PH and photo-thermal dual-responsive polydopamine nano drug-loaded microsphere as well as preparation method and application of pH and photo-thermal dual-responsive polydopamine nano drug-loaded microsphere

Abstract

The invention provides a pH and photo-thermal dual-responsive polydopamine nano drug-loaded microsphere as well as a preparation method and application thereof, and relates to the technical field of medicines. The pH (Potential of Hydrogen) and photo-thermal dual-responsive polydopamine nano drug-loaded microsphere provided by the invention is prepared from a hydrophobic drug, polydopamine coating the hydrophobic drug and poly (N, N-dimethylaminoethyl methacrylate) grafted on the surface of the polydopamine. The drug-loaded microsphere is designed by utilizing the pH difference between the acidity of a tumor microenvironment and a normal physiological condition, and does not depend on a tumor specific antigen, so that the drug-loaded microsphere has wide applicability and relatively low off-target risk. The drug-loaded microsphere provided by the invention has excellent pH and photo-thermal responsiveness and good biocompatibility, can gather at a tumor site with an acidic microenvironment and rapidly release drugs to achieve the purpose of targeted therapy, and has general applicability of targeted tumors; meanwhile, under local near-infrared light irradiation, a photo-thermal treatment effect is achieved, and the tumor inhibition effect is enhanced in cooperation with drugs.

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to a pH and photothermal dual-responsive polydopamine nano drug-loaded microsphere and a preparation method and application thereof. Background Art

[0002] Cancer treatment is one of the major challenges facing the world. Due to its high recurrence rate and easy metastasis, the prognosis is poor. The research and development and application of chemotherapy drugs have always been the direction of continuous efforts of scientific researchers. Although many newly developed drugs have significant anti-tumor effects, their practical application in clinical practice is limited due to problems such as hydrophobicity and bioavailability. In addition, the use of drugs alone also has problems such as insufficient targeting and potential toxic side effects.

[0003] In order to improve the therapeutic effect, various drug nano-delivery platforms are being developed, but their accuracy in tumor cell or tissue recognition still needs to be further optimized. Polydopamine (PDA) has high adhesion, biocompatibility, photothermal conversion and drug binding ability. It has potential in drug delivery, photothermal therapy, imaging and biosensing, and is widely used in the biomedical field. PDA can load anti-tumor drugs and target tumors, improving efficacy and reducing adverse reactions. However, previous active targeted drug delivery systems must be combined with specific targets of tumor cells, that is, they rely on binding to specific proteins of specific tumors to achieve targeting effects. This requires the design of different carriers according to different tumors, but in fact many "cold tumors" lack specific proteins and have no specific targets, making them difficult to target, which seriously limits the applicability of drug delivery systems. Summary of the invention

[0004] In view of this, the purpose of the present invention is to provide a pH and photothermal dual-responsive polydopamine nanoparticle drug-loaded microsphere and its preparation method and application. The pH and photothermal dual-responsive polydopamine nanoparticle drug-loaded microsphere provided by the present invention has universal applicability for targeting tumors.

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

[0006] The invention provides a pH and photothermal dual-responsive polydopamine nano drug-loaded microsphere, comprising a hydrophobic drug, polydopamine coating the hydrophobic drug, and polymethacrylate N,N-dimethylaminoethyl grafted on the surface of the polydopamine.

[0007] Preferably, the hydrophobic drug comprises evodiamine.

[0008] Preferably, the particle size of the pH and photothermal dual-responsive polydopamine nano-drug-loaded microspheres is 180 to 225 nm.

[0009] The present invention provides a method for preparing the pH and photothermal dual-responsive polydopamine nano-drug-loaded microspheres described in the above technical solution, comprising the following steps:

[0010] The dispersion of the hydrophobic drug is mixed with dopamine hydrochloride to perform a coating reaction to obtain drug-coated polydopamine;

[0011] The drug-coated polydopamine is mixed with N,N-dimethylaminoethyl methacrylate to carry out graft polymerization reaction to obtain the pH and photothermal dual-responsive polydopamine nano drug-loaded microspheres.

[0012] Preferably, the dispersion of the hydrophobic drug comprises the hydrophobic drug, Tris-HCl buffer and an alcohol solvent; the pH value of the Tris-HCl buffer is 8.0-8.5; and the volume ratio of the Tris-HCl buffer to the alcohol solvent is 4:1-12:1.

[0013] Preferably, the concentration of the hydrophobic drug in the dispersion of the hydrophobic drug is 0.01-0.8 g / L; the mass ratio of the hydrophobic drug to dopamine hydrochloride in the dispersion of the hydrophobic drug is 10-20:100-200.

[0014] Preferably, the coating reaction is carried out at room temperature for 12 to 24 hours; the coating reaction is carried out under stirring at a rate of 300 to 500 rpm.

[0015] Preferably, the dosage ratio of the drug-coated polydopamine to N,N-dimethylaminoethyl methacrylate is (2-5) mg: (2-3) mL.

[0016] Preferably, the graft polymerization reaction temperature is room temperature, and the time is 0.5 to 8 hours; the graft polymerization reaction is carried out in darkness, under ultraviolet irradiation and stirring conditions, the power of the ultraviolet irradiation is 8 to 20 W, and the stirring rate is 200 to 500 rpm.

[0017] The present invention provides the use of the pH and photothermal dual-responsive polydopamine nano-drug-loaded microspheres described in the above technical scheme or the pH and photothermal dual-responsive polydopamine nano-drug-loaded microspheres prepared by the preparation method described in the above technical scheme in the preparation of drugs for treating tumors.

[0018] The present invention provides a pH and photothermal dual-responsive polydopamine nano drug-loaded microsphere, comprising a hydrophobic drug, polydopamine coated with the hydrophobic drug, and polymethacrylate N,N-dimethylaminoethyl ester grafted on the surface of the polydopamine. The tumor cell microenvironment is acidic, which is different from the weak alkalinity of normal physiological environment body fluids. The present invention designs a drug-loading system based on the commonality of the acidic microenvironment of tumor cells, making it universal. Specifically, the present invention coats the hydrophobic drug with polydopamine, which can achieve a high encapsulation rate and make the drug-loaded microspheres photothermal responsive; polymethacrylate N,N-dimethylaminoethyl ester is grafted on the surface of the polydopamine coated with the hydrophobic drug, so that the drug-loaded microspheres have excellent pH responsiveness. When the pH is reduced, polymethacrylate N,N-dimethylaminoethyl ester changes from non-water-soluble particles to a relatively stretched random coil state due to different solubility, which can promote the release of the drug. The drug-loaded microspheres provided by the present invention have excellent pH responsiveness and photothermal responsiveness and good biocompatibility. The drug-loaded microspheres can aggregate and rapidly release drugs at tumor sites with an acidic microenvironment to achieve the purpose of targeted therapy, and have a universally applicable targeting effect on all types of tumors; at the same time, under local near-infrared light irradiation, photothermal therapy is exerted, and the synergistic drug enhances the tumor inhibition effect. The present invention provides a new approach for the delivery of hydrophobic drugs and photothermal combined therapy, and has wide applicability and excellent practical application value.

[0019] Furthermore, when the hydrophobic drug includes evodiamine, evodiamine is a multi-target anticancer drug that can promote tumor ferroptosis. The present invention uses polydopamine as a carrier material for evodiamine. Polydopamine has excellent photothermal conversion performance and good biocompatibility, and synergistically promotes ferroptosis. The present invention integrates chemotherapy, photothermal therapy (PTT) and ferroptosis to achieve multimodal synergistic treatment of tumors.

[0020] The present invention provides a method for preparing the pH and photothermal dual-responsive polydopamine nano-drug-loaded microspheres described in the above technical solution, which has a simple process and an environmentally friendly preparation process. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 The pH response mechanism of the pH and photothermal dual-responsive polydopamine nano-drug-loaded microspheres provided by the present invention;

[0022] Figure 2 A schematic diagram of the process for preparing the pH and photothermal dual-responsive polydopamine nano-drug-loaded microspheres according to an embodiment of the present invention and the macroscopic pH response mechanism of the dual-responsive polydopamine nano-drug-loaded microspheres;

[0023] Figure 3The SEM, TEM and particle size distribution diagrams of the intermediate product A2 prepared in Example 1 and the dual-responsive polydopamine nano-drug-loaded microspheres B2 prepared in Example 2 are shown in FIG. Figure 3 A is the SEM image of the intermediate product A2, the inset in A is the TEM image of the intermediate product A2, B is the particle size distribution diagram of the intermediate product A2, C is the SEM image of the dual-responsive polydopamine nano-drug-loaded microspheres B2, the inset in C is the TEM image of the dual-responsive polydopamine nano-drug-loaded microspheres B2, and D is the particle size distribution diagram of the dual-responsive polydopamine nano-drug-loaded microspheres B2;

[0024] Figure 4 The photothermal properties of the intermediate product A2 prepared in Example 1 and the dual-responsive polydopamine nano-drug-loaded microspheres B2 prepared in Example 2 are shown in FIG. Figure 4 A is water, A2 and B2 are at 1.5W / cm 2 Temperature rise curve within 5 minutes of near-infrared light irradiation. B is the temperature change of B2 within 5 cycles of near-infrared light irradiation-natural cooling. DETAILED DESCRIPTION

[0025] The invention provides a pH and photothermal dual-responsive polydopamine nano drug-loaded microsphere, comprising a hydrophobic drug, polydopamine (PDA) coating the hydrophobic drug, and poly (N,N-dimethylaminoethyl methacrylate) (PDMAEMA) grafted on the surface of the polydopamine.

[0026] In the present invention, the hydrophobic drug is a poorly soluble drug. The present invention has general applicability to hydrophobic drugs. The groups of hydrophobic drugs are mainly alkyl chains, aromatic rings, olefin groups, ester groups, amide groups, and aromatic hydrocarbon groups. In the present invention, the hydrophobic drug preferably includes evodiamine (hydrophobic small molecule drug). In the present invention, the polydopamine (PDA) has a high photothermal conversion efficiency, has high-performance light absorption in the entire ultraviolet, visible and near-infrared bands, and the interference of light scattering is almost negligible, so it is an efficient broadband photothermal conversion material, and polydopamine can achieve a high encapsulation rate for hydrophobic drugs. In the present invention, the polymethacrylate N, N-dimethylaminoethyl ester (PDMAEMA) is transformed from non-water-soluble particles to a relatively stretched random coil state due to different solubility when the pH is reduced, which promotes the release of the drug; at the same time, the polymethacrylate N, N-dimethylaminoethyl ester is an amphiphilic component, which can improve the solubility of the drug-loaded microspheres as a whole.

[0027] In the present invention, the particle size of the pH and photothermal dual-responsive polydopamine nano-drug-loaded microspheres is preferably 180 to 225 nm.

[0028] The tumor cell microenvironment is acidic, which is different from the weak alkalinity of normal physiological body fluids (the pH value of normal tissues and blood is 7.4, while the tumor microenvironment is acidic due to active metabolism, mostly between 6.5 and 7.0, and the lowest can reach 6.0. The pH value of the inner membrane and lysosome in the subcellular organelle is lower, between 4.5 and 5.5). The present invention designs a drug delivery system based on the pH commonality of the tumor cell microenvironment. The pH-responsive drug delivery system does not require prior knowledge of tumor-specific markers, thereby having a wider range of applicability, lower off-target effects and higher therapeutic efficiency. The dual-responsive drug-loaded microspheres provided by the present invention are constructed by polydopamine (PDA) coated with hydrophobic drugs and polymethacrylate N, N-dimethylaminoethyl ester (PDMAEMA) grafted on its surface. The constructed drug-loaded microspheres not only have a high encapsulation rate for hydrophobic drugs, but also exhibit excellent pH responsiveness, photothermal responsiveness and good biocompatibility. After intravenous injection, the drug-loaded microspheres can aggregate at the tumor site with an acidic microenvironment and rapidly release the drug, thereby achieving the purpose of targeted therapy; at the same time, under local near-infrared light irradiation, they exert a photothermal therapeutic effect and synergistically enhance the tumor inhibition effect. Figure 1 The pH response mechanism of the pH and photothermal dual-responsive polydopamine nano-drug-loaded microspheres provided by the present invention, wherein the state of PDA at pH <7 is because PDA will gradually hydrolyze in an acidic and high temperature environment, and the generation of positive charge at pH <7 is due to the protonation of PDMAEMA in the acidic environment.

[0029] In addition, when the hydrophobic drug includes evodiamine, evodiamine (EVO) is a multi-target anticancer drug that can promote ferroptosis. Ferroptosis is an important frontier in the study of cell death regulation and has become a promising cancer treatment strategy in recent years. In the process of ferroptosis, Fe 3+ Reduced to Fe 2+, consuming glutathione (GSH), and generating reactive oxygen species (ROS) through the Fenton reaction, leading to toxic lipid peroxide (LPO) accumulation and cell damage. The cysteine / glutamate transporter (SLC7A11) and glutathione peroxidase 4 (GPX4) are key to this process, promoting ferroptosis through GSH reduction. The multi-target anti-tumor drug evodiamine can promote tumor ferroptosis by inhibiting the SLC7A11 / GPX4 axis. However, due to the complex mechanisms of tumor resistance and differences in sensitivity to ferroptosis in different tumor types, the application of ferroptosis alone is still limited. Photothermal therapy (PTT) uses near-infrared light to generate heat and induce tumor cell death. PTT can accurately induce tumor thermal damage, while destroying cell membranes (enhancing drug absorption) and mitochondria (amplifying ROS, consuming GSH, inhibiting GPX4 and accumulating LPO), synergistically promoting ferroptosis. The present invention uses the photothermal properties of the polydopamine carrier and the death mechanism of the drug to synergistically treat tumors. This strategy can significantly improve the effectiveness of cancer therapy based on ferroptosis.

[0030] The present invention designs a drug delivery system based on the acidic property of the tumor cell microenvironment, which solves the problem that previous antigen-antibody binding targeted drug delivery systems are difficult to achieve universal applicability in targeting tumors, while exerting multifunctional anti-tumor capabilities in a limited drug delivery platform structure.

[0031] The present invention provides a method for preparing the pH and photothermal dual-responsive polydopamine nano-drug-loaded microspheres described in the above technical solution, comprising the following steps:

[0032] The dispersion of the hydrophobic drug is mixed with dopamine hydrochloride to perform a coating reaction to obtain drug-coated polydopamine;

[0033] The drug-coated polydopamine is mixed with N,N-dimethylaminoethyl methacrylate to carry out graft polymerization reaction to obtain the pH and photothermal dual-responsive polydopamine nano drug-loaded microspheres.

[0034] In the present invention, unless otherwise specified, the raw materials involved are commercially available products well known in the art.

[0035] Figure 2 The schematic diagram of the process of preparing the pH and photothermal dual-responsive polydopamine nanoparticles and the macroscopic pH response mechanism of the dual-responsive polydopamine nanoparticles is shown in the following. The preparation method of the pH and photothermal dual-responsive polydopamine nanoparticles is described in detail.

[0036] The invention mixes the dispersion of the hydrophobic drug with dopamine hydrochloride to carry out coating reaction, thereby obtaining the drug-coated polydopamine.

[0037] In the present invention, the dispersion of the hydrophobic drug preferably includes a hydrophobic drug, a Tris (tris (hydroxymethyl) aminomethane) -HCl buffer and an alcohol solvent; the pH value of the Tris-HCl buffer is preferably 8.0-8.5, and the concentration is preferably 0.01 mol / L; the alcohol solvent is preferably anhydrous ethanol; the volume ratio of the Tris-HCl buffer to the alcohol solvent is preferably 4:1-12:1, more preferably 4.5-9:1, and may be 4.5:1, 6:1, 7:1, 8:1 or 9:1. In the present invention, the alcohol solvent is used to uniformly disperse the hydrophobic drug, thereby facilitating dopamine to be encapsulated with these dispersed drugs as the core. In the present invention, the concentration of the hydrophobic drug in the dispersion of the hydrophobic drug is preferably 0.01-0.8 g / L, and may be 0.05, 0.09, 0.1 or 0.5 g / L. In the present invention, the preparation method of the dispersion of the hydrophobic drug is preferably: mixing Tris-HCl buffer and alcohol solvent to obtain a mixed solution; adding the mixed solution to the hydrophobic drug, and ultrasonically dispersing to obtain the dispersion of the hydrophobic drug; the ultrasonic dispersion time can be 2 minutes.

[0038] In the present invention, the mass ratio of the hydrophobic drug to dopamine hydrochloride in the dispersion of the hydrophobic drug is preferably 10-20:100-200, more preferably 10:100-200; when the amount of dopamine hydrochloride added is too much, the encapsulation rate of the product increases, but the drug loading amount will decrease; and when the amount of dopamine hydrochloride added is too little, the generated polydopamine is difficult to completely encapsulate the drug, resulting in a decrease in the drug loading amount.

[0039] In the present invention, the coating reaction temperature is preferably room temperature, the time is preferably 12 to 24 hours, and can be 12, 15, 20 or 24 hours; the coating reaction is preferably carried out under stirring conditions, and the stirring rate is preferably 300 to 500 rpm, and can be 300, 400 or 500 rpm. In the present invention, the dopamine hydrochloride is preferably added to the dispersion of the hydrophobic drug, and the coating reaction is carried out at room temperature and under magnetic stirring conditions. During the coating reaction, dopamine is oxidized and self-polymerized into polydopamine (PDA). Polydopamine contains active catechol groups and amino groups, which can interact with various groups of hydrophobic drugs to achieve the encapsulation of hydrophobic drugs: (1) Hydrogen bonding: The amino groups of PDA can form hydrogen bonds with drug molecules containing carboxylic acid, ester or amide groups to enhance the interaction between molecules; (2) π-π stacking: The aromatic ring structure of PDA enables it to interact with other drug aromatic rings through π-π stacking to form a stable complex; (3) Hydrophobic interaction: The hydrophobic characteristics of PDA enable it to combine with hydrophobic drug molecules through hydrophobic interactions to form a stable complex, thereby improving the solubility and bioavailability of the drug. Hydrophobic drugs dispersed in the solution can basically be combined with polydopamine and loaded.

[0040] After the coating reaction is completed, the present invention preferably centrifuges and washes the obtained reaction solution and dries it in sequence to obtain the drug-coated polydopamine (i.e., the intermediate product). In the present invention, the washing reagent used in the centrifugal washing is preferably deionized water, the centrifugal speed of the centrifugal washing can be 1000rpm, the number of centrifugations can be 3 times, and the time of a single centrifugation can be 10min; the drying is preferably vacuum drying, the temperature of the vacuum drying can be 25°C, and the time can be 12h. In an embodiment of the present invention, the drug-coated polydopamine is expressed as "drug@PDA". In the present invention, the particle size of the drug-coated polydopamine is preferably 150-205nm.

[0041] After obtaining the drug-coated polydopamine, the present invention mixes the drug-coated polydopamine with N,N-dimethylaminoethyl methacrylate (DMAEMA) to carry out graft polymerization reaction to obtain the pH and photothermal dual-responsive polydopamine nano drug-loaded microspheres.

[0042] In the present invention, the dosage ratio of the drug-coated polydopamine to N,N-dimethylaminoethyl methacrylate is preferably (2-5) mg: (2-3) mL, more preferably (2-4) mg: 2 mL, and can be 3 mg: 2 mL. When the amount of N,N-dimethylaminoethyl methacrylate added is too much, the outermost graft layer of the product is too thick, and the total cumulative drug release of the product will decrease, making it difficult to reach an effective drug concentration at the tumor site; when the amount of N,N-dimethylaminoethyl methacrylate added is too little, the outermost graft layer of the product is too thin, and there is almost no pH response effect, and the total cumulative drug release of the product will increase, and a large amount of drug will be released in the blood vessel after intravenous injection. In the present invention, the method of mixing the drug-coated polydopamine with N,N-dimethylaminoethyl methacrylate is preferably: adding N,N-dimethylaminoethyl methacrylate to the drug-coated polydopamine, and ultrasonically dispersing. Since N,N-dimethylaminoethyl methacrylate (DMAEMA) is a monomer containing an unsaturated double bond, it is easy to undergo free radical polymerization under the catalysis of light, heat or impurities. In order to prevent it from polymerizing during storage, transportation or processing, commercial DMAEMA usually needs to be added with an inhibitor. Therefore, before using commercial DMAEMA, it needs to be passed through a column filled with basic alumina to remove the inhibitor.

[0043] In the present invention, the temperature of the graft polymerization reaction is preferably room temperature, and the time is preferably 0.5 to 8 hours, which can be 0.5, 1, 2, 3 or 5 hours. If the graft polymerization reaction time is too long, the outer graft layer is too thick, and the cumulative release amount at the tumor site is lower; if the graft polymerization time is too short, the graft layer is too thin, the pH responsiveness is poor, and a large amount of drugs will be released immediately after injection into the blood vessel. In the present invention, the graft polymerization reaction is preferably carried out under the conditions of darkness (i.e., avoiding light), ultraviolet irradiation and stirring, that is, stirring is carried out while ultraviolet irradiation; the power of the ultraviolet irradiation is preferably 8 to 20W, which can be 8, 10, 15 or 20W, and the ultraviolet irradiation is preferably irradiated using an ultraviolet lamp close to the outer wall of the reaction container; the stirring rate is preferably 200 to 500rpm, which can be 300rpm. The present invention preferably mixes the drug-coated polydopamine with N,N-dimethylaminoethyl methacrylate, exhausts the air in the reaction container with argon, seals the reaction container (to prevent air from entering), and then performs the graft polymerization reaction. In the present invention, PDA generates carbon free radicals as initiators under ultraviolet irradiation, attacks the carbon-carbon double bonds of the electron-poor monomer DMAEMA, and then gradually polymerizes through a free radical chain reaction (chain initiation → chain growth) to form a polymer chain. The graft polymerization reaction performed under ultraviolet irradiation conditions of the present invention is self-initiated photografting and photopolymerization (SIPGP), which does not require the introduction of catalysts or inducers, and can react only by ultraviolet irradiation. The process is simple, green, and pollution-free, and the toxicity of the product can be reduced, which is conducive to large-scale production.

[0044] After the graft polymerization reaction is completed, the present invention preferably centrifuges and washes the obtained reaction solution and dries it in sequence to obtain the pH and photothermal dual-responsive polydopamine nano-drug-loaded microspheres. In the present invention, the washing agent used in the centrifugal washing is preferably deionized water, the centrifugal speed of the centrifugal washing can be 1000rpm, the number of centrifugations can be 3 times, and the time of a single centrifugation can be 10min; the drying is preferably vacuum drying, the temperature of the vacuum drying can be 25°C, and the time can be 12h.

[0045] In an embodiment of the present invention, the pH and photothermal dual-responsive polydopamine nano-drug-loaded microspheres are expressed as "drug@PDA@PDMAEMA".

[0046] The present invention does not add any additional initiator or catalyst during the entire process of preparing the pH and photothermal dual-responsive polydopamine nano-drug-loaded microspheres, which conforms to the synthesis concept of green chemistry and greatly reduces the toxicity of the product. The present invention provides a new solution for the development of an environmentally friendly intelligent drug delivery system, a solution for overcoming cancer drug resistance, and support for the application of functional polymer materials in the biomedical field.

[0047] The present invention provides the use of the pH and photothermal dual-responsive polydopamine nano-drug-loaded microspheres described in the above technical solution or the pH and photothermal dual-responsive polydopamine nano-drug-loaded microspheres prepared by the preparation method described in the above technical solution in the preparation of drugs for treating tumors. The dual-responsive polydopamine nano-drug-loaded microspheres provided by the present invention have obvious pH responsiveness and photothermal effects, good biocompatibility, and are expected to solve various insoluble drug loading and targeting problems, and achieve the purpose of multidisciplinary combined treatment of tumors.

[0048] In order to further illustrate the present invention, the pH and photothermal dual-responsive polydopamine nano-drug-loaded microspheres provided by the present invention and their preparation methods and applications are described in detail below in conjunction with examples, but they should not be construed as limiting the scope of protection of the present invention.

[0049] In the examples, dopamine hydrochloride and DMAEMA (N,N-dimethylaminoethyl methacrylate) were purchased from Merck KGaA, Darmstadt, Germany, with product numbers H8502 and 234907, respectively.

[0050] Example 1

[0051] 10 mg of the poorly soluble drug Evodiamine (EVO) was weighed, added into a mixed solution consisting of 10 mL of anhydrous ethanol and 90 mL of pH = 8.5 Tris-HCl buffer (concentration of 0.01 mol / L), and dispersed for 2 min using an ultrasonic disperser to obtain a drug dispersion. 100 mg of dopamine hydrochloride was weighed and added into the above drug dispersion, stirred at 300 rpm with a magnetic stirrer at room temperature for 1 day, washed by centrifugation with deionized water for 3 times (centrifugal speed of 1000 rpm, single centrifugation time of 10 min), and dried in vacuo at 25 °C for 12 h to obtain a polydopamine intermediate (EVO@PDA) coated with the poorly soluble drug Evodiamine, denoted as A1.

[0052] Referring to the preparation method of A1, the mass of dopamine hydrochloride added was adjusted to 200 mg, and the volume of anhydrous ethanol added was 10 mL to obtain an intermediate product, which was recorded as A2.

[0053] Referring to the preparation method of A1, the mass of dopamine hydrochloride added was adjusted to 200 mg, and the volume of anhydrous ethanol added was 15 mL to obtain an intermediate product, which was recorded as A3.

[0054] Referring to the preparation method of A1, the mass of dopamine hydrochloride added was adjusted to 200 mg, and the volume of anhydrous ethanol added was 20 mL to obtain an intermediate product, which was recorded as A4.

[0055] The polydopamine intermediates A1, A2, A3, and A4 coated with the poorly soluble drug evodiamine were subjected to a calculation experiment of drug loading and encapsulation efficiency. First, the poorly soluble drug evodiamine was measured by the standard curve, and then all the liquids in the deionized water washing process were collected. The collected liquid was diluted by a suitable multiple and placed in an ultraviolet visible spectrophotometer (Japan Shimadzu Corporation, UV-2600) to calculate the free drug concentration, and the drug loading and encapsulation efficiency were calculated according to the input drug mass and the dopamine hydrochloride mass. Take 3 portions of each of the intermediates A1, A2, A3, and A4, and calculate the drug loading and encapsulation efficiency according to the above conditions, and the results are shown in Table 1.

[0056] Table 1 Drug loading and encapsulation efficiency of the intermediate product prepared in Example 1

[0057]

[0058] It can be seen that, under the same conditions, only the mass of dopamine hydrochloride added was increased from 100 mg to 200 mg, the drug loading changed from 16.7% to 14.0%, and the encapsulation rate increased significantly, from 40.7% to 80.8%, which was due to the fact that 100 mg of dopamine hydrochloride was not enough to cover the drug added; after the mass of dopamine hydrochloride added was increased to 200 mg, when the volume of anhydrous ethanol in the mixed solution was 10 mL, 15 mL and 20 mL, the drug loading increased insignificantly, which were 14.0%, 14.6% and 15.3%, respectively; while the encapsulation rate decreased successively, which were 80.8%, 72.3% and 61.2%, respectively. Considering the encapsulation rate and drug loading, the intermediate product A2 was the best, so the experiment was continued with the feed ratio and synthesis method of A2.

[0059] Example 2

[0060] Select a suitable basic alumina column, load the basic alumina into the column, ensure that there are no bubbles inside the column, pour the commercial DMAEMA (N, N-dimethylaminoethyl methacrylate) solution into the top of the column, let it flow naturally through the basic alumina, collect the eluent, pay attention to the color and clarity of the eluent, and obtain DMAEMA after removing the inhibitor. Weigh 3 mg of the vacuum-dried intermediate product A2 (the intermediate product prepared in Example 1) in a glass tube, add 2 mL of DMAEMA after removing the inhibitor, and ultrasonically disperse. Pass argon gas for 10 minutes to exhaust the air in the glass tube, use a sealing plug to seal the test tube mouth, ensure that the plug fits tightly with the test tube mouth to prevent air from infiltrating.

[0061] In a dark environment, an 8W ultraviolet lamp was used to irradiate close to the tube wall for 0.5 h. During this period, the magnetic stirrer was kept stirring at 300 rpm at room temperature. Then, it was centrifuged and washed three times with deionized water (the centrifugal speed was 1000 rpm, and the single centrifugation time was 10 min). It was vacuum dried at 25°C for 12 h to obtain dual-responsive polydopamine nanodrug-loaded microspheres (EVO@PDA@PDMAEMA), which were recorded as the final product B1.

[0062] Referring to the preparation method of the final product B1, the ultraviolet irradiation time was adjusted to 1 h and 2 h, respectively, to obtain the final products, which were recorded as B2 and B3, respectively.

[0063] The dual-responsive polydopamine nanoparticles B1, B2 and B3 prepared in Example 2 were subjected to drug cumulative release experiments under different pH conditions (pH=6.5, 7.4). The results are shown in Tables 2 and 3.

[0064] Table 2 Cumulative release changes of dual-responsive polydopamine nanoparticles prepared in Example 2 under pH 6.5 environment

[0065]

[0066] Table 3 Cumulative release changes of dual-responsive polydopamine nanoparticles prepared in Example 2 under pH 7.4 environment

[0067]

[0068] It can be seen that, whether under the conditions of pH 6.5 or pH 7.4, as the UV irradiation time increases, the drug release rate and the total amount of cumulative release of the final product will decrease. In addition, when the UV irradiation time is the same, the cumulative release of the final product drug at pH 6.5 is always higher than that at pH 7.4. In summary, these results show that the dual-responsive polydopamine drug-loaded microspheres are pH-responsive.

[0069] Figure 3 The SEM, TEM and particle size distribution diagrams of the intermediate product A2 prepared in Example 1 and the dual-responsive polydopamine nano-drug-loaded microspheres B2 prepared in Example 2 are shown in FIG. Figure 3A in the figure is the SEM image of the intermediate product A2, the inset in A is the TEM image of the intermediate product A2, B is the particle size distribution of the intermediate product A2, C is the SEM image of the dual-responsive polydopamine nanoparticles B2, the inset in C is the TEM image of the dual-responsive polydopamine nanoparticles B2, and D is the particle size distribution of the dual-responsive polydopamine nanoparticles B2. It can be seen that the intermediate product A2 and the dual-responsive polydopamine nanoparticles B2 are both spherical, and the aggregation degree of B2 is lower than that of A2, which is due to the grafting of PDMAEMA on the surface; the particle size of A2 is 179.53±21.68nm, while the particle size of B2 is 204.86±18.07nm, both of which are within the range of enhanced permeability and retention effect (EPR) of tumors, proving that the dual-responsive polydopamine nanoparticles (B2) with appropriate particle size have been successfully prepared.

[0070] Example 3

[0071] Select a 2 mL centrifuge tube, prepare deionized water and the same concentration of A2 and B2 (200 μg / mL), take 1.5 mL of each into a centrifuge tube, and disperse evenly, wherein A2 is the intermediate product prepared in Example 1, and B2 is the dual-responsive polydopamine nano-drug microspheres prepared in Example 2. Prepare an 808 nm near-infrared laser, and adjust the power of the laser through a laser power device.

[0072] At room temperature, use 808nm near-infrared to irradiate the centrifuge tube wall, allowing the light to pass through the sample solution, and adjust the laser power to 1.5W / cm 2 , time 5min, during which the solution temperature change is recorded every 20s and a heating curve is drawn, such as Figure 4 As shown in A.

[0073] Take another portion of B2 and use 808nm near-infrared irradiation to irradiate the centrifuge tube wall at room temperature with the same concentration, so that the light passes through the sample solution and adjust the laser power to 1.5W / cm 2 , time 5min. After 5min, turn off the laser, wait for the solution to cool down to near room temperature, then turn on the laser again, which is counted as one cycle. Repeat the same cycle 5 times. During this period, record the solution temperature change every 20s and draw a temperature rise curve, such as Figure 4 As shown in B.

[0074] It can be seen that under the irradiation of 808nm near-infrared laser with the same power density, the maximum temperature of A2 and B2 with the same concentration is slightly lower than that of the former, but it can still rise by about 30℃ ( Figure 4 In addition, after 5 cycles of near-infrared laser irradiation and cooling ( Figure 4In B), the maximum temperature of B2 did not decrease significantly, and the photothermal performance did not decay significantly, indicating that B2 has excellent photothermal conversion effect and good photothermal stability, and can be used as a photothermal material for photothermal treatment of tumors.

[0075] The above description is only a preferred embodiment of the present invention and does not limit the present invention in any form. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be regarded as the protection scope of the present invention.

Claims

1. A pH and photothermal dual-responsive polydopamine nanoparticle drug-loaded microsphere, characterized in that: The invention comprises a hydrophobic drug, polydopamine coating the hydrophobic drug and polymethacrylate N,N-dimethylaminoethyl ester grafted on the surface of the polydopamine.

2. The pH and photothermal dual-responsive polydopamine nanoparticles according to claim 1, characterized in that: The hydrophobic drugs include evodiamine.

3. The pH and photothermal dual-responsive polydopamine nanoparticles according to claim 1 or 2, characterized in that: The particle size of the pH and photothermal dual-responsive polydopamine nano drug-loaded microspheres is 180-225 nm.

4. The method for preparing the pH and photothermal dual-responsive polydopamine nanoparticles according to any one of claims 1 to 3, characterized in that: The following steps are involved: The dispersion of the hydrophobic drug is mixed with dopamine hydrochloride to perform a coating reaction to obtain drug-coated polydopamine; The drug-coated polydopamine is mixed with N,N-dimethylaminoethyl methacrylate to carry out graft polymerization reaction to obtain the pH and photothermal dual-responsive polydopamine nano drug-loaded microspheres.

5. The preparation method according to claim 4, characterized in that: The dispersion of the hydrophobic drug comprises the hydrophobic drug, Tris-HCl buffer and alcohol solvent; the pH value of the Tris-HCl buffer is 8.0-8.5; and the volume ratio of the Tris-HCl buffer to the alcohol solvent is 4:1-12:

1.

6. The preparation method according to claim 4 or 5, characterized in that: The concentration of the hydrophobic drug in the dispersion of the hydrophobic drug is 0.01-0.8 g / L; the mass ratio of the hydrophobic drug to dopamine hydrochloride in the dispersion of the hydrophobic drug is 10-20:100-200.

7. The preparation method according to claim 4, characterized in that: The coating reaction is carried out at room temperature for 12 to 24 hours; the coating reaction is carried out under stirring at a stirring rate of 300 to 500 rpm.

8. The preparation method according to claim 4, characterized in that: The dosage ratio of the drug-coated polydopamine to N,N-dimethylaminoethyl methacrylate is (2-5) mg: (2-3) mL.

9. The preparation method according to claim 4 or 8, characterized in that: The graft polymerization reaction is carried out at room temperature for 0.5 to 8 hours. The graft polymerization reaction is carried out in darkness, under ultraviolet irradiation and stirring. The power of the ultraviolet irradiation is 8 to 20 W, and the stirring rate is 200 to 500 rpm.

10. Use of the pH and photothermal dual-responsive polydopamine nanoparticles drug-loaded microspheres according to any one of claims 1 to 3 or the pH and photothermal dual-responsive polydopamine nanoparticles drug-loaded microspheres prepared by the preparation method according to any one of claims 4 to 9 in the preparation of drugs for treating tumors.

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