A photosensitizer carrier, a preparation method and application thereof

Abstract

The application discloses a photosensitizer carrier and a preparation method and application thereof, relates to the technical field of photosensitizer carriers, and discloses the following preparation method of the photosensitizer carrier: 4-carboxyphenyl boronic acid is activated by EDC / NHS, and then is subjected to amidation reaction with PEI to construct an intermediate containing a phenyl boronic acid group; then, under alkaline conditions, the phenyl boronic acid group on the intermediate forms a borate ester bond with ortho diols of 1,2,4-trihydroxyanthraquinone, and a photosensitizer carrier with pH responsiveness is obtained; the prepared photosensitizer carrier has good water solubility, can be combined with a hydrophobic photosensitizer through pi-pi conjugation, and can significantly improve the water solubility of the photosensitizer; the carrier is connected through a pH-sensitive borate ester bond, and can be specifically released in a tumor micro-acidic environment; meanwhile, the boronic acid group can be specifically combined with tumor cells overexpressing sialic acid, and the targeting and safety of drug delivery are improved.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of photosensitizer carriers, in particular to a photosensitizer carrier and a preparation method and application thereof. BACKGROUND

[0002] Photodynamic therapy (PDT) is a new type of tumor treatment method, which achieves therapeutic effect by using photosensitizer, oxygen and light. The mechanism of action of PDT mainly includes two types of photodynamic reactions: type I reaction is that the triplet excited state of photosensitizer transfers hydrogen atom or electron to the substrate to form free radicals or free radical ions, which interact with activated oxygen in the tissue to produce oxidants, thereby killing target cells; type II reaction is that the triplet excited state of photosensitizer directly transfers energy to oxygen molecules to form singlet oxygen, which can quickly make cancer cells atrophy and die due to its strong oxidizing property. During the PDT treatment, the photosensitizer needs 3-96 hours to reach the maximum concentration in the tumor tissue after being injected into the human body, which depends on the nature of the photosensitizer. When the photosensitizer reaches a certain concentration in the tumor tissue, a specific wavelength of light is used to irradiate the tumor site, thereby activating the photosensitizer to produce a photodynamic effect.

[0003] PDT has double selectivity, one is based on the concentration difference of photosensitizer in normal tissue and target tissue, and the other is through spatial limitation and focusing to make light directly irradiate the target tissue. In addition, PDT has good tissue selectivity and can specifically act on target tissue; the effective depth of action is basically below 0.01 μm; the damage to microvascular tissue is strong, especially suitable for the treatment of microvascular diseases; the whole body has few side effects and can be used repeatedly; it can be used flexibly with other therapies such as surgery, chemotherapy, radiotherapy, etc.

[0004] The development of photosensitizer has experienced several stages. The first generation of photosensitizer is represented by hematoporphyrin derivative (HpD), but it has the disadvantages of complex components, unclear structure, unsatisfactory action spectrum, poor tissue penetration ability, slow excretion, and easy occurrence of phototoxicity. The second generation of photosensitizer has improved in structure, component, and absorption wavelength, including porphyrin, benzoporphyrin, and other types, which have more operability in clinical application. Among them, purpurin imide as a new type of second-generation photosensitizer has characteristic absorption at 418 nm and 707 nm, and can form strong combination with serum proteins. In addition, other types of photosensitizer such as aminolevulinic acid (5-ALA) and other endogenous photosensitizers have simple structure and are easy to synthesize, and can improve bioavailability through acetylation and phthalamidation.

[0005] CN102370980A discloses a preparation method of a nano graphene oxide carrier for photodynamic therapy, which comprises the following steps: modifying the surface of nano graphene oxide with a hydrophilic polymer, and then loading a hydrophobic photosensitizer with a large pi conjugated structure through pi-pi stacking and hydrophobic-hydrophobic interaction to obtain a product for photodynamic therapy.

[0006] However, the existing photosensitizer still has the following problems: the photosensitizer has a complex molecular structure and a complicated synthesis process; the photosensitizer has poor water solubility and unsatisfactory biocompatibility; the photosensitizer has insufficient targeting in the body and is difficult to be specifically enriched in tumor tissues; and the photodynamic activity and stability of the photosensitizer need to be improved.

[0007] Therefore, it is of great significance to develop a new photosensitizer carrier system which has a simple structure, a simple preparation process, good water solubility and biocompatibility. By combining the photosensitizer molecule with a specific carrier material, the water solubility and biocompatibility of the photosensitizer can be improved, and the targeting in tumor tissues and the photodynamic activity of the photosensitizer can be enhanced, so that the treatment effect of PDT can be improved. SUMMARY

[0008] In order to solve the problems in the prior art, the purpose of the present application is to provide a photosensitizer carrier and a preparation method and application thereof. The prepared photosensitizer carrier has good water solubility, can be combined with a hydrophobic photosensitizer through pi-pi conjugation, significantly improves the water solubility of the photosensitizer, is connected through a pH-sensitive borate ester bond, can be specifically released in the micro-acidic environment of tumor, and the boronic acid group can be specifically combined with tumor cells overexpressing sialic acid, thereby improving the targeting and safety of drug delivery.

[0009] In order to achieve the above purpose, the technical scheme adopted by the present application is as follows:

[0010] A preparation method of a photosensitizer carrier, characterized in that the method comprises the following steps:

[0011] (1) 4-carboxyphenylboronic acid, EDC and NHS are added to anhydrous methanol, stirred and mixed to obtain a 4-carboxyphenylboronic acid activation solution, a methanol solution of polyethyleneimine is added dropwise, stirred and reacted, and the product is precipitated, washed, dialyzed and freeze-dried to obtain an intermediate;

[0012] Carboxyl activation coupling: 4-carboxyphenylboronic acid (PSA) is first generated into an O-acylisourea intermediate under the activation of EDC, and then NHS attacks to form an active ester structure, thereby improving the reactivity of the carboxyl group; the active ester intermediate reacts with the primary / secondary amino groups on the polyethyleneimine (PEI) molecule through nucleophilic substitution, and the phenylboronic acid group is connected to the PEI skeleton through the formation of a stable amide bond to obtain a PEI-PSA intermediate.

[0013] Preferably, in step (1), the stirring mixing condition is dark, stirring mixing for 30-60 min at room temperature; the stirring reaction condition is stirring reaction for 12-24 h at room temperature.

[0014] Preferably, in step (1), the amount ratio of 4-carboxyphenylboronic acid, EDC, NHS, anhydrous methanol is 150-200 mg: 230-280 mg: 140-170 mg: 8-15 mL; the amount ratio of polyethyleneimine, methanol solution of polyethyleneimine is 180-220 mg: 5-10 mL; the amount ratio of 4-carboxyphenylboronic acid, polyethyleneimine is 150-200 mg: 180-220 mg.

[0015] Preferably, in step (1), the molecular weight of the polyethyleneimine is MW=1500-2000.

[0016] The PEI with a molecular weight of 1500-2000 is selected as the skeleton, which not only ensures good water solubility and biocompatibility of the carrier, but also avoids cytotoxicity caused by too high molecular weight, and provides an appropriate amount of amino groups for modification.

[0017] In step (1), the product is added dropwise into ice ether for precipitation, centrifugal separation, washed with ice ether for three times, dissolved in ultrapure water, then transferred to a dialysis bag (MWCO=1000 Da) for dialysis for 48-72 h, the dialysis solution is replaced every 12 h, after dialysis, the solution is freeze-dried for 48-72 h to obtain a white viscous intermediate, which is sealed, protected from light, and stored at 4°C.

[0018] (2) The intermediate is dissolved in a phosphate buffer, molecular sieves are added, and a methanol solution of 1,2,4-trihydroxyanthraquinone is added dropwise under stirring, and the product is obtained by stirring reaction in the dark, filtration, dialysis, and freeze-drying.

[0019] Boronic esterification: the benzene boronic acid groups in the PEI-PSA intermediate are converted into tetrahedral borate in a phosphate buffer at pH 8.0-8.5, and a reversible coordination reaction occurs with the 1,2-ortho diol in the 1,2,4-trihydroxyanthraquinone (purpurin) molecule to form a cyclic borate structure, and the presence of molecular sieves promotes the movement of the dehydration equilibrium to the product direction, obtaining a pH-responsive photosensitizer carrier.

[0020] Preferably, in step (2), the pH of the phosphate buffer is 8.0-8.5, and the molecular sieves are

[0021]

[0022] Preferably, in step (2), the amount ratio of the intermediate, the phosphate buffer, and the molecular sieve is 100-150 mg: 15-20 mL: 1-2 g; the amount ratio of the intermediate and 1,2,4-trihydroxyanthraquinone is 100-150 mg: 80-100 mg; and the amount ratio of 1,2,4-trihydroxyanthraquinone and the methanol solution of 1,2,4-trihydroxyanthraquinone is 80-100 mg: 5 mL.

[0023] Preferably, in step (2), the light-shielded stirring reaction condition is that the reaction is stirred at 20-30 ℃ and 300-500 r / min for 12-20 h in the dark, and the pH is checked every 4 h during the reaction to maintain the pH of the solution at 8.0-8.5.

[0024] In step (2), the molecular sieve is removed by suction filtration, the filtrate is transferred into a dialysis bag (MWCO = 2000 Da), and the dialysis is performed with the phosphate buffer with a pH of 8.0 for 24 h and with ultrapure water for 48 h, the dialysis solution is replaced every 12 h, the solution is freeze-dried for 48-72 h to obtain the photosensitizer carrier, and the photosensitizer carrier is sealed, shielded from light, and stored at -20 ℃.

[0025] The application also claims a photosensitizer carrier prepared by the preparation method.

[0026] The application also claims the use of the photosensitizer carrier in the preparation of a photodynamic therapy drug.

[0027] Preferably, the photosensitizer is a hydrophobic photosensitizer containing a large pi conjugated structure, including porphyrin, phthalocyanine, and chlorophyll derivatives.

[0028] Preferably, the photosensitizer is a purpurin imide.

[0029] Preferably, the method comprises the following steps: the photosensitizer carrier and the hydrophobic photosensitizer are added to an organic solvent, the mixture is ultrasonically treated to obtain a mixed solution, the mixed solution is added dropwise into an aqueous solution of a surfactant, the organic solvent is volatilized after stirring, and then the mixture is centrifuged, washed, and dried to obtain the drug for photodynamic therapy.

[0030] Compared with the prior art, the application has the following beneficial effects:

[0031] 1. The application provides a preparation method of a photosensitizer carrier, which realizes the preparation of the carrier through two-step reactions: first, 4-carboxyphenylboronic acid is activated by EDC / NHS and then subjected to amidation reaction with PEI to construct an intermediate containing a phenylboronic acid group; and then, under alkaline conditions, the phenylboronic acid group on the intermediate forms a borate ester bond with the ortho diol of 1,2,4-trihydroxyanthraquinone to obtain a photosensitizer carrier with pH responsiveness. The whole preparation process has mild conditions, simple operation, controllable reaction, and is easy to scale up.

[0032] 2、The photosensitizer carrier provided by the application has the following advantages: ① the anthraquinone groups in the carrier structure can be combined with visible light photosensitizers through π-π conjugation, and the singlet oxygen generated under light can cause the anthraquinone groups to break, thereby realizing light-controlled release; ② the carrier is connected through a pH-sensitive borate ester bond, and can be released specifically in a tumor micro-acidic environment; ③ the introduction of the PEI skeleton significantly improves the solubility of the hydrophobic photosensitizer in an aqueous environment, avoids the aggregation of the photosensitizer, and improves the efficiency of active oxygen generation; ④ the boronic acid groups in the carrier structure can specifically recognize tumor cells overexpressing sialic acid, thereby improving the targeting property; and ⑤ the carrier has light-controlled and pH dual responsiveness, and can realize precise delivery of drugs, thereby having a broad application prospect in photodynamic therapy. BRIEF DESCRIPTION OF DRAWINGS

[0033] In order to more clearly illustrate the technical solutions of the embodiments of the application, the following will briefly introduce the drawings needed to be used in the embodiments. It should be understood that the following drawings only show some embodiment schematic diagrams of the application, and therefore should not be regarded as a limitation on the scope. For those skilled in the art, other related drawings can also be obtained without creative labor on the basis of these drawings.

[0034] Figure 1 SEM diagram of the drug for photodynamic therapy prepared by the photosensitizer carrier of Example 1;

[0035] Figure 2 Absorption curve of the drug for photodynamic therapy prepared by the photosensitizer carrier of Example 1;

[0036] Figure 3 Appearance of the drug for photodynamic therapy and PVA aqueous solution.

[0037] Figure 3 In the figure, the appearance of the drug colloidal solution for photodynamic therapy prepared by the photosensitizer carrier of Example 1 is shown in the colorimetric cuvettes 1 and 2, and the appearance of the PVA aqueous solution is shown in the colorimetric cuvette 3. DETAILED DESCRIPTION

[0038] In order to make the objects, technical solutions and advantages of the application more clear, the following will combine the embodiments to make further detailed description of the application. Of course, the specific embodiments described here are only used to explain the application, and are not used to limit the application.

[0039] Unless otherwise specified, the chemical reagents and materials in the application are purchased through market channels or synthesized through raw materials purchased through market channels.

[0040] A preparation method of a photosensitizer carrier, characterized in that the method comprises the following steps:

[0041] (1) Add 150-200 mg of 4-carboxyphenylboronic acid, 230-280 mg of EDC, and 140-170 mg of NHS to 8-15 mL of anhydrous methanol. Stir and mix in the dark at room temperature for 30-60 min to obtain an activated 4-carboxyphenylboronic acid solution. Add 5-10 mL of a methanol solution of polyethyleneimine (polyethyleneimine content 180-220 mg, molecular weight MW = 1500-2000) dropwise to the solution. Stir and react at room temperature for 12-24 h. Add the product dropwise to ice-cold ether for precipitation. Centrifuge and wash three times with ice-cold ether. Dissolve the precipitate in ultrapure water and then transfer it to a dialysis bag (MWCO = 1000 Da) for dialysis for 48-72 h. Change the dialysis solution every 12 h. After dialysis, freeze-dry the solution for 48-72 h to obtain a white viscous intermediate. Seal, protect from light, and store at 4 °C.

[0042] (2) Dissolve 100-150 mg of the intermediate in 15-20 mL of phosphate buffer (pH 8.0-8.5), and add 1-2 g of [the solution]. Molecular sieves were used to add 5 mL of a methanol solution of 1,2,4-trihydroxyanthraquinone (containing 80–100 mg of 1,2,4-trihydroxyanthraquinone) dropwise under stirring. The mixture was stirred at 20–30 °C and 300–500 rpm for 12–20 h in the dark. The pH was checked every 4 h during the reaction to maintain the solution pH at 8.0–8.5. The product was filtered to remove the molecular sieves, and the filtrate was transferred to a dialysis bag (MWCO = 2000 Da). The solution was dialyzed with phosphate buffer (pH 8.0) for 24 h and then with ultrapure water for 48 h, with the dialysate being changed every 12 h. The solution was then freeze-dried for 48–72 h to obtain the photosensitizer carrier, which was then sealed, protected from light, and stored at -20 °C.

[0043] The application of a photosensitizer carrier in the preparation of a drug for photodynamic therapy includes the following steps:

[0044] A photosensitizer carrier and a hydrophobic photosensitizer are added to an organic solvent and ultrasonically treated to obtain a mixture. The mixture is then added dropwise to an aqueous solution of a surfactant. After stirring to evaporate the organic solvent, the mixture is centrifuged, washed, and dried to obtain the drug for photodynamic therapy.

[0045] The present invention will be further described below through specific embodiments.

[0046] Example 1

[0047] A method for preparing a photosensitizer carrier, characterized by comprising the following steps:

[0048] (1) 200 mg of 4-carboxyphenylboronic acid, 280 mg of EDC, and 170 mg of NHS were added to 15 mL of anhydrous methanol and stirred for 60 min in the dark at room temperature to obtain an activated solution of 4-carboxyphenylboronic acid. The solution was then added dropwise to 10 mL of a methanol solution of polyethyleneimine (polyethyleneimine content 220 mg, molecular weight MW = 1700). The solution was stirred for 24 h at room temperature. The product was then added dropwise to ice-cold ether for precipitation. The product was separated by centrifugation and washed three times with ice-cold ether. The precipitate was dissolved in ultrapure water and then transferred to a dialysis bag (MWCO = 1000 Da) for dialysis for 72 h. The dialysis solution was changed every 12 h. After dialysis, the solution was freeze-dried for 72 h to obtain a white viscous intermediate. The intermediate was sealed, protected from light, and stored at 4 °C.

[0049] (2) Dissolve 150 mg of the intermediate in 20 mL of phosphate buffer (pH 8.5), and add 2 g of [unclear text - possibly a continuation of the previous sentence]. Molecular sieves were used to add 5 mL of a methanol solution of 1,2,4-trihydroxyanthraquinone (100 mg of 1,2,4-trihydroxyanthraquinone) dropwise under stirring. The mixture was stirred at 30°C and 500 rpm for 12 h in the dark. The pH was checked every 4 h during the reaction to maintain the pH at 8.5. The product was filtered to remove the molecular sieves, and the filtrate was transferred to a dialysis bag (MWCO = 2000 Da). The solution was dialyzed with phosphate buffer (pH 8.0) for 24 h and then with ultrapure water for 48 h, with the dialysate being changed every 12 h. The solution was then freeze-dried for 48–72 h to obtain the photosensitizer carrier, which was then sealed, protected from light, and stored at -20°C.

[0050] Example 2

[0051] A method for preparing a photosensitizer carrier, characterized by comprising the following steps:

[0052] (1) 180 mg of 4-carboxyphenylboronic acid, 260 mg of EDC, and 160 mg of NHS were added to 12 mL of anhydrous methanol and stirred for 50 min in the dark at room temperature to obtain an activated solution of 4-carboxyphenylboronic acid. The solution was then added dropwise to 8 mL of a methanol solution of polyethyleneimine (polyethyleneimine content 200 mg, molecular weight MW = 2000). The solution was stirred for 20 h at room temperature. The product was then added dropwise to ice-cold ether for precipitation. The product was separated by centrifugation and washed three times with ice-cold ether. The precipitate was dissolved in ultrapure water and then transferred to a dialysis bag (MWCO = 1000 Da) for dialysis for 60 h. The dialysis solution was changed every 12 h. After dialysis, the solution was freeze-dried for 60 h to obtain a white viscous intermediate. The intermediate was sealed, protected from light, and stored at 4 °C.

[0053] (2) Dissolve 130 mg of the intermediate in 18 mL of phosphate buffer (pH 8.5), and add 2 g of [unclear text - possibly a continuation of the previous sentence]. Molecular sieve, 5 mL of 1,2,4-trihydroxyanthraquinone methanol solution (1,2,4-trihydroxyanthraquinone content is 90 mg) was added dropwise under stirring, stirring at 28℃, 400 r / min in the dark for 18 h, the pH was checked every 4 h during the reaction, and the solution pH was maintained at 8.5, the product was suction filtered to remove the molecular sieve, the filtrate was transferred to a dialysis bag (MWCO = 2000 Da), and was dialyzed with a phosphate buffer solution with a pH of 8.0 for 24 h and ultrapure water for 48 h, the dialysis liquid was replaced every 12 h, and the solution was freeze-dried for 60 h to obtain the photosensitizer carrier, which was sealed, kept away from light, and stored at -20℃.

[0054] Example 3

[0055] A preparation method of a photosensitizer carrier, characterized in that the method comprises the following steps:

[0056] (1) 160 mg of 4-carboxyphenylboronic acid, 240 mg of EDC, and 150 mg of NHS were added to 10 mL of anhydrous methanol, stirred and mixed in the dark at room temperature for 40 min to obtain 4-carboxyphenylboronic acid activation solution, 6 mL of polyethyleneimine methanol solution (polyethyleneimine content 200 mg, molecular weight MW = 1800) was added dropwise, and stirred at room temperature for 16 h, the product was added dropwise into ice ethyl ether for precipitation, centrifuged, washed with ice ethyl ether three times, dissolved in ultrapure water, and then transferred to a dialysis bag (MWCO = 1000 Da) for dialysis for 60 h, the dialysis liquid was replaced every 12 h, and the solution was freeze-dried for 60 h after dialysis to obtain a white viscous intermediate, which was sealed, kept away from light, and stored at 4℃;

[0057] (2) 120 mg of the intermediate was dissolved in 16 mL of a phosphate buffer (pH 8.0), 1 g of 1,2,4-trihydroxyanthraquinone was added, and stirred at 24℃, 400 r / min in the dark for 14 h, the pH was checked every 4 h during the reaction, and the solution pH was maintained at 8.0, the product was suction filtered to remove the molecular sieve, the filtrate was transferred to a dialysis bag (MWCO = 2000 Da), and was dialyzed with a phosphate buffer solution with a pH of 8.0 for 24 h and ultrapure water for 48 h, the dialysis liquid was replaced every 12 h, and the solution was freeze-dried for 48 h to obtain the photosensitizer carrier, which was sealed, kept away from light, and stored at -20℃. Example 4

[0058] A preparation method of a photosensitizer carrier, characterized in that the method comprises the following steps:

[0059]

[0060] ​(1) 150 mg 4-carboxyphenylboronic acid, 230 mg EDC, 140 mg NHS were added into 8 mL anhydrous methanol, the mixture was stirred at room temperature for 30 min in the dark to obtain 4-carboxyphenylboronic acid activation solution, 5 mL methanol solution of polyethyleneimine (polyethyleneimine content 180 mg, molecular weight MW = 1500) was added dropwise, and the reaction was stirred at room temperature for 12 h. The product was added dropwise into ice ethyl ether for precipitation, centrifugal separation, washed with ice ethyl ether for three times, dissolved in ultrapure water, then transferred to a dialysis bag (MWCO = 1000 Da) for dialysis for 48 h, the dialysate was replaced every 12 h. After dialysis, the solution was freeze-dried for 48 h to obtain a white viscous intermediate, which was sealed, protected from light and stored at 4°C;

[0061] (2) 100 mg of the intermediate was dissolved in 15 mL phosphate buffer (pH 8.0), 1 g molecular sieve was added, 5 mL 1,2,4-trihydroxyanthraquinone methanol solution (1,2,4-trihydroxyanthraquinone content 80 mg) was added dropwise under stirring, and the reaction was stirred at 20°C for 12 h in the dark. The pH of the solution was checked every 4 h during the reaction to maintain the pH at 8.0. The product was filtered to remove the molecular sieve, and the filtrate was transferred to a dialysis bag (MWCO = 2000 Da) for dialysis with phosphate buffer (pH 8.0) for 24 h and ultrapure water for 48 h, respectively. The dialysate was replaced every 12 h. The solution was freeze-dried for 48 h to obtain the photosensitizer carrier, which was sealed, protected from light and stored at -20°C.

[0062] Comparative Example 1

[0063] A preparation method of a photosensitizer carrier, characterized in that it comprises the following steps:

[0064] (1) 200 mg 4-carboxyphenylboronic acid, 280 mg EDC, 170 mg NHS were added into 15 mL anhydrous methanol, the mixture was stirred at room temperature for 60 min in the dark to obtain 4-carboxyphenylboronic acid activation solution, 10 mL methanol solution of polyethyleneimine (polyethyleneimine content 220 mg, molecular weight MW = 1700) was added dropwise, and the reaction was stirred at room temperature for 24 h. The product was added dropwise into ice ethyl ether for precipitation, centrifugal separation, washed with ice ethyl ether for three times, dissolved in ultrapure water, then transferred to a dialysis bag (MWCO = 1000 Da) for dialysis for 72 h, the dialysate was replaced every 12 h. After dialysis, the solution was freeze-dried for 72 h to obtain the photosensitizer carrier, which was sealed, protected from light and stored at 4°C;

[0065] Comparative Example 2

[0066] The polyethyleneimine with a molecular weight MW = 1700 was used as the photosensitizer carrier.

[0067] The photosensitizer carriers obtained in Examples 1-4 and Comparative Examples 1-2 were used in the preparation of drugs for photodynamic therapy, and the steps are as follows:

[0068] 40 mg of photosensitizer carrier and 6 mg of erythropoietin-18-imide were added to a 20 mL mixture of dichloromethane and methanol (volume ratio 1:1). The mixture was sonicated for 120 s to obtain a solution. The solution was then added dropwise to 75 mL of 2 wt% PVA aqueous solution. After stirring in a 25 °C water bath for 8 h to evaporate the organic solvent, the solution was centrifuged at 5000 r / min, washed with deionized water, and repeated 3 times. The solution was then vacuum dried to obtain the drug for photodynamic therapy.

[0069] The prepared drug for photodynamic therapy was first washed three times by centrifugation with deionized water, then dispersed by ultrasonication. A small amount was then dropped onto a clean gold-plated silicon wafer, dried at room temperature, and then sputter-coated with gold. The wafer was then placed in a vacuum drying oven for 24 hours and observed by scanning electron microscopy (SEM). SEM observation showed that the microspheres had smooth and flat surfaces, an average particle size of 0.25±0.05μm, good sphericity, and mostly uniform particle size and morphology. Figure 1 SEM image of the drug for photodynamic therapy prepared using the photosensitizer carrier of Example 1.

[0070] Weigh an appropriate amount of the prepared drug for photodynamic therapy, dissolve it in a mixed solvent of dichloromethane and methanol, and measure its absorption curve and maximum absorption wavelength. Figure 2 (The absorption curve of the drug for photodynamic therapy prepared from the photosensitizer carrier in Example 1) was obtained by preparing mixed solutions of dichloromethane and methanol with different concentrations of rutin-18-imide, and measuring the absorbance at the maximum absorption wavelength to obtain a standard curve of absorbance and solution concentration. Then, 1 mg of the drug for photodynamic therapy was weighed and dissolved in 5 mL of a mixed organic solvent of dichloromethane and methanol (volume ratio 1:1). After complete dissolution... Figure 3 The absorbance of the drug at the same absorption wavelength as the standard curve was measured using a UV spectrophotometer. The concentration of erythropoietin-18-imide was then determined by comparing it with the standard curve. The drug loading and encapsulation efficiency in the drug used for photodynamic therapy were calculated using the formula (see Table 1 for specific data).

[0071] Drug loading = [Weight of photosensitizer in the drug / Weight of the drug] × 100%;

[0072] Encapsulation efficiency = [weight of photosensitizer in the drug / amount of drug administered] × 100%.

[0073] The 5ml (20mg / ml) of the drug for photodynamic therapy prepared in Examples 1-4 and Comparative Examples 1-2 was poured into a dialysis bag, and the dialysis bag was placed in a flask containing 20ml of acetic acid buffer solution (pH=5.0, simulating the weak acid environment of tumor tissue), and the dialysis bag and the flask were placed at 37°C and stirred. After 12h, a small amount of solution was taken from the flask for UV detection. After detection, the solution in the cuvette was poured back into the flask to ensure the constant total volume of the flask. The in vitro drug release concentration was recorded (see Table 1 for specific data).

[0074] Table 1 Performance test of the drug for photodynamic therapy prepared by the photosensitizer carrier

[0075]

[0076] The above description is merely preferred specific embodiments of the present application, but the protection scope of the present application is not limited thereto. Any person skilled in the art, according to the technical range disclosed in the present application and the inventive concept, should be covered within the protection scope of the present application.

Claims

1. A method of preparing a photosensitizer carrier, characterized by, It comprises the following steps: (1) adding 4-carboxyphenylboronic acid, EDC, NHS into anhydrous methanol, stirring and mixing to obtain 4-carboxyphenylboronic acid activation solution, adding methanol solution of polyethyleneimine dropwise, stirring and reacting, precipitating, washing, dialysis, and freeze-drying to obtain an intermediate; (2) dissolving the intermediate in phosphate buffer, adding molecular sieve, stirring and adding 1,2,4-trihydroxyanthraquinone methanol solution dropwise, stirring and reacting in the dark, and then extracting, dialysis, and freeze-drying to obtain the photosensitizer carrier.

2. The method of claim 1, wherein the photosensitizer carrier is prepared by the steps of: In step (1), the stirring and mixing condition is stirring and mixing for 30-60 min in the dark at room temperature; and the stirring and reacting condition is stirring and reacting for 12-24 h at room temperature.

3. The method of claim 1, wherein the photosensitizer carrier is prepared by the steps of: In step (1), the amount ratio of 4-carboxyphenylboronic acid, EDC, NHS, and anhydrous methanol is 150-200 mg:230-280 mg:140-170 mg:8-15 mL; the amount ratio of polyethyleneimine and methanol solution of polyethyleneimine is 180-220 mg:5-10 mL; and the amount ratio of 4-carboxyphenylboronic acid and polyethyleneimine is 150-200 mg:180-220 mg.

4. The method for preparing the photosensitizer carrier according to claim 1, characterized in that, In step (1), the molecular weight of the polyethyleneimine is 1500-2000.

5. The method of claim 1, wherein the photosensitizer carrier is prepared by the steps of: In step (2), the phosphate buffer has a pH of 8.0 to 8.5 and the molecular sieve is 6. The method of claim 1, wherein the photosensitizer carrier is prepared by the steps of: In step (2), the amount ratio of the intermediate, phosphate buffer, and molecular sieve is 100-150 mg:15-20 mL:1-2 g; the amount ratio of the intermediate and 1,2,4-trihydroxyanthraquinone is 100-150 mg:80-100 mg; and the amount ratio of 1,2,4-trihydroxyanthraquinone and 1,2,4-trihydroxyanthraquinone methanol solution is 80-100 mg:5 mL.

7. The method of claim 1, wherein the photosensitizer carrier is prepared by the steps of: In step (2), the light-avoiding stirring and reacting condition is stirring and reacting for 12-20 h at 20-30℃ and 300-500 r / min in the dark, and the pH is checked every 4 h during the reaction to maintain the pH of the solution at 8.0-8.

5.

8. A photosensitizer carrier prepared by the preparation method of any one of claims 1-7.

9. Use of the photosensitizer carrier of claim 8 in the preparation of a photodynamic therapy drug.

10. Use according to claim 9, characterized in that, The photosensitizer is a hydrophobic photosensitizer containing a large π conjugated structure, including porphyrin, phthalocyanine, and chlorophyll derivatives.

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