A kind of highly cross-linked amphiphilic embolic microsphere and preparation method thereof

Abstract

The present invention discloses a highly cross-linked amphiphilic embolic microsphere and a preparation method thereof. The hydroxyl groups of polyvinyl alcohol embolic microspheres are used as grafting sites, and a benzene ring-containing cross-linking agent is grafted between the polymer main chains of the polyvinyl alcohol embolic microspheres to form a bridging structure containing a benzene ring, thereby obtaining the highly cross-linked amphiphilic embolic microspheres. The cross-linked structure formed between the polyvinyl alcohol polymer chains helps to prop the polymer chains apart from each other, forming more pore structures, thereby improving the utilization rate of the ionic functional groups in the microspheres and further improving the loading efficiency of the microspheres for drugs. On the other hand, by modifying the polymer skeleton of the polyvinyl alcohol embolic microspheres using the method of the present invention, the obtained highly cross-linked amphiphilic embolic microspheres can have functional characteristics such as drug loading and development of various properties, which can benefit more patients in actual clinical applications.

Description

Technical Field

[0001] The invention belongs to the technical field of medical materials, and in particular relates to a highly cross-linked amphiphilic embolic microsphere and a preparation method thereof. Background Art

[0002] Drug-eluting microsphere transcatheter arterial chemoembolization (D-TACE) is a different interventional treatment for mid-to-late stage liver cancer than traditional transcatheter arterial chemoembolization (c-TACE). It has become a hot spot in the current research and application of hepatocellular carcinoma (HCC) treatment due to its good therapeutic effect and lower side effects. D-TACE mainly uses drug-loaded microspheres as embolic agents. The drug-loaded microspheres have functional groups that can combine with chemotherapy drugs, allowing the drug-loaded microspheres to stably load chemotherapy drugs. Therefore, in addition to the physical embolization effect of D-TACE, the drug-loaded microspheres can also slowly release chemotherapy drugs, allowing chemotherapy drugs to act on tumors more accurately for a longer time, achieving a therapeutic effect of 1+1>2.

[0003] In the clinical application scenario of drug-eluting embolic microspheres, before using them for hepatic artery embolization treatment, blank microspheres need to be used to load the drug first. Currently, embolic microspheres with drug-loading functions on the market, such as DC Bead, Callisphere, Embosphere and Hepasphere, are all combined by the mutual attraction of anionic groups in the microsphere polymer skeleton and cationic groups in the chemotherapy drugs through Coulomb force. Because the polymer network structure of the embolic microspheres is relatively loose, on the one hand, the drug loading performance of the microspheres is unstable, and the drug loading efficiency fluctuates greatly with changes in ambient temperature or time. On the other hand, the loose structure will make the loading of the microspheres on the drug mainly rely on the binding of the anionic groups on the polymer chain segments to the drugs. Moreover, the single drug binding mechanism makes the microspheres have limited loading efficiency for drugs, which leads to the need to spend more time preparing drug-loaded embolic microspheres during clinical use. At the same time, due to the presence of a large number of hydroxyl structures on the surface of the embolic microspheres, a large number of adhesions will occur between the microspheres, which will not only significantly reduce the drug loading efficiency of the microspheres, but also the adhesion and aggregation of multiple drug-loaded microspheres will significantly reduce the suspension and catheter passability of the microspheres. Summary of the invention

[0004] The present invention aims to solve at least one of the technical problems existing in the above-mentioned prior art. To this end, the present invention provides a highly cross-linked amphiphilic embolic microsphere and a preparation method thereof, wherein the prepared highly cross-linked amphiphilic embolic microsphere not only has improved drug loading efficiency, stability, monodispersity, suspension and catheter permeability, but also has a multi-property drug efficient loading and imaging function.

[0005] The first aspect of the present invention provides a method for preparing highly cross-linked amphiphilic embolic microspheres, comprising: using the hydroxyl groups of polyvinyl alcohol embolic microspheres as grafting sites, grafting a benzene ring-containing cross-linking agent between the polymer main chains of the polyvinyl alcohol embolic microspheres to form a bridging structure containing benzene rings, and obtaining the highly cross-linked amphiphilic embolic microspheres.

[0006] In some embodiments of the present invention, the polyvinyl alcohol embolic microspheres are prepared by cross-linking a polyvinyl alcohol derivative with a monomer containing a sulfonic acid or carboxylic acid group through a reverse suspension polymerization method, and the polyvinyl alcohol derivative is prepared by reacting a water-soluble cross-linking agent with a polyvinyl alcohol polymer.

[0007] In some embodiments of the present invention, the water-soluble cross-linking agent is a dimethoxy-type small molecule containing a double bond.

[0008] In some embodiments of the present invention, the benzene ring-containing cross-linking agent is a multi-aldehyde or alkoxy small molecule containing at least one benzene ring.

[0009] In some embodiments of the present invention, the multi-aldehyde or alkoxy small molecule containing at least one benzene ring includes at least one of 1,4-phthalaldehyde, terephthalaldehyde bis(diethyl acetal), 4,4'-biphenyldicarboxaldehyde or tris(4-formylphenyl)amine, N-(5-methylisophthalaldehyde)iodopyrrole, 5-imino-tert-butylcalix[6]arene isophthalaldehyde, 5-imino-tert-butylcalix[5]arene isophthalaldehyde, 5-imino-tert-butylcalix[4]arene isophthalaldehyde, N-β-cyclodextrin-5-iminoisophthalaldehyde, N-α-cyclodextrin-5-iminoisophthalaldehyde or N-γ-cyclodextrin-5-iminoisophthalaldehyde.

[0010] In some embodiments of the present invention, the benzene ring-containing crosslinking agent is crosslinked with the polyvinyl alcohol microspheres through any one of the following structures:

[0011]

[0012] In some embodiments of the present invention, the water-soluble crosslinking agent includes at least one of N-(2,2-dimethoxyethyl)-2-acrylamide, 8,8-dimethoxy-2,6-dimethyl-2-octene, methylvinyldimethoxysilane, 8,8-dimethoxy-2,6-dimethyl-2-octene, 1,1-dimethoxy-3,7-dimethyl-2,6-octadiene, 1,1-dimethoxyethylene or 4-(1-propenyl)-1,2-dimethoxybenzene.

[0013] In some embodiments of the present invention, the monomer containing a sulfonic acid or carboxylic acid group includes at least one of 2-acrylamide-2-methylpropane sulfonic acid sodium salt, 2-methyl-2-propylene-1-sulfonate sodium salt, 4-styrene sulfonate sodium salt, 3-allyloxy-2-hydroxy-1-propane sulfonic acid sodium salt, acrylic acid, 5-hexenoic acid, 7-octenoic acid or 8-nonenoic acid.

[0014] In some embodiments of the present invention, in the preparation method, a water-soluble cross-linking agent is used to modify the grafting site on the polyvinyl alcohol polymer, and a polymerization reaction is carried out with a water-soluble monomer with an ionic functional group to generate polyvinyl alcohol embolic microspheres, and a benzene ring-containing cross-linking agent is used to modify the hydroxyl site of the polyvinyl alcohol embolic microspheres to form a bridging structure containing a benzene ring. The obtained highly cross-linked amphiphilic embolic microspheres have functional characteristics such as efficient drug loading and development of various properties.

[0015] In some embodiments of the present invention, during the preparation of highly cross-linked amphiphilic embolic microspheres, the benzene ring-containing cross-linking agent can undergo a cross-linking reaction with the prepared polyvinyl alcohol embolic microspheres to obtain highly cross-linked amphiphilic embolic microspheres, or can react together in the polyvinyl alcohol embolic microsphere synthesis system to obtain highly cross-linked amphiphilic embolic microspheres.

[0016] In some embodiments of the present invention, polyvinyl alcohol embolic microspheres and a benzene ring-containing crosslinking agent are mixed, an organic solvent and a catalyst are added, and a crosslinking reaction occurs at 40° C. to 80° C. to obtain the highly crosslinked amphiphilic embolic microspheres.

[0017] In some embodiments of the present invention, the obtained polyvinyl alcohol embolic microspheres are dried to obtain powdered dry polyvinyl alcohol embolic microspheres, which are then mixed with a benzene ring-containing cross-linking agent.

[0018] In some embodiments of the present invention, during the preparation of the powdered dry polyvinyl alcohol embolic microspheres, 1 part of polyvinyl alcohol embolic microspheres is mixed with 1-3 parts of deionized water, 2-4 parts of acetone are added for the first time, the mixture is allowed to stand for the first time, and the supernatant is removed, 3-5 parts of acetone are added for the second time, the mixture is allowed to stand for the second time, and the supernatant is removed, the addition of acetone is repeated, the mixture is allowed to stand, and the supernatant is removed, excess acetone is poured out, and the microspheres are placed in a vacuum oven at 40°C-80°C and dried overnight to obtain dry polyvinyl alcohol embolic microsphere powder. Preferably, the number of repetitions is 2-4 times.

[0019] In some embodiments of the present invention, the organic solvent is dimethyl sulfoxide and the catalyst is methanesulfonic acid. Preferably, dimethyl sulfoxide is added to the mixed solution of polyvinyl alcohol embolic microspheres and the benzene ring-containing crosslinking agent. Preferably, the mixing method is stirring, the stirring temperature includes 40°C-80°C, and the stirring time is 8h-14h. Further, methanesulfonic acid is added to the polyvinyl alcohol embolic microspheres containing dimethyl sulfoxide and the reaction solution of the benzene ring crosslinking agent, and the mixing is continued. The mixing method is stirring, and the stirring time is 45h-50h.

[0020] In some embodiments of the present invention, the ratio of the benzene ring-containing crosslinking agent to the polyvinyl alcohol embolic microspheres is 1*10 -4 mol / g-5*10 -3 mol / g.

[0021] In some embodiments of the present invention, comprising:

[0022] A water-soluble crosslinking agent and a polyvinyl alcohol polymer are mixed and reacted under acidic conditions, and after the reaction is completed, the mixture is placed in an alkaline solution for neutralization to obtain a polyvinyl alcohol derivative;

[0023] The polyvinyl alcohol derivative, the monomer containing sulfonic acid or carboxylic acid group, the benzene ring-containing cross-linking agent and the catalyst are mixed to prepare an aqueous phase material;

[0024] Adding an oil-soluble dispersant to an organic solvent to prepare an oil phase material;

[0025] The aqueous phase material is added to the oil phase material, and a polymerization reaction occurs at 60° C. to 100° C. to obtain the highly cross-linked amphiphilic embolic microspheres.

[0026] In some embodiments of the present invention, the preparation of the acidic condition comprises adding an acidic solution such as hydrochloric acid.

[0027] In some embodiments of the present invention, the alkaline solution includes sodium hydroxide solution or the like.

[0028] In some embodiments of the present invention, the water-soluble cross-linking agent and the polyvinyl alcohol polymer are mixed and / or reacted by stirring.

[0029] In some embodiments of the present invention, the mass ratio of the polyvinyl alcohol derivative to the monomer containing sulfonic acid or carboxylic acid groups is (3-8): 1. Optionally, the polyvinyl alcohol derivative is first mixed with water to obtain a polyvinyl alcohol derivative solution. Optionally, the mixing method adopts a stirring method, and the stirring speed includes 300r / min-500r / min; further, the monomer containing sulfonic acid or carboxylic acid groups and the catalyst are mixed evenly and then mixed with the polyvinyl alcohol derivative solution, and optionally, the mixing time with the polyvinyl alcohol derivative solution is 30min-1h.

[0030] In some embodiments of the present invention, the catalyst is hydrochloric acid and / or ammonium persulfate.

[0031] In some embodiments of the present invention, the mixed solution generated after the polyvinyl alcohol derivative, the monomer containing sulfonic acid or carboxylic acid group, the benzene ring-containing crosslinking agent and the catalyst are mixed is placed in an ultrasonic cleaning machine for 3-5 minutes.

[0032] In some embodiments of the present invention, the oil-soluble dispersant is cellulose acetate butyrate. Preferably, the organic solvent is butyl acetate. Preferably, the oil-soluble dispersant is added to the organic solvent for mixing and heating to obtain an oil phase. Preferably, the mixing method is stirring, and the stirring speed is 300r / min-500r / min. Preferably, the heating temperature includes 40°C-100°C.

[0033] In some embodiments of the present invention, the aqueous phase solution is slowly added dropwise to the oil phase using a syringe, and after the aqueous phase and the oil phase are mixed, the temperature is raised to 60° C. to 100° C. and stirring is continued for 20 min to 50 min.

[0034] In some embodiments of the present invention, during the preparation of highly cross-linked amphiphilic embolic microspheres, after adding the aqueous phase material to the oil phase material, tetramethylethylenediamine is added to the mixed solution of the aqueous phase and the oil phase, and the mixture is stirred for the first time, heated, stirred for the second time, taken out, and cooled to obtain highly cross-linked amphiphilic embolic microspheres. Preferably, the first stirring time is 2h-5h. Preferably, the heating temperature includes 60°C-100°C. Preferably, the second stirring time includes 0.5h-3h.

[0035] In some embodiments of the present invention, the generated highly cross-linked amphiphilic embolic microspheres are washed multiple times with dimethyl sulfoxide and deionized water respectively to obtain the final usable highly cross-linked amphiphilic embolic microspheres. The washing times are preferably 2-5 times.

[0036] In some embodiments of the present invention, the benzene ring-containing crosslinking agent is 5-imino-tert-butylcalix[6]arene isophthalaldehyde. Preferably, the preparation method of 5-imino-tert-butylcalix[6]arene isophthalaldehyde comprises the following steps:

[0037] S11, mixing 4-tert-butylcalix[6]arene with deionized water, adding toluenesulfonyl chloride and reacting, the reaction time is preferably 16h-24h;

[0038] S12, adding a deionized water solution containing sodium hydroxide into the reaction system, and continuing the reaction, the reaction time is preferably 20 min-40 min;

[0039] S13, filtering the solution after the second reaction, taking out the clarified solution, adding ammonium chloride, and storing at a low temperature, preferably at 2° C.-6° C.;

[0040] S14, filtering the solution after low temperature storage to obtain white crystals, and vacuum drying the white crystals;

[0041] S15, mixing the dried white crystals, 5-aminoisophthalaldehyde and deionized water to react, the reaction temperature is preferably 60° C.-80° C., and the reaction time is preferably 2 h-6 h;

[0042] S16, adding acetone to the reaction solution obtained in step S25, filtering to obtain a white precipitate, collecting the white solid and drying it.

[0043] In some embodiments of the present invention, the benzene ring-containing crosslinking agent is N-β-cyclodextrin-5-iminoisophthalaldehyde. Preferably, the preparation method of N-β-cyclodextrin-5-iminoisophthalaldehyde comprises the following steps:

[0044] S21, mixing β-cyclodextrin with deionized water, adding toluenesulfonyl chloride and reacting, and the reaction time is preferably 16h-24h;

[0045] S22, adding a deionized water solution containing sodium hydroxide into the reaction system, and continuing the reaction, the reaction time is preferably 20 min-40 min;

[0046] S23, filtering the solution after the second reaction, taking out the clarified solution, adding ammonium chloride, and storing at a low temperature, preferably at 2° C.-6° C.;

[0047] S24, filtering the solution after low temperature storage to obtain white crystals, and vacuum drying the white crystals;

[0048] S25, mixing the dried white crystals, 5-aminoisophthalaldehyde and deionized water to react, the reaction temperature is preferably 60° C.-80° C., and the reaction time is preferably 2 h-6 h;

[0049] S26, adding acetone to the reaction solution obtained in step S25, filtering to obtain a white precipitate, collecting the white solid and drying it.

[0050] In some embodiments of the present invention, the highly cross-linked amphiphilic embolic microspheres have any one of the following molecular structures:

[0051]

[0052] The main chain wavy line cross-linked with R represents the main chain structure of the polyvinyl alcohol chain molecule, and the other main chain wavy line represents the main chain of the molecule containing sulfonic acid or carboxylic acid groups. R can be selected from any one of the following structures:

[0053]

[0054] Among them, benzene rings and hydrophobic cavities, as structures that interact with benzene ring-containing or hydrophobic molecules, are grafted onto the polymer skeleton of polyvinyl alcohol embolization microspheres (with hydroxyl groups as sites), so that the originally hydrophilic polymer skeleton of the microspheres is transformed into an amphiphilic structure; at the same time, the efficient attraction mechanism of hydrophobic molecules in the water system is increased, thereby improving the loading efficiency of the microspheres for drugs.

[0055] The second aspect of the present invention provides a highly cross-linked amphiphilic embolic microsphere, which is prepared by the method for preparing highly cross-linked amphiphilic embolic microspheres provided in the first aspect. In the structure of the highly cross-linked amphiphilic embolic microsphere, hydroxyl groups are used as grafting sites between the main chains of polyvinyl alcohol polymers, at least a portion is cross-linked with polymer chains with ionic functional groups through cross-linking molecules, and at least another portion is bridged with hydrophobic benzene ring molecules to form a supporting structure containing benzene rings between the main chains of polyvinyl alcohol polymers, so that the highly cross-linked amphiphilic embolic microspheres have the function of adsorbing and loading drugs.

[0056] In some embodiments of the present invention, the particle size of the highly cross-linked amphiphilic embolic microspheres ranges from 50 μm to 1500 μm.

[0057] In addition, the microspheres prepared by the method for preparing highly cross-linked amphiphilic embolic microspheres provided in the first aspect of the present invention can be used as chemotherapeutic embolic materials such as embolic agents for treating liver cancer.

[0058] The microspheres prepared by the method for preparing highly cross-linked amphiphilic embolic microspheres provided by the first aspect of the present invention can also be used for efficient loading of drugs of various properties and / or medical imaging.

[0059] Compared with the prior art, the beneficial effects of the present invention are as follows: the hydroxyl groups of polyvinyl alcohol embolic microspheres are used as grafting sites, and the benzene ring-containing crosslinking agent is grafted between the polymer main chains of the polyvinyl alcohol embolic microspheres to form a bridging structure containing benzene rings, thereby obtaining the highly crosslinked amphiphilic embolic microspheres. The benzene rings and hydrophobic cavities are used as structures that interact with the benzene ring-containing or hydrophobic molecules, and are crosslinked between the polymer skeletons of the polyvinyl alcohol embolic microspheres (with the hydroxyl groups as sites), so that the originally hydrophilic microsphere polymer skeleton is transformed into an amphiphilic structure; by increasing the efficient attraction mechanism of hydrophobic molecules in the water system, the loading efficiency of the microspheres for drugs is improved. In addition, the crosslinking structure formed between the polyvinyl alcohol polymer chains helps to open the polymer chains from each other to form more pore structures, thereby improving the utilization rate of the ionic functional groups in the microspheres and further improving the loading efficiency of the microspheres for drugs. Thus, the application of the microspheres in the clinic becomes more convenient, saving more time for the preparation before surgery. Moreover, the presence of the crosslinking structure makes the originally loose polymer microsphere structure more stable, thereby improving its performance stability. The improvement of stability helps to prolong the performance retention time of the microspheres, so that the microspheres have a longer shelf life. The cross-linking process uses the hydroxyl groups on the polyvinyl alcohol polymer chain segments as grafting sites, which reduces the number of hydroxyl groups on the surface of the microspheres to a certain extent, thereby improving the adhesion phenomenon between the microspheres. Fewer adhesion phenomena can effectively improve the catheter passability and clinical embolization effect of the microspheres. It can also be verified through experiments that the highly cross-linked amphiphilic embolic microspheres obtained by modifying the polymer skeleton of the polyvinyl alcohol embolic microspheres by the method of the present invention can have functional characteristics such as drug loading and development of various properties. For example, for the loading of hydrophobic targeting or chemotherapy drugs that cannot be achieved by the prior art, including but not limited to oxaliplatin, sorafenib or lenvatinib, the drug loading effect can be achieved, so that more patients can benefit in actual clinical applications.

[0060] It is to be understood that the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive of the present disclosure. BRIEF DESCRIPTION OF THE DRAWINGS

[0061] The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate embodiments consistent with the present invention and, together with the description, serve to explain the principles of the present invention;

[0062] Figure 1 Schematic diagram of the preparation of highly cross-linked amphiphilic embolic microspheres from polyvinyl alcohol embolic microspheres in an embodiment of the present invention;

[0063] Figure 2 This is a diagram showing the development effect of the unmodified embolic microspheres and the modified embolic microspheres in Example 5 of the present invention;

[0064] Figure 3This is a microscope image of the highly cross-linked amphiphilic embolic microspheres in Example 1 of the present invention. DETAILED DESCRIPTION

[0065] The following will be combined with the embodiments to clearly and completely describe the concept of the present invention and the technical effects produced, so as to fully understand the purpose, characteristics and effects of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, other embodiments obtained by those skilled in the art without creative work are all within the scope of protection of the present invention.

[0066] The experimental methods in the following examples without specifying specific conditions are generally carried out according to conventional conditions in the art or conditions recommended by the manufacturers; the raw materials and reagents used, unless otherwise specified, are all raw materials and reagents that can be obtained from commercial channels such as conventional markets.

[0067] Some embodiments of the present application provide a method for preparing highly cross-linked amphiphilic embolic microspheres, comprising:

[0068] The hydroxyl groups of the polyvinyl alcohol embolic microspheres are used as grafting sites, and the benzene ring-containing crosslinking agent is grafted between the polymer main chains of the polyvinyl alcohol embolic microspheres to form a bridging structure containing a benzene ring, thereby obtaining highly crosslinked amphiphilic embolic microspheres.

[0069] In a specific embodiment, the benzene ring-containing crosslinking agent can undergo a crosslinking reaction with the prepared polyvinyl alcohol embolic microspheres to obtain highly crosslinked amphiphilic embolic microspheres, or can react together in the polyvinyl alcohol embolic microsphere synthesis system to obtain highly crosslinked amphiphilic embolic microspheres.

[0070] In a specific embodiment, the grafting sites are modified on the polyvinyl alcohol polymer using a water-soluble cross-linking agent, and then a polymerization reaction is carried out with a water-soluble monomer with an ionic functional group to generate polyvinyl alcohol embolic microspheres. The hydroxyl sites of the polyvinyl alcohol embolic microspheres are then modified with a benzene ring-containing cross-linking agent to form a bridging structure containing a benzene ring. The obtained highly cross-linked amphiphilic embolic microspheres have functional properties such as efficient drug loading and development of various properties.

[0071] In a specific embodiment, the ratio of the benzene ring-containing crosslinking agent to the polyvinyl alcohol embolic microspheres is 1*10 - 4 mol / g-5*10 -3 mol / g.

[0072] In a specific implementation manner, the polyvinyl alcohol may be polyvinyl alcohol 67000, polyvinyl alcohol 1788, polyvinyl alcohol 0588, or the like.

[0073] Example 1

[0074] Disclosed is a highly cross-linked amphiphilic embolic microsphere, the preparation method of which comprises the following steps:

[0075] Step 1: Prepare the reverse suspension polymerization aqueous phase:

[0076] Weigh 50g of 15% polyvinyl alcohol solution into a 250mL flask, set the speed to 200r / min, add 0.125mL of N-(2,2-dimethoxyethyl)-2-acrylamide, and stir for 10-15min. Then, drop 5mL of hydrochloric acid solution into the 250mL flask and stir overnight. Place the 250mL flask in an ice bath and stir, and slowly drop 10% sodium hydroxide solution to adjust its pH value to 7 to obtain a polyvinyl alcohol derivative.

[0077] Weigh 20g of polyvinyl alcohol derivative and place it in a 50mL beaker and stir (speed: 300r / min). Weigh 4g of 2-acrylamide-2-methylpropane sulfonic acid sodium salt and add it to a vial, then add 2mL of deionized water, 400μL of hydrochloric acid and 1000μL of 10% ammonium persulfate solution in sequence and mix thoroughly. Add the prepared mixture to a 50mL beaker and continue stirring for 30min. After stirring, place the beaker in an ultrasonic cleaner and ultrasonicate for 3-5min to obtain an aqueous phase material.

[0078] Step 2: Prepare the reverse suspension polymerization oil phase:

[0079] Weigh 200 mL of butyl acetate and 6 g of cellulose acetate butyrate into a 500 mL three-necked flask, place it in an oil bath, and stir at 500 r / min until fully mixed. Raise the temperature to 56°C and continue stirring for 20 minutes to obtain an oil phase material.

[0080] Step 3: Reverse suspension polymerization:

[0081] The aqueous solution obtained in step 1 was slowly added to the oil phase obtained in step 2 using a 10 mL syringe. The addition time was 10-20 min. After the addition was completed, stirring was continued for 30 min. 800 μL of tetramethylethylenediamine (TEDMA) was added to the above solution and stirring was continued for 3 h. The system was heated to 80 ° C, stirred for 1 h, and then the 500 mL three-necked flask was removed from the oil bath and cooled. Polyethanol embolic microspheres were obtained.

[0082] Step 4: Modification of polymer skeleton of embolization microspheres:

[0083] Take 100mL of polyvinyl alcohol embolic microspheres and put them into a 500mL three-necked flask, add 100mL of deionized water and stir. Slowly add 200mL of acetone and let it stand for 30 minutes. Remove the supernatant, slowly add 300mL of acetone, let it stand, and remove the supernatant. Repeat the above steps twice. After pouring out the excess acetone, place the microspheres in a 60℃ vacuum oven and dry overnight. Obtain dry polyvinyl alcohol embolic microsphere powder. Weigh 0.2g of polyvinyl alcohol embolic microspheres and 2*10 -4 mol 1,4-benzenedialdehyde was added to a 250mL three-necked flask. 30mL dimethyl sulfoxide was added and stirred at 60℃ for 12h. 1mL methanesulfonic acid (CH3SO3H) was added to the reaction system and stirred for 48h. After the reaction, the microspheres were washed three times with DMSO and deionized water respectively. Highly cross-linked amphiphilic embolic microspheres were obtained.

[0084] like Figure 1 As shown in the figure, it is a schematic diagram of the preparation of highly cross-linked amphiphilic embolic microspheres formed from polyvinyl alcohol embolic microspheres in this embodiment. In the reaction between the grafting agent and the polyvinyl alcohol polymer, cross-linked molecules between the main chains of polyvinyl alcohol molecules are formed on the main chain of the polyvinyl alcohol molecules, and react with the monomer containing the sulfonic acid group to form a main chain of the sulfonic acid group-containing molecule, thereby generating polyvinyl alcohol embolic microspheres. The hydroxyl groups of the polyvinyl alcohol embolic microspheres are used as grafting sites to graft small organic molecules with R functional groups onto the polymer skeleton of the polyvinyl alcohol embolic microspheres. The organic small molecules with R functional groups grafted in this embodiment are specifically 1,4-benzenedicarboxaldehyde, and the structure of its R functional group is: The benzene ring and the hydrophobic cavity are structures that interact with benzene rings or hydrophobic molecules, and are cross-linked between the polymer skeletons of the polyvinyl alcohol embolic microspheres (with hydroxyl groups as sites), so that the originally hydrophilic polymer skeleton of the microspheres is transformed into an amphiphilic structure, and the loading efficiency of the microspheres for drugs is improved by increasing the efficient attraction mechanism of hydrophobic molecules in the water system. In addition, the cross-linked structure formed between the polyvinyl alcohol polymer chains helps to stretch the polymer chains apart, forming more pore structures, thereby improving the utilization rate of the sulfonic acid groups in the microspheres and further improving the loading efficiency of drugs.

[0085] Example 2

[0086] The preparation method of the highly cross-linked amphiphilic embolic microspheres disclosed in this embodiment is different from that of Example 1 only in that an equal molar amount of terephthalaldehyde bis(diethyl acetal) is used to replace the 1,4-phthalaldehyde in Example 1.

[0087] like Figure 1 As shown, the organic small molecule with R functional group grafted in this embodiment is specifically terephthalaldehyde bis(diethyl acetal), and the structure of its R functional group is: The benzene ring and the hydrophobic cavity are structures that interact with benzene ring-containing or hydrophobic molecules, increasing the sites for loading positively charged drugs through anion-cation interactions, and grafting them onto the polymer skeleton of polyvinyl alcohol embolic microspheres (with hydroxyl groups as sites), so that the originally hydrophilic polymer skeleton of the microspheres is transformed into an amphiphilic structure, and the loading efficiency of the microspheres for drugs is improved by increasing the efficient attraction mechanism of hydrophobic molecules in the water system. In addition, the cross-linked structure formed simultaneously between the polyvinyl alcohol polymer chains helps to prop the polymer chains apart from each other, forming more pore structures, thereby improving the utilization rate of the sulfonic acid groups in the microspheres and further improving the loading efficiency of the drugs.

[0088] Example 3

[0089] This example discloses a highly cross-linked amphiphilic embolic microsphere, and the preparation method thereof is different from that of Example 1 only in that an equal molar amount of 4,4'-biphenyldicarboxaldehyde is used to replace the 1,4-benzenedicarboxaldehyde in Example 1.

[0090] like Figure 1 As shown, in this embodiment, an organic small molecule with an R functional group is grafted, specifically 4,4'-biphenyldicarboxaldehyde, and the structure of its R functional group is: The benzene ring and the hydrophobic cavity are structures that interact with benzene ring-containing or hydrophobic molecules, increase the sites for loading positively charged drugs through anion-cation interactions, and provide more sites for loading cationic drugs. They are grafted onto the polymer skeleton of polyvinyl alcohol embolization microspheres (with hydroxyl groups as sites), so that the originally hydrophilic microsphere polymer skeleton is transformed into an amphiphilic structure, and the loading efficiency of the microspheres for drugs is improved by increasing the efficient attraction mechanism of hydrophobic molecules in the water system. In addition, the cross-linked structure formed simultaneously between the polyvinyl alcohol polymer chains helps to prop the polymer chains apart from each other, forming more pore structures, thereby improving the utilization rate of the sulfonic acid groups in the microspheres and further improving the loading efficiency of the drugs.

[0091] Example 4

[0092] This example discloses a highly cross-linked amphiphilic embolic microsphere, and the preparation method thereof is different from that of Example 1 only in that an equimolar amount of tris(4-formylphenyl)amine is used to replace 1,4-benzenedicarboxaldehyde in Example 1.

[0093] like Figure 1 As shown, in this embodiment, an organic small molecule with an R functional group is grafted, specifically tris(4-formylphenyl)amine, and the structure of its R functional group is: The benzene ring and the hydrophobic cavity are structures that interact with benzene ring-containing or hydrophobic molecules, increase the sites of carboxyl groups for loading cationic drugs, and graft them onto the polymer skeleton of polyvinyl alcohol embolic microspheres (with hydroxyl groups as sites), so that the originally hydrophilic polymer skeleton of the microspheres is transformed into an amphiphilic structure, and the loading efficiency of the microspheres for drugs is improved by increasing the efficient attraction mechanism of hydrophobic molecules in the water system. In addition, the cross-linked structure formed simultaneously between the polyvinyl alcohol polymer chains helps to prop the polymer chains apart from each other, forming more pore structures, thereby improving the utilization rate of the sulfonic acid groups in the microspheres and further improving the loading efficiency of the drugs.

[0094] Example 5

[0095] This embodiment discloses a highly cross-linked amphiphilic embolic microsphere, and the preparation method thereof is different from that of embodiment 1 only in that an equimolar amount of N-(4-methylbenzaldehyde) iodine pyrrole is used to replace 1,4-benzenedicarbaldehyde in embodiment 1, wherein the preparation process of N-(5-methylbenzaldehyde) iodine pyrrole is specifically as follows:

[0096] Take 10.2g of iodine pyrrole and 20mL of deionized water and add them to a 100mL three-necked flask, and stir rapidly for 30min. Prepare 40mL of 10% sodium hydroxide solution and slowly add it to the above solution, and continue stirring for 30min. Weigh 3.3g of 5-(chloromethyl)benzene-1,3-dicarboxaldehyde and slowly add it to the above solution. Stir the reaction at 70°C for 6 hours. After the reaction is completed, concentrate the solution by rotary evaporation, and use methanol to precipitate the product. The obtained product is dried in a vacuum oven at 40°C.

[0097] like Figure 1 As shown, the organic small molecule with R functional group grafted in this embodiment is specifically N-(5-methylisophthalaldehyde) iodine pyrrole, and its R functional group structure is: This is a molecule with developing properties, which can provide X-ray developing properties for microspheres, and because of the benzene ring structure carried by this molecule, it can also promote the adsorption of benzene ring-containing or hydrophobic molecules by microspheres. It is grafted onto the polymer skeleton of polyvinyl alcohol embolization microspheres (with hydroxyl groups as sites), so that the originally hydrophilic polymer skeleton of the microspheres is transformed into an amphiphilic structure, and the loading efficiency of the microspheres for drugs is improved by increasing the efficient attraction mechanism of hydrophobic molecules in the water system. In addition, the cross-linked structure formed simultaneously between the polyvinyl alcohol polymer chains helps to prop the polymer chains apart from each other, forming more pore structures, thereby improving the utilization rate of the sulfonic acid groups in the microspheres and further improving the loading efficiency of drugs.

[0098] Example 6

[0099] This embodiment discloses a highly cross-linked amphiphilic embolic microsphere, the preparation method of which is different from that of Example 1 only in that an equal molar amount of any one of 5-imino-tert-butylcalix[6]arene isophthalaldehyde, 5-imino-tert-butylcalix[5]arene isophthalaldehyde, and 5-imino-tert-butylcalix[4]arene isophthalaldehyde is used to replace the 1,4-phthalaldehyde in Example 1.

[0100] The preparation process of 5-imino-tert-butylcalix[6]arene isophthalaldehyde is specifically as follows:

[0101] Weigh 21.5g of 4-tert-butylcalix[6]arene and 200mL of deionized water into a 500mL three-necked flask and stir. Weigh 7.5g of p-toluenesulfonyl chloride and slowly add it into the three-necked flask, keep it at room temperature, and stir it for 21h. Prepare 20mL of deionized water solution containing 5g of sodium hydroxide, slowly add it into the above reaction system, and continue stirring for 30min. Filter the above solution, take the clear solution, and add 6g of ammonium chloride into the solution to terminate the reaction. Store it in a refrigerator at 4℃ overnight. Filter the above solution to obtain white crystals. Place the white crystals in a vacuum oven until they are completely dry. Weigh 2.5g of white crystals, 7.3g of 5-aminoisophthalaldehyde and 150mL of deionized water into a 500mL three-necked flask, and stir it at 75℃ for 4h. After the reaction is completed, add acetone to the reaction solution to obtain a white precipitate. The white solid was collected by filtration and dried in a vacuum oven to obtain a small molecule containing a hydrophobic microporous structure, namely 5-imino-tert-butylcalix[6]arene isophthalaldehyde.

[0102] The preparation process of 5-imino-tert-butylcalix[5]arene isophthalaldehyde is different from the preparation process of 5-imino-tert-butylcalix[6]arene isophthalaldehyde in that an equal weight of 4-tert-butylcalix[5]arene is used to replace 4-tert-butylcalix[6]arene.

[0103] The preparation process of 5-imino-tert-butylcalix[4]arene isophthalaldehyde is different from the preparation process of 5-imino-tert-butylcalix[6]arene isophthalaldehyde in that 4-tert-butylcalix[4]arene is substituted for 4-tert-butylcalix[6]arene by an equal weight.

[0104] like Figure 1 As shown, the organic small molecule with R functional group grafted in this embodiment is specifically any one of 5-imino-tert-butylcalix[6]arene isophthalaldehyde, 5-imino-tert-butylcalix[5]arene isophthalaldehyde, and 5-imino-tert-butylcalix[4]arene isophthalaldehyde. The R functional group structure of 5-imino-tert-butylcalix[6]arene isophthalaldehyde is: The R functional group structure of 5-imino-tert-butylcalix[5]arene isophthalaldehyde is: The R functional group structure of 5-imino-tert-butylcalix[4]arene isophthalaldehyde is: Benzene rings and hydrophobic cavities, as structures that interact with benzene ring-containing or hydrophobic molecules, can promote the adsorption of hydrophobic molecules by the microspheres, and graft them onto the polymer skeleton of polyvinyl alcohol embolization microspheres (with hydroxyl groups as sites), so that the originally hydrophilic polymer skeleton of the microspheres is transformed into an amphiphilic structure; at the same time, the efficient attraction mechanism of hydrophobic molecules in the water system is increased, thereby improving the loading efficiency of the microspheres for drugs.

[0105] In the test example, the benzene ring-containing cross-linking agent used in Example 6 was tested using 5-imino-tert-butylcalix[6]arene isophthalaldehyde as an example. The effects of 5-imino-tert-butylcalix[5]arene isophthalaldehyde and 5-imino-tert-butylcalix[4]arene isophthalaldehyde can be predicted based on the test results of 5-imino-tert-butylcalix[6]arene isophthalaldehyde.

[0106] Example 7

[0107] This embodiment discloses a highly cross-linked amphiphilic embolic microsphere, and the preparation method thereof is different from that of Example 1 only in that an equimolar amount of any one of N-β-cyclodextrin-5-iminoisophthalaldehyde, N-α-cyclodextrin-5-iminoisophthalaldehyde, and N-γ-cyclodextrin-5-iminoisophthalaldehyde is used to replace the 1,4-phthalaldehyde in Example 1.

[0108] The preparation process of N-β-cyclodextrin-5-iminoisophthalaldehyde is specifically as follows:

[0109] Weigh 25g of β-cyclodextrin and 200mL of deionized water into a 500mL three-necked flask and stir. Weigh 7.5g of p-toluenesulfonyl chloride and slowly add it to the three-necked flask, keep it at room temperature, and stir it for 21h. Prepare 20mL of deionized water solution containing 5g of sodium hydroxide, slowly add it to the above reaction system, and continue stirring for 30min. Filter the above solution, take the clear solution, and add 6g of ammonium chloride to the solution to terminate the reaction. Store it in a refrigerator at 4℃ overnight. Filter the above solution to obtain white crystals. Place the white crystals in a vacuum oven until completely dry. Weigh 2.5g of white crystals, 7.3g of 5-aminoisophthalaldehyde and 150mL of deionized water into a 500mL three-necked flask and stir it at 75℃ for 4h. After the reaction is completed, acetone is added to the reaction solution to obtain a white precipitate. The white solid was collected by suction filtration and dried in a vacuum oven to obtain a small molecule containing a hydrophobic microporous structure, namely N-β-cyclodextrin-5-iminoisophthalaldehyde.

[0110] The difference between the preparation process of N-α-cyclodextrin-5-iminoisophthalaldehyde and the preparation process of N-β-cyclodextrin-5-iminoisophthalaldehyde is that β-cyclodextrin is replaced by an equal weight of α-cyclodextrin.

[0111] The difference between the preparation process of N-γ-cyclodextrin-5-iminoisophthalaldehyde and the preparation process of N-β-cyclodextrin-5-iminoisophthalaldehyde is that β-cyclodextrin is replaced by an equal weight of γ-cyclodextrin.

[0112] like Figure 1 As shown, the organic small molecule with an R functional group grafted in this embodiment is specifically any one of N-β-cyclodextrin-5-iminoisophthalaldehyde, N-α-cyclodextrin-5-iminoisophthalaldehyde, and N-γ-cyclodextrin-5-iminoisophthalaldehyde, and the R functional group structure of N-β-cyclodextrin-5-iminoisophthalaldehyde is: The R functional group structure of N-α-cyclodextrin-5-iminoisophthalaldehyde is: The R functional group structure of N-γ-cyclodextrin-5-iminoisophthalaldehyde is: The benzene ring and the hydrophobic cavity are structures that interact with the benzene ring-containing or hydrophobic molecules. The presence of a hydrophobic cavity can promote the adsorption of hydrophobic molecules by the microspheres, and graft them onto the polymer skeleton of the polyvinyl alcohol embolization microspheres (with hydroxyl groups as sites), thereby transforming the originally hydrophilic polymer skeleton of the microspheres into an amphiphilic structure; at the same time, the efficient attraction mechanism of hydrophobic molecules in the water system is increased, thereby improving the loading efficiency of the microspheres for drugs.

[0113] In the test example, the benzene ring-containing cross-linking agent used in Example 7 was tested using N-β-cyclodextrin-5-iminoisophthalaldehyde as an example. The effects of N-α-cyclodextrin-5-iminoisophthalaldehyde and N-γ-cyclodextrin-5-iminoisophthalaldehyde can be predicted based on the test effect of N-β-cyclodextrin-5-iminoisophthalaldehyde.

[0114] Example 8

[0115] This embodiment discloses a highly cross-linked amphiphilic embolic microsphere. The preparation method thereof is different from that of the embodiment 1 only in that the highly cross-linked amphiphilic embolic microsphere is prepared by reacting together in a polyvinyl alcohol embolic microsphere synthesis system. The specific steps are as follows:

[0116] Step 1: Prepare the reverse suspension polymerization aqueous phase:

[0117] Weigh 50g of 15% polyvinyl alcohol solution into a 250mL flask, set the speed to 200r / min, add 0.125mL of N-(2,2-dimethoxyethyl)-2-acrylamide, and stir for 10-15min. Then, drop 5mL of hydrochloric acid solution into the 250mL flask and stir overnight. Place the 250mL flask in an ice bath and stir, and slowly drop 10% sodium hydroxide solution to adjust its pH to 7.

[0118] Weigh 20g of polyvinyl alcohol grafted with N-(2,2-dimethoxyethyl)-2-acrylamide and place it in a 50mL beaker and stir (speed: 300r / min). Weigh 4g of 2-acrylamide-2-methylpropane sulfonic acid sodium salt and add it to the vial, then add 2*10 -4 mol 1,4-benzenedialdehyde, 2mL deionized water, 400μL hydrochloric acid and 1000μL 10% ammonium persulfate solution and mix thoroughly. Add the prepared mixture into a 50mL beaker and continue stirring for 30 minutes. After stirring, place the beaker in an ultrasonic cleaner and ultrasonicate for 3-5 minutes.

[0119] Step 2: Prepare the reverse suspension polymerization oil phase:

[0120] Weigh 200 mL of butyl acetate and 6 g of cellulose acetate butyrate into a 500 mL three-necked flask, place it in an oil bath, and stir at 500 r / min until fully mixed. Raise the temperature to 56°C and continue stirring for 20 min.

[0121] Step 3: Reverse suspension polymerization:

[0122] The aqueous solution obtained in step 1 was slowly added to the oil phase obtained in step 2 using a 10 mL syringe. The addition time was 10-20 min. After the addition was completed, stirring was continued for 30 min. 800 μL of TEDMA was added to the above solution and stirring was continued for 3 h. The system was heated to 80 ° C, stirred for 1 h, and then the 500 mL three-necked flask was removed from the oil bath and cooled. Finally, highly cross-linked amphiphilic embolic microspheres were obtained.

[0123] Example 9

[0124] This embodiment discloses a highly cross-linked amphiphilic embolic microsphere. The preparation method thereof is different from that of the embodiment 1 in that the molar amount of 1,4-benzenedicarbaldehyde used is different, which is 2*10 -5 mol.

[0125] Example 10

[0126] This embodiment discloses a highly cross-linked amphiphilic embolic microsphere. The preparation method thereof is different from that of the embodiment 1 in that the molar amount of 1,4-benzenedicarbaldehyde used is different, which is 1*10 -3 mol.

[0127] Embodiment 11

[0128] This example discloses a highly cross-linked amphiphilic embolic microsphere, and the preparation method thereof is different from that of Example 1 only in that an equal mass of 5-hexenoic acid is used to replace the 2-acrylamide-2-methylpropane sulfonic acid sodium salt in Example 1.

[0129] Comparative Example 1

[0130] The comparative example is the embolic microsphere product DC Bead (size: 100-300 μm) produced by Biocompatibles UK Limited.

[0131] Comparative Example 2

[0132] Step 1: Prepare the reverse suspension polymerization aqueous phase:

[0133] Weigh 50g of 15% polyvinyl alcohol solution into a 250mL flask, set the speed to 200r / min, add 0.125mL of N-(2,2-dimethoxyethyl)-2-acrylamide, and stir for 10-15min. Then, drop 5mL of hydrochloric acid solution into the 250mL flask and stir overnight. Place the 250mL flask in an ice bath and stir, and slowly drop 10% sodium hydroxide solution to adjust its pH value to 7 to obtain a polyvinyl alcohol derivative.

[0134] Weigh 20g of polyvinyl alcohol derivative and place it in a 50mL beaker and stir (speed: 300r / min). Weigh 4g of 2-acrylamide-2-methylpropane sulfonic acid sodium salt and add it to a vial, then add 2mL of deionized water, 400μL of hydrochloric acid and 1000μL of 10% ammonium persulfate solution in sequence and mix thoroughly. Add the prepared mixture to a 50mL beaker and continue stirring for 30min. After stirring, place the beaker in an ultrasonic cleaner and ultrasonicate for 3-5min to obtain an aqueous phase material.

[0135] Step 2: Prepare the reverse suspension polymerization oil phase:

[0136] Weigh 200 mL of butyl acetate and 6 g of cellulose acetate butyrate into a 500 mL three-necked flask, place it in an oil bath, and stir at 500 r / min until fully mixed. Raise the temperature to 56°C and continue stirring for 20 minutes to obtain an oil phase material.

[0137] Step 3: Reverse suspension polymerization:

[0138] The aqueous phase solution obtained in step 1 was slowly added to the oil phase obtained in step 2 using a 10 mL syringe. The addition time was 10-20 min. After the addition was completed, stirring was continued for 30 min. 800 μL of TEDMA was added to the above solution and stirring was continued for 3 h. The system was heated to 80 ° C, stirred for 1 h, and then the 500 mL three-necked flask was removed from the oil bath and cooled. Polyethanol embolic microspheres were obtained.

[0139] Test Case

[0140] Test Examples The performance tests were conducted on the highly cross-linked amphiphilic embolic microspheres obtained in Examples 1 to 11, and the embolic microspheres obtained in Comparative Example 1 and Comparative Example 2, which are commercially available products. Specifically, the following tests were conducted:

[0141] (1) Drug adsorption test

[0142] Take 0.5 mL of each of the embolic microspheres with a particle size of 50 to 1500 μm prepared in Examples 1 to 11, the embolic microspheres prepared in Comparative Example 2, and the commercially available product of Comparative Example 1, remove excess water, and add 1 mL of a 20 mg / mL doxorubicin hydrochloride aqueous solution. Invert up and down 5 times every 5 to 15 minutes to temporarily disperse the microspheres; take 20 μL of the solution at 5 minutes, 10 minutes, 15 minutes, 20 minutes, 30 minutes, 45 minutes, and 60 minutes, respectively, and dilute it. Use an ultraviolet spectrophotometer to test the concentration of the doxorubicin hydrochloride solution at 483 nm, and calculate the drug adsorption amount and drug loading rate of the embolic microspheres. The drug loading rate data are shown in Table 1.

[0143] Table 1 Adsorption test table of doxorubicin by various embolic microspheres

[0144]

[0145] As can be seen from the table of doxorubicin adsorption rate, compared with the commercially available drug-loaded microspheres in comparative example 1 and the unmodified drug-loaded microspheres in comparative example 2, the drug loading rate of the highly cross-linked amphiphilic embolic microspheres prepared in Examples 1-7 and 11 is significantly improved, and the drug loading rate of the microspheres in Examples 2, 5-7 and 11 can reach more than 96% within 10 minutes. This shows that the highly cross-linked amphiphilic embolic microspheres of the present invention have excellent drug loading performance. The reason is that the benzene ring and the hydrophobic cavity are structures that interact with the benzene ring-containing or hydrophobic molecules, increase the sites for loading positively charged drugs through anion-cation interactions, and graft them onto the polymer skeleton of the polyvinyl alcohol embolic microspheres (with hydroxyl groups as sites), so that the originally hydrophilic polymer skeleton of the microspheres is transformed into an amphiphilic structure; at the same time, the efficient attraction mechanism of hydrophobic molecules in the water system is increased, and the loading efficiency of the microspheres for drugs is improved. In addition, the cross-linked structure formed between the polyvinyl alcohol polymer chains helps to prop the polymer chains apart from each other, forming more pore structures, thereby improving the utilization rate of the sulfonic acid groups in the microspheres, thereby improving the loading efficiency of the drugs. Example 7 Compared with other embodiments, the drug loading rate of the microspheres reached 92.9% within 5 minutes. The reason is that the benzene ring and the hydrophobic cavity are structures that interact with the benzene ring or hydrophobic molecules, and contain a hydrophobic cavity at the same time, which can promote the adsorption of hydrophobic molecules by the microspheres, and graft them on the polymer skeleton of the polyvinyl alcohol embolization microspheres (with hydroxyl groups as sites), so that the effect of converting the originally hydrophilic microsphere polymer skeleton into an amphiphilic structure is more obvious. Compared with Example 1, Example 8 shows that the drug loading efficiency of the microspheres is not much different, indicating that the highly cross-linked amphiphilic embolic microspheres are prepared by reacting together in the polyvinyl alcohol embolic microsphere synthesis system, which has no effect on the performance of the highly cross-linked amphiphilic embolic microspheres. The benzene ring-containing cross-linking agent can be cross-linked with the prepared polyvinyl alcohol embolic microspheres to obtain highly cross-linked amphiphilic embolic microspheres, and can also be reacted together in the polyvinyl alcohol embolic microsphere synthesis system to obtain highly cross-linked amphiphilic embolic microspheres. Example 9 shows that compared with Example 1, although the amount of 1,4-benzenedicaldehyde used is too small, it may lead to insufficient structural modification of the microspheres, and the drug loading efficiency of the microspheres is slowed down compared with Example 1, but the drug loading rate of 99.8% is still achieved in 60 minutes; Example 10 shows that compared with Example 1, although the amount of 1,4-benzenedicaldehyde used is too large, the cross-linked structure of the microspheres is too dense. During the drug loading process, the drug filling in the skeleton of the microspheres caused the pores to be blocked, resulting in a decrease in drug loading efficiency, but the drug loading rate still reached 99.3% in 60 minutes. Compared with Example 1, the drug loading efficiency of the microspheres in Example 11 was basically unchanged, indicating that the cross-linking reaction of monomers containing sulfonic acid or carboxylic acid groups to prepare polyvinyl alcohol embolic microspheres is suitable for the preparation of highly cross-linked amphiphilic embolic microspheres.

[0146] (2) Drug adsorption stability test

[0147] Take 0.5 mL of the embolic microspheres with a particle size of 50 to 1500 μm prepared in Example 1 and the embolic microspheres prepared in Comparative Example 2, remove excess water, and add 1 mL of a 20 mg / mL doxorubicin hydrochloride aqueous solution. Invert up and down 5 times every 5 to 15 minutes to temporarily disperse the microspheres; take 20 μL of solution at 5 minutes, 10 minutes, 15 minutes, 20 minutes, 30 minutes, 45 minutes and 60 minutes, respectively, and dilute it. Use an ultraviolet spectrophotometer to test the concentration of the doxorubicin hydrochloride solution at 483 nm, and calculate the drug adsorption and drug loading rate of the embolic microspheres. Repeat the above test three times, and the drug loading rate data are shown in Table 2.

[0148] Table 2 Adsorption test table of doxorubicin by microspheres of Example 1 and Comparative Example 2

[0149]

[0150]

[0151] As can be seen from the above table, in the three repeated tests of Example 1 and Comparative Example 2, the difference between the drug loading data of Example 1 is smaller than that of Comparative Example 2. The drug adsorption stability of the highly cross-linked amphiphilic embolic microspheres prepared by the method of the present invention is verified, because the presence of the cross-linked structure makes the originally loose polymer microsphere structure more stable, thereby improving its performance stability.

[0152] (3) Hydrophobic drug adsorption test

[0153] Take 1 mL of the embolic microspheres with a particle size of 50 to 1500 μm prepared in Example 6 and Example 7 and the commercially available product of Comparative Example 1, remove excess water, and add a mixed solution of oxaliplatin (0.037 mmol), sorafenib (0.073 mmol) or lenvatinib (0.073 mmol) in water and a trace amount of DMSO. Shake the solution 5 times per minute to fully disperse the microspheres; take 20 μL of the solution for dilution at 5 min, 10 min, 15 min, 20 min, 30 min, 45 min and 60 min, respectively, and use an ultraviolet spectrophotometer to test the concentration of the drug solution, calculate the drug adsorption amount and drug loading rate of the embolic microspheres, and the drug loading rate data are shown in Table 2.

[0154] Table 2 Adsorption test table of hydrophobic drugs by embolic microspheres in Example 6 and Example 7

[0155]

[0156] As can be seen from the above table, from the adsorption test table of hydrophobic drugs, compared with the drug loading rate of commercially available products, the highly cross-linked amphiphilic embolic microspheres prepared in Examples 6 and 7 of the present invention have drug loading properties for sorafenib and lenvatinib, and the drug loading rate for oxaliplatin is also significantly improved. The organic small molecules with R functional groups in Examples 6 and 7, whose benzene rings and hydrophobic cavities serve as structures that interact with benzene rings or hydrophobic molecules, and contain a hydrophobic cavity, can promote the adsorption of hydrophobic molecules by the microspheres, and graft them on the polymer skeleton of the polyvinyl alcohol embolic microspheres, so that the originally hydrophilic polymer skeleton of the microspheres is transformed into an amphiphilic structure; at the same time, the efficient attraction mechanism of hydrophobic molecules in the water system is increased, thereby improving the loading efficiency of the microspheres for drugs.

[0157] (4) Visual development test

[0158] The embolic microspheres with a particle size of 50 to 150 μm prepared in Example 5 were taken and fully swollen and balanced with physiological saline. A certain amount of the developable embolic microspheres were placed in a centrifuge tube and observed under X-ray fluoroscopy using a digital X-ray imaging system (iPET-400, Shenzhen Yihua). The test results are shown in the following photos: Figure 2 As shown: Figure 2 (a) and Figure 2 (b) is a diagram showing the development effect of polyethanol embolic microspheres in Example 5; Figure 2 (c) and Figure 2 (d) is a diagram showing the development effect of the highly cross-linked amphiphilic embolic microspheres in Example 5. It can be seen that the highly cross-linked amphiphilic embolic microspheres obtained by modifying the polymer skeleton of the polyvinyl alcohol embolic microspheres have obvious imaging images under X-ray fluoroscopy, and the effect is remarkable, which can meet the visualization requirements under X-ray.

[0159] (5) Suspension time test of microspheres after drug loading

[0160] The microspheres (drug loading of 37.5 mg / mL) prepared in Examples 1 to 8, Example 11 and Comparative Example 2 were mixed with iohexol saline solution at a ratio of 1:5 to form a uniformly dispersed suspension, and placed in a 5 mL syringe. Expel air, then gently flip the syringe several times to observe the suspension. Start timing after the flip suspension is stable, keep the syringe vertically placed, and the suspension stop judgment standard is that 2 / 3 of the volume of the syringe contains microspheres.

[0161] Table 4 Suspension schedule of microspheres after drug loading in each embodiment and comparative example 2

[0162] sample Suspension time Example 1 32min Example 2 29min Example 3 22min Example 4 25min Example 5 20min Example 6 15min Example 7 18min Example 8 12min Embodiment 11 31min Comparative Example 2 2min

[0163] As can be seen from the above table, the suspension performance of the highly cross-linked amphiphilic embolic microspheres prepared in Examples 1 to 8 and 11 after drug loading is significantly improved, as shown in the figure below. Figure 3 As shown, this is because the cross-linking process uses the hydroxyl groups on the polyvinyl alcohol polymer chain segments as grafting sites, which reduces the number of hydroxyl groups on the surface of the microspheres to a certain extent, thereby improving the adhesion between the microspheres, thereby improving the suspension performance of the drug-loaded microspheres prepared by the present invention.

[0164] In summary, a highly cross-linked amphiphilic embolic microsphere and a preparation method thereof disclosed in an embodiment of the present invention use the hydroxyl group of the polyvinyl alcohol embolic microsphere as a grafting site, and graft a benzene ring-containing cross-linking agent between the polymer main chains of the polyvinyl alcohol embolic microsphere to form a bridging structure containing a benzene ring, thereby obtaining the highly cross-linked amphiphilic embolic microsphere. Benzene rings and hydrophobic cavities are structures that interact with benzene rings or hydrophobic molecules, cross-linked between the polymer skeletons of the polyvinyl alcohol embolic microspheres (using hydroxyl groups as sites), so that the originally hydrophilic microsphere polymer skeleton is transformed into an amphiphilic structure; by increasing the efficient attraction mechanism of hydrophobic molecules in the water system, the loading efficiency of the microspheres for drugs is improved. In addition, the cross-linked structure formed between the polyvinyl alcohol polymer chains helps to open the polymer chains from each other, forming more pore structures, thereby improving the utilization rate of the ionic functional groups in the microspheres, and further improving the loading efficiency of the microspheres for drugs. This makes the clinical application of microspheres more convenient, saving more time for preoperative preparations. Moreover, the presence of the cross-linked structure makes the originally loose polymer microsphere structure more stable, thereby improving its performance stability. The improvement of stability helps to prolong the performance retention time of the microspheres, so that the microspheres have a longer shelf life. The cross-linking process uses the hydroxyl groups on the polyvinyl alcohol polymer chain segments as grafting sites, which reduces the number of hydroxyl groups on the surface of the microspheres to a certain extent, thereby improving the adhesion phenomenon between the microspheres. Less adhesion can effectively improve the catheter passability and clinical embolization effect of the microspheres. It can also be verified through experiments that the high cross-linked amphiphilic embolic microspheres obtained by modifying the polymer skeleton of the polyvinyl alcohol embolic microspheres by the method of the present invention can have functional characteristics such as drug loading and development of various properties. For example, for the loading of hydrophobic targeting or chemotherapeutic drugs that cannot be achieved by the prior art, including but not limited to oxaliplatin, sorafenib or lenvatinib, the drug loading effect can be achieved, so that more patients can benefit in actual clinical applications.

[0165] The technical features of the above-described embodiments may be arbitrarily combined. To make the description concise, not all possible combinations of the technical features in the above-described embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0166] The above-mentioned embodiments only express several implementation methods of the present invention, and the descriptions thereof are relatively specific and detailed, but they cannot be understood as limiting the scope of the invention patent. It should be pointed out that, for ordinary technicians in this field, several variations and improvements can be made without departing from the concept of the present invention, and these all belong to the protection scope of the present invention. Therefore, the protection scope of the patent of the present invention shall be subject to the attached claims.

Claims

1. A method for preparing highly cross-linked amphiphilic embolic microspheres, characterized in that: include: Using the hydroxyl groups of the polyvinyl alcohol embolic microspheres as grafting sites, grafting a benzene ring-containing crosslinking agent between the polymer main chains of the polyvinyl alcohol embolic microspheres to obtain the highly crosslinked amphiphilic embolic microspheres; The polyvinyl alcohol embolic microspheres are prepared by cross-linking a polyvinyl alcohol derivative with a monomer containing a sulfonic acid or carboxylic acid group through a reverse suspension polymerization method. The polyvinyl alcohol derivative is prepared by reacting a water-soluble cross-linking agent with a polyvinyl alcohol high molecular polymer. The dosage ratio of the benzene ring-containing cross-linking agent to the polyvinyl alcohol embolic microspheres is 1*10 -4 mol / g-5*10 -3 mol / g; The water-soluble crosslinking agent includes at least one of N-(2,2-dimethoxyethyl)-2-acrylamide, 8,8-dimethoxy-2,6-dimethyl-2-octene, methylvinyldimethoxysilane, 8,8-dimethoxy-2,6-dimethyl-2-octene, 1,1-dimethoxy-3,7-dimethyl-2,6-octadiene, 1,1-dimethoxyethylene or 4-(1-propenyl)-1,2-dimethoxybenzene; The monomer containing sulfonic acid or carboxylic acid group includes at least one of 2-acrylamide-2-methylpropane sulfonic acid sodium salt, 2-methyl-2-propylene-1-sulfonate sodium salt, 4-styrene sulfonate sodium salt, 3-allyloxy-2-hydroxy-1-propane sulfonic acid sodium salt, acrylic acid, 5-hexenoic acid, 7-octenoic acid or 8-nonenoic acid; The benzene ring-containing crosslinking agent is crosslinked with the polyvinyl alcohol microspheres through any one of the following structures:

2. The method for preparing highly cross-linked amphiphilic embolic microspheres according to claim 1, characterized in that: The benzene ring-containing crosslinking agent includes at least one of 1,4-phthalaldehyde, terephthalaldehyde bis(diethyl acetal), 4,4'-biphenyldicarboxaldehyde, tri(4-formylphenyl)amine, N-(5-methylisophthalaldehyde)iodopyrrole, 5-imino-tert-butylcalix[6]areneisophthalaldehyde, 5-imino-tert-butylcalix[5]areneisophthalaldehyde, 5-imino-tert-butylcalix[4]areneisophthalaldehyde, N-β-cyclodextrin-5-iminoisophthalaldehyde, N-α-cyclodextrin-5-iminoisophthalaldehyde or N-γ-cyclodextrin-5-iminoisophthalaldehyde.

3. The method for preparing highly cross-linked amphiphilic embolic microspheres according to claim 1, characterized in that: The preparation process of the highly cross-linked amphiphilic embolic microspheres includes: cross-linking the benzene ring-containing cross-linking agent with the prepared polyvinyl alcohol embolic microspheres to obtain the highly cross-linked amphiphilic embolic microspheres, or adding the benzene ring-containing cross-linking agent to a reaction system in which a polyvinyl alcohol derivative is used as a matrix and cross-linking the benzene ring-containing cross-linking agent with a monomer containing a sulfonic acid or carboxylic acid group is performed by reverse suspension polymerization to obtain the highly cross-linked amphiphilic embolic microspheres.

4. The method for preparing highly cross-linked amphiphilic embolic microspheres according to claim 3, characterized in that: include: The polyvinyl alcohol embolic microspheres and the benzene ring-containing crosslinking agent are mixed, and an organic solvent and a catalyst are added to carry out a crosslinking reaction at 40° C. to 80° C. to obtain the highly crosslinked amphiphilic embolic microspheres.

5. The method for preparing highly cross-linked amphiphilic embolic microspheres according to claim 4, characterized in that: The generated highly cross-linked amphiphilic embolic microspheres are washed 2-5 times with dimethyl sulfoxide and deionized water respectively to obtain the final usable highly cross-linked amphiphilic embolic microspheres.

6. The method for preparing highly cross-linked amphiphilic embolic microspheres according to claim 3, characterized in that: include: A water-soluble crosslinking agent and a polyvinyl alcohol polymer are mixed and reacted under acidic conditions, and after the reaction is completed, the mixture is placed in an alkaline solution for neutralization to obtain a polyvinyl alcohol derivative; The polyvinyl alcohol derivative, the monomer containing sulfonic acid or carboxylic acid group, the benzene ring-containing cross-linking agent and the catalyst are mixed to prepare an aqueous phase material; Adding an oil-soluble dispersant to an organic solvent to prepare an oil phase material; The aqueous phase material is added to the oil phase material, and a polymerization reaction occurs at 60° C. to 100° C. to obtain the highly cross-linked amphiphilic embolic microspheres.

7. The method for preparing highly cross-linked amphiphilic embolic microspheres according to claim 6, characterized in that: The mass ratio of the polyvinyl alcohol derivative to the monomer containing sulfonic acid or carboxylic acid groups is (3-8):

1.

8. The method for preparing highly cross-linked amphiphilic embolic microspheres according to claim 6, characterized in that: After adding the aqueous phase material to the oil phase material, tetramethylethylenediamine is added to the mixture of the aqueous phase and the oil phase, stirred for 2h-5h for the first time, heated at 60°C-100°C, stirred for 0.5h-3h for the second time, taken out, cooled, and then highly cross-linked amphiphilic embolic microspheres are obtained.

9. A highly cross-linked amphiphilic embolic microsphere, characterized in that: The method is prepared by any one of claims 1 to 8.

10. The highly cross-linked amphiphilic embolic microspheres according to claim 9, characterized in that: In the highly cross-linked amphiphilic embolic microsphere structure, hydroxyl groups are used as grafting sites between the main chains of polyvinyl alcohol polymers, at least a portion of which is cross-linked with polymer chains with ionic functional groups through cross-linking molecules, and at least another portion of which is bridged with hydrophobic benzene ring molecules to form a supporting structure containing benzene rings between the main chains of polyvinyl alcohol polymers, so that the highly cross-linked amphiphilic embolic microspheres have drug-carrying functions; 11. The highly cross-linked amphiphilic embolic microspheres according to claim 10, characterized in that: The particle size of the highly cross-linked amphiphilic embolic microspheres ranges from 50 μm to 1500 μm.

12. The highly cross-linked amphiphilic embolic microspheres according to claim 11, characterized in that: The particle size of the highly cross-linked amphiphilic embolic microspheres ranges from 50 μm to 150 μm.

13. The highly cross-linked amphiphilic embolic microspheres according to claim 10, characterized in that: The highly cross-linked amphiphilic embolic microspheres have any one of the following molecular structures: The main chain wavy line cross-linked with R represents the main chain structure of the polyvinyl alcohol chain molecule, the other main chain wavy line represents the main chain of the molecule containing sulfonic acid or carboxylic acid groups, and R represents the benzene ring-containing cross-linking agent cross-linked with the polyvinyl alcohol microspheres.

14. The highly cross-linked amphiphilic embolic microspheres according to claim 13, characterized in that: R is selected from any one of the following structures:

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