of freezing with liquid nitrogen, pumping, and thawing to remove oxygen and backfilling with nitrogen, the mixture was stirred at 80 °C for 3 hours under nitrogen protection. After cooling to room temperature, the solvent was removed by rotary evaporation under reduced pressure. The crude product was separated by silica gel column chromatography, and the eluent was a mixed solvent of methanol and dichloromethane (v / v = 1:20), yielding a dark green solid powder of DTP-T-Indol (144.5 mg, yield: 68%). 1 H NMR (400 MHz, DMSO-d6) δ 9.22 (d, J = 7.3 Hz, 1H), 9.00 (d, J = 15.4 Hz, 1H), 8.60 (d, J = 7.7 Hz, 1H), 8.25–8.14 (m, 3H), 8.11 (t, J = 7.7 Hz, 1H), 7.96 (s, 1H), 7.87 (t, J = 7.8 Hz, 1H), 7.70 (d, J = 4.0 Hz, 1H), 7.52 (d, J = 5.3 Hz, 1H), 7.45 (d, J = 15.3 Hz, 1H), 7.28 (d, J = 5.3 Hz, 1H), 4.76 (q, J = 6.6 Hz, 2H), 4.28 (t, J = 6.8 Hz, 2H), 1.84–1.76 (m, 2H), 1.51 (t, J = 7.2 Hz, 3H), 1.28–1.15 (m, 10H), 0.82 (t, J = 6.8 Hz, 3H). 13 C NMR (101 MHz, DMSO) δ 160.67, 151.74, 148.82, 146.60, 145.57, 141.88, 140.42, 140.35, 137.12, 134.90, 133.47, 132.09, 131.28, 130.35, 129.98, 129.03, 124.76, 118.62, 118.42, 115.69, 113.70, 113.41, 112.42, 48.07, 32.65, 31.16, 30.10, 30.01, 27.69, 23.52, 16.73, 15.42. MS (ESI+): m / z calc. for C 35 H 35 N2S3 + : 579.20, found: 579.10 [M-I] + 。

[0078] 1H NMR spectrum of DTP-T-Indol 1 is as shown in Figure 1 the following figure 13 13C NMR spectrum is as shown in Figure 2 the following figure Figure 3 and ESI-MS spectrum is as shown in

[0079] Preparation of Photosensitizer DTP-TT-Indol in Example 2

[0080]

[0081] The preparation method of the above photosensitizer is as follows:

[0082] The synthesis of compounds 2, 3, 5a and 6a is the same as that in Example 1.

[0083]

[0084] Synthesis of compound 7b-II: Add compound 6a (208.7 mg, 0.50 mmol), 5-bromo-2,2'-bithiophene-5'-carbaldehyde (177.6 mg, 0.65 mmol), potassium carbonate (207.3 mg, 1.50 mmol), toluene (16 mL), ethanol (4 mL) and distilled water (4 mL) into a 50 mL Schlenk reaction flask. After two cycles of liquid nitrogen freezing - pumping - thawing to remove oxygen and nitrogen supplementation, quickly add tetrakis(triphenylphosphine)palladium (Pd(PPh3)4, 57.8 mg, 0.05 mmol). After three cycles of liquid nitrogen freezing - pumping - thawing to remove oxygen and backfill with nitrogen, stir the reaction at 90 °C for 24 hours under nitrogen protection. Cool to room temperature, add DCM to extract the reaction solution, wash twice with saturated NaCl solution and once with distilled water, combine the organic phases, dry with anhydrous sodium sulfate, filter off the solids, concentrate the solvent under reduced pressure, and separate the crude product by silica gel column chromatography. The eluent is a mixed solvent of petroleum ether and dichloromethane (v / v = 1:1) to obtain an orange - red solid compound 7a-II (169.3 mg, yield: 70%). 1 1HNMR(400MHz,Chloroform - d)δ7.12(d,J = 5.3Hz,2H),7.00(d,J = 5.3Hz,2H),4.19(t,J = 7.1Hz,2H),1.91–1.81(m,2H),1.34–1.21(m,10H),0.86(t,J = 6.8Hz,3H).MS(ESI+):m / z calc.for C 25 H 25 NOS4:483.08;found:506.12[M+Na] + .

[0085]

[0086] Synthesis of DTP-TT-Indol: Compound 3 (97.0 mg, 0.3 mmol), compound 7a-II (159.6 mg, 0.33 mmol), and anhydrous ethanol (10 mL) were added to a 25 mL Schlenk flask. After three cycles of freezing with liquid nitrogen, pumping, and thawing to remove oxygen and backfilling with nitrogen, the mixture was stirred at 80 °C for 3 hours under nitrogen protection. It was cooled to room temperature, the solvent was removed by rotary evaporation under reduced pressure, the residue was dissolved in DCM, washed twice with saturated NaCl solution and once with distilled water, the organic phases were combined, dried over anhydrous sodium sulfate, the solid was filtered off, the solvent was removed by concentration under reduced pressure, and the crude product was separated by silica gel column chromatography. The eluent was a mixed solvent of methanol and dichloromethane (v / v = 1:15), and dark green solid powder DTP-TT-Indol (147 mg, yield: 62%) was obtained. 1 HNMR (400 MHz, DMSO-d6) δ 9.13 (s, 2H), 8.57 (s, 1H), 8.16 (s, 3H), 7.86 (d, J = 7.4 Hz, 1H), 7.58 (d, J = 25.0 Hz, 3H), 7.46–7.28 (m, 3H), 7.21 (d, J = 5.3 Hz, 2H), 4.75 (s, 2H), 4.20 (t, J = 6.7 Hz, 2H), 1.79–1.72 (m, 2H), 1.50 (t, J = 6.9 Hz, 3H), 1.24–1.16 (m, 10H), 0.82 (t, J = 6.7 Hz, 3H). MS (ESI+): m / z calc. for C 39 H 37 N2S4 + : 661.18, found: 661.05 [M-I] + 。

[0087] Preparation of Photosensitizer DTP-OMe-TT-Indol of Example 3, Type I

[0088]

[0089] The preparation method of the above photosensitizer is as follows:

[0090] The synthesis of compounds 2 and 3 was the same as in Example 1.

[0091]

[0092] Synthesis of Compound 5b: Compound 4 (5.0 g, 15.4 mmol), anisidine (2.28 g, 18.5 mmol), 1,1'-binaphthalene-2,2'-bis(diphenylphosphine) (BINAP, 0.96 g, 1.54 mmol), sodium tert-butoxide (3.71 g, 38.5 mmol) and 80 mL of anhydrous toluene were added to a 200 mL Schlenk flask. After two cycles of freezing with liquid nitrogen - pumping - thawing to remove oxygen and nitrogen supplementation, tris(dibenzylideneacetone)dipalladium(0) (Pd2(dba)3, 0.36 g, 0.39 mmol) was added. After three more cycles of freezing with liquid nitrogen - pumping - thawing to remove oxygen and backfilling with nitrogen, the reaction mixture was stirred at 110 °C for 24 hours under nitrogen protection. After cooling to room temperature, the reaction solution was extracted with DCM, washed twice with saturated NaCl solution and once with distilled water. The organic phases were combined, dried over anhydrous sodium sulfate, filtered to remove solids, and the solvent was removed by concentration under reduced pressure. The crude product was separated by silica gel column chromatography with a mixed solvent of petroleum ether and dichloromethane (v / v = 15:1) to obtain a pale yellow transparent liquid compound 5b (3.64 g, yield: 85%). 1 1H NMR (400 MHz, Chloroform - d) δ 7.49 (d, J = 9.0 Hz, 2H), 7.16 (d, J = 5.3 Hz, 2H), 7.09 (d, J = 5.3 Hz, 2H), 7.05 (d, J = 9.0 Hz, 2H), 3.88 (s, 3H).

[0093]

[0094] Synthesis of Compound 6b: Compound 5b (2.0 g, 7.02 mmol) and anhydrous THF (20 mL) were added to a 50 mL Schlenk reaction flask. After two cycles of freezing with liquid nitrogen - pumping - thawing to remove oxygen and nitrogen supplementation, it was placed in a -78 °C low - temperature reaction bath and stirred for 0.5 hour. n - Butyllithium (n - BuLi, 3.3 mL, 2.5 mmol / L in hexane) was slowly added dropwise to the reaction flask with a syringe and stirring was continued for 1 hour. Subsequently, isopropoxyboronic acid pinacol ester (1.44, 7.72 mmol) was slowly added to the reaction flask with a syringe. After 4 hours, it was transferred to room temperature and stirred for 4 hours. The reaction was quenched by slowly adding saturated ammonium chloride solution, extracted with DCM, washed twice with saturated NaCl solution and twice with distilled water. The organic phases were combined, dried over anhydrous sodium sulfate, filtered to remove solids. The solvent was removed by concentration under reduced pressure to obtain a black oily liquid compound 6b. Due to its instability, it could be used directly in the next step without further purification and characterization.

[0095]

[0096] Synthesis of Compound 7b: Compound 6b (1.00 mmol), 5-bromo-2,2'-dithiophene-5'-carbaldehyde (355.1 mg, 1.30 mmol), potassium carbonate (207.3 mg, 3.00 mmol), toluene (32 mL), ethanol (8 mL), and distilled water (8 mL) were added to a 100 mL Schlenk reaction flask. After two cycles of freezing with liquid nitrogen - pumping - thawing and nitrogen supplementation, tetrakis(triphenylphosphine)palladium (Pd(PPh3)4, 115.6 mg, 0.10 mmol) was quickly added. After three more cycles of freezing with liquid nitrogen - pumping - thawing to remove oxygen and backfilling with nitrogen, the reaction mixture was stirred at 90 °C for 24 hours under nitrogen protection. After cooling to room temperature, the reaction solution was extracted with DCM, washed twice with saturated NaCl solution and once with distilled water. The organic phases were combined, dried over anhydrous sodium sulfate, filtered to remove solids, and the solvent was removed by rotary evaporation under reduced pressure. The crude product was separated by silica gel column chromatography, and the eluent was a mixed solvent of petroleum ether and dichloromethane (v / v = 1:1), obtaining an orange - red solid compound 7b (244.4 mg, yield: 51%). 1 1H NMR (400 MHz, DMSO - d6) δ 9.88 (s, 1H), 8.00 (s, 1H), 7.72–7.39 (m, 7H), 7.42–7.09 (m, 3H), 3.86 (s, 3H).

[0097]

[0098] Synthesis of DTP - OMe - TT - Indol: Compound 3 (97.0 mg, 0.3 mmol), compound 7b (157.6 mg, 0.33 mmol), and anhydrous ethanol (10 mL) were added to a 25 mL Schlenk flask. After three cycles of freezing with liquid nitrogen - pumping - thawing to remove oxygen and backfilling with nitrogen, the reaction mixture was stirred at 80 °C for 3 hours under nitrogen protection. After cooling to room temperature, the solvent was removed by rotary evaporation under reduced pressure. The residue was dissolved in DCM, washed twice with saturated NaCl solution and once with distilled water. The organic phases were combined, dried over anhydrous sodium sulfate, filtered to remove solids, and the solvent was removed by rotary evaporation under reduced pressure. The crude product was separated by silica gel column chromatography, and the eluent was a mixed solvent of methanol and dichloromethane (v / v = 1:15), obtaining a dark green solid powder DTP - OMe - TT - Indol (176 mg, yield: 75%). 11H NMR (400 MHz, DMSO-d6) δ 9.38–7.92 (m, 4H), 7.89–7.68 (m, 2H), 7.61–7.51 (m, 3H), 7.50–7.34 (m, 4H), 7.32 - 7.25 (m, 1H), 7.19–7.08 (m, 5H), 4.71 (s, 2H), 3.88 (s, 3H), 1.50 (t, J = 6.8 Hz, 3H). MS (ESI+): m / z calc. for C 38 H 27 N2OS4 + : 655.10, found: 655.50 [M-I] + 。

[0099] Preparation of Comparative Example 1 DTP-vinyl-Indol

[0100]

[0101] The preparation method of the above photosensitizer is as follows:

[0102] The synthesis of compounds 2, 3, and 5a is the same as that in Example 1.

[0103]

[0104] Synthesis of compound 8: Under an ice-water bath, anhydrous DMF (2.5 mL) and anhydrous DCM (10 mL) were added to a 150 mL round-bottom flask. Phosphorus oxychloride (POCl3, 2 mL, 21.5 mmol) was dissolved in 5 mL of anhydrous DCM and slowly added dropwise to the round-bottom flask, and stirred in an ice-water bath for 1 hour. Subsequently, compound 5a (4.77 g, 16.37 mmol) was dissolved in 20 mL of anhydrous DCM and slowly added dropwise to the above reaction solution. After the addition, the reaction was carried out at 40 °C overnight. After the reaction was completed, under an ice-water bath, saturated sodium bicarbonate (NaHCO3) solution was slowly added and stirred for 2 hours. DCM was added for extraction, and washed twice with 1 M hydrochloric acid solution, saturated NaCl solution, and distilled water respectively. The organic phases were combined, dried over anhydrous sodium sulfate, and the solid was filtered off. The solvent was removed by reduced pressure concentration, and the crude product was separated by silica gel column chromatography. The eluent was a mixed solvent of petroleum ether and dichloromethane (v / v = 1:2), and 4.76 g of yellow-green solid compound 8 was obtained (yield: 91%). 11H NMR (400 MHz, Chloroform-d) δ 9.87 (s, 1H), 7.65 (s, 1H), 7.37 (d, J = 5.4 Hz, 1H), 7.02 (d, J = 5.4 Hz, 1H), 4.22 (t, J = 7.1 Hz, 2H), 1.88 (dd, J = 8.9, 4.7 Hz, 2H), 1.35–1.21 (m, 10H), 0.90–0.82 (m, 3H). MS (ESI+): m / z calc. for C 17 H 21 NOS2: 319.11; found: 358.13 [M+K] + 。

[0105]

[0106] Synthesis of DTP-vinyl-Indol: Compound 3 (97.0 mg, 0.3 mmol), compound 8 (105.4 mg, 0.33 mmol), and absolute ethanol (10 mL) were added to a 25 mL Schlenk flask. After three cycles of freezing with liquid nitrogen, pumping, and thawing to remove oxygen and backfilling with nitrogen, the mixture was stirred at 80 °C for 3 hours under nitrogen protection. After cooling to room temperature, the solvent was removed by rotary evaporation under reduced pressure. The residue was dissolved in DCM, washed twice with saturated NaCl solution and once with distilled water. The organic phases were combined, dried over anhydrous sodium sulfate, filtered to remove solids, and concentrated under reduced pressure to remove the solvent. The crude product was separated by silica gel column chromatography, and the eluent was a mixed solvent of methanol and dichloromethane (v / v = 1:15) to obtain blue-green solid powder DTP-vinyl-Indol (127 mg, yield: 68%). 1 1H NMR (400 MHz, DMSO-d6) δ 9.20 (d, J = 9.0 Hz, 1H), 9.08 (d, J = 15.9 Hz, 1H), 8.56 (d, J = 7.8 Hz, 1H), 8.44 (s, 1H), 8.22–8.00 (m, 3H), 7.91–7.78 (m, 2H), 7.46 (d, J = 14.3 Hz, 1H), 7.38 (d, J = 5.3 Hz, 1H), 4.72 (d, J = 6.5 Hz, 2H), 4.34 (t, J = 6.4 Hz, 2H), 1.92–1.79 (m, 2H), 1.50 (t, J = 7.3 Hz, 3H), 1.31–1.19 (m, 10H), 0.82 (t, J = 6.5 Hz, 3H). MS (ESI+): m / z calc. for C 31 H 33 N2S2 + : 497.21, found: 497.15 [M-I] + 。

[0107] Preparation of Comparative Example 2 DTP-OMe-divinyl-Indol

[0108]

[0109] The preparation method of the above photosensitizer is as follows:

[0110] The synthesis of Compounds 2, 3, and 5b is the same as that in Example 3.

[0111]

[0112] Synthesis of Compound 9: Add Compound 5b (1.5 g, 5.26 mmol) and anhydrous THF (20 mL) into a 50 mL Schlenk reaction flask. After two cycles of freezing with liquid nitrogen - pumping - thawing to remove oxygen and replenishing nitrogen, place it in a -78 °C low - temperature reaction bath and stir for 0.5 hour. Slowly drip n - butyllithium (n - BuLi, 2.6 mL, 2.5 mmol / L in hexane) into the reaction flask with a syringe and continue stirring for 1 hour. Subsequently, slowly add 3 - dimethylaminopropenal (0.625 g, 6.31 mmol) into the reaction flask with a syringe. After 4 hours, transfer it to room temperature and stir for 4 hours. Slowly add saturated ammonium chloride solution to quench the reaction, extract with DCM, wash twice with saturated NaCl solution and distilled water respectively, combine the organic phases, dry with anhydrous sodium sulfate, and filter off the solids. Concentrate under reduced pressure to remove the solvent. The crude product is separated by silica gel column chromatography, and the eluent is a mixed solvent of petroleum ether and dichloromethane (v / v = 1:2) to obtain 1.21 g of orange - yellow solid Compound 9 (yield: 68%). 1 H NMR(400MHz,Chloroform - d)δ9.60(d,J=7.7Hz,1H),7.56(d,J=15.4Hz,1H),7.46(d,J=8.9Hz,2H),7.34–7.28(m,2H),7.10–7.04(m,3H),6.50(dd,J=15.4,7.7Hz,1H),3.90(s,3H).

[0113]

[0114] Synthesis of DTP-OMe-divinyl-Indol: Compound 3 (97.0 mg, 0.3 mmol), compound 9 (112.0 mg, 0.33 mmol), and absolute ethanol (10 mL) were added to a 25 mL Schlenk flask. After three cycles of freezing with liquid nitrogen, pumping, and thawing to remove oxygen and backfilling with nitrogen, the mixture was stirred at 80 °C for 3 hours under nitrogen protection. After cooling to room temperature, the solvent was removed by rotary evaporation under reduced pressure. The residue was dissolved in DCM, washed twice with saturated NaCl solution and once with distilled water. The organic phases were combined, dried over anhydrous sodium sulfate, filtered to remove solids, and concentrated under reduced pressure to remove the solvent. The crude product was separated by silica gel column chromatography, and the eluent was a mixed solvent of methanol and dichloromethane (v / v = 1:15) to obtain a blue-black solid powder DTP-OMe-divinyl-Indol (131 mg, yield: 68%). 1 H NMR (400 MHz, DMSO-d6) δ 9.11 (s, 1H), 8.76–8.49 (m, 2H), 8.24–7.94 (m, 4H), 7.90–7.78 (m, 1H), 7.74–7.65 (m, 2H), 7.60 (d, J = 8.9 Hz, 2H), 7.55–7.27 (m, 2H), 7.18 (dd, J = 7.1, 1.7 Hz, 3H), 4.68–4.55 (m, 2H), 3.87 (s, 3H), 1.49 (t, J = 7.2 Hz, 3H). MS (ESI+): m / z calc. for C 32 H 25 N2OS2 + : 517.14, found: 517.45 [M-I] + 。

[0115] Example 4

[0116] The compounds prepared in Examples 1-3 of the present invention and Comparative Examples 1 and 2 were respectively dissolved in dichloromethane, and their absorption spectra and fluorescence spectra in dichloromethane were measured using an ultraviolet-visible absorption spectrophotometer and a fluorescence spectrometer. The normalization results are as follows Figure 4As shown, the maximum absorption peaks and maximum emission peaks of DTP-T-Indol, DTP-TT-Indol, and DTP-OMe-TT-Indol in dichloromethane are located at 791 / 994 nm, 802 / 1105 nm, and 777 / 1180 nm, respectively. For the comparative examples DTP-vinyl-Indol and DTP-OMe-divinyl-Indol, the maximum absorption peaks and maximum emission peaks in dichloromethane are located at 702 / 925 nm and 700 / 883 nm, respectively. The test data show that the photosensitizers with mono / dithienyl as the π-bridge described in the present invention have long absorption in the near-infrared region (>700 nm) and long emission in the second near-infrared region, which is beneficial for penetrating deep biological tissues. However, the maximum emission peak of the photosensitizers with mono / divinyl as the π-bridge far from reaches the second near-infrared region, which is not conducive to fluorescence imaging of deep biological tissues.

[0117] Example 5

[0118] Test and analysis of the ROS generation ability of DTP-T-Indol and DTP-TT-Indol prepared in Examples 1 and 2 of the present invention are specifically implemented as follows:

[0119] Dihydrodichlorofluorescein (DCFH) was used to test the total ROS generation ability of DTP-T-Indol and DTP-TT-Indol. A methanol solution (3 mL) containing 40 μM DCFH and 10 μM DTP-T-Indol or DTP-TT-Indol was prepared, and the solution was irradiated with an 808 nm laser (0.8 W / cm 2 ). The fluorescence spectrum was recorded with a fluorescence spectrometer every 30 seconds of irradiation (excitation wavelength: 488 nm, emission wavelength range: 500 - 600 nm), and the change in fluorescence intensity of DCFH at 525 nm was analyzed. The test results are as Figure 5 shown. As the irradiation time of the 808 nm laser increased, the fluorescence intensity of DCFH at 525 nm in both groups of DTP-T-Indol and DTP-TT-Indol gradually increased, indicating that both DTP-T-Indol and DTP-TT-Indol have the ability to generate ROS, and the ROS generation ability of DTP-T-Indol is higher than that of DTP-TT-Indol.

[0120] 1,3-Diphenylbenzofuran (DPBF) was used to test the 1 O2 generation ability of DTP-T-Indol and DTP-TT-Indol. A methanol solution (3 mL) containing 50 μM DPBF and 10 μM DTP-T-Indol or DTP-TT-Indol was prepared, and the solution was irradiated with an 808 nm laser (0.8 W / cm 2)Irradiate the solution, record the absorption spectrum with an absorption spectrometer every 30 seconds (absorption wavelength range: 300 - 500 nm), and analyze the change in absorbance of DPBF at 418 nm. The test results are as Figure 6 shown. As the irradiation time of the 808 nm laser increases, the absorbance of DPBF in the DTP-T-Indol group at 418 nm gradually decreases, indicating that DTP-T-Indol can effectively generate 1 O2, while the absorbance of DPBF in DTP-TT-Indol at 418 nm hardly changes, indicating that DTP-TT-Indol has no 1 O2 generation ability.

[0121] Use hydroxyphenyl fluorescein (HPF) to test the ·OH generation ability of DTP-T-Indol and DTP-TT-Indol. Prepare a methanol solution (3 mL) containing 10 μM HPF and 10 μM DTP-T-Indol or DTP-TT-Indol, and irradiate the solution with an 808 nm laser (0.8 W / cm 2 ). Record the fluorescence spectrum with a fluorescence spectrometer every 30 seconds (excitation wavelength: 490 nm, emission wavelength range: 500 - 600 nm), and analyze the change in fluorescence intensity of HPF at 515 nm. The test results are as Figure 7 shown. As the irradiation time of the 808 nm laser increases, the fluorescence intensity of HPF in the DTP-T-Indol group at 515 nm gradually increases, indicating that DTP-T-Indol can effectively generate ·OH, while the fluorescence intensity of HPF in DTP-TT-Indol at 515 nm only changes slightly, indicating that the ·OH generation ability of DTP-TT-Indol is very weak.

[0122] Use dihydrorhodamine 123 (DHR123) to test the O2· - generation ability of DTP-T-Indol and DTP-TT-Indol. Prepare a methanol solution (3 mL) containing 10 μM HPF and 10 μM DTP-T-Indol or DTP-TT-Indol, and irradiate the solution with an 808 nm laser (0.8 W / cm 2 ). Record the fluorescence spectrum with a fluorescence spectrometer every 30 seconds (excitation wavelength: 488 nm, emission wavelength range: 500 - 625 nm), and analyze the change in fluorescence intensity of DHR123 at 527 nm. The test results are as Figure 8 shown. As the irradiation time of the 808 nm laser increases, the fluorescence intensity of DHR123 in both the DTP-T-Indol and DTP-TT-Indol groups at 527 nm gradually increases, indicating that both DTP-T-Indol and DTP-TT-Indol have O2·- generating ability, and the O2· of DTP-T-Indol - generating ability is higher than that of DTP-TT-Indol.

[0123] Example 6

[0124] To improve the water solubility and biocompatibility of DTP-T-Indol and DTP-TT-Indol of the present invention, the amphiphilic polymer DSPE-PEG 2000 is used as a coating agent to prepare DTP-T-Indol and DTP-TT-Indol into water-dispersible nanoparticles respectively. The specific implementation is as follows: Take 1 mg of DTP-T-Indol or DTP-TT-Indol and 10 mg of DSPE-PEG 2000 and dissolve them in 0.5 mL of DMSO. Under ultrasonic conditions, slowly drop the above mixed solution into 10 mL of deionized water, continue ultrasonic treatment for 10 minutes. After the ultrasonic treatment, filter with a 220 nm filter membrane and transfer it into an ultrafiltration tube, and ultrafiltrate and concentrate at a speed of 4500 rmp to finally obtain water-dispersible nanoparticles, which are named DTP-T-Indol NPs and DTP-TT-Indol NPs respectively.

[0125] Use a nanoparticle size analyzer and a transmission electron microscope to test the size and morphology of DTP-T-Indol NP and DTP-TT-Indol NPs. The test results are as Figure 9 shown. The test results of the nanoparticle size analyzer show that the hydrodynamic diameters of DTP-T-Indol NPs and DTP-TT-Indol NPs are 8.3 and 7.2 nm respectively. The test results of the transmission electron microscope show that both DTP-T-Indol NPs and DTP-TT-Indol NPs are typical spherical particles, and the diameters are about 6.2 and 6.5 nm respectively.

[0126] Use an ultraviolet-visible absorption spectrophotometer and a fluorescence spectrometer to test the absorption spectrum and fluorescence spectrum of DTP-T-Indol NPs and DTP-TT-Indol NPs in aqueous solution respectively. The normalization results are as Figure 10 shown. The maximum absorption peak and maximum emission peak of DTP-T-Indol NPs and DTP-TT-Indol NPs in water are located at 701 / 1067 nm and 705 / 1120 nm respectively. The test data show that both DTP-T-Indol NPs and DTP-TT-Indol NPs have long absorption in the near-infrared region (>700 nm) and long emission in the second near-infrared region, and are expected to achieve photodynamic therapy for deep tumors.

[0127] Example 7

[0128] Analysis of the ROS generation ability of the DTP-T-Indol NPs prepared in Example 6 of the present invention in aqueous solution was carried out as follows:

[0129] DCFH was used to test the total ROS generation ability of DTP-T-Indol NPs in aqueous solution. An aqueous solution (3 mL) containing 20 μM DCFH and 10 μM DTP-T-Indol NPs was prepared and irradiated with an 808 nm laser (0.8 W / cm 2 ). The fluorescence spectrum was recorded with a fluorescence spectrometer every 30 seconds (excitation wavelength: 488 nm, emission wavelength range: 500 - 600 nm), and the change in fluorescence intensity of DCFH at 525 nm was analyzed. The test results are as Figure 11 shown. As the irradiation time of the 808 nm laser increased, the fluorescence intensity of DCFH at 525 nm gradually increased, indicating that the DTP-T-Indol NPs prepared in Example 6 have ROS generation ability.

[0130] DPBF was used to test the 1 O2 generation ability of DTP-T-Indol NPs in aqueous solution. An aqueous solution (3 mL) containing 80 μM DPBF and 10 μM DTP-T-Indol was prepared and irradiated with an 808 nm laser (0.8 W / cm 2 ). The absorption spectrum was recorded with an absorption spectrometer every 30 seconds (absorption wavelength range: 300 - 550 nm), and the change in absorbance of DPBF at 418 nm was analyzed. The test results are as Figure 12 shown. As the irradiation time of the 808 nm laser increased, the absorbance of DPBF at 418 nm gradually decreased, indicating that the DTP-T-Indol NPs prepared in Example 6 have 1 O2 generation ability.

[0131] HPF was used to test the ·OH generation ability of DTP-T-Indol NPs in aqueous solution. An aqueous solution (3 mL) containing 10 μM HPF and 10 μM DTP-T-Indol was prepared and irradiated with an 808 nm laser (0.8 W / cm 2 ). The fluorescence spectrum was recorded with a fluorescence spectrometer every 30 seconds (excitation wavelength: 490 nm, emission wavelength range: 500 - 600 nm), and the change in fluorescence intensity of DCFH at 515 nm was analyzed. The test results are as Figure 13 shown. As the irradiation time of the 808 nm laser increased, the fluorescence intensity of HPF at 515 nm gradually increased, indicating that the DTP-T-Indol NPs prepared in Example 6 have ·OH generation ability.

[0132] Using DHR123 to test the O2· generation ability of DTP-T-Indol NPs in aqueous solution - Prepare an aqueous solution (3 mL) containing 10 μM HPF and 10 μM DTP-T-Indol, and irradiate the solution with an 808 nm laser (0.8 W / cm 2 ²). Record the fluorescence spectrum every 30 seconds with a fluorescence spectrometer (excitation wavelength: 488 nm, emission wavelength range: 500 - 625 nm), and analyze the change in the fluorescence intensity of DHR123 at 527 nm. The test results are as Figure 14 shown. As the irradiation time of the 808 nm laser increases, the fluorescence intensity of DHR123 at 527 nm gradually increases, indicating that DTP-T-Indol NPs have the O2· - generation ability.

[0133] Example 8

[0134] The photothermal performance test and analysis of the DTP-T-Indol NPs of the present invention are specifically implemented as follows:

[0135] Photothermal effect test: Prepare an aqueous solution of DTP-T-Indol NPs with a concentration of 50 μg / mL, and continuously irradiate the solution with an 808 nm laser (0.8 W / cm 2 ²) for 6 minutes. Monitor the solution temperature with an infrared thermal imager, record the solution temperature every 20 seconds, and take an infrared thermal image every minute. The test results are as Figure 15 shown. As the irradiation time of the 808 nm laser increases, the solution temperature rises rapidly. The solution temperature rises to 49.4 °C after 2 minutes of irradiation and reaches 60.2 °C after 6 minutes of irradiation, indicating that DTP-T-Indol NPs have good photothermal conversion performance.

[0136] Concentration-dependent test: Prepare aqueous solutions of DTP-T-Indol NPs with concentrations of 25, 50, 75, and 100 μg / mL respectively, and continuously irradiate the solutions with an 808 nm laser (0.8 W / cm 2 ²) for 6 minutes. Monitor the solution temperature with an infrared thermal imager and record the solution temperature every 20 seconds. The test results are as Figure 16 (a) shown. As the irradiation time of the 808 nm laser increases, the solution temperatures at all concentrations gradually increase; and as the concentration of DTP-T-Indol NPs increases, the increase in the solution temperature gradually becomes larger, indicating that the photothermal effect of DTP-T-Indol NPs is positively correlated with the particle concentration.

[0137] Power dependence test: An aqueous solution of DTP-T-Indol NPs with a concentration of 50 μg / mL was prepared. The solution was continuously irradiated with an 808 nm laser at power densities of 0.4, 0.6, 0.8, and 1 W / cm 2 for 6 minutes. The temperature of the solution was monitored by an infrared thermal imager, and the solution temperature was recorded every 20 seconds. The test results are shown in Figure 16 (b). As the irradiation time of the 808 nm laser increased, the solution temperature at all laser power densities gradually increased; moreover, as the laser power density increased, the amplitude of the solution temperature increase gradually became larger, indicating that the photothermal effect of DTP-T-Indol NPs was positively correlated with the laser power density.

[0138] Photothermal stability test: An aqueous solution of DTP-T-Indol NPs with a concentration of 50 μg / mL was prepared. After continuously irradiating the solution with an 808 nm laser (0.8 W / cm 2 ) for 6 minutes, the laser light source was turned off, and the solution was cooled for 10 minutes. This cycle was repeated 4 times, and the change in the solution temperature was monitored throughout the process by an infrared thermal imager. The test results are shown in Figure 17 . The DTP-T-Indol NPs solution could still maintain a stable temperature increase after 4 cycles of light irradiation, indicating that DTP-T-Indol NPs had excellent photothermal stability.

[0139] Example 9

[0140] The test and analysis of the photodynamic therapy effect of the DTP-T-Indol NPs of the present invention at the cellular level were specifically implemented as follows:

[0141] Cell dark toxicity test: NIH 3T3 mouse embryonic fibroblasts, MCF-7 human breast cancer cells, and 4T1 mouse breast cancer cells were selected as experimental cells for the cell dark toxicity test in the present invention. The above three types of cells were inoculated in a 96-well culture plate at a density of about 5×10 3 cells per well, and each well contained 100 μL of culture medium. They were placed in a cell culture incubator at 37 °C and 5% CO2 for adherent culture for 24 hours. Then, fresh culture medium containing different concentrations of DTP-T-Indol NPs was added, and co-culture continued for 24 hours. Subsequently, the culture medium was aspirated, and 100 μL of fresh culture medium and 20 μL of PBS solution containing 5 mg / mL MTT were added. After culturing for another four hours, the culture medium was aspirated, and 100 μL of DMSO was added to fully dissolve the formazan crystals by shaking. Finally, the absorbance of the well plate at 562 nm was detected using an enzyme-linked immunosorbent assay (ELISA) reader, and the cell survival rate was calculated according to the following formula.

[0142]

[0143] The experimental results are shown inFigure 18 As shown, even when the concentration of DTP-T-Indol NPs reaches 1000 μg / mL, the cell viabilities of NIH 3T3, MCF-7 and 4T1 cells are all above 85%, indicating that DTP-T-Indol NPs have no obvious cytotoxicity and have good biocompatibility.

[0144] Cell phototoxicity test: In this invention, 4T1 murine breast cancer cells are selected as the experimental cells for cell phototoxicity test. After the 4T1 cells are inoculated in a 96-well culture plate and adherently cultured for 24 hours, different concentrations of DTP-T-Indol NPs are added into the culture plate. After 12 hours, they are continuously irradiated with an 808 nm laser (0.8 W / cm 2 ) for 5 minutes respectively, and then placed in a cell incubator at 37°C and 5% CO2 and continue to be cultured for 12 hours. Other operation steps are the same as those in the cell dark toxicity experiment. The experimental results are as Figure 19 shown. Under the irradiation of an 808 nm laser (0.8 W / cm 2 ), the viability of 4T1 cells shows a concentration-dependent decreasing trend. When the concentration is 1000 μg / mL, only 24% of the cells survive after 5 minutes of light irradiation, indicating that in the presence of DTP-T-Indol NPs, tumor cells can be killed by 808 nm laser irradiation.

[0145] Calcein AM / PI live-dead staining: After the 4T1 cells are inoculated in a 24-well culture plate and adherently cultured for 24 hours, culture media containing PBS and DTP-T-Indol NPs (where the concentration of DTP-T-Indol NPs is 1 mg / mL) are added into the well plates respectively. After continuing to be cultured for 12 hours, light irradiation (808 nm laser, 0.8 W / cm 2 , continuously irradiated for 5 minutes) or non-light irradiation treatment is given, and they are marked as "PBS", "PBS+Laser", "DTP-T-Indol NPs" and "DTP-T-Indol NPs+Laser" groups. Subsequently, calcein (Calcein AM) and propidium iodide (PI) are added to stain the cells of different groups for 20 minutes under darkroom conditions. After rinsing three times with PBS, they are placed under an inverted fluorescence microscope for imaging. The experimental results are as Figure 20As shown, a large amount of red fluorescence signals (PI signals) were observed in the "DTP-T-Indol NPs+Laser" group. For the "PBS", "PBS+Laser" and "DTP-T-Indol NPs" groups, rich green fluorescence signals (Calcein AM signals) were observed throughout the field of view, indicating that neither the administration of DTP-T-Indol NPs alone nor laser irradiation would cause damage to cells. However, when DTP-T-Indol NPs and laser irradiation coexist, they can effectively kill cancer cells.

[0146] Example 10

[0147] The test analysis of the ROS generation ability of the DTP-T-Indol NPs of the present invention in cells is specifically implemented as follows:

[0148] DCFH-DA (2,7-dichlorofluorescein diacetate) was used to test the total ROS generation ability of DTP-T-Indol NPs in cells. After culturing 4T1 cells in a glass-bottom cell culture dish (Ф20mm, NEST) for 12 hours, culture media containing PBS and DTP-T-Indol NPs (where the concentration of DTP-T-Indol NPs is 1mg / mL) were added to the culture dish respectively. After continuing to culture for 12 hours, the culture media were aspirated, and fresh serum-free cell culture media containing 10 μM DCFH-DA were added respectively, and incubated at 37 °C for 20 minutes. The culture media were aspirated, washed three times with PBS, 1 mL of PBS was added to each, and light irradiation (808nm laser, 0.8W / cm 2 , continuously irradiated for 5 minutes) or non-light irradiation treatment was given, and they were denoted as the "PBS", "PBS+Laser", "DTP-T-Indol NPs" and "DTP-T-Indol NPs+Laser" groups, and then imaged under a laser confocal microscope (excitation wavelength: 488nm, emission wavelength range: 500-600nm). The experimental results are as Figure 21 (a) shown. The "PBS", "PBS+Laser" and "DTP-T-Indol NPs" groups did not show the characteristic green fluorescence of this indicator. In contrast, obvious green fluorescence signals were detected in the cells of the "DTP-T-Indol NPs+Laser" group, indicating that under laser irradiation, DTP-T-Indol NPs can efficiently generate reactive oxygen species in living cells.

[0149] SOSG (singlet oxygen green fluorescent probe), HPF and DHE (dihydroethidium) were used to test the 1 O2, ·OH and O2· in the DTP-T-Indol NPs prepared in Example 4 in cells respectively- Generating ability. The experimental procedures were the same as those for the detection of total intracellular ROS. When detecting 1 O2, the fresh serum-free cell culture medium containing 10 μM of SOSG was used to replace the fresh serum-free cell culture medium containing 10 μM of DCFH-DA, and the imaging parameters of the laser confocal microscope were: excitation wavelength: 504 nm, emission wavelength range: 510 - 600 nm; when detecting ·OH, the fresh serum-free cell culture medium containing 10 μM of HPF was used to replace the fresh serum-free cell culture medium containing 10 μM of DCFH-DA, and the imaging parameters of the laser confocal microscope were: excitation wavelength: 490 nm, emission wavelength range: 500 - 600 nm; when detecting O2· - , the fresh serum-free cell culture medium containing 10 μM of HPF was used to replace the fresh serum-free cell culture medium containing 10 μM of DHE, and the imaging parameters of the laser confocal microscope were: excitation wavelength: 535 nm, emission wavelength range: 550 - 700 nm. The experimental results are as shown in Figure 21 (b)(c)(d). In the "PBS", "PBS+Laser" and "DTP-T-Indol NPs" groups, the characteristic green fluorescence of the two indicators SOSG and HPF and the characteristic red fluorescence of the DHE indicator were not presented. In contrast, obvious green and red fluorescence signals were detected in the cells of the "DTP-T-Indol NPs+Laser" group, indicating that under laser irradiation, DTP-T-Indol NPs can efficiently generate 1 O2, ·OH and O2· - .

[0150] Example 11

[0151] The application test and analysis of the DTP-T-Indol NPs of the present invention in the NIR-II fluorescence imaging detection of blood vessels and lymphatic vessels in mice are specifically implemented as follows:

[0152] A 100 μL PBS solution containing 2 mg / mL of DTP-T-Indol NPs was injected into the tail vein of healthy female BALB / c mice, and then placed in a NIR-II fluorescence imaging system. Using an 808 nm laser as the excitation light source (70 mW / cm 2 ), the abdominal blood vessels and lymphatic vessels of the mice were imaged under 1000 and 1300 nm long-pass (LP) filters respectively. The imaging results of the abdominal blood vessels are as shown in Figure 22 . Under the 1000LP and 1300LP filters, the full width at half maximum of the abdominal blood vessels of the mice were 0.564 mm and 0.362 mm respectively, and the signal-to-noise ratio increased from 1.20 to 1.88. The imaging results of the lymphatic vessels are as shown in Figure 23As shown, under 1000LP and 1300LP filters, the full width at half maximum of mouse lymphatic vessels was 0.393 mm and 0.391 mm respectively, and the signal-to-noise ratio increased from 3.64 to 5.22. The above experimental results indicate that DTP-T-Indol NPs can perform high-quality NIR-II fluorescence imaging on mouse blood vessels and lymphatic vessels, and the images taken under the 1300LP filter have higher resolution and lower signal-to-noise ratio than those under the 1000LP filter.

[0153] Example 12

[0154] The test analysis of the application of the DTP-T-Indol NPs of the present invention in the NIR-II fluorescence imaging detection of mouse tumors is specifically implemented as follows:

[0155] Suspend 2×10 6 4T1 cells in PBS buffer solution and inject them subcutaneously into the right hind limb of BALB / c mice to construct a subcutaneous 4T1 tumor model in mice. It can be used when the tumor volume reaches about 100 mm 3 . Inject 100 μL of PBS solution containing DTP-T-Indol NPs at a concentration of 2 mg / mL into the tail vein of 4T1 tumor-bearing mice, and then perform NIR-II fluorescence imaging (808 nm laser, 70 mW / cm 2 , 1000LP) on the tumor-bearing mice at different set time points. The imaging results are as Figure 24 shown. Obvious fluorescence signals were detected in the tumor one hour after injection, and the fluorescence signals in the tumor gradually became stronger with time and began to weaken after 7 hours. The signal intensity ratio (T / N) between the tumor site and normal muscle reached 6.0 after 7 hours (higher than the ROSE gold standard value of 5.0), indicating that DTP-T-Indol NPs can not only be efficiently enriched in the tumor, but also be used for tumor detection with high sensitivity and high signal-to-noise ratio.

[0156] Example 13

[0157] The test analysis of the application of the DTP-T-Indol NPs of the present invention in the combined photodynamic and photothermal therapy of mouse tumors. The specific implementation is as follows:

[0158] 4T1 tumor-bearing mice were randomly divided into 4 groups, denoted as "PBS", "PBS+Laser", "DTP-T-Indol NPs", and "DTP-T-Indol NPs+Laser", with 5 mice in each group. Among them, mice in the "PBS" and "PBS+Laser" groups were injected with 100 μL of PBS via the tail vein, and mice in the "DTP-T-Indol NPs" and "DTP-T-Indol NPs+Laser" groups were injected with 100 μL of PBS solution containing 2 mg / mL of DTP-T-Indol NPs via the tail vein. 7 hours after injection, the tumor sites of mice in the "PBS+Laser" and "DTP-T-Indol NPs+Laser" groups were continuously irradiated with 808 nm laser (0.8 W / cm 2 ) for 5 minutes, and during the irradiation process, the temperature change of the tumor was monitored in real time with an infrared thermal imager. The thermal imaging maps and temperature change of the mouse tumors are as shown in Figure 25 . In the "DTP-T-Indol NPs+Laser" group, the tumor temperature rapidly increased from 36.6 °C to 50.7 °C within 5 minutes (ΔT = 14.1 °C), while only a slight temperature increase (ΔT = 3.5 °C) occurred in the "PBS+Laser" group, indicating that in the absence of DTP-T-Indol NPs, the temperature change of the tumor caused by laser can be ignored, but the tumor temperature can be significantly increased when DTP-T-Indol NPs and laser irradiation coexist. Therefore, DTP-T-Indol NPs have good photothermal effects in mice.

[0159] After laser treatment, the body weights of the tumor-bearing mice in each group were weighed every 2 days, and the tumor volumes were measured with a vernier caliper. The relative tumor volumes of the tumor-bearing mice are as shown in Figure 26 (a). Among them, the tumor volumes of the "PBS", "PBS+Laser", and "DTP-T-Indol NPs" groups showed an increasing trend throughout the monitoring period, while the tumor volume of the mice in the "DTP-T-Indol NPs+Laser" group gradually decreased after laser treatment and was completely ablated on the 14th day. The body weight changes of the tumor-bearing mice are as shown in Figure 26 (b). There was no significant difference in the body weights of the mice in the "DTP-T-Indol NPs", "PBS+Laser", and "DTP-T-Indol NPs+Laser" groups compared with those in the "PBS" group, indicating that the systemic toxicity of DTP-T-Indol NPs can be ignored and the laser tolerance is good.

[0160] 14 days after treatment, the mice in each group were euthanized, and the tumors were dissected out, weighed, and photographed. The masses and photos of the excised tumors of the mice are as shown in Figure 27As shown, the average tumor weights of the "PBS", "PBS+Laser", and "DTP-T-Indol NPs" groups were 0.402 g, 0.374 g, and 0.376 g, respectively, while the tumors in the mice of the "DTP-T-Indol NPs+Laser" group completely disappeared. The above results indicate that the combination of DTP-T-Indol NPs and laser irradiation has excellent anti-tumor effects and can ultimately eradicate tumors.

[0161] In addition, 14 days after treatment, the mice in each group were euthanized, and then the important organs of the heart, liver, spleen, lungs, and kidneys were removed for H&E tissue section analysis. The analysis results are as Figure 28 shown. Compared with the "PBS" group, no pathological abnormalities or inflammatory lesions were found in the other three groups, further demonstrating the good biosafety of DTP-T-Indol NPs.

[0162] Based on the above test data, it can be seen that the type I photosensitizer provided by the present invention with NIR-I absorption and NIR-II emission characteristics has simple and feasible conditions, strong controllability, and high type I reactive oxygen species generation ability. The photosensitizer provided by the present invention can not only achieve high signal-to-noise ratio bio-tissue fluorescence imaging detection in the NIR-II region, but also perform efficient combined photodynamic and photothermal therapy on tumors under hypoxic conditions, and has great potential application value in the biomedical field, especially in tumor diagnosis and treatment.

[0163] It should be understood that the application of the present invention is not limited to the above examples. For those of ordinary skill in the art, improvements or transformations can be made according to the above description, and all such improvements and transformations should fall within the protection scope of the appended claims of the present invention.

Claims

1. A type I photosensitizer, characterized in that, It has the structure shown in Formula I: Among them, R1 is a straight-chain or branched-chain alkyl group with 1 to 10 carbon atoms or a straight-chain or branched-chain alkynyl group with 3 to 6 carbon atoms, R2 is a straight-chain or branched-chain alkyl group with 1 to 20 carbon atoms or R3 is a straight-chain or branched-chain alkyl group with 1 to 10 carbon atoms, n is 1 or 2, X - represents an anion.

2. The photosensitizer of type I according to claim 1, characterized in that, R1 is methyl, ethyl, isopropyl or R2 is n is 1 or 2.

3. The photosensitizer of type I according to claim 1, wherein X - is Cl - , Br - , I - , NO3 - or PF4 - .

4. The preparation method of the type I photosensitizer according to any one of claims 1-3, characterized in that, It includes the following steps: (1) React compound 1 with IR1 under the action of sodium hydride to obtain compound 2; under nitrogen protection, add the Grignard reagent CH3ClMg to the solution of compound 2, heat the reaction, cool to room temperature, and react with hydrochloric acid solution and saturated KX solution to obtain compound 3, where R1 is a straight-chain or branched-chain alkyl group with 1-10 carbon atoms or a straight-chain or branched-chain alkynyl group with 3-6 carbon atoms; (2) Under nitrogen protection, compound 4 and R2NH2 are subjected to Buchwald-Harwig coupling reaction under the action of a palladium catalyst to obtain compound 5; under nitrogen protection, n-butyllithium is added to compound 5 and reacted with 2-isopropoxy-4,4,5,5-tetramethyl-1,3,2-dioxaborolane to generate compound 6, where R2 is a straight-chain or branched-chain alkyl group with 1 to 20 carbon atoms or R3 is a straight-chain or branched-chain alkyl group with 1 to 10 carbon atoms; (3) Under nitrogen protection, carry out a Suzuki coupling reaction between compound 6 and 5-bromothiophene-2-carbaldehyde under the action of a palladium catalyst to obtain compound 7; n is 1 or 2; (4) Under nitrogen protection, carry out a Knoevenagel condensation reaction between compound 3 and compound 7 to obtain the photosensitizer; n is 1 or 2.

5. The synthesis method according to claim 4, wherein In the said step (1), the molar ratio of compound 1 to R1X is 1:1-2, the molar ratio of compound 1 to NaH is 1:1.5-2, the reaction temperature is 0-40 °C, and the reaction time is 6-24 hours; the molar ratio of compound 2 to CH3ClMg is 1:1-2, the reaction temperature is 40-100 °C, and the reaction time is 1-6 hours; in the said step (2), the molar ratio of compound 4 to R2NH2 is 1:1-2, the palladium catalyst is one or more of tris(dibenzylideneacetone)dipalladium, palladium acetate, tetrakis(triphenylphosphine)palladium, (diphenylphosphino)ferrocene dichloropalladium, the reaction temperature is 90-150 °C, and the reaction time is 12-48 hours; the molar ratio of compound 5 to n-BuLi is 1:1-1.5, the molar ratio of compound 5 to 2-isopropoxy-4,4,5,5-tetramethyl-1,3,2-dioxaborolane is 1:1-2, the reaction temperature is -78 °C, and the reaction time is 1-6 hours; in the said step (3), the molar ratio of compound 6 to 5-bromothiophene-2-carbaldehyde or 5-bromo-2,2'-dithiophene-5'-carbaldehyde is 1:1-3, the palladium catalyst is one or more of tris(dibenzylideneacetone)dipalladium, palladium acetate, tetrakis(triphenylphosphine)palladium, (diphenylphosphino)ferrocene dichloropalladium, the reaction temperature is 60-150 °C, and the reaction time is 12-48 hours; in the said step (4), the molar ratio of compound 3 to compound 7 is 1:0.5-2, the solvent used is one or more of acetonitrile, methanol, ethanol, isopropanol, and n-butanol, the reaction temperature is 50-100 °C, and the reaction time is 1-24 hours.

6. A nanoparticle, characterized in that, It is prepared from the type I photosensitizer described in any one of claims 1-3.

7. The nanoparticle according to claim 6, wherein After mixing the one described in any one of claims 1-3 with a type I photosensitizer coating agent and an organic solvent, it is dropped into deionized water and ultrasonicated, and then ultrafiltered and concentrated to remove the organic solvent to obtain water-dispersed nanoparticles.

8. Use of the type I photosensitizer described in any one of claims 1-3 or the nanoparticles described in claim 6 or 7 in the preparation of a cell or tissue fluorescence imaging detection reagent.

9. Use of the type I photosensitizer described in any one of claims 1-3 or the nanoparticles described in claim 6 or 7 in the preparation of a photodynamic and photothermal therapy drug.

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