A near-infrared second-region crotonate dye and its preparation method and application

Abstract

The present invention belongs to the technical field of crotonate dyes, and in particular to a near-infrared second-region crotonate dye and its preparation method and application. The near-infrared second-region crotonate dye having a structure shown in Formula 1 provided by the present invention introduces an electron-donating group into the crotonate structure to expand the π conjugation degree of the system, and has a donor-acceptor-donor structure to form a rigid resonant plane structure that stabilizes the zwitterion, effectively regulating the absorption and emission wavelengths of the dye, and red-shifting it to a near-infrared region with a longer wavelength. Thus, the crotonate dye having a structure shown in Formula 1 provided by the present invention has good absorption properties in the near-infrared second region and can be used for photoacoustic imaging of tumor detection; at the same time, it has good fluorescence quantum yield and can be used for fluorescence imaging of tumor detection; and the near-infrared second-region crotonate dye provided by the present invention also has good photothermal conversion efficiency and can be used for photothermal therapy for tumor treatment.

Description

Technical Field

[0001] The invention belongs to the technical field of croconic acid dyes, and particularly relates to a near-infrared second-region croconic acid dye and a preparation method and application thereof. Background Art

[0002] Accurate detection, diagnosis, and resection of tumors rely on precise imaging systems. Compared to traditional bioimaging techniques such as radiation imaging, ultrasound imaging, and magnetic resonance imaging, fluorescence imaging and photoacoustic imaging show great application potential in diagnosis and guided surgical resection due to their high sensitivity and specificity, low cost, non-invasiveness, and improved biosafety. Many fluorescent materials have also been widely used in fluorescence imaging research, but most of them emit visible light and the near-infrared region I (NIR-I, 750-900nm). When entering biological tissue, they are absorbed and scattered to varying degrees and attenuated, thereby reducing imaging depth and contrast. The near-infrared region II (NIR-II) has become a hot topic in biomedical research due to its longer emission wavelength (1000-1700nm), stronger penetration into biological tissues, deeper detection depth, and higher spatial resolution.

[0003] Some existing inorganic materials such as rare earth down-conversion nanoparticles, carbon nanotubes, quantum dots, etc. can achieve near-infrared second region emission, but their emission wavelengths are mostly in the near-infrared first region, and the poor biosafety of heavy metals and slow metabolism after entering the living body limit their application. Summary of the Invention

[0004] The purpose of the present invention is to provide a near-infrared second-zone crotonate dye and its preparation method and application. The near-infrared second-zone crotonate dye provided by the present invention can not only be used for tumor photoacoustic and fluorescence imaging, but also display the tumor tissue boundary under the excitation of near-infrared light, providing a more objective reference for the complete resection of the tumor. In addition, it can also be used for photothermal therapy of tumors to effectively ablate tumor tissue.

[0005] In order to achieve the above object, the present invention provides the following technical solutions:

[0006] The present invention provides a near-infrared second-region crezone dye having a structure shown in Formula 1:

[0007]

[0008] The present invention provides a method for preparing the near-infrared second-region crezone dye described in the above technical solution, comprising the following steps:

[0009] Mixing a compound of formula 2, croconic acid, and a first organic solvent, and heating to perform a dehydration condensation reaction to obtain a near-infrared second-region croconic acid dye of formula 1;

[0010]

[0011] Preferably, the preparation method of the compound of the structure shown in Formula 2 comprises the following steps:

[0012] Mixing the compound of formula 3, a second organic solvent, and a Grignard reagent to perform a Grignard reaction to obtain a compound of formula 2;

[0013]

[0014] Preferably, the preparation method of the compound of the structure shown in Formula 3 comprises the following steps:

[0015] Mixing the compound of formula 4, sodium hydride, n-butyl iodide, and a third organic solvent to perform a nucleophilic substitution reaction to obtain a compound of formula 3;

[0016]

[0017] Preferably, the mass ratio of the compound of formula 2 to crotonic acid is 1:0.1 to 1.0;

[0018] The first organic solvent is 2-methoxyethanol; the ratio of the volume of the first organic solvent to the mass of the compound having the structure represented by Formula 2 is 100-200 mL:1 g.

[0019] Preferably, the Grignard reagent is methylmagnesium chloride; the ratio of the volume of the Grignard reagent to the mass ratio of the compound of the structure represented by Formula 3 is 1-10 mL:1 g;

[0020] The second organic solvent is tetrahydrofuran; the ratio of the volume of the second organic solvent to the mass ratio of the compound of the structure represented by Formula 3 is 5 to 20 mL:1 g.

[0021] Preferably, the mass ratio of the compound of the structure shown in Formula 4 to sodium hydride is 1:0.1 to 1.0;

[0022] The mass ratio of the compound of formula 4 to n-butane iodide is 1:1 to 5;

[0023] The third organic solvent is N,N-dimethylformamide; the ratio of the volume of the third organic solvent to the mass ratio of the compound of the structure represented by Formula 4 is 10-30 mL:1 g.

[0024] The present invention provides the use of the near-infrared second-region croconic acid dye described in the above technical solution or the near-infrared second-region croconic acid dye prepared by the preparation method described in the above technical solution in the preparation of a tumor imaging reagent; the tumor imaging reagent is used for photoacoustic imaging or fluorescence imaging of tumor detection.

[0025] The present invention provides the use of the near-infrared second-region crotonate dye described in the above technical solution or the near-infrared second-region crotonate dye prepared by the preparation method described in the above technical solution in the preparation of tumor cell photothermal therapy drugs.

[0026] Preferably, the tumor cells include one or more of breast cancer cells, oral cancer cells, liver cancer cells, lung cancer cells, gastric cancer cells, pancreatic cancer cells, colorectal cancer cells, bladder cancer cells and prostate cancer cells.

[0027] The present invention provides a near-infrared second-zone crotonate dye having a structure shown in Formula 1. The near-infrared second-zone crotonate dye having a structure shown in Formula 1 provided by the present invention has a symmetrical structure. The present invention expands the π conjugation degree of the system by introducing an electron-donating group into the crotonate structure, and has a donor-acceptor-donor structure to form a rigid resonant plane structure of a stable zwitterion, effectively regulating the absorption and emission wavelengths of the dye, and red-shifting it to a near-infrared region with a longer wavelength. Therefore, the near-infrared second-zone crotonate dye having a structure shown in Formula 1 provided by the present invention has good absorption properties in the near-infrared region and can be used for photoacoustic imaging for tumor detection; at the same time, it has good fluorescence quantum yield and can be used for fluorescence imaging for tumor detection; and the crotonate dye provided by the present invention also has good photothermal conversion efficiency and can be used for photothermal therapy for tumor treatment.

[0028] In summary, the near-infrared second-zone crotonate dye with a structure shown in Formula 1, which has absorption in the near-infrared region, provided by the present invention, can realize the combination of fluorescence imaging, photoacoustic imaging and photothermal therapy, which can not only realize the complementary advantages between imaging technologies and provide richer biological information for disease diagnosis, but also achieve the purpose of diagnosis and treatment at the same time, and avoid the risks and burdens brought by multiple injections of different contrast agents, and has broad application prospects.

[0029] The present invention provides a method for preparing a near-infrared second-range croconic acid dye according to the above technical solution, comprising the steps of mixing a compound represented by Formula 2, croconic acid, and a first organic solvent, and heating to carry out a dehydration condensation reaction to obtain a near-infrared second-range croconic acid dye represented by Formula 1. The preparation method provided by the present invention is simple, has a distinct reaction site, and is easy to control the reaction progress. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 This is the H NMR spectrum of the near-infrared second region crezone dye (IR1024) prepared in Example 2;

[0031] Figure 2 This is the NMR carbon spectrum of the near-infrared second region crezone dye (IR1024) prepared in Example 2;

[0032] Figure 3is the UV-visible absorption spectrum of the near-infrared second region crezone dye in Example 3;

[0033] Figure 4 is the fluorescence emission spectrum of the near-infrared second region crotonate dye in Example 3;

[0034] Figure 5 The photoacoustic signal spectrum of the near-infrared second region crotonate dye in Example 3;

[0035] Figure 6 This is a photothermal heating effect diagram of the near-infrared second region crotonate dye in Example 4;

[0036] Figure 7 This is a graph of the effect of near-infrared second-region crotonate dye on cancer cell survival rate in Example 5. DETAILED DESCRIPTION

[0037] The present invention provides a near-infrared second-region crezone dye having a structure shown in Formula 1:

[0038]

[0039] The near-infrared second-region crezone dye with the structure shown in Formula 1 provided by the present invention has a good molar absorption coefficient and photothermal conversion efficiency, can be used for photoacoustic imaging and photothermal therapy for tumor detection, has a good fluorescence quantum yield, and can be used for fluorescence imaging for tumor detection.

[0040] In the present invention, unless otherwise specified, all preparation raw materials / components are commercially available products well known to those skilled in the art.

[0041] The present invention provides a method for preparing the near-infrared second-region crezone dye described in the above technical solution, comprising the following steps:

[0042] Mixing a compound of formula 2, croconic acid, and a first organic solvent, and heating to perform a dehydration condensation reaction to obtain a near-infrared second-region croconic acid dye of formula 1;

[0043]

[0044] In the present invention, the method for preparing the compound of the structure shown in Formula 2 preferably comprises the following steps:

[0045] Mixing the compound of formula 3, a second organic solvent, and a Grignard reagent to perform a Grignard reaction to obtain a compound of formula 2;

[0046]

[0047] In the present invention, the method for preparing the compound of the structure shown in Formula 3 preferably comprises the following steps:

[0048] Mixing the compound of formula 4, sodium hydride, n-butyl iodide, and a third organic solvent to perform a nucleophilic substitution reaction to obtain a compound of formula 3;

[0049]

[0050] In the present invention, the mass ratio of the compound of the structure shown in Formula 4 to sodium hydride is preferably 1:0.1 to 1.0, more preferably 1:0.4. The mass ratio of the compound of the structure shown in Formula 4 to n-butane iodide is preferably 1:1 to 5, more preferably 1:1.67. The third organic solvent is preferably N,N-dimethylformamide; the ratio of the volume of the third organic solvent to the mass ratio of the compound of the structure shown in Formula 4 is preferably 10 to 30 mL:1 g, more preferably 19.5 mL:1 g. In the present invention, the N,N-dimethylformamide is preferably anhydrous N,N-dimethylformamide. The sodium hydride serves as a catalyst for the nucleophilic substitution reaction.

[0051] In the present invention, the order of mixing the compound of Formula 4, sodium hydride, n-butane iodide and the third organic solvent is preferably: dissolving the compound of Formula 4 in the third organic solvent, and then adding sodium hydride and n-butane iodide.

[0052] In the present invention, the temperature of the nucleophilic substitution reaction is preferably room temperature, the reaction time is preferably 2 hours, and the nucleophilic substitution reaction is carried out under stirring.

[0053] In the present invention, after the nucleophilic substitution reaction is completed, a nucleophilic substitution reaction solution is obtained. The nucleophilic substitution reaction solution is preferably washed and extracted sequentially to obtain an extracted organic phase; the extracted organic phase is desolventized to obtain a crude product; and the crude product is purified by column chromatography to obtain a pure compound having the structure represented by Formula 3. In the present invention, the washing reagent is preferably saturated saline. The extraction reagents are preferably ethyl acetate and water, with the volume ratio of ethyl acetate to water preferably being 1:1. The extracted organic phase is ethyl acetate. A specific embodiment of desolventization is preferably rotary evaporation. The crude product is preferably dissolved in ethyl acetate to obtain a sample solution for column chromatography purification. The column chromatography purification is preferably performed using a silica gel column, the eluent is preferably ethyl acetate, the elution rate is preferably 1 mL / min, and the elution volume is preferably 5 to 7 column volumes. The present invention preferably uses a mixed solvent of ethyl acetate and petroleum ether as a developing solvent to perform thin layer chromatography monitoring on the eluate, then collects components with an Rf value of 0.4-0.6, and vacuum rotary evaporates the ethyl acetate to obtain a pure product of the compound with the structure represented by Formula 3; the volume ratio of ethyl acetate to petroleum ether in the mixed solvent of ethyl acetate and petroleum ether is preferably 4:1.

[0054] In the present invention, the Grignard reagent is preferably methylmagnesium chloride (CH3MgCl); the ratio of the volume of the Grignard reagent to the mass of the compound represented by Formula 3 is preferably 1-10 mL:1 g, more preferably 5 mL:1 g. The second organic solvent is preferably tetrahydrofuran; the ratio of the volume of the second organic solvent to the mass of the compound represented by Formula 3 is preferably 5-20 mL:1 g, more preferably 10 mL:1 g. The tetrahydrofuran is preferably anhydrous tetrahydrofuran.

[0055] In the present invention, the order of mixing the compound represented by Formula 3, the second organic solvent and the Grignard reagent is preferably as follows: dissolving the compound represented by Formula 3 in tetrahydrofuran (THF), and then adding the Grignard reagent in an ice-water bath under nitrogen protection.

[0056] In the present invention, the Grignard reaction temperature is preferably 50-60°C, the moisturizing time is preferably 2 hours, and the Grignard reaction is preferably carried out under nitrogen protection.

[0057] In the present invention, after the Grignard reaction is completed, a Grignard reaction solution is obtained. Preferably, after the Grignard reaction solution is cooled to room temperature, perchloric acid is added to quench it to obtain a quenched reaction solution; the quenched reaction solution is precipitated with water, and solid-liquid separation is performed to obtain a solid-phase product; and the solid-phase product is dried to obtain a compound with the structure represented by Formula 2. In the present invention, the ratio of the volume of the perchloric acid to the mass of the compound with the structure represented by Formula 3 is preferably 1-10 mL:1 g, more preferably 2 mL:1 g. The drying is preferably vacuum freeze-drying; the vacuum freeze-drying temperature is preferably -40°C.

[0058] In the present invention, the structural formula of crezolic acid is:

[0059] In the present invention, the mass ratio of the compound represented by Formula 2 to crotonic acid is preferably 1:0.1 to 1.0, preferably 1:0.26. The first organic solvent is preferably 2-methoxyethanol; and the ratio of the volume of the first organic solvent to the mass of the compound represented by Formula 2 is preferably 100 to 200 mL:1 g, more preferably 148 mL / g.

[0060] In the present invention, the dehydration condensation reaction is preferably carried out under heating reflux conditions, and the heating reflux temperature is preferably 110-120° C.; the reaction insulation time is preferably 15 minutes.

[0061] In the present invention, after the dehydration condensation reaction is completed, a dehydration condensation reaction solution is obtained. The dehydration condensation reaction solution is preferably desolventized to obtain a concentrate. The concentrate is then purified by column chromatography to obtain a pure product of the croconic acid dye having the structure represented by Formula 1. In the present invention, the desolventization is preferably performed by rotary evaporation. The concentrate is preferably dissolved in dichloromethane to obtain a sample solution for column chromatography purification. The column chromatography purification is preferably performed using a silica gel column, the eluent is preferably dichloromethane, the elution rate is preferably 1 mL / min, and the elution volume is preferably 6 to 8 column volumes. The eluate is preferably subjected to thin-layer chromatography monitoring using a mixed solvent of dichloromethane and methanol as a developing solvent, and fractions having an Rf value preferably of 0.2-0.4 are collected. The dichloromethane is then rotary evaporated to dryness to obtain a pure product of the croconic acid dye having the structure represented by Formula 1. The volume ratio of dichloromethane to methanol in the mixed solvent of dichloromethane and methanol is preferably 10:1.

[0062] The preparation method of the near-infrared second-region crezone dye with the structure shown in Formula 1 provided by the present invention is simple, the reaction site is obvious, and the reaction process is easy to control.

[0063] The present invention provides the use of the near-infrared second-range croconate dye described in the above technical solution or the near-infrared second-range croconate dye prepared by the preparation method described in the above technical solution in the preparation of a tumor imaging reagent. The tumor imaging reagent is used for photoacoustic imaging or fluorescence imaging of tumor detection.

[0064] The present invention provides the use of the near-infrared second-region crotonate dye described in the above technical solution or the near-infrared second-region crotonate dye prepared by the preparation method described in the above technical solution in the preparation of tumor cell photothermal therapy drugs.

[0065] In the present invention, the tumor cells preferably include one or more of breast cancer cells, oral cancer cells, liver cancer cells, lung cancer cells, gastric cancer cells, pancreatic cancer cells, colorectal cancer cells, bladder cancer cells and prostate cancer cells; more preferably include one or more of breast cancer cells 4T1, human tongue squamous cell carcinoma cells CAL27 and human tongue squamous cell carcinoma cells UM1.

[0066] The near-infrared second-zone structure cresonate dye shown in Formula 1, which has absorption in the near-infrared region, provided by the present invention, can realize the combination of fluorescence imaging technology, photoacoustic imaging technology and photothermal therapy, not only realizing the complementary advantages between imaging technologies, but also providing richer biological information for disease diagnosis, achieving the purpose of diagnosis and treatment at the same time, and avoiding the risks and burdens brought about by multiple injections of different contrast agents, and has broad application prospects.

[0067] In order to further illustrate the present invention, the technical solutions provided by the present invention are described in detail below in conjunction with the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0068] In the following examples, IR1024 is recorded as the near-infrared second-region crotonate dye of the target product of Formula 1 of the present invention; intermediate C is recorded as the compound of the structure of Formula 2 of the present invention; compound B is recorded as the compound of the structure of Formula 3 of the present invention; and compound A is recorded as the compound of the structure of Formula 4 of the present invention.

[0069] Example 1 Synthesis of Intermediate C

[0070]

[0071] (1) Compound A (2.56 g) was placed in a round-bottom flask containing 50 mL of DMF. After complete dissolution, sodium hydride (1.09 g) was added in an ice-water bath. After stirring at room temperature for 5 minutes, n-butyl iodide (4.27 g) was added. After stirring at room temperature for 2 hours, the reactant was washed with sodium chloride saturated water (2×50 mL) and then extracted with ethyl acetate and water (volume ratio of 1:1). The ethyl acetate layer was rotary evaporated to remove the solvent to obtain a crude product. The crude product was dissolved in 1 mL of ethyl acetate and the entire amount was used as the sample solution for silica gel column chromatography. A chromatography column (3 cm × 30 cm) was filled with 50 g of 100-200 mesh column chromatography silica gel and 50 mL of ethyl acetate. After releasing the ethyl acetate, the sample was slowly loaded along the wall of the tube. Ethyl acetate was used as the eluent for elution at a rate of 1 mL / min for 6 column volumes. Thin layer chromatography was performed using a mixed solvent of ethyl acetate and petroleum ether (volume ratio of 4:1) as the developing solvent. Components with Rf values ​​of 0.4-0.6 were collected. The ethyl acetate was evaporated under vacuum to obtain 1.41 g of compound B.

[0072] NMR data of compound B: 1 H NMR (400MHz, DMSO) δ8.10(d,J=8.0Hz,1H),8.00(d,J=8.0Hz,1H),7.74(t,J=8Hz,1H),7.56(d,J=8Hz,1H),7.48(t ,J=8Hz,1H),7.08(t,J=8.0Hz,1H),3.82(t,J=8Hz,2H),1.66-1.58(m,2H),1.32-1.23(m,2H),0.84(t,J=8Hz,3H). 13C NMR (101MHz, DMSO) δ167.25,139.33,131.27,129.22,129.05,126.33,124.6 3,124.25,120.28,105.87,40.16,39.95,39.74,39.62,30.68,19.93,13.91.

[0073] (2) Compound B (300 mg) was placed in a round-bottom flask containing 3 mL of anhydrous THF. Methylmagnesium chloride (1.5 mL) was added in an ice-water bath under nitrogen protection. After reacting at room temperature for 30 minutes, the mixture was transferred to a 60°C water bath and reacted for 2 hours. After cooling to 0°C in an ice-water bath, perchloric acid (0.6 mL) was added to quench the mixture. Water (50 mL) was added for precipitation, and the mixture was washed with water (7 × 50 mL). After freeze-drying in a vacuum at -40°C, 280 mg of intermediate C was obtained.

[0074] NMR data of compound C:

[0075] 1 H NMR(400MHz, CDCl3)δ8.02(d,J=6.5Hz,2H),7.95(d,J=5.2Hz,2H),7.88–7.79(m,2H),7.69(d,J=6.8Hz,2H),7.61–7.52(m,2H),7 .24(d,J=5.4Hz,2H),7.06(s,2H),4.31(s,4H),1.94(dt,J=14.8,7.5Hz,4H),1.52(td,J=14.7,7.3Hz,4H),1.01(t,J=7.3Hz,6H).

[0076] Example 2 Synthesis of ketoacid dye IR1024.

[0077]

[0078] Intermediate C (50 mg) prepared by the method of Example 1 and crotonic acid (9.7 mg) were placed in a round-bottom flask containing 2-methoxyethanol (30 mL). The mixture was heated under reflux at 110°C for 1 h. The reaction solution was rotary evaporated to remove the solvent, and the concentrate was dissolved in 1 mL of dichloromethane and the entire solution was used as the sample for silica gel column chromatography. A chromatography column (3 cm × 30 cm) was filled with 50 g of 100-200 mesh column chromatography silica gel and 50 mL of dichloromethane. After releasing dichloromethane, the sample was slowly loaded along the tube wall. Dichloromethane was used as the eluent for elution at a rate of 1 mL / min and an elution volume of 7 column volumes. Thin layer chromatography monitoring was performed using a mixed solvent of dichloromethane and methanol (volume ratio of 10:1) as the developing solvent. Components with Rf values ​​of 0.2-0.4 were collected. After rotary evaporation of dichloromethane to dryness, 20.1 mg of a near-infrared second-region keratin dye (denoted as IR1024) with the structure shown in Formula 1 was obtained. H NMR spectrum is shown in Figure 1 , NMR carbon spectrum see Figure 2 .

[0079] IR1024 NMR data:

[0080] 1 H NMR(400MHz, CDCl3)δ8.02(d,J=6.5Hz,2H),7.95(d,J=5.2Hz,2H),7.88–7.79(m,2H),7.69(d,J=6.8Hz,2H),7.61–7.52(m,2H),7 .24(d,J=5.4Hz,2H),7.06(s,2H),4.31(s,4H),1.94(dt,J=14.8,7.5Hz,4H),1.52(td,J=14.7,7.3Hz,4H),1.01(t,J=7.3Hz,6H).

[0081] 13 C NMR (100MHz, CDCl3) δ152.05,140.95,133.06,131.45,130.77,129.52,12 8.38,125.15,123.50,109.29,101.27,44.62,31.39,29.64,20.40,13.73.

[0082] Example 3 Detection of Absorption and Emission Spectra of Crotonate Dye in the Near-Infrared Region II

[0083] UV-visible absorption spectrum test: The UV-visible absorption spectrum of the keratin dye IR1024 prepared by the method of Example 2 was tested using an UV-visible spectrophotometer (UV-5500PC, Shanghai Yuanxi Instrument Co., Ltd.): 300 μL of 0.15 mg / mL IR1024 DMF solution was placed in a quartz cuvette, DMF was set to zero as the baseline, and the spectrum was scanned in the wavelength range of 600 nm to 1600 nm. The results are shown in FIG. Figure 3 As shown, IR1024 has strong absorption between 800nm ​​and 1200nm.

[0084] Fluorescence emission spectrum test: The fluorescence emission spectrum of the keratin dye IR1024 prepared by the method of Example 2 was tested using a fluorescence spectrophotometer (PerkinElmer Lambda 750): 3 mL of 10 μM IR1024 DMF solution was taken and the spectrum was scanned in the wavelength range of 1000 nm to 1600 nm. The results are shown in FIG. Figure 4 As shown, IR1024 has a broad emission between 1000nm and 1400nm.

[0085] Photoacoustic signal test: The photoacoustic signal of the cyanine dye IR1024 prepared by the method of Example 2 was detected using a photoacoustic imager (Vevo LAZR, FUJIFILM VisualSonics): the photoacoustic spectra of IR1024 DMF solutions of different concentrations (400 μM, 100 μM, 50 μM, 25 μM, 10 μM) were detected using 1064 nm excitation light. The results are shown in Figure 5 As shown, the photoacoustic signal increases with increasing concentration.

[0086] The results show that the absorption and emission spectra of IR1024 are both in the near-infrared region, the emission spectrum extends to the second near-infrared region, and has a strong photoacoustic signal.

[0087] Example 4 Photothermal heating effect of keratin dye IR1024 in the near-infrared region II

[0088] The keratin dye IR1024 prepared by the method of Example 2 was dissolved in dimethyl sulfoxide to prepare solutions of different concentrations (5 μg / mL, 10 μg / mL, 20 μg / mL, 40 μg / mL, and 80 μg / mL). The solutions were irradiated with a 1064 nm laser for 600 s, and the temperature changes of the IR1024 solutions at different concentrations were recorded with an infrared camera. Figure 6 As shown in the figure, with the increase of concentration, the warming effect of IR1024 increases. At a concentration of 80 μg / mL, the temperature rises to 55°C after 1064 nm laser irradiation for 600 seconds, showing good photothermal conversion efficiency.

[0089] Example 5 Anti-4T1 Tumor Cell Proliferation Activity of Near-Infrared Region II Crozonate Dye IR1024 This example verifies the photothermal therapeutic effect of the crozonate dye with the structure shown in Formula 1 on tumor cells.

[0090] 1. The near-infrared second-region keratin dye IR1024 prepared by the method of Example 2 was prepared into a nanoformulation by a thin film dispersion method to improve bioavailability and biosafety. The specific operation was as follows: 2 mg of IR1024 and 10 mg of distearoylphosphatidyl acetamide-methoxy polyethylene glycol 5000 (DSPE-mPEG5000) were co-dissolved in dimethylformamide-water (v:v = 1:5), and ultrasonic dispersion was performed using a 100 Hz frequency ultrasonic probe to obtain 10 mg of nanoparticles with a particle size of 120 nm, which were recorded as IR1024 NPs.

[0091] 2. The metabolic activity of IR1024NPs on breast cancer 4T1 cells was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method. The specific steps are as follows:

[0092] Breast cancer 4T1 cells (purchased from Hunan Fenghui Biotechnology Co., Ltd.) were inoculated into RPMI 1640 complete medium and incubated in an incubator (37°C, 5% CO2). After culturing for three generations, cells in the logarithmic growth phase were inoculated into 96-well plates at a density of 5 × 10 3 cells / well and incubated overnight to allow the cells to fully adhere to the wall. The old culture medium was discarded and divided into the experimental group (IR1024NPs+L) and the blank control group (IR1024 NPs), with 4 parallels for each group (calculation error). The experimental group (IR1024 NPs+L) was added with 100μL of RPMI 1640 complete culture medium containing IR1024 NPs at different concentrations (0.5μg / mL, 1μg / mL, 2.5μg / mL, 5μg / mL, 10μg / mL, 20μg / mL, 40μg / mL), and the blank control group (IR1024 NPs) was added with 100μL of RPMI 1640 complete culture medium. After further culture for 4 hours, the experimental groups were given 1W / cm 2 The cells were irradiated with 1064 nm laser for 5 min, while the blank control group was not irradiated. After 24 h of incubation, the cells were analyzed using a microplate reader (Thermo Scientific TM Multiskan TM FC) detects the absorbance of each group at 490nm, and calculates the effect of the drug on the survival rate of tumor cells by the ratio of the absorbance of the experimental group to the blank control group. Figure 7 As shown. Figure 7The results show that the near-infrared second-region crotonate dye with the structure shown in Formula 1 exhibits dose-dependent photocytotoxicity, and cell survival rate decreases with increasing dye concentration under illumination conditions, indicating that the near-infrared second-region crotonate dye with the structure shown in Formula 1 has high cell killing efficiency under these conditions. At the same time, the present invention found that the near-infrared second-region crotonate dye with the structure shown in Formula 1 has a high cell survival rate in the dark, indicating that the near-infrared second-region crotonate dye with the structure shown in Formula 1 is non-toxic in the absence of light. These results demonstrate the application prospects of the near-infrared second-region crotonate dye with the structure shown in Formula 1 in photothermal therapy.

[0093] Although the above embodiment provides a detailed description of the present invention, it is only a part of the embodiments of the present invention, not all of the embodiments. Other embodiments can be obtained based on this embodiment without creativity, and these embodiments all fall within the scope of protection of the present invention.

Claims

1. Application of a near-infrared second-region crezone dye in the preparation of a tumor imaging agent, characterized in that: The tumor imaging agent is used for photoacoustic imaging or fluorescence imaging for tumor detection. The near-infrared second-region crotonate dye has a structure shown in Formula 1: Formula 1.

2. Application of a near-infrared second-region crezone dye in the preparation of a drug for photothermal therapy of tumor cells, characterized in that: The near-infrared second region cretonate dye has a structure shown in Formula 1: Formula 1.

3. The use according to claim 2, characterized in that The tumor cells include one or more of breast cancer cells, oral cancer cells, liver cancer cells, lung cancer cells, gastric cancer cells, pancreatic cancer cells, colorectal cancer cells, bladder cancer cells and prostate cancer cells.

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