Photosensitizer carrier as well as preparation method and application thereof

Abstract

The invention discloses a photosensitizer carrier and a preparation method and application thereof, and relates to the technical field of photosensitizer carriers, the preparation method of the photosensitizer carrier is as follows: 4-carboxyphenylboronic acid is activated by EDC / NHS and then is amidated with PEI to construct an intermediate containing phenylboronic acid groups; then, under the alkaline condition, a phenylboronic acid group on the intermediate and ortho-diol of 1, 2, 4-trihydroxy anthraquinone form a borate bond, and a photosensitizer carrier with pH responsiveness is obtained; the prepared photosensitizer carrier has good water solubility and can be combined with a hydrophobic photosensitizer through a pi-pi conjugation effect, the water solubility of the photosensitizer carrier is remarkably improved, the carrier is connected through a pH-sensitive boric acid ester bond and can be specifically released in a tumor subacid environment, meanwhile, a boric acid group can be specifically combined with tumor cells of overexpression sialic acid, and the carrier has a good application prospect. The targeting property and the safety of drug delivery are improved.

Description

Technical Field

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

[0002] Photodynamic therapy (PDT) is a new type of tumor treatment method that achieves therapeutic effects by combining photosensitizers, oxygen and light. The mechanism of action of PDT mainly includes two types of photodynamic reactions: Type I reaction is that the photosensitizer in the triplet excited state transfers hydrogen atoms or electrons with the substrate to form free radicals or free radical ions. These free radicals interact with activated oxygen in the tissue to produce oxides, thereby killing target cells; Type II reaction is that the photosensitizer in the triplet excited state directly transfers energy to oxygen molecules to form singlet oxygen. Since singlet oxygen has strong oxidizing properties, it can quickly cause cancer cells to shrink and die. During PDT treatment, it takes 3 to 96 hours for the photosensitizer to reach the maximum enrichment concentration in the tumor tissue after being injected into the human body. This time depends on the properties of the photosensitizer. When the photosensitizer is enriched to a certain concentration in the tumor tissue, light of a specific wavelength is used to irradiate the tumor site, thereby activating the photosensitizer to produce a photodynamic effect.

[0003] PDT has dual selectivity, one is based on the concentration difference of photosensitizer in normal tissue and target tissue, the other is to directly irradiate the target tissue through spatial restriction and focusing. In addition, PDT has good tissue selectivity and can act specifically on the target tissue; the effect is superficial, and the effective action depth is basically below 0.01μm; it has a strong damaging effect on microvascular tissue, and is particularly suitable for the treatment of microvascular diseases; it has few systemic side effects and can be used repeatedly; it can be flexibly used in combination with other therapies such as surgery, chemotherapy, and radiotherapy.

[0004] The development of photosensitizers has gone through several stages. The first generation of photosensitizers is represented by hematoporphyrin derivatives (HpD), but they have disadvantages such as complex components, unclear structures, unsatisfactory action spectra, poor tissue penetration, slow excretion, and easy phototoxic reactions. The second generation of photosensitizers has improved in terms of structural clarity, component uniformity, and absorption wavelength, including porphyrins, benzoporphyrins and other types, and has more operability in clinical applications. Among them, purpurin imide, as a new type of second-generation photosensitizer, has characteristic absorption at 418nm and 707nm and can have a strong binding effect with serum proteins. In addition, other types of photosensitizers, such as endogenous photosensitizers such as aminolevulinic acid (5-ALA), have simple structures and are easy to synthesize. The bioavailability can be improved by esterification and phthalimide.

[0005] CN102370980A discloses a method for preparing a nanographene oxide carrier for photodynamic therapy. In this method, the surface of the nanographene oxide is modified with a hydrophilic polymer, and then a hydrophobic photosensitizer containing a large π conjugated structure is loaded through π-π stacking and hydrophobic-hydrophobic interaction to obtain a product that can be used for photodynamic therapy.

[0006] However, existing photosensitizers still have the following problems: the molecular structure of photosensitizers is complex and the synthesis process is cumbersome; the water solubility of photosensitizers is poor and the biocompatibility is not ideal; the targeting of photosensitizers in the body is insufficient and it is difficult to specifically enrich in tumor tissues; the photodynamic activity and stability of photosensitizers need to be improved.

[0007] Therefore, it is of great significance to develop a new photosensitizer carrier system with simple structure, simple preparation process, good water solubility and biocompatibility. By combining photosensitizer molecules with specific carrier materials, not only can the water solubility and biocompatibility of photosensitizers be improved, but also their targeting and photodynamic activity in tumor tissues can be enhanced, thereby improving the therapeutic effect of PDT. Summary of the invention

[0008] In order to solve the deficiencies in the prior art, the purpose of the present invention is to provide a photosensitizer carrier and a preparation method and application thereof. The prepared photosensitizer carrier has good water solubility and can be combined with a hydrophobic photosensitizer through π-π conjugation, significantly improving its water solubility. The carrier is connected through a pH-sensitive boronate bond and can be specifically released in the slightly acidic environment of the tumor. At the same time, the boric acid group can specifically bind to tumor cells overexpressing sialic acid, thereby improving the targeting and safety of drug delivery.

[0009] In order to achieve the above object, the present invention adopts the following technical solution:

[0010] A method for preparing a photosensitizer carrier, characterized in that it comprises the following steps:

[0011] (1) adding 4-carboxyphenylboronic acid, EDC, and NHS to anhydrous methanol, stirring and mixing to obtain a 4-carboxyphenylboronic acid activation solution, adding dropwise to a methanol solution of polyethyleneimine, stirring for reaction, precipitating the product, washing, dialyzing, and freeze-drying to obtain an intermediate;

[0012] Carboxyl activation coupling: 4-Carboxylphenylboronic acid (PSA) first generates an O-acylisourea intermediate under EDC activation, and NHS then attacks to form an active ester structure, thereby increasing the reactivity of the carboxyl group. The active ester intermediate undergoes a nucleophilic substitution reaction with the primary / secondary amino group on the polyethyleneimine (PEI) molecule, and the phenylboronic acid group is connected to the PEI backbone by forming a stable amide bond, thereby obtaining a PEI-PSA intermediate.

[0013] Preferably, in step (1), the stirring and mixing conditions are stirring and mixing in the dark at room temperature for 30 to 60 minutes; and the stirring reaction conditions are stirring and reacting at room temperature for 12 to 24 hours.

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

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

[0016] PEI with a molecular weight of 1500 to 2000 is selected as the backbone. This molecular weight range not only ensures good water solubility and biocompatibility of the carrier, but also avoids cytotoxicity caused by too high molecular weight, and at the same time provides an appropriate amount of amino groups for modification.

[0017] In step (1), the product is added dropwise to icy ether for precipitation, centrifuged, washed three times with icy ether, dissolved in ultrapure water, and then transferred to a dialysis bag (MWCO = 1000Da) for dialyzing for 48 to 72 hours, and the dialysate is replaced every 12 hours. After the dialysis is completed, the solution is freeze-dried for 48 to 72 hours 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, a molecular sieve is added, and a methanol solution of 1,2,4-trihydroxyanthraquinone is added dropwise under stirring, and the reaction is stirred in the dark, and the product is filtered, dialyzed, and freeze-dried to obtain the photosensitizer carrier.

[0019] Boronation: In a phosphate buffer solution at pH 8.0-8.5, the phenylboronic acid group of the PEI-PSA intermediate is converted into a tetrahedral borate, which undergoes a reversible coordination reaction with the 1,2-vicinal diol in the 1,2,4-trihydroxyanthraquinone (purpurin) molecule and dehydrates to form a cyclic borate structure. The presence of the molecular sieve promotes the dehydration equilibrium to move toward the product, thereby obtaining a photosensitizer carrier with pH responsiveness.

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

[0021]

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

[0023] Preferably, in step (2), the reaction conditions are: stirring the reaction at 20-30° C. and 300-500 r / min in a dark environment for 12-20 hours, and checking the pH every 4 hours during the reaction to maintain the solution pH at 8.0-8.5.

[0024] In step (2), the product is filtered to remove the molecular sieve, and the filtrate is transferred to a dialysis bag (MWCO = 2000Da), and dialyzed with a phosphate buffer solution with a pH of 8.0 for 24 hours and ultrapure water for 48 hours, and the dialysate is replaced every 12 hours. The solution is freeze-dried for 48 to 72 hours to obtain the photosensitizer carrier, which is sealed, protected from light, and stored at -20°C.

[0025] The present invention also claims to protect a photosensitizer carrier prepared by the preparation method.

[0026] The present invention also claims to protect the use of the photosensitizer carrier in the preparation of photodynamic therapy drugs.

[0027] Preferably, the photosensitizer is a hydrophobic photosensitizer containing a large π-conjugated structure, including porphyrins, phthalocyanines, and chlorophyll derivatives.

[0028] Preferably, the photosensitizer is purpurin imide.

[0029] Preferably, the method comprises the following steps: adding a photosensitizer carrier and a hydrophobic photosensitizer to an organic solvent, ultrasonically treating the mixture to obtain a mixed solution, dropping the mixed solution into an aqueous solution of a surfactant, stirring to volatilize the organic solvent, centrifuging, washing, and drying to obtain the drug for photodynamic therapy.

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

[0031] 1. The present invention provides a method for preparing a photosensitizer carrier, which is prepared by a two-step reaction: first, 4-carboxyphenylboronic acid is activated by EDC / NHS and then reacted with PEI to undergo an amidation reaction to construct an intermediate containing a phenylboronic acid group; 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 entire preparation process has mild conditions, simple operation, controllable reaction, and is easy to scale up.

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

[0033] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings required for use in the embodiments are briefly introduced below. It should be understood that the following drawings only show schematic diagrams of certain embodiments of the present invention and therefore should not be regarded as limiting the scope. For ordinary technicians in this field, other related drawings can be obtained based on these drawings without paying creative work.

[0034] Figure 1 This is a SEM image of the drug for photodynamic therapy prepared by the photosensitizer carrier in Example 1;

[0035] Figure 2 This is the absorption curve of the drug for photodynamic therapy prepared by the photosensitizer carrier in Example 1;

[0036] Figure 3 This is the appearance of the drug and PVA aqueous solution used in photodynamic therapy.

[0037] Figure 3 In the figure, cuvettes No. 1 and No. 2 show the appearance of the drug colloid solution for photodynamic therapy prepared by the photosensitizer carrier in Example 1, and cuvette No. 3 shows the appearance of the PVA aqueous solution. DETAILED DESCRIPTION

[0038] In order to make the purpose, technical solution and advantages of the present invention clearer, the present invention is further described in detail below in conjunction with embodiments. Of course, the specific embodiments described here are only used to explain the present invention and are not used to limit the present invention.

[0039] Unless otherwise specified, the chemical reagents and materials in the present invention are purchased from commercial sources or synthesized from raw materials purchased from commercial sources.

[0040] A method for preparing a photosensitizer carrier, characterized in that it comprises the following steps:

[0041] (1) 150-200 mg 4-carboxyphenylboronic acid, 230-280 mg EDC, and 140-170 mg NHS are added to 8-15 mL anhydrous methanol, and the mixture is stirred and mixed for 30-60 min in the dark at room temperature to obtain a 4-carboxyphenylboronic acid activation solution, which is then added dropwise to 5-10 mL of a methanol solution of polyethyleneimine (polyethyleneimine content: 180-220 mg, molecular weight: MW=1500-2000), and the mixture is stirred and reacted at room temperature for 12-24 h. The product is then added dropwise to glacial ether for precipitation, centrifuged, and washed three times with glacial ether. The precipitate is dissolved in ultrapure water, and then transferred to a dialysis bag (MWCO=1000 Da) for dialysis for 48-72 h, and the dialysis solution is replaced every 12 h. After the dialysis is completed, 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.

[0042] (2) Dissolve 100-150 mg of the intermediate in 15-20 mL of phosphate buffer (pH 8.0-8.5), add 1-2 g The product was filtered to remove the molecular sieve, and the filtrate was transferred to a dialysis bag (MWCO = 2000Da), and dialyzed with a phosphate buffer of pH 8.0 for 24 hours and ultrapure water for 48 hours, and the dialyzate was replaced every 12 hours. The solution was freeze-dried for 48 to 72 hours to obtain the photosensitizer carrier, which was sealed and stored at -20 ° C in a dark place.

[0043] A use of a photosensitizer carrier in preparing a drug for photodynamic therapy comprises the following steps:

[0044] The photosensitizer carrier and the hydrophobic photosensitizer are added to an organic solvent, and ultrasonic treatment is performed to obtain a mixed solution. The mixed solution is added dropwise to an aqueous solution of a surfactant, and after stirring to volatilize 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 in that it comprises 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 the mixture was stirred and mixed for 60 min in the dark at room temperature to obtain a 4-carboxyphenylboronic acid activation solution, which was added dropwise to 10 mL of a methanol solution of polyethyleneimine (polyethyleneimine content: 220 mg, molecular weight: MW=1700), and the mixture was stirred and reacted at room temperature for 24 h. The product was added dropwise to icy ether for precipitation, and the mixture was centrifuged and washed three times with icy 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 replaced every 12 h. After the dialysis was completed, the solution was freeze-dried for 72 h to obtain a white viscous intermediate, which was sealed and stored in a dark place at 4° C.

[0049] (2) Dissolve 150 mg of the intermediate in 20 mL of phosphate buffer (pH 8.5) and add 2 g The product was filtered to remove the molecular sieve, and the filtrate was transferred to a dialysis bag (MWCO = 2000Da), and dialyzed with a phosphate buffer of pH 8.0 for 24 hours and ultrapure water for 48 hours, and the dialyzate was replaced every 12 hours. The solution was freeze-dried for 48 to 72 hours to obtain the photosensitizer carrier, which was sealed and stored at -20 ° C in a dark place.

[0050] Example 2

[0051] A method for preparing a photosensitizer carrier, characterized in that it comprises 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 the mixture was stirred and mixed for 50 min in the dark at room temperature to obtain a 4-carboxyphenylboronic acid activation solution, which was added dropwise to 8 mL of a methanol solution of polyethyleneimine (polyethyleneimine content: 200 mg, molecular weight: MW=2000), and the mixture was stirred and reacted at room temperature for 20 h. The product was added dropwise to icy ether for precipitation, and the mixture was centrifuged and washed three times with icy ether. The precipitate was dissolved in ultrapure water, and then transferred to a dialysis bag (MWCO=1000 Da) for 60 h of dialysis, and the dialysis solution was replaced every 12 h. After the dialysis was completed, the solution was freeze-dried for 60 h to obtain a white viscous intermediate, which was sealed and stored in a dark place at 4° C.

[0053] (2) Dissolve 130 mg of the intermediate in 18 mL of phosphate buffer (pH 8.5) and add 2 g The product was filtered to remove the molecular sieve, and the filtrate was transferred to a dialysis bag (MWCO = 2000Da), and dialyzed with a phosphate buffer solution of pH 8.0 for 24 hours and ultrapure water for 48 hours, and the dialyzate was replaced every 12 hours. The solution was freeze-dried for 60 hours to obtain the photosensitizer carrier, which was sealed and stored at -20°C in a dark place.

[0054] Example 3

[0055] A method for preparing a photosensitizer carrier, characterized in that it 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, and the mixture was stirred and mixed for 40 min in the dark at room temperature to obtain a 4-carboxyphenylboronic acid activation solution, which was added dropwise to 6 mL of a methanol solution of polyethyleneimine (polyethyleneimine content: 200 mg, molecular weight: MW=1800), and the mixture was stirred and reacted at room temperature for 16 h. The product was added dropwise to icy ether for precipitation, and centrifuged and washed three times with icy ether. The precipitate was dissolved in ultrapure water, and then transferred to a dialysis bag (MWCO=1000 Da) for 60 h of dialysis, and the dialysis solution was replaced every 12 h. After the dialysis was completed, the solution was freeze-dried for 60 h to obtain a white viscous intermediate, which was sealed and stored in a dark place at 4° C.

[0057] (2) Dissolve 120 mg of the intermediate in 16 mL of phosphate buffer (pH 8.0) and add 1 g The product was filtered to remove the molecular sieve, and the filtrate was transferred to a dialysis bag (MWCO = 2000Da), and dialyzed with a phosphate buffer solution of pH 8.0 for 24 hours and ultrapure water for 48 hours, and the dialyzate was replaced every 12 hours. The solution was freeze-dried for 48 hours to obtain the photosensitizer carrier, which was sealed and stored at -20°C in a dark place.

[0058] Example 4

[0059] A method for preparing a photosensitizer carrier, characterized in that it comprises the following steps:

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

[0061] (2) Dissolve 100 mg of the intermediate in 15 mL of phosphate buffer (pH 8.0) and add 1 g The product was filtered to remove the molecular sieve, and the filtrate was transferred to a dialysis bag (MWCO = 2000Da), and dialyzed with a phosphate buffer solution of pH 8.0 for 24 hours and ultrapure water for 48 hours, and the dialyzate was replaced every 12 hours. The solution was freeze-dried for 48 hours to obtain the photosensitizer carrier, which was sealed and stored at -20°C in a dark place.

[0062] Comparative Example 1

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

[0064] (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 the mixture was stirred and mixed for 60 min in the dark at room temperature to obtain a 4-carboxyphenylboronic acid activation solution, which was added dropwise to 10 mL of a methanol solution of polyethyleneimine (polyethyleneimine content: 220 mg, molecular weight: MW=1700), and the mixture was stirred and reacted at room temperature for 24 h. The product was added dropwise to icy ether for precipitation, and the mixture was centrifuged and washed three times with icy ether. The precipitate was dissolved in ultrapure water, and then transferred to a dialysis bag (MWCO=1000Da) for dialysis for 72 h, and the dialysate was replaced every 12 h. After the dialysis was completed, 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] Polyethyleneimine with a molecular weight of MW=1700 was used as the photosensitizer carrier.

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

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

[0069] Take the prepared drug for photodynamic therapy, wash it with deionized water for 3 times by centrifugation, disperse it with ultrasound, and then use a dropper to drop a small amount on a clean gold-coated silicon wafer, dry it at room temperature, spray gold on the surface, and then put it in a vacuum drying oven to dry for 24 hours, and then observe it with a scanning electron microscope (SEM). According to SEM observation, the surface of the microspheres is smooth and flat, the average particle size is 0.25±0.05μm, the spherical shape is relatively good, and the particle morphology and particle size are mostly uniform. Figure 1 This is a 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 by the photosensitizer carrier of Example 1) was prepared, and the mixed solution of dichloromethane and methanol of purpurin-18-imide with different concentrations was prepared, and its absorbance was measured at the maximum absorption wavelength to obtain the standard curve of its 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), and after it was completely dissolved ( Figure 3 ), use an ultraviolet spectrophotometer to measure its absorbance at the same absorption wavelength as the standard curve, then compare the standard curve to find out its purpurin-18-imide concentration, and calculate the drug loading and encapsulation rate in the drug used for photodynamic therapy according to the formula (specific data see Table 1):

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

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

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

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

[0075]

[0076] The above description is only a preferred specific implementation manner of the present invention, but the protection scope of the present invention is not limited thereto. Any technician familiar with the technical field can make equivalent replacements or changes according to the technical scheme and inventive concept of the present invention within the technical scope disclosed by the present invention, which should be covered by the protection scope of the present invention.

Claims

1. A method for preparing a photosensitizer carrier, characterized in that: The steps include: (1) adding 4-carboxyphenylboronic acid, EDC, and NHS to anhydrous methanol, stirring and mixing to obtain a 4-carboxyphenylboronic acid activation solution, adding dropwise to a methanol solution of polyethyleneimine, stirring for reaction, precipitating the product, washing, dialyzing, and freeze-drying to obtain an intermediate; (2) The intermediate is dissolved in a phosphate buffer, a molecular sieve is added, and a methanol solution of 1,2,4-trihydroxyanthraquinone is added dropwise under stirring, and the reaction is stirred in the dark, and the product is filtered, dialyzed, and freeze-dried to obtain the photosensitizer carrier.

2. The method for preparing a photosensitizer carrier according to claim 1, characterized in that: In step (1), the stirring and mixing conditions are stirring and mixing in the dark at room temperature for 30 to 60 minutes; and the stirring reaction conditions are stirring and reacting at room temperature for 12 to 24 hours.

3. The method for preparing a photosensitizer carrier according to claim 1, characterized in that: In step (1), the usage ratio of 4-carboxyphenylboric acid, EDC, NHS and anhydrous methanol is 150-200 mg: 230-280 mg: 140-170 mg: 8-15 mL; the usage ratio of polyethyleneimine and the methanol solution of polyethyleneimine is 180-220 mg: 5-10 mL; the usage ratio of 4-carboxyphenylboric acid and polyethyleneimine is 150-200 mg: 180-220 mg.

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

5. The method for preparing a photosensitizer carrier according to claim 1, characterized in that: In step (2), the pH of the phosphate buffer is 8.0 to 8.5, and the molecular sieve is 6. The method for preparing a photosensitizer carrier according to claim 1, characterized in that: In step (2), the ratio of the intermediate, phosphate buffer and molecular sieve is 100-150 mg: 15-20 mL: 1-2 g; the ratio of the intermediate to 1,2,4-trihydroxyanthraquinone is 100-150 mg: 80-100 mg; and the ratio of 1,2,4-trihydroxyanthraquinone to the methanol solution of 1,2,4-trihydroxyanthraquinone is 80-100 mg: 5 mL.

7. The method for preparing a photosensitizer carrier according to claim 1, characterized in that: In step (2), the reaction conditions are: stirring the reaction at 20-30° C. and 300-500 r / min in a dark environment for 12-20 hours. During the reaction, the pH value is checked every 4 hours to maintain the pH value of the solution at 8.0-8.

5.

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

9. Use of the photosensitizer carrier according to claim 8 in the preparation of photodynamic therapy drugs.

10. The use according to claim 9, characterized in that: The photosensitizer is a hydrophobic photosensitizer containing a large π conjugated structure, including porphyrins, phthalocyanines, and chlorophyll derivatives.

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