High-molecular-weight poly(p-dioxanone) microsphere, preparation method therefor and use thereof

Abstract

Disclosed in the present invention are a high-molecular-weight poly(p-dioxanone) (PPDO) microsphere, a preparation method therefor, and a use thereof. By using a double emulsion-solvent evaporation method, a double-micelle protective layer is formed using an amphiphilic polymer, such as oligomeric p-dioxanone (PDO) or a copolymer of PDO and a hydrophilic fragment (such as mPEG or mPPG), under the amphiphilicity of the amphiphilic polymer and the co-action of the amphiphilic polymer and polyvinyl alcohol (PVA) by means of self-assembly at a two-phase interface formed by emulsification. The protective layer effectively blocks the degradation effect of water on PPDO during the formation of the microsphere, and a high-molecular-weight PPDO microsphere having stable performance is finally obtained, and thus, the high-molecular-weight PPDO microsphere has long-term stability. The preparation method has high operability and mild requirements, and is suitable for mass production, and the obtained high-molecular-weight PPDO microsphere has wide application prospects in tissue fillers, drug carriers, tissue scaffolds, etc.

Description

High molecular weight polydioxanone microspheres and their preparation method and application Technical Field

[0001] The present invention relates to the technical field of biomedical polymer materials, and in particular to high molecular weight polydioxanone microspheres, a preparation method and application thereof. Background Art

[0002] Polydioxanone (PPDO) is a commonly used biodegradable polymer in medical applications. It belongs to the aliphatic polyester family and holds a significant position in the field of medical biodegradable materials. This type of material exhibits excellent biocompatibility and minimal tissue reaction upon implantation. This is due to the ester bonds within its polymer chain, which impart excellent biodegradability, and the ether bonds, which impart unique flexibility. In recent years, the field of medical aesthetics has rapidly developed, with numerous biomedical polymers being used in facial contouring applications. PPDO, due to its excellent properties, has been widely used in absorbable sutures and suture implants for medical aesthetics. While PPDO suture implants are relatively mature, PPDO microspheres are currently not commercially available in China. Existing microsphere products are primarily based on materials such as poly(L-lactic acid) and polyvinyl alcohol. PPDO microspheres are spherical particles dissolved in a solvent and then dispersed in an emulsifier. Due to the inherent properties of PPDO, PPDO microspheres also exhibit excellent biocompatibility and biodegradability. However, since PPDO is easily degraded when exposed to water, its molecular weight is easily reduced during its preparation process, which affects the performance of the final microspheres. This also limits the development and large-scale production of PPDO microspheres.

[0003] The main methods for preparing microspheres include phase separation, emulsion-solvent drying, freeze-drying, and spray drying. Emulsion-solvent drying is convenient and most suitable for large-scale production. Currently, the predominant methods used to prepare PPDO microspheres are emulsification and spray drying, both of which have been reported in previous studies. However, these methods have limitations that affect the molecular weight and particle size of PPDO microspheres. In particular, the molecular weight of PPDO is easily reduced by high temperatures during the spray drying process, while the rapid spraying process can result in excessively small particle sizes. Furthermore, most reported PPDO microspheres are composite microspheres of polydioxanone and another polymer, or copolymers of polydioxanone (PDO) with other materials, which can also affect the in vivo degradability of PPDO microspheres.

[0004] Wang Tianqiang previously reported in his doctoral dissertation "Synthesis and Application of Polymer Microparticles Based on 4-dioxanone in Supercritical CO2" a method for preparing PPDO microspheres by suspension polymerization in a solution. The method mainly relies on the crystal growth of PPDO in the emulsion. However, because the polymerization temperature has a great influence on the growth rate of the crystals and the operation requirements are extremely harsh, the application range of this method is extremely narrow and it is not suitable for actual production use. Patent document CN106727422A discloses a core-shell double-layer microsphere with poly-4-dioxanone as the core prepared by emulsification and spray drying, as well as its preparation method and application. Because it uses hexafluoroisopropanol, a good solvent for poly-4-dioxanone, in the preparation process, the volatilization of the solution after emulsification needs to be carried out under negative pressure conditions. The system pressure affects the volatilization rate of the solution, which has a greater impact on the microspheres and is not conducive to large-scale preparation. Patent document CN116589710A discloses a poly-4-dioxanone-based polymer microsphere, as well as its preparation method and application. The polydioxanone used has a low molecular weight, which may cause the prepared microspheres to degrade quickly, affecting their subsequent use effects. In addition, the patent document mainly focuses on the preparation of microspheres using copolymers of polydioxanone and other materials, which will also affect the degradability of PPDO microspheres in the body to a certain extent. Patent document CN106492284A discloses a method for preparing a biodegradable filling material and its application. However, during the preparation process of this invention, drying treatment at 110-120°C is required, which exacerbates the degradation of PPDO and will affect the properties of the microspheres. Summary of the Invention

[0005] To address the defects and problems of the above-mentioned prior art, the present application provides a high molecular weight polydioxanone microsphere, a preparation method and application thereof, which adopts the double-emulsion solvent evaporation technology (DESE method). By adding an amphiphilic polymer during the preparation of the microspheres, the amphiphilic polymer is utilized to form a double micelle protective layer at the two-phase interface formed by emulsification under the combined action of PVA, effectively blocking the degradation of PPDO by water during the microsphere formation process, so that the high molecular weight PPDO microspheres have long-term stability, effectively solving the problem of low molecular weight and complex preparation process of polydioxanone microspheres in the prior art.

[0006] To achieve the purpose of the present invention, the technical solution provided by the present invention is as follows:

[0007] In a first aspect of the present invention, high molecular weight polydioxanone microspheres are provided. High molecular weight polydioxanone microspheres are obtained by adding an amphiphilic polymer to a high molecular weight PPDO raw material. The amphiphilic polymer is an oligomeric PDO or a copolymer of PDO and a hydrophilic segment.

[0008] Furthermore, the hydrophilic segment comprises a polyethylene glycol or polypropylene glycol segment, for example, polyethylene glycol monomethyl ether (mPEG) or polypropylene glycol monomethyl ether (mPPG) and other hydrophilic polyether polymers.

[0009] Furthermore, the weight average molecular weight of the high molecular weight PPDO raw material is 20×10 4 -80×10 4 Da(such as 20×10 4 , 30×10 4 , 40×10 4 , 50×10 4 , 60×10 4 , 70×10 4 ,80×10 4 Da), preferably 20×10 4 -40×10 4 Da.

[0010] Furthermore, the weight average molecular weight of the oligomeric PDO is 500-2000 Da (e.g., 500, 600, 700, 800, 900, 1000, 1200, 1400, 1500, 1600, 1800, 2000 Da).

[0011] Furthermore, the weight average molecular weight of the copolymer is 800-5000 Da (e.g., 800, 1000, 1200, 1400, 1600, 1800, 2000, 2200, 2400, 2600, 2800, 3000 Da, 3200, 3400, 3600, 3800, 4000 Da, 4200, 4400, 4600, 4800, 5000 Da), preferably 1000-4000 Da.

[0012] Furthermore, the weight average molecular weight of the hydrophilic segment is 500-2000 Da (e.g., 500, 600, 700, 800, 900, 1000 Da, 1100 Da, 1200 Da, 1300 Da, 1400 Da, 1500 Da, 1600 Da, 1700 Da, 1800 Da, 1900 Da, 2000 Da).

[0013] The second aspect of the present invention provides a method for preparing high molecular weight PPDO microspheres, the method comprising the following steps:

[0014] (1) adding a high molecular weight PPDO raw material and an amphiphilic polymer into solvent A to obtain a mixed solution;

[0015] (2) adding the mixed solution obtained in step (1) to a polyvinyl alcohol (PVA) aqueous solution and emulsifying to obtain an emulsion;

[0016] (3) removing the solvent A from the emulsion obtained in step (2), performing solid-liquid separation, and obtaining the high molecular weight PPDO microspheres.

[0017] Furthermore, the weight average molecular weight of the raw material in step (1) is 20×10 4 -80×10 4 Da(such as 20×10 4 , 30×10 4 , 40×10 4 , 50×10 4 , 60×10 4 , 70×10 4 ,80×10 4 Da), preferably 20×10 4 -40×10 4 Da.

[0018] Furthermore, the amphiphilic polymer is oligomeric PDO or a copolymer of PDO and a hydrophilic segment.

[0019] Furthermore, the hydrophilic segment comprises a polyethylene glycol or polypropylene glycol segment, for example, polyethylene glycol monomethyl ether (mPEG) or polypropylene glycol monomethyl ether (mPPG) and other hydrophilic polyether polymers.

[0020] Furthermore, the weight average molecular weight of the oligomeric PDO is 500-2000 Da (e.g., 500, 600, 700, 800, 900, 1000, 1200, 1400, 1500, 1600, 1800, 2000 Da), preferably 500-1000 Da.

[0021] Furthermore, the weight average molecular weight of the hydrophilic segment is 500-2000 Da (e.g., 500, 600, 700, 800, 900, 1000 Da, 1100 Da, 1200 Da, 1300 Da, 1400 Da, 1500 Da, 1600 Da, 1700 Da, 1800 Da, 1900 Da, 2000 Da).

[0022] Furthermore, the weight average molecular weight of the copolymer of PDO and hydrophilic segments (such as PPDO-b-mPEG or PPDO-b-mPPG) is 800-5000 Da (such as 800, 1000, 1200, 1400, 1600, 1800, 2000, 2200, 2400, 2600, 2800, 3000 Da, 3200, 3400, 3600, 3800, 4000 Da, 4200, 4400, 4600, 4800, 5000 Da), preferably 1000-4000 Da.

[0023] By controlling the molecular weight of oligomeric PDO within an appropriate range, the amphiphilicity of lower molecular weight PDO oligomers allows for the formation of a micellar protective layer at the interface between the two phases. PDO oligomers within this range, due to their low molecular weight and amphiphilic nature, remain at the interface, protecting the high-molecular-weight PPDO microspheres without participating in their formation. When the molecular weight of oligomeric PDO is too high, its hydrophilicity is almost eliminated, and it remains in the oil phase, participating in microsphere formation and thus failing to form a micellar protective layer at the interface. Conversely, when the molecular weight of oligomeric PDO is too low, its hydrophilicity increases, and it remains in the aqueous phase, making it difficult to form a micellar protective layer at the interface between the two phases, thus failing to provide effective protection.

[0024] Furthermore, PDO can also be copolymerized with a hydrophilic segment, such as mPEG or mPPG, to form a PPDO-b-mPEG or PPDO-b-mPPG copolymer. By adjusting the amount of PDO, the chain length (or molecular weight) of the PPDO in the copolymer is controlled to be equivalent to the chain length (or molecular weight) of the hydrophilic segment, thereby acting as a hydrophobic segment. The resulting copolymer achieves a balance between hydrophilicity and hydrophobicity, making it easier to form a micellar protective layer at the interface between the two phases, thereby achieving a better protective effect. If the proportion of PDO in the copolymer is too high, the chain length (or molecular weight) of the hydrophobic segment PPDO is higher than that of the hydrophilic segment, which disrupts the hydrophilic-hydrophobic balance of the copolymer, making the copolymer more hydrophobic. Due to the increased hydrophobicity, it is difficult for the copolymer to stay at the water-oil interface, and it stays more in the oil phase, thus failing to form a protective layer at the water-oil interface. Conversely, if the proportion of PDO is too low, the copolymer becomes more hydrophilic, and due to the increased hydrophilicity, it is difficult for the copolymer to stay at the water-oil interface, and it stays more in the water phase, similarly failing to form a protective layer at the water-oil interface.

[0025] In some preferred embodiments of the present invention, the copolymer is PPDO-b-mPEG.

[0026] Furthermore, the oligomeric PDO and copolymers of PDO and hydrophilic segments can be prepared in-house or purchased. The homemade method is to prepare them by conventional bulk polymerization in the laboratory.

[0027] Furthermore, the oligomeric PDO preparation process includes: polymerizing PDO monomers at 70-90° C. (such as 80° C.) for 8-16 hours (such as 12 hours) to obtain the oligomeric PDO.

[0028] Furthermore, the oligomeric PDO preparation process further includes the step of adding a catalyst (such as stannous octoate).

[0029] Furthermore, the oligomeric PDO preparation process further includes the step of adding an initiator (such as water).

[0030] Furthermore, the preparation process of the copolymer of PDO and hydrophilic segment includes: mixing PDO monomer with a hydrophilic segment (such as mPEG or mPPG), and polymerizing at 70-90° C. (such as 80° C.) for 8-16 hours (such as 12 hours) to obtain the copolymer.

[0031] Furthermore, the process for preparing the copolymer of PDO and hydrophilic segments further includes the step of adding a catalyst (such as stannous octoate).

[0032] Furthermore, the mass ratio of the PDO monomer to the hydrophilic segment is 1:1-2.5 (e.g., 1:2.5, 1:2.2, 1:2.0, 1:1.8, 1:1.6, 1:1.4, 1:1.2, 1:1).

[0033] Furthermore, the amount of the amphiphilic polymer added in step (1) is 0.5-5 wt% (e.g., 0.5 wt%, 1 wt%, 1.5 wt%, 2 wt%, 2.5 wt%, 3 wt%, 4 wt%, 5 wt%) of the high molecular weight PPDO raw material, preferably 1-2 wt%.

[0034] Furthermore, the solvent A in step (1) is one or more of dichloromethane, chloroform or carbon tetrachloride, especially dichloromethane.

[0035] Furthermore, the concentration of the high molecular weight PPDO raw material in step (1) is 1-10 wt% (e.g., 1 wt%, 1.5 wt%, 2 wt%, 2.5 wt%, 3 wt%, 3.5 wt%, 4 wt%, 4.5 wt%, 5 wt%, 5.5 wt%, 6 wt%, 6.5 wt%, 7 wt%, 7.5 wt%, 8 wt%, 8.5 wt%, 9 wt%, 9.5 wt%, 10 wt%).

[0036] Furthermore, step (1) includes: adding a high molecular weight PPDO raw material and an amphiphilic polymer to solvent A, heating and refluxing under the protection of an inert gas to obtain a mixed solution.

[0037] Furthermore, the inert gas is nitrogen or helium.

[0038] Furthermore, step (2) comprises: adding the mixed solution obtained in step (1) to the polyvinyl alcohol aqueous solution at a constant rate, stirring, and emulsifying the mixed solution in the polyvinyl alcohol aqueous solution to obtain an emulsion;

[0039] Furthermore, the rate is 5-50 ml / min (5 ml / min, 10 ml / min, 15 ml / min, 20 ml / min, 25 ml / min, 30 ml / min, 35 ml / min, 40 ml / min, 45 ml / min, 50 ml / min), preferably 30-50 ml / min.

[0040] Furthermore, the alcoholysis degree of the PVA in step (2) is 80-90% (such as 80%, 82%, 84%, 86%, 88%, 90%); preferably 86-88%.

[0041] Furthermore, the viscosity of the PVA in step (2) is 3.0-6.5 mPa·s (such as 3.0 mPa·s, 3.5 mPa·s, 4.0 mPa·s, 4.5 mPa·s, 5.0 mPa·s, 5.5 mPa·s, 6.0 mPa·s), preferably 4.5-5.5 mPa·s.

[0042] Furthermore, the mass volume concentration of the PVA aqueous solution in step (2) is 0.5%-10% (such as 0.5%, 1%, 2%, 3%, 5%, 7%, 9%, 10%, w / v unit is g / ml), preferably 1-3%.

[0043] Furthermore, the emulsification temperature in step (2) is 0-30°C (such as 0°C, 5°C, 10°C, 15°C, 20°C, 25°C, 30°C), preferably 10-25°C.

[0044] Furthermore, the emulsification in step (2) is carried out by mechanically stirring the mixture; further, the mechanical stirring speed is 100-500 rpm (such as 100 rpm, 150 rpm, 200 rpm, 250 rpm, 300 rpm, 350 rpm, 400 rpm, 450 rpm, 500 rpm), preferably 200-300 rpm.

[0045] Furthermore, when a high-molecular-weight polydioxanone dichloromethane solution containing an amphiphilic polymer is added to a polyvinyl alcohol aqueous solution, a double emulsification phenomenon occurs. Due to the emulsification effect, the hydrophobic end segments of the amphiphilic polymer will contact the oil phase, and the hydrophilic end segments will contact the water phase, self-assembling to form a micellar layer near the oil phase end at the interface between the two phases. At the same time, polyvinyl alcohol will also self-assemble to form a micellar layer near the water phase end of the two-phase interface through water-in-oil emulsification, thereby forming a double micellar protective layer at the water-oil interface. The preparation of microspheres by the double emulsification method effectively blocks the degradation of high-molecular-weight PPDO by water during the formation process, thereby improving the stability of the microspheres.

[0046] Furthermore, step (3) comprises: stirring the emulsion obtained in step (2) to volatilize the solvent A, and performing solid-liquid separation to obtain the microspheres.

[0047] Furthermore, the volatilization temperature of the solvent A is 0-70°C (such as 0°C, 5°C, 10°C, 15°C, 20°C, 25°C, 30°C, 35°C, 40°C, 45°C, 50°C, 55°C, 60°C, 65°C, 70°C), preferably 15-30°C.

[0048] Furthermore, the solid-liquid separation method may be a combination of one or more of precipitation, filtration and centrifugation.

[0049] Furthermore, the solid-liquid separation method includes: adding water to the solid-liquid mixture system. Since the microspheres are solid spheres with high density, the microspheres will precipitate after adding water, making it easy to further separate.

[0050] Preferably, in the present invention, the solid-liquid separation is carried out by precipitation and filtration; specifically, the method comprises: adding 1-5 times the volume of purified water to the microsphere system from which solvent A has been removed, precipitating the microspheres, and separating by suction filtration to obtain microspheres. Furthermore, the volume of the purified water added is preferably 3-5 times.

[0051] Furthermore, the obtained microspheres may be washed after separation.

[0052] Furthermore, the method further comprises:

[0053] (4) Screening the microspheres obtained in step (3) (to obtain high molecular weight polydioxanone microspheres with a certain particle size).

[0054] Furthermore, the sieve used for screening is 50-2000 mesh (such as 50, 100, 200, 500, 100, 200 mesh), and the screening can obtain high molecular weight polydioxanone microspheres with a particle size of 5-100 μm; preferably, the obtained microspheres have a particle size of 10-50 μm.

[0055] Furthermore, the method further comprises:

[0056] (5) vacuum drying the microspheres obtained in step (3) or (4) (preferably the microspheres obtained in step (4)) to obtain a high molecular weight polydioxanone microsphere product.

[0057] Furthermore, the vacuum drying temperature is 20-50°C (such as 20°C, 25°C, 30°C, 35°C, 40°C, 45°C, 50°C), preferably 30-40°C.

[0058] The third aspect of the present invention provides the use of the high molecular weight PPDO microspheres described in the first aspect or the high molecular weight PPDO microspheres prepared by the method described in the second aspect.

[0059] Furthermore, the application is the use of the high molecular weight PPDO microspheres in the preparation of tissue fillers (such as mixed gels), drug carriers and tissue scaffolds.

[0060] Specifically, the tissue filler (mixed gel) can be obtained by mixing high molecular weight PPDO microspheres with one or more hydrogels. The resulting mixed gel can be used for medical cosmetic filling (such as nasolabial folds, forehead, temples, chin, hands, etc.) and anti-aging.

[0061] Furthermore, the raw materials of the hydrogel are selected from one or more of the following: hyaluronic acid or its salts (such as sodium hyaluronate), chitosan (such as carboxymethyl chitosan), cellulose (such as sodium carboxymethyl cellulose, hydroxypropyl methylcellulose, hydroxyethyl cellulose), collagen, poly-L-lactic acid, etc.

[0062] Specifically, the drug is loaded with high molecular weight PPDO microspheres as a carrier to achieve the purposes of sustained release, controlled release, targeted drug delivery, etc.

[0063] Specifically, the tissue scaffold can be used as a scaffold material in bone tissue repair and liver tissue repair. The holes and wrinkles on the high molecular weight PPDO microspheres can provide support for cell growth and promote tissue repair.

[0064] The present invention has the following beneficial effects:

[0065] The present invention utilizes a double emulsion-solvent evaporation method to prepare high-molecular-weight polydioxanone microspheres. The amphiphilic polymer (particularly oligomeric PDO or PPDO-b-mPEG copolymer) introduced during the preparation process synergizes with PVA to produce a double emulsion. The hydrophobic end segments of the amphiphilic polymer contact the oil phase, while the hydrophilic end segments contact the water phase. These segments self-assemble near the oil phase at the interface between the two phases to form a micellar layer. Simultaneously, polyvinyl alcohol (PVA) also self-assembles near the water phase through oil-in-water emulsification, forming a micellar layer. This creates a double micellar protective layer at the water-oil interface, effectively preventing water degradation of the high-molecular-weight PPDO during microsphere formation and significantly improving microsphere stability. Compared to conventional freeze-drying and spray-drying methods for preparing polydioxanone microspheres, the present method is highly operable, requires mild preparation conditions, and is suitable for mass production of high-molecular-weight polydioxanone microspheres. It has broad application prospects in tissue fillers, drug carriers, and tissue scaffolds. BRIEF DESCRIPTION OF THE DRAWINGS

[0066] Figure 1 shows a schematic structural diagram of a double micellar protective layer formed by PVA and an amphiphilic polymer (oligomeric PDO or PPDO-b-mPEG copolymer) at the two-phase interface of a high molecular weight PPDO dichloromethane solution droplet: the spherical structure inside the figure is a dichloromethane solution droplet of a high molecular weight PPDO, which is the oil phase; the outermost blank portion is a PVA aqueous solution, which is the water phase; at the two-phase interface, the black and white sticks near the oil phase end are the micellar protective layer formed by the amphiphilic polymer, where the white end is the hydrophobic end and the black end is the hydrophilic end; the gray sticks near the water phase end at the two-phase interface are the micellar protective layer formed by PVA.

[0067] FIG2 is a scanning electron microscope image of microspheres obtained by the double emulsion-solvent evaporation method in Example 4. DETAILED DESCRIPTION

[0068] Unless otherwise defined, all scientific and technical terms used in the present invention have the same meanings as commonly understood by one of ordinary skill in the art to which this invention relates.

[0069] The disclosures of various publications, patents, and published patent specifications cited herein are incorporated by reference in their entirety.

[0070] It should be noted that the experimental methods without specific conditions specified in the examples are generally based on conventional conditions and conditions described in the manual, or conditions recommended by the manufacturer; the materials, reagents, etc. used, unless otherwise specified, can be obtained from commercial channels.

[0071] The following will clearly and completely describe the technical solutions of the present invention in conjunction with the embodiments of the present invention. Obviously, the embodiments described are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making any creative efforts shall fall within the scope of protection of the present invention.

[0072] Synthesis Example 1: Synthesis of PPDO-b-mPEG500 copolymer

[0073] 5 g of PDO monomer and 8.6 g of mPEG500 were added to a polymerization tube and mixed. 1 mg of stannous octoate was added as a catalyst. Polymerization was carried out at 80°C for 12 h to obtain a PPDO-b-mPEG500 copolymer. The copolymer was dissolved in hexafluoroisopropanol and precipitated with ethanol. The precipitate was dried and set aside for use. GPC analysis showed a weight-average molecular weight of 1050 Da.

[0074] Synthesis Example 2: Synthesis of PPDO-b-mPEG1000 copolymer

[0075] 5 g of PDO monomer and 9.5 g of mPEG1000 were added to a polymerization tube, and 1 mg of stannous octoate was added as a catalyst. Polymerization was carried out at 80°C for 12 h to obtain PPDO-b-mPEG1000 copolymer. The copolymer was dissolved in hexafluoroisopropanol and precipitated with ethanol. The precipitate was dried and set aside for use. GPC analysis showed a weight-average molecular weight of 2100 Da.

[0076] Synthesis Example 3: Synthesis of PPDO-b-mPEG2000 copolymer

[0077] 5 g of PDO monomer and 11.7 g of mPEG2000 were added to a polymerization tube, and 1 mg of stannous octoate was added as a catalyst. Polymerization was carried out at 80°C for 12 h to obtain PPDO-b-mPEG2000 copolymer. The copolymer was dissolved in hexafluoroisopropanol and precipitated with ethanol. The precipitate was dried and set aside for use. GPC analysis showed a weight-average molecular weight of 3640 Da.

[0078] Synthesis Example 4: Synthesis of oligomeric p-dioxanone

[0079] 5 g of PDO monomer was added to a polymerization tube, 1 mg of stannous octoate was added as a catalyst, and 0.1 g of water was added as an initiator. The oligomeric PDO was polymerized at 80°C for 12 h. The oligomeric PDO was dissolved in hexafluoroisopropanol and precipitated with ethanol. The precipitate was dried and set aside for use. GPC analysis showed a weight-average molecular weight of 1080 Da.

[0080] Synthesis Example 5: Synthesis of PPDO-b-mPEG1000 copolymer

[0081] 5 g of PDO monomer and 1.6 g of mPEG1000 were added to a polymerization tube, and 1 mg of stannous octoate was added as a catalyst. Polymerization was carried out at 80°C for 12 h to obtain PPDO-b-mPEG1000 copolymer. The copolymer was dissolved in hexafluoroisopropanol and precipitated with ethanol. The precipitate was dried and set aside for use. GPC analysis showed a weight-average molecular weight of 7270 Da.

[0082] Example 1: High molecular weight polydioxanone microspheres A1

[0083] Weigh 2 g of high molecular weight polydioxanone raw material (Mw is 20×10 4 Da) and 20 mg of the amphiphilic polymer PPDO-b-mPEG500 prepared in Synthesis Example 1 were placed in a round-bottom flask. 100 ml of dichloromethane was added to the flask, nitrogen was introduced, and the mixture was heated under reflux under nitrogen protection until completely dissolved to obtain a mixed solution. 4 g of polyvinyl alcohol was weighed and added to 200 ml of purified water. After complete dissolution, the mixture was cooled for later use. The mixed solution was added to the polyvinyl alcohol solution at a rate of 20 ml / min and fully emulsified at 23°C to form droplets of a high molecular weight PPDO dichloromethane solution. The PVA and amphiphilic polymer jointly formed a double micellar protective layer at the interface between the two phases of the high molecular weight PPDO dichloromethane solution droplets. The specific structure is shown in Figure 1. As can be seen from the figure, the amphiphilic polymer PPDO-b-mPEG500 forms a micellar protective layer near the oil phase at the water-oil interface, with the hydrophobic segment (such as PPDO in this example) closer to the oil phase (the white portion of the black and white sticks) and the hydrophilic segment (such as mPEG500 in this example) closer to the water phase (the black portion of the black and white sticks). Polyvinyl alcohol forms a micellar protective layer near the water phase at the oil-water interface, forming a double micellar protective layer together with the amphiphilic polymer. Subsequently, the solvent dichloromethane is completely evaporated at 30°C to obtain high molecular weight PPDO microspheres. An appropriate amount of purified water is then added to precipitate the high molecular weight PPDO microspheres. The supernatant is discarded, the microspheres are filtered, washed, sieved, and dried to obtain high molecular weight PPDO microspheres A1.

[0084] Example 2: High molecular weight polydioxanone microspheres A2

[0085] Weigh 2 g of high molecular weight polydioxanone raw material (Mw is 30×10 4Da) and 20 mg of the PPDO-b-mPEG500 copolymer prepared in Synthesis Example 1 were placed in a round-bottom flask. 100 ml of dichloromethane was added to the flask, nitrogen was introduced, and the mixture was heated under reflux under nitrogen protection until completely dissolved to obtain a mixed solution. 4 g of polyvinyl alcohol was weighed and added to 200 ml of purified water. After complete dissolution, the mixture was cooled for later use. The mixed solution was added to the polyvinyl alcohol solution at a rate of 20 ml / min and fully emulsified at 23°C. The solvent dichloromethane was then completely evaporated at 30°C to obtain high molecular weight PPDO microspheres. An appropriate amount of purified water was then added to precipitate the high molecular weight PPDO microspheres. The supernatant was discarded, and the microspheres were filtered, washed, sieved, and dried to obtain high molecular weight PPDO ketone microspheres A2.

[0086] Example 3: High molecular weight polydioxanone microspheres A3

[0087] Weigh 3 g of high molecular weight polydioxanone raw material (Mw is 60×10 4 Da) and 60 mg of the PPDO-b-mPEG500 copolymer prepared in Synthesis Example 1 were placed in a round-bottom flask. 100 ml of dichloromethane was added to the flask, nitrogen was introduced, and the mixture was heated under reflux under nitrogen protection to completely dissolve it to obtain a mixed solution. 6 g of polyvinyl alcohol was weighed and added to 200 ml of purified water. After complete dissolution, it was cooled for use. The mixed solution was added to the polyvinyl alcohol solution at a rate of 40 ml / min and fully emulsified at 23°C. The solvent dichloromethane was then completely evaporated at 30°C to obtain high molecular weight PPDO microspheres. An appropriate amount of purified water was then added to precipitate the high molecular weight PPDO microspheres. The supernatant was discarded, and the microspheres were filtered, washed, sieved, and dried to obtain high molecular weight PPDO microspheres A3.

[0088] Example 4: High molecular weight polydioxanone microspheres A4

[0089] Weigh 3 g of high molecular weight polydioxanone raw material (Mw is 30×10 4 Da) and 60 mg of the PPDO-b-mPEG1000 copolymer prepared in Synthesis Example 2 were placed in a round-bottom flask. 100 ml of dichloromethane was added to the flask, nitrogen was introduced, and the mixture was heated under reflux under nitrogen protection to completely dissolve it to obtain a mixed solution. 6 g of polyvinyl alcohol was weighed and added to 200 ml of purified water. After complete dissolution, it was cooled for use. The mixed solution was added to the polyvinyl alcohol solution at a rate of 40 ml / min and fully emulsified at 23°C. The solvent dichloromethane was then completely evaporated at 30°C to obtain high molecular weight PPDO microspheres. An appropriate amount of purified water was then added to precipitate the high molecular weight PPDO microspheres. The supernatant was discarded, and the microspheres were filtered, washed, sieved, and dried to obtain high molecular weight PPDO microspheres A4.

[0090] Example 5: High molecular weight polydioxanone microspheres A5

[0091] Weigh 3 g of high molecular weight polydioxanone raw material (Mw is 30×10 4 Da) and 60 mg of oligomeric paradioxanone (Mw of 1080 Da) prepared in Synthesis Example 4 were placed in a round-bottom flask, and 100 ml of dichloromethane was added to the flask, nitrogen was introduced, and the mixture was heated under reflux under nitrogen protection to completely dissolve it to obtain a mixed solution; 6 g of polyvinyl alcohol was weighed, 200 ml of purified water was added, and after complete dissolution, it was cooled for use. The mixed solution was added to the polyvinyl alcohol solution at a rate of 40 ml / min, fully emulsified at 23 ° C, and then the solvent dichloromethane was completely evaporated at 30 ° C to obtain high molecular weight PPDO microspheres; an appropriate amount of purified water was added to precipitate the high molecular weight PPDO microspheres, the supernatant was discarded, and the microspheres were filtered and washed, then sieved and dried to obtain high molecular weight PPDO microspheres A5.

[0092] Example 6: High molecular weight polydioxanone microspheres A6

[0093] Weigh 1.5 g of high molecular weight polydioxanone raw material (Mw is 40×10 4 Da) and 30 mg of the PPDO-b-mPEG2000 copolymer prepared in Synthesis Example 3 were placed in a round-bottom flask. 100 ml of dichloromethane was added to the flask, nitrogen was introduced, and the mixture was heated under reflux under nitrogen protection to completely dissolve it to obtain a mixed solution. 3 g of polyvinyl alcohol was weighed and added to 200 ml of purified water. After complete dissolution, it was cooled for use. The mixed solution was added to the polyvinyl alcohol solution at a rate of 40 ml / min and fully emulsified at 23°C. The solvent dichloromethane was then completely evaporated at 30°C to obtain high molecular weight PPDO microspheres. An appropriate amount of purified water was then added to precipitate the high molecular weight PPDO microspheres. The supernatant was discarded, and the microspheres were filtered, washed, sieved, and dried to obtain high molecular weight PPDO microspheres A6.

[0094] Example 7: High molecular weight polydioxanone microspheres A7

[0095] Weigh 2 g of high molecular weight polydioxanone raw material (Mw is 40×10 4Da) and 40 mg of the PPDO-b-mPEG500 copolymer prepared in Synthesis Example 1 were placed in a round-bottom flask. 100 ml of dichloromethane was added to the flask, nitrogen was introduced, and the mixture was heated under reflux under nitrogen protection to completely dissolve it to obtain a mixed solution. 4 g of polyvinyl alcohol was weighed and added to 200 ml of purified water. After complete dissolution, it was cooled for use. The mixed solution was added to the polyvinyl alcohol solution at a rate of 40 ml / min and fully emulsified at 23°C. The solvent dichloromethane was then completely evaporated at 30°C to obtain high molecular weight PPDO microspheres. An appropriate amount of purified water was then added to precipitate the high molecular weight PPDO microspheres. The supernatant was discarded, and the microspheres were filtered, washed, sieved, and dried to obtain high molecular weight PPDO microspheres A7.

[0096] Example 8: High molecular weight polydioxanone microspheres A8

[0097] Weigh 2 g of high molecular weight polydioxanone raw material (Mw is 80×10 4 Da) and 40 mg of the PPDO-b-mPEG1000 copolymer prepared in Synthesis Example 2 were placed in a round-bottom flask. 100 ml of dichloromethane was added to the flask, nitrogen was introduced, and the mixture was heated under reflux under nitrogen protection to completely dissolve it to obtain a mixed solution. 6 g of polyvinyl alcohol was weighed and added to 200 ml of purified water. After complete dissolution, it was cooled for use. The mixed solution was added to the polyvinyl alcohol solution at a rate of 40 ml / min and fully emulsified at 23°C. The solvent dichloromethane was then completely evaporated at 30°C. An appropriate amount of purified water was added to precipitate high molecular weight PPDO microspheres. The supernatant was discarded, and the microspheres were filtered, washed, sieved, and dried to obtain high molecular weight PPDO microspheres A8.

[0098] Comparative Example 1: High molecular weight polydioxanone microspheres B1

[0099] The preparation process was the same as that of Example 4, except that “60 mg of the PPDO-b-mPEG1000 copolymer prepared in Synthesis Example 2” was not added to obtain high molecular weight polydioxanone microspheres B1.

[0100] Comparative Example 2: High molecular weight polydioxanone microspheres B2

[0101] The preparation process was the same as that in Example 4, except that "60 mg of PPDO-b-mPEG1000 copolymer prepared in Synthesis Example 2" was replaced with "60 mg of mPEG1000" to obtain high molecular weight polydioxanone microspheres B2.

[0102] Comparative Example 3: High molecular weight polydioxanone microspheres B3

[0103] The preparation process was the same as that in Example 4, except that “60 mg of the PPDO-b-mPEG1000 copolymer prepared in Synthesis Example 2” was replaced with “60 mg of the PPDO-b-mPEG1000 copolymer prepared in Synthesis Example 5” to obtain high molecular weight polydioxanone microspheres B3.

[0104] Performance example 1:

[0105] The weight average molecular weight Mw of the microsphere samples prepared in the examples and comparative examples was measured by GPC. The test results are shown in Table 1.

[0106] Table 1 Mw results of microspheres of Examples and Comparative Examples

[0107] As can be seen from Table 1, the molecular weights of the microspheres prepared in Examples 1-8 are close to that of the high molecular weight PPDO raw material, and the loss rates are all within 15%, indicating that the preparation process of the present invention can maintain the high molecular weight of the microspheres and the molecular weight loss rate is low by using amphiphilic polymers for preparing microspheres; while the molecular weight of the microspheres in Comparative Example 1, which uses the same high molecular weight PPDO raw material as in Example 4, is greatly reduced compared with that in Example 4 due to the lack of the introduction of amphiphilic polymers, indicating that it is difficult to prepare high molecular weight PPDO microspheres by the conventional emulsification-solvent evaporation method; Comparative Example 2 introduces mPEG during the emulsification process, although the molecular weight of the microspheres is increased to a certain extent (20×10 4 ), but it is still significantly lower than the molecular weight of the microspheres in Example 4 (30×10 4 ), and the molecular weight loss rate of the microspheres in Comparative Example 2 was 33.3%, far higher than that of the microspheres in Example 4, which showed almost no molecular weight loss. This further demonstrates that the introduction of the amphiphilic polymer can increase the molecular weight of the microspheres. The molecular weight of the microspheres in Comparative Example 3 also decreased significantly, with a loss rate of 23.3%. This is primarily due to the excessive proportion of hydrophobic PPDO in the amphiphilic copolymer. The hydrophilic mPEG at the hydrophilic end is not sufficiently hydrophilic to retain it at the interface between the two phases, instead remaining in the oil phase and failing to form an effective micellar protective layer. This indicates that when the proportion of hydrophobic PPDO in the copolymer is too high during preparation, it is difficult to achieve an ideal protective effect for high-molecular-weight PPDO microspheres.

[0108] Performance example 2:

[0109] The high molecular weight PPDO microspheres obtained in Example 4 were observed using a scanning electron microscope, and the results are shown in Figure 2. As can be seen from Figure 2, the prepared high molecular weight PPDO microspheres have a regular surface shape, a microsphere particle size ranging from 10 to 50 μm, an average particle size of 38 μm, and a uniform distribution, indicating that uniformly distributed high molecular weight PPDO microspheres can be obtained using the process of the present invention.

[0110] Performance example 3:

[0111] One gram of each of the microspheres obtained in Example 4 and Comparative Examples 1-3 (after thorough drying) was subjected to accelerated degradation experiments at 70°C. Five parallel groups were collected from each sample and tested at five different time points (days) 1, 2, 3, 4, and 5. The samples after the accelerated degradation experiments were tested for mass and Mw, using constant weight determination and GPC for Mw. The results are shown in Table 2.

[0112] Table 2 Accelerated degradation test results of microspheres of Example 4 and Comparative Examples 1-3 at 70°C

[0113] As can be clearly seen from Table 2, after the introduction of the PPDO-b-mPEG1000 copolymer in Example 4, the high molecular weight polydioxanone microspheres prepared lost mass and molecular weight more slowly in the accelerated degradation experiment. This is because the introduced copolymer can self-assemble at the interface of the two phases to form a protective layer during the emulsification and spheroidization process, thereby effectively isolating the microspheres from the degradation effect of moisture. Therefore, the molecular weight of the prepared microspheres is high, thereby extending the degradation cycle of the microspheres. In Comparative Examples 1 and 2, although the same PPDO raw material ratio is used, the conventional emulsification-solvent evaporation method is used to prepare polydioxanone microspheres. The molecular weight of the microspheres obtained is much lower than that of Example 4. The degradation quality and degradation rate of the microspheres are significantly higher than those of the microspheres in Example 4, indicating that the microspheres without the introduction of the amphiphilic polymer are easily affected by water and thus degrade quickly. The microspheres in Comparative Example 3 also introduce the amphiphilic polymer under the premise of using the same PPDO raw material ratio, and the molecular weight of the microspheres is improved, and the degradation quality and degradation rate are slowed down to a certain extent. However, since the hydrophobic end PPDO accounts for too high a proportion of the amphiphilic polymer and the mPEG hydrophilic end accounts for too low a proportion, the hydrophobicity is enhanced and the hydrophilicity is weakened, resulting in the amphiphilic polymer being unable to stay at the interface between the two phases, so that the amphiphilic polymer stays more in the oil phase, making it difficult to form an effective protective layer, resulting in the molecular weight of the microspheres being lower than that of the microspheres.

[0114] Example 4, and the degradation quality and degradation rate are also significantly faster than Example 4. The above results further demonstrate that the double micelle protective layer formed by the double emulsification method of PPDO-b-mPEG1000 and PVA introduced in Example 4 can effectively prolong the degradation time of high molecular weight polydioxanone microspheres. Because 3 days of accelerated degradation experiment is approximately equivalent to 100 days of conventional degradation at 37°C, 5 days is equivalent to 150 days of conventional degradation, and 7 days of accelerated degradation is equivalent to 200 days of conventional degradation, the double micelle protective layer formed by double emulsification can show a more obvious advantage in protecting the molecular weight of microspheres during actual clinical application, and the stability effect of the microspheres can be sustained longer.

[0115] Application Example 1:

[0116] Take 10g of the high-molecular-weight polydioxanone microspheres A4 obtained in Example 4 and sterilize with ethylene oxide before use. Dissolve 0.2g of sodium hyaluronate in 10ml of water for injection, add 7μl of 1,4-butanediol diglycidyl ether to crosslink under alkaline conditions, adjust the pH of the gel to 6.5-7.5 with hydrochloric acid solution, and adjust the osmotic pressure to 250-350mOsm / l with PBS buffer to obtain 10ml of a 20% sodium hyaluronate gel. Sterilize with moist heat at 121°C for 15 minutes before use. Add 2.5g of microspheres to 10ml of the sodium hyaluronate gel, mix thoroughly, and aseptically dispense into a 1ml prefilled syringe to obtain a mixed gel containing high-molecular-weight polydioxanone microspheres.

[0117] Application Example 2:

[0118] Take 10g of the high molecular weight polydioxanone microspheres A4 obtained in Example 4 and sterilize with ethylene oxide before use. Dissolve 0.75g of carboxymethyl chitosan in 10ml of PBS buffer (pH 7.0), add 250μl of a 1.5% solution of 1,4-butanediol diglycidyl ether, and crosslink at room temperature for 24h. Adjust the pH of the gel to 6.5-7.5 with hydrochloric acid solution, and adjust the osmotic pressure to 250-350mOsm / l with PBS buffer to obtain 10ml of a 7.5% carboxymethyl chitosan gel. Sterilize with moist heat at 121°C for 15 minutes before use. Add 2.5g of the microspheres to 10ml of the carboxymethyl chitosan gel, mix thoroughly, and aseptically fill into a 1ml prefilled syringe to obtain a mixed gel containing high molecular weight polydioxanone microspheres.

[0119] Application Example 3:

[0120] Take 10g of the high-molecular-weight polydioxanone microspheres B1 obtained in Comparative Example 1 and sterilize with ethylene oxide before use. Dissolve 0.2g of sodium hyaluronate in 10ml of water for injection, add 7μl of 1,4-butanediol diglycidyl ether to crosslink under alkaline conditions, adjust the pH of the gel to 6.5-7.5 with hydrochloric acid solution, and adjust the osmotic pressure to 250-350mOsm / l with PBS buffer to obtain 10ml of a 20% sodium hyaluronate gel. Sterilize with moist heat at 121°C for 15 minutes before use. Add 2.5g of microspheres to 10ml of the sodium hyaluronate gel, mix thoroughly, and aseptically fill into a 1ml prefilled syringe to obtain a mixed gel containing polydioxanone microspheres.

[0121] The mixed gels obtained in Examples 1-3 were subjected to osmotic pressure and rheological tests, and the obtained data are shown in Table 3.

[0122] Table 3 Osmotic pressure and rheological test results of the mixed gel obtained in Application Examples 1-3

[0123] As can be seen from the test data, the elastic modulus G' of the gel prepared using the microspheres of Comparative Example 1 in Application Example 3 is only 321 Pa, which is much lower than the gel formed using the microspheres of Example 4 in Application Examples 1-2 (546 Pa and 587 Pa). Therefore, the gel prepared from the PPDO microspheres of Comparative Example 1 has a low elastic modulus and poor mechanical properties during use, which may result in poor filling effect and inability to provide ideal support, which is not conducive to clinical application. The gel prepared from the PPDO microspheres of Example 4 of the present invention has ideal viscosity and elastic modulus, good mechanical properties, and can provide ideal support in clinical filling applications. It can meet the clinical performance requirements for filling mixed gels and is suitable for filling and plasticity in medical cosmetic applications.

[0124] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, etc. made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

[0125] The aforementioned embodiments and methods described in the present invention may be varied based on the ability, experience, and preference of those skilled in the art.

[0126] In the present invention, merely listing the steps of the method in a certain order does not constitute any limitation on the order of the method steps.

Claims

1. A high molecular weight polydioxanone microsphere, characterized in that: High molecular weight polydioxanone microspheres are obtained by adding an amphiphilic polymer to a high molecular weight PPDO raw material, wherein the amphiphilic polymer is an oligomeric PDO or a copolymer of PDO and a hydrophilic segment.

2. The microsphere according to claim 1, wherein The hydrophilic segment comprises a polyethylene glycol or polypropylene glycol segment; preferably, the hydrophilic segment is selected from polyethylene glycol monomethyl ether (mPEG) or polypropylene glycol monomethyl ether (mPPG).

3. The microsphere according to claim 1 or 2, wherein The weight average molecular weight of the high molecular weight PPDO raw material is 20×10 4 -80×10 4 Da; the weight average molecular weight of the oligomeric PDO is 500-2000Da; the weight average molecular weight of the copolymer is 800-5000Da; the weight average molecular weight of the hydrophilic segment is 500-2000Da.

4. A method for preparing high molecular weight PPDO microspheres according to any one of claims 1 to 3, characterized in that: The following steps are involved: (1) adding a high molecular weight PPDO raw material and an amphiphilic polymer into solvent A to obtain a mixed solution; (2) adding the mixed solution obtained in step (1) to a polyvinyl alcohol aqueous solution and emulsifying to obtain an emulsion; (3) removing the solvent A from the emulsion obtained in step (2), performing solid-liquid separation, and obtaining the microspheres.

5. The method according to claim 4, wherein The amount of the amphiphilic polymer added is 0.5-5 wt% of the high molecular weight PPDO raw material; the concentration of the high molecular weight PPDO raw material is 1-10 wt%.

6. The method according to claim 4, wherein The step (1) comprises: adding a high molecular weight PPDO raw material and an amphiphilic polymer into a solvent A, and heating and refluxing the solvent under the protection of an inert gas to obtain a mixed solution; Preferably, the inert gas is nitrogen or helium; Preferably, the solvent A is one or more of dichloromethane, chloroform or carbon tetrachloride.

7. The method according to claim 4, wherein The step (2) comprises: adding the mixed solution obtained in the step (1) to the polyvinyl alcohol aqueous solution at a constant rate, stirring, and emulsifying the mixed solution in the polyvinyl alcohol aqueous solution to obtain an emulsion; Preferably, the rate is 5-50 ml / min; Preferably, the mass volume concentration of the polyvinyl alcohol aqueous solution is 0.5%-10%; Preferably, the emulsification temperature is 0-30°C; Preferably, the step (3) comprises: stirring the emulsion obtained in the step (2) to volatilize the solvent A, and performing solid-liquid separation to obtain the microspheres.

8. The method according to any one of claims 4 to 7, wherein: The microspheres obtained after solid-liquid separation in step (3) are vacuum dried at a temperature of 20-50° C.; optionally, screening treatment is performed before vacuum drying.

9. The method according to any one of claims 4 to 7, wherein: The oligomeric PDO preparation process comprises: polymerizing PDO monomers at 70-90° C. for 8-16 hours to obtain the oligomeric PDO; Preferably, the preparation process of the copolymer of PDO and hydrophilic segment comprises: mixing PDO monomer and hydrophilic segment, and polymerizing at 70-90° C. for 8-16 hours to obtain the copolymer; Preferably, the mass ratio of the PDO monomer to the hydrophilic segment is 1:1-2.

5.

10. Use of the microspheres according to any one of claims 1 to 3 or the microspheres prepared by the method according to any one of claims 4 to 9 in the preparation of tissue fillers, drug carriers or tissue scaffolds.

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