Preparation method of new near-infrared photodiagnosis and treatment agent based on anthracene ketone purple derivative

Abstract

The application discloses a preparation method of a novel near-infrared photodiagnosis and phototherapy agent based on anthraquinone violet derivative, which comprises the following steps: under heating conditions, K2CO3 is added into a N,N-dimethylformamide (DMF) solution of 16,17-dihydroxyanthraquinone violet to carry out dehydrogenation, [1-(4-bromomethylphenyl)-1,1,2-triphenyl] ethylene is added into the reaction solution, the temperature is increased, and reaction is carried out for 12 h; the S1 reaction product is settled with methanol and water, and filtered; the crude product is used as an eluent of dichloromethane and ethyl acetate, and is separated through a silica gel column chromatography; after rotary evaporation and drying, a dark blue tetraphenyl ethylene modified anthraquinone violet derivative is obtained; and the water solubility of the molecule is realized through a mesoporous silica or amphiphilic polymer wrapping mode, and a corresponding photodiagnosis and phototherapy agent is obtained. The method is simple in operation, the obtained material can emit 700-1200 nm fluorescence under excitation of a near-infrared region one laser, simultaneously generates singlet oxygen and hydroxyl radicals for photodynamic therapy, and the mesoporous silica wrapping can also be used for drug loading and transportation.

Description

Technical Field

[0001] This invention belongs to the technical field of near-infrared luminescence and phototherapy materials, specifically relating to a method for preparing a novel near-infrared phototherapy agent based on anthrone violet derivatives. Background Technology

[0002] Near-infrared fluorescence imaging is a rapid, high-resolution detection technology that has emerged in the biomedical field in recent years and has been quickly applied to the research of diseases such as tumors. The development of optical imaging technology is inseparable from the design and development of high-performance optical contrast agents.

[0003] Common optical contrast agents mainly include organic small molecules, inorganic nanomaterials, and conjugated polymers. Among them, organic small molecules are the most promising class of optical contrast agents for clinical application due to their fixed structure, good metabolizability, and low toxicity. Currently, clinically used organic small molecules such as ICG and MB have shortcomings such as poor photostability and low fluorescence quantum yield. Meanwhile, therapeutic integration is at the forefront of research in related fields, and the efficiency of molecules such as ICG and MB in photodynamic therapy is also relatively poor. Therefore, there is a need to develop novel organic small molecule materials with high photostability, high near-infrared II fluorescence intensity, and outstanding photodynamic therapeutic effects, which is of great significance to the field of optical diagnosis and therapy.

[0004] Anthrone violet is a commonly used vat dye in the textile industry, possessing outstanding photostability and low toxicity. In particular, methylated dimethyloxanthrone violet, also known as vat brilliant green B, accounts for a large proportion in yarn dyeing. However, these dyes have poor water solubility, low fluorescence intensity, and no significant reactive oxygen species production capacity, making them unsuitable for direct application in the biomedical field.

[0005] In view of this, it is necessary to design a method for preparing a novel near-infrared phototherapy agent based on anthrone violet derivatives to solve the above problems. Summary of the Invention

[0006] The purpose of this invention is to provide a method for preparing a novel near-infrared phototherapy agent based on anthrone violet derivatives.

[0007] To achieve the above objectives, the present invention adopts the following technical solution:

[0008] S1: Prepare a 0.125 mol / L solution of 16,17-dihydroxyanthrone purple in dimethylformamide, add K2CO3 at a molar ratio of 4 times that of the dimethylformamide solution as a catalyst, heat to 80℃ and react for 30 min; then add [1-(4-bromomethylphenyl)-1,1,2-triphenyl]ethylene at a molar ratio of 3 times that of the dimethylformamide solution, raise the temperature to 100℃ and continue the reaction for 12 h;

[0009] S2: The product of reaction S1 was precipitated with methanol and water. After precipitation for 24 hours, the solid crude product was obtained by filtration. The product was separated by silica gel column chromatography using dichloromethane and ethyl acetate as eluents. After rotary evaporation and drying, a deep blue tetraphenylethylene-modified anthrone purple derivative was obtained.

[0010] S3: The deep blue tetraphenylethylene-modified anthrone violet derivative is water-soluble by encapsulation with mesoporous silica or an amphiphilic polymer to obtain a phototherapy agent.

[0011] As a further improvement of the present invention: in S2, the volume ratio of methanol to water is 3:2, and the volume ratio of dichloromethane to ethyl acetate is 50:1.

[0012] As a further improvement of the present invention: the step of obtaining the phototherapy agent by means of mesoporous silica in S3 includes:

[0013] A chloroform solution of 1 mg / mL tetraphenylethylene-modified anthrone violet derivative was prepared and mixed with a 20 mg / mL hexadecyltrimethylammonium bromide aqueous solution. After ultrasonic emulsification, the chloroform was removed by heating at 60 °C to obtain a first aqueous solution. The first aqueous solution was mixed with water, tetraethoxysilane, ethyl acetate, and 2 mol / L sodium hydroxide aqueous solution. After reacting at 70 °C for 30 min, triethoxysilane-modified polyethylene glycol was added and the reaction was continued for 2.5 h. After purification by ultrafiltration, a water-soluble anthrone violet-mesoporous silica nanoparticle phototherapy agent was obtained.

[0014] As a further improvement of the present invention: in step S3, the step of obtaining the phototherapy agent by means of an amphiphilic polymer includes:

[0015] The tetraphenylethylene-modified anthrone violet derivative obtained in S2 and the amphiphilic polymer surfactant were dissolved in a mixed solvent of chloroform and tetrahydrofuran. The concentration of the chloroform mixed solvent was 1 mg / mL and the concentration of the tetrahydrofuran mixed solvent was 20 mg / mL. Water with 10 times the volume of the mixed solvent was added under ultrasonic conditions. The organic solvent was blown off with nitrogen under heating conditions at 50°C to obtain a polymer-encapsulated water-soluble anthrone violet nanoparticle phototherapy agent.

[0016] As a further improvement of the present invention: the volume ratio of the S2 chloroform solution and the hexadecyltrimethylammonium bromide aqueous solution is 1:10, and the volume ratio of the first aqueous solution to the water, tetraethoxysilane, ethyl acetate, and 2 mol / L sodium hydroxide aqueous solution is 1:9:0.04:0.2:0.06.

[0017] As a further improvement of the present invention: the volume ratio of chloroform to tetrahydrofuran is 1:1.

[0018] As a further improvement of the present invention: the amphiphilic polymeric surfactant includes at least one of Pluronic F-127, DSPE-PEG or polyoxyethylene (100) octadecyl ether.

[0019] Anthrone violet, a common dye, boasts high stability and good biocompatibility. However, its planar molecular packing is severe, leading to significant molecular aggregation and greatly reducing its fluorescence and photodynamic properties. Tetraphenylene units, with their unique twisted structure, can restrict molecular motion, reduce non-radiative transitions, and improve luminescence efficiency. After encapsulation with amphiphilic polymeric surfactants or mesoporous silica, the resulting water-soluble material can simultaneously achieve high-intensity fluorescence emission in the near-infrared region and photodynamic therapeutic properties, making it valuable in optical diagnostics and treatment.

[0020] Beneficial effects:

[0021] A novel material with anthrone violet unit as its core was synthesized, and a novel near-infrared phototherapy reagent was obtained through nanotechnology. This reagent has strong stability and good biocompatibility. When excited by near-infrared laser, it generates a large number of singlet oxygen and hydroxyl radicals, which can be used for fluorescence imaging-guided photodynamic therapy and drug delivery. Attached Figure Description

[0022] Figure 1 The absorption spectra of the raw materials and the materials obtained in Example 1 in organic solvents;

[0023] Figure 2 The fluorescence emission spectra of the raw materials and the materials obtained in Example 1 in organic solvents are shown.

[0024] Figure 3 Transmission electron microscope images of the material obtained in Example 2;

[0025] Figure 4 The absorption spectra of the raw materials and the materials obtained in Example 2 are shown below.

[0026] Figure 5 The fluorescence emission spectra of the raw materials and the materials obtained in Example 2 are shown below.

[0027] Figure 6 This describes the singlet oxygen production in Example 2;

[0028] Figure 7 The generation of hydroxyl radicals in Example 2;

[0029] Figure 8 The absorption spectrum of the material obtained in Example 3;

[0030] Figure 9 The fluorescence emission spectrum of the material obtained in Example 3 is shown below.

[0031] Figure 10 The singlet oxygen generation of the material obtained in Example 3;

[0032] Figure 11 The generation of hydroxyl radicals in the material obtained in Example 3;

[0033] Figure 12 The reaction circuit diagram is for anthrone purple derivatives. Detailed Implementation

[0034] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be described in detail below with reference to the accompanying drawings and specific embodiments.

[0035] Please see Figures 1 to 11 As shown, the preparation method of a novel near-infrared phototherapy agent based on anthrone violet derivatives includes the following steps:

[0036] S1: Prepare a 0.125 mol / L solution of 16,17-dihydroxyanthrone purple in dimethylformamide, add K2CO3 at a molar ratio of 4 times that of the dimethylformamide solution as a catalyst, heat to 80℃ and react for 30 min; then add [1-(4-bromomethylphenyl)-1,1,2-triphenyl]ethylene at a molar ratio of 3 times that of the dimethylformamide solution, raise the temperature to 100℃ and continue the reaction for 12 h;

[0037] S2: The product of reaction S1 was precipitated with methanol and water. After precipitation for 24 hours, the solid crude product was obtained by filtration. The product was separated by silica gel column chromatography using dichloromethane and ethyl acetate as eluents. After rotary evaporation and drying, a deep blue tetraphenylethylene-modified anthrone purple derivative was obtained.

[0038] S3: The deep blue tetraphenylethylene-modified anthrone violet derivative is water-soluble by encapsulation with mesoporous silica or amphiphilic polymer to obtain a phototherapy agent;

[0039] (1) Prepare a chloroform solution of 1 mg / mL tetraphenylethylene-modified anthrone violet derivative and mix it with 20 mg / mL hexadecyltrimethylammonium bromide aqueous solution at a volume ratio of 1:10. After ultrasonic emulsification, remove the chloroform under heating at 60°C to obtain the first aqueous solution. Mix the first aqueous solution with water, tetraethoxysilane, ethyl acetate and 2 mol / L sodium hydroxide aqueous solution at a volume ratio of 1:9:0.04:0.2:0.06. After reacting at 70°C for 30 min, add triethoxysilane-modified polyethylene glycol and continue to react for 2.5 h. After ultrafiltration purification, obtain water-soluble anthrone violet-mesoporous silica nanoparticle phototherapy agent.

[0040] (2) The tetraphenylethylene-modified anthrone violet derivative obtained in S2 and the amphiphilic polymer surfactant were dissolved in a mixed solvent of chloroform and tetrahydrofuran with a volume ratio of 1:1. The concentration of the chloroform mixed solvent was 1 mg / mL and the concentration of the tetrahydrofuran mixed solvent was 20 mg / mL. Water with a volume of 10 times that of the mixed solvent was added under ultrasonic conditions. The organic solvent was blown off with nitrogen under heating conditions at 50°C to obtain a polymer-encapsulated water-soluble anthrone violet nanoparticle phototherapy agent.

[0041] The specific implementation method is as follows:

[0042] Example 1:

[0043] 1) Prepare a 0.125 mol / L solution of 16,17-dihydroxyanthrone purple in dimethylformamide, add K2CO3 at a molar ratio of 4 times that of the dimethylformamide solution as a catalyst, heat to 80℃ and react for 30 min; then add [1-(4-bromomethylphenyl)-1,1,2-triphenyl]ethylene at a molar ratio of 3 times that of the dimethylformamide solution, raise the temperature to 100℃ and continue the reaction for 12 h;

[0044] 2) The reaction product was precipitated with methanol and water in a volume ratio of 3:2. After precipitation for 24 hours, the product was filtered to obtain a solid crude product. The product was separated by silica gel column chromatography using dichloromethane and ethyl acetate in a volume ratio of 50:1 as the eluent. After rotary evaporation and drying, a deep blue tetraphenylethylene-modified anthrone purple derivative was obtained.

[0045] 3) Prepare a chloroform solution of the obtained product and measure its absorption spectrum and fluorescence emission spectrum.

[0046] Experimental results show that, Figure 1 As shown, the derivative has two absorption peaks near 400 nm and 650 nm, while the original material has an absorption peak around 780 nm; Figure 2 As shown, the tetraphenylethylene-modified anthrone violet derivative exhibits strong fluorescence at 760 nm under 635 nm light excitation, which extends to 1150 nm, with an emission wavelength reaching the near-infrared II region, while the raw material shows almost no fluorescence. This indicates that the fluorescence performance of the derivative is significantly improved.

[0047] Example 2:

[0048] 1) Prepare a 0.125 mol / L solution of 16,17-dihydroxyanthrone purple in dimethylformamide, add K2CO3 at a molar ratio of 4 times that of the dimethylformamide solution as a catalyst, heat to 80℃ and react for 30 min; then add [1-(4-bromomethylphenyl)-1,1,2-triphenyl]ethylene at a molar ratio of 3 times that of the dimethylformamide solution, raise the temperature to 100℃ and continue the reaction for 12 h;

[0049] 2) The reaction product was precipitated with methanol and water in a volume ratio of 3:2. After precipitation for 24 hours, the product was filtered to obtain a solid crude product. The product was separated by silica gel column chromatography using dichloromethane and ethyl acetate in a volume ratio of 50:1 as the eluent. After rotary evaporation and drying, a deep blue tetraphenylethylene-modified anthrone purple derivative was obtained.

[0050] 3) Prepare a 1 mg / mL anthrone violet derivative chloroform solution, mix it with a 20 mg / mL hexadecyltrimethylammonium bromide aqueous solution at a volume ratio of 1:10, emulsify by sonication, and remove the chloroform by heating at 60℃ to obtain the first aqueous solution; mix the first solution with water, tetraethoxysilane, ethyl acetate, and 2 mol / L sodium hydroxide aqueous solution at a volume ratio of 1:9:0.04:0.2:0.06, react at 70℃ for 30 min, add triethoxysilane-modified polyethylene glycol and continue the reaction for 2.5 h, and obtain a water-soluble anthrone violet-mesoporous silica nanoparticle phototherapy agent after ultrafiltration purification.

[0051] Experimental results show that, Figure 3 As shown, the material has an average particle size of approximately 50 nm, exhibits uniform particle size, and good dispersibility; Figure 4 As shown, the material exhibits a novel absorption peak at 780 nm, which may correspond to J-aggregation of the molecules. Figure 5 As shown, the material still exhibits a strong fluorescence emission signal at 750 nm in water, indicating that the tetraphenylethylene group modification successfully improved the optical properties of the nanomaterial. Figure 6 As shown, this material can degrade diphenylbenzofuran (DPBF) under 660nm laser irradiation, indicating that the material has the ability to generate singlet oxygen. Figure 7 As shown, under 660nm laser irradiation, this material can oxidize 3,3',5,5'-tetramethylbenzidine (TMB) to produce corresponding characteristic absorption peaks, indicating that the material has the ability to generate hydroxyl radicals. In summary, this material has good fluorescence and photodynamic properties and can be effectively applied to near-infrared diagnostics and therapy.

[0052] Example 3:

[0053] 1) Prepare a 0.125 mol / L solution of 16,17-dihydroxyanthrone purple in dimethylformamide, add K2CO3 at a molar ratio of 4 times that of the dimethylformamide solution as a catalyst, heat to 80℃ and react for 30 min; then add [1-(4-bromomethylphenyl)-1,1,2-triphenyl]ethylene at a molar ratio of 3 times that of the dimethylformamide solution, raise the temperature to 100℃ and continue the reaction for 12 h;

[0054] 2) The reaction product was precipitated with methanol and water in a volume ratio of 3:2. After precipitation for 24 hours, the product was filtered to obtain a solid crude product. The product was separated by silica gel column chromatography using dichloromethane and ethyl acetate in a volume ratio of 50:1 as the eluent. After rotary evaporation and drying, a deep blue tetraphenylethylene-modified anthrone purple derivative was obtained.

[0055] 3) The tetraphenylethylene-modified anthrone violet derivative and the amphiphilic polymer surfactant F127 were dissolved in a mixed solvent of chloroform and tetrahydrofuran (volume ratio 1:1). The concentration of the chloroform mixed solvent was 1 mg / mL and the concentration of the tetrahydrofuran mixed solvent was 20 mg / mL. Water with 10 times the volume of the mixed solvent was added under ultrasonic conditions. The organic solvent was blown off with nitrogen under heating conditions at 50°C to obtain a polymer-encapsulated water-soluble anthrone violet nanoparticle phototherapy agent.

[0056] Experimental results show that, Figure 8 As shown, the absorption peak of this material is at 660 nm, and the absorption at 780 nm is low, indicating that the diagnostic reagents obtained by this method do not show obvious J aggregation. Figures 9-11 As shown, the fluorescence intensity and hydroxyl radical generation capacity of this material are lower than those of Example 2, but the singlet oxygen generation capacity is higher than that of Example 2.

[0057] This invention uses 16,17-dihydroxyanthrone violet as a raw material and modifies it with tetraphenylethylene substituents to obtain anthrone violet derivatives with near-infrared fluorescence emission. These derivatives can also generate singlet oxygen and hydroxyl radicals under near-infrared light irradiation for photodynamic therapy. The specific reaction pathway is described below. Figure 12 .

[0058] In summary, this invention synthesizes a novel material with anthrone violet unit as its core and obtains a novel near-infrared phototherapy reagent through nanotechnology. This reagent has strong stability and good biocompatibility. When excited by near-infrared laser, it generates a large number of singlet oxygen and hydroxyl radicals, which can be used for fluorescence imaging-guided photodynamic therapy and drug delivery.

[0059] The above embodiments are only used to illustrate the present invention and are not intended to limit the technical solutions described in the present invention. The understanding of this specification should be based on those skilled in the art. Although the present invention has been described in detail with reference to the above embodiments, those skilled in the art should understand that they can still make modifications or equivalent substitutions to the present invention. All technical solutions and improvements that do not depart from the spirit and scope of the present invention should be covered within the scope of the claims of the present invention.

Claims

1. A method for preparing a novel near-infrared phototherapy agent based on anthrone violet derivatives, characterized in that, Includes the following steps: S1: Prepare a 0.125 mol / L solution of 16,17-dihydroxyanthrone purple in dimethylformamide, add K2CO3 at a molar ratio of 4 times that of the dimethylformamide solution as a catalyst, heat to 80℃ and react for 30 min; then add [1-(4-bromomethylphenyl)-1,1,2-triphenyl]ethylene at a molar ratio of 3 times that of the dimethylformamide solution, raise the temperature to 100℃ and continue the reaction for 12 h; S2: The product of reaction S1 was precipitated with methanol and water. After precipitation for 24 hours, the solid crude product was obtained by filtration. The product was separated by silica gel column chromatography using dichloromethane and ethyl acetate as eluents. After rotary evaporation and drying, a deep blue tetraphenylethylene-modified anthrone purple derivative was obtained. S3: The deep blue tetraphenylethylene-modified anthrone violet derivative is water-soluble by encapsulation with mesoporous silica or an amphiphilic polymer to obtain a phototherapy agent.

2. The method for preparing the novel near-infrared phototherapy agent based on anthrone violet derivatives according to claim 1, characterized in that: In step S2, the volume ratio of methanol to water is 3:2, and the volume ratio of dichloromethane to ethyl acetate is 50:

1.

3. The method for preparing the novel near-infrared phototherapy agent based on anthrone violet derivatives according to claim 1, characterized in that: In step S3, the step of obtaining the phototherapy agent using mesoporous silica includes: A chloroform solution of 1 mg / mL tetraphenylethylene-modified anthrone violet derivative was prepared and mixed with a 20 mg / mL hexadecyltrimethylammonium bromide aqueous solution. After ultrasonic emulsification, the chloroform was removed by heating at 60 °C to obtain a first aqueous solution. The first aqueous solution was mixed with water, tetraethoxysilane, ethyl acetate, and 2 mol / L sodium hydroxide aqueous solution. After reacting at 70 °C for 30 min, triethoxysilane-modified polyethylene glycol was added and the reaction was continued for 2.5 h. After purification by ultrafiltration, a water-soluble anthrone violet-mesoporous silica nanoparticle phototherapy agent was obtained.

4. The method for preparing the novel near-infrared phototherapy agent based on anthrone violet derivatives according to claim 1, characterized in that: In step S3, the step of obtaining the phototherapy agent using an amphiphilic polymer method includes: The tetraphenylethylene-modified anthrone violet derivative obtained in S2 and the amphiphilic polymer surfactant were dissolved in a mixed solvent of chloroform and tetrahydrofuran. Water with a volume of 10 times that of the mixed solvent was added under ultrasonic conditions. The organic solvent was blown off with nitrogen under heating conditions at 50°C to obtain a polymer-encapsulated water-soluble anthrone violet nanoparticle phototherapy agent.

5. The method for preparing the novel near-infrared phototherapy agent based on anthrone violet derivatives according to claim 3, characterized in that: The volume ratio of the first aqueous solution to the water, tetraethoxysilane, ethyl acetate, and 2 mol / L sodium hydroxide aqueous solution is 1:9:0.04:0.2:0.

06.

6. The method for preparing the novel near-infrared phototherapy agent based on anthrone violet derivatives according to claim 4, characterized in that: The volume ratio of chloroform to tetrahydrofuran is 1:

1.

7. The method for preparing the novel near-infrared phototherapy agent based on anthrone violet derivatives according to claim 4, characterized in that: The amphiphilic polymeric surfactant includes at least one of Pluronic F-127 and DSPE-PEG.

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