Method for preparing macromolecular hydrogel microspheres by using low-concentration reaction solution

Abstract

The invention provides a method for preparing high-molecular hydrogel microspheres by using a low-concentration reaction solution. The method comprises the following steps: (1) dissolving components including a high-molecular material and a cross-linking agent into water to form an internal phase solution; dissolving a surfactant in the oil phase to form an external phase solution; preparing a water-in-oil emulsion from the internal phase solution and the external phase solution; (2) removing part of water in the water-in-oil emulsion liquid drops by adopting an in-liquid drying method or a volatilization drying method, and concentrating all components in the internal phase solution of the water-in-oil emulsion liquid drops to obtain concentrated water-in-oil emulsion liquid drops; and (3) applying cross-linking reaction conditions to enable the high polymer material in the liquid drops of the concentrated water-in-oil emulsion to be subjected to cross-linking reaction with the cross-linking agent so as to cure the concentrated water-in-oil emulsion. According to the invention, the preparation of the high-molecular hydrogel microspheres can be realized under the condition of relatively low concentrations of the high-molecular material and the cross-linking agent, the modification degree of the high-molecular material is reduced, the biocompatibility of the hydrogel microspheres is improved, and the operation difficulty of the preparation of the hydrogel microspheres is reduced.

Description

Technical Field

[0001] The invention belongs to the field of polymer hydrogel microsphere preparation, and relates to a method for preparing polymer hydrogel microspheres by using a low-concentration reaction solution. Background Art

[0002] Hydrogel microparticles prepared by cross-linking and then cutting have the problem of an uneven surface. After injection into the body, the extremely irregular shape of the hydrogel microparticles can easily irritate tissues, causing redness and swelling. If the surface of the microspheres were smooth, the irritation to tissues after injection would be much less. Currently, there are many methods for preparing microspheres, such as homogenous emulsification, membrane emulsification, spray drying, and droplet microfluidics.

[0003] Hydrogel microspheres used in vivo for various applications, such as embolization and filling, must meet certain requirements for durability, support, and hardness. Currently, both physical and chemical methods can achieve this goal. While polymer chains can be physically entangled, a certain amount of stabilizer or crosslinker can be added to stabilize the polymer chains. Although most polymer materials are inherently biocompatible, such as gelatin, chitosan, and hyaluronic acid, the crosslinkers used in the preparation of hydrogel microspheres are often toxic and can cause biocompatibility. For example, commonly used crosslinkers such as 1,4-butanediol diglycidyl ether, 1-bromo-3,4-epoxybutane, 1-chloro-2,3-epoxypropane, 1-bromo-2,3-epoxypropane, 2,5-dibromopentanol, 2,4-dibromobutanethiol, 2,5-dibromopentane-thiol epichlorohydrin, dimethylaminopropylcarbodiimide, and divinyl sulfone all exhibit these issues to some extent. However, in order to achieve a more ideal cross-linking effect, such as to achieve the required elastic modulus or hardness, it is often necessary to add a large amount of cross-linking agent. However, a high concentration of cross-linking agent will inevitably cause a high degree of modification (MoD) of the original polymer material, resulting in the inability of the organism to recognize it, triggering a foreign body reaction, and causing adverse reactions such as granulomas or delayed swelling.

[0004] At the same time, high concentrations of polymer materials are also helpful for the reaction, but the higher the content of polymer materials in the reaction solution, the higher the viscosity of the reaction solution, which will cause the difficulty of preparing hydrogel microspheres to increase sharply. For example, for sodium hyaluronate with a molecular weight of more than 1 million Daltons, the highest concentration that can be prepared is a solution in the range of 10wt% to 20wt%. If the concentration continues to increase, the solution will become gel-like and cannot be stirred evenly, resulting in uneven dispersion of ingredients such as cross-linking agents in the reaction solution, which in turn leads to differences in the chemical composition of gel microspheres prepared in the same batch. Therefore, in order to avoid the problem of high modification degree of the original polymer material due to excessive cross-linking agent concentration in the reaction solution, or to avoid the problem of increased difficulty in operation due to excessive concentration of polymer materials in the reaction solution, it is currently urgent to develop a cross-linking method for gel microspheres to prepare gel microspheres under the conditions of low concentration of polymer materials and low concentration of cross-linking agents and produce a cross-linking effect comparable to or even better than that of the prior art. Summary of the Invention

[0005] In response to the problems existing in the preparation methods of hydrogel microspheres, such as excessively high modification degree of the original polymer material due to the high concentration of the cross-linking agent in the reaction solution, or increased difficulty in operation due to the high concentration of the polymer material in the reaction solution, the present invention provides a method for preparing polymer hydrogel microspheres using a low-concentration reaction solution, so as to achieve the preparation of polymer hydrogel microspheres under conditions of lower polymer material and cross-linking agent concentrations, reduce the modification degree of the original polymer material, increase the biocompatibility of the hydrogel microspheres, and reduce the operational difficulty of preparing the hydrogel microspheres.

[0006] To achieve the above-mentioned purpose, the present invention provides the following technical solutions:

[0007] A method for preparing polymer hydrogel microspheres using a low-concentration reaction solution comprises the following steps:

[0008] (1) dissolving components including a polymer material and a cross-linking agent in water to form an internal phase solution; dissolving an oil-soluble surfactant in the oil phase to form an external phase solution; and preparing a water-in-oil emulsion using the internal phase solution and the external phase solution;

[0009] The concentrations of the polymer material and the crosslinking agent in the internal phase solution should be such that, when the crosslinking reaction conditions are applied, the polymer material and the crosslinking agent in the water-in-oil emulsion droplets cannot undergo a crosslinking reaction;

[0010] (2) removing part of the water in the water-in-oil emulsion droplets by a liquid drying method or a volatile drying method, thereby concentrating the components in the internal phase solution of the water-in-oil emulsion droplets to obtain concentrated water-in-oil emulsion droplets; the degree of concentration of the components in the internal phase solution of the water-in-oil emulsion droplets should ensure that when the crosslinking reaction conditions are applied, the polymer material and the crosslinking agent in the concentrated water-in-oil emulsion droplets can undergo a crosslinking reaction;

[0011] (3) applying cross-linking reaction conditions to allow the polymer material in the concentrated oil-in-water emulsion droplets to undergo a cross-linking reaction with the cross-linking agent to solidify the concentrated oil-in-water emulsion, and washing to remove unreacted components to obtain polymer hydrogel microspheres.

[0012] In the above technical solution, the operation of removing part of the water in the oil-in-water emulsion droplets by the liquid drying method is as follows: dissolving an oil-soluble surfactant in a concentrator to obtain a collection solution, wherein the concentrator is an organic solvent that is miscible with water but does not dissolve the polymer material in the internal phase solution; placing the oil-in-water emulsion droplets prepared in step (1) in the collection solution, and the volume of the collection solution is at least 10 times the volume of the internal phase solution used in step (1) when preparing the oil-in-water emulsion. The water in the internal phase solution in the oil-in-water emulsion droplets will diffuse into the collection solution, so that the components in the internal phase solution of the oil-in-water emulsion droplets are concentrated to obtain concentrated oil-in-water emulsion droplets.

[0013] Furthermore, in the above technical solution, when the sub-liquid drying method is used to remove part of the water in the water-in-oil emulsion droplets, the water-in-oil emulsion droplets prepared in step (1) are placed in a collecting solution until the concentration of water in the water-in-oil emulsion droplets and the collecting solution reaches equilibrium, thereby obtaining concentrated water-in-oil emulsion droplets.

[0014] Furthermore, in the above technical solution, when the sub-liquid drying method is used to remove a portion of the water in the water-in-oil emulsion droplets, the concentrating agent is one or more of ethyl acetate, isobutanol, ethanol, acetone, and methyl isobutyl ketone. In practical applications, the type of concentrating agent is mainly selected based on the specific polymer material used.

[0015] In the above technical solution, when the liquid drying method is used to remove part of the water in the water-in-oil emulsion droplets, the volume of the collection solution can be 10 to 100 times the volume of the internal phase solution used in step (1) when preparing the water-in-oil emulsion. By adjusting the ratio of the volume of the collection solution to the volume of the internal phase solution used in step (1) when preparing the water-in-oil emulsion, the concentration degree of each component in the water-in-oil emulsion droplets can be adjusted.

[0016] In the above technical solution, the operation of removing part of the water in the oil-in-water emulsion droplets by the volatile drying method is as follows: the oil-in-water emulsion prepared in step (1) is placed in an open container, the oil-in-water emulsion droplets will naturally sink, and the liquid level of the oil phase solution in the oil-in-water emulsion is controlled to be 1 to 3 mm higher than the upper surface of the uppermost oil-in-water emulsion droplets. After standing at room temperature, the water in the oil-in-water emulsion droplets will evaporate into the air, so that the components in the internal phase solution of the oil-in-water emulsion droplets are concentrated to obtain concentrated oil-in-water emulsion droplets.

[0017] In the above technical solution, when the water in the water-in-oil emulsion droplets is partially removed by evaporative drying, the time of standing at room temperature is controlled so that the diameter of the resulting concentrated water-in-oil emulsion droplets is (0.3 to 0.8) times the diameter of the water-in-oil emulsion droplets prepared in step (1). Typically, standing at room temperature for 24 to 72 hours can bring the diameter of the concentrated water-in-oil emulsion droplets to the aforementioned range.

[0018] In the above technical solution, when a portion of the water in the water-in-oil emulsion droplets is removed by evaporative drying, the rate at which the water in the water-in-oil emulsion droplets evaporate into the air can be increased by promoting air flow in the environment of the open container while the emulsion is still at room temperature. For example, a fan can be used to blow air into the open container to increase the rate at which the water in the water-in-oil emulsion droplets evaporate into the air.

[0019] In the above technical solution, the polymer material includes at least one of hyaluronic acid, sodium hyaluronate, agarose, gelatin, chitosan, polyvinyl alcohol, and sodium alginate.

[0020] In the above technical solution, since the components in the internal phase solution of the oil-in-water emulsion droplets are concentrated in step (2), step (3) is crosslinked based on the concentrated oil-in-water emulsion droplets, while the prior art directly crosslinks based on a water-in-oil emulsion containing a high concentration of crosslinking agent, or a high concentration of crosslinking agent and polymer material. Therefore, in step (1), the concentration of the crosslinking agent and polymer material in the prepared internal phase solution is relatively low, and the concentration of the crosslinking agent and polymer material in the internal phase solution is lower than the concentration of the crosslinking agent and polymer material in the oil-in-water emulsion droplets used for crosslinking in the prior art. Generally speaking, the concentration of the crosslinking agent in the internal phase solution is 0.001wt% to 10wt%, and the concentration of the polymer material in the internal phase solution should be such that the viscosity of the internal phase solution does not exceed 100mPa·s. In practical applications, the concentration of the polymer material in the internal phase solution is determined according to the type and molecular weight of the polymer material, with the viscosity of the internal phase solution not exceeding 100 mPa·s (for example, between 0.1 and 100 mPa·s) as a principle. Generally speaking, for the different polymer materials listed above, the concentration of the polymer material in the internal phase solution can be selected in the range of 0.5 wt% to 15 wt%.

[0021] In step (1) of the above technical solution, the cross-linking agent is a reagent that can undergo a cross-linking reaction with a polymer material. In practical applications, the type of cross-linking agent can be selected mainly according to the specific polymer material used. Generally, the cross-linking agent includes any one of 1,4-butanediol glycidyl ether, divinyl sulfone, 1-bromo-3,4-epoxybutane, 1-chloro-2,3-epoxypropane, 1-bromo-2,3-epoxypropane, 2,5-dibromopentanol, 2,4-dibromobutyl mercaptan, 2,5-dibromopentane-thiol epichlorohydrin, dimethylaminopropylcarbodiimide, formaldehyde and glutaraldehyde.

[0022] In step (1) of the above technical solution, the internal phase solution also contains at least one of an acid-base regulator, a catalyst, and an initiator. Whether to use an acid-base regulator, a catalyst, and an initiator, and what type of acid-base regulator, catalyst, and initiator to use, mainly depends on the selected polymer material and cross-linking agent. In practical applications, the selection can be made based on the specific polymer material and cross-linking agent selected. For example, when the polymer material is hyaluronic acid and the cross-linking agent is 1,4-butanediol glycidyl ether, the internal phase solution should also contain an acid-base regulator (such as sodium hydroxide or hydrochloric acid). Specifically, when it is necessary to prepare cross-linked hyaluronic acid gel microspheres, a feasible method for preparing the internal phase solution is: dissolving the polymer material hyaluronic acid, the cross-linking agent 1,4-butanediol glycidyl ether, and the acid-base regulator sodium hydroxide in water to obtain an internal phase solution; in the internal phase solution, the concentration of the polymer material is 0.5wt% to 5wt%, the concentration of the cross-linking agent does not exceed 1wt%, and the concentration of the acid-base regulator is 0.1wt% to 3wt%.

[0023] In the above technical solution, the role of the oil-soluble surfactant is to maintain the stability of the water-in-oil emulsion. Possible oil-soluble surfactants include any one of Span 20, Span 60, Span 80, glyceryl monostearate, polyglyceryl ricinoleate, and cetyl polyethylene glycol / polypropylene glycol-10 / 1 dimethicone.

[0024] In step (1) of the above technical solution, the concentration of the oil-soluble surfactant in the external phase solution is preferably 0.1 wt% to 5 wt%.

[0025] In step (1) of the above technical solution, the oil phase used to prepare the external phase solution is immiscible with water.

[0026] In the above technical solution, the concentration of the oil-soluble surfactant in the collected solution is preferably 0.1 wt% to 5 wt%.

[0027] In step (1) of the above technical solution, the internal phase solution and the external phase solution are prepared into a water-in-oil emulsion by a method in the prior art. For example, common methods include mechanical stirring, ultrasonic emulsification, high-pressure emulsification, membrane emulsification, microfluidics, etc.

[0028] The technical principles of the present invention are mainly as follows:

[0029] In order to solve the problem that the original polymer material has a high degree of modification due to the high concentration of the cross-linking agent in the reaction solution in the existing preparation method of hydrogel microspheres, or to solve the problem that the operation difficulty is increased due to the high concentration of the polymer material in the reaction solution, the present invention first prepares the components including the polymer material and the cross-linking agent into an internal phase solution with low polymer material concentration and low cross-linking agent concentration using water as a solvent, uses an oil phase containing an oil-soluble surfactant as an external phase solution, and prepares an oil-in-oil emulsion with the internal phase solution and the external phase solution; then, removes part of the water in the oil-in-oil emulsion droplets to concentrate the components in the internal phase solution of the oil-in-oil emulsion droplets, so that the concentration of the components (such as the polymer material and the cross-linking agent) in the oil-in-oil emulsion droplets is significantly improved, and concentrated oil-in-oil emulsion droplets are obtained; finally, cross-linking reaction conditions are applied to cause the polymer material in the concentrated oil-in-oil emulsion droplets to undergo a cross-linking reaction with the cross-linking agent to solidify the concentrated oil-in-oil emulsion, thereby obtaining polymer hydrogel microspheres.

[0030] Since the concentrations of polymer materials and cross-linking agents in the concentrated water-in-oil emulsion droplets have been effectively increased, the polymer materials that could not undergo cross-linking reactions under low concentration conditions can be successfully cross-linked with the cross-linking agents, thereby preparing cross-linked polymer hydrogel microspheres under conditions of low cross-linking agent dosage, or low cross-linking agent and low polymer material concentrations. This can avoid the problem of excessive cross-linking agent modification of the cross-linked polymer hydrogel microspheres due to excessive cross-linking agent dosage, thereby reducing the biocompatibility of the cross-linked polymer hydrogel microspheres. It can also avoid the problem of excessive viscosity of the reaction system due to excessive concentration of polymer materials in the reaction solution, resulting in increased difficulty in operation and a large difference in the chemical composition of the cross-linked products of the same batch.

[0031] Compared with the prior art, the technical solution of the present invention produces the following beneficial technical effects:

[0032] 1. The present invention provides a method for preparing polymer hydrogel microspheres using a low-concentration reaction solution. The method first dissolves components including a polymer material and a crosslinking agent in water to form an internal phase solution, dissolves an oil-soluble surfactant in an oil phase to form an external phase solution, and uses the internal phase solution and the external phase solution to prepare a water-in-oil emulsion; then, a liquid-in-drying method or a volatile drying method is used to remove part of the water in the water-in-oil emulsion droplets, thereby concentrating the components in the internal phase solution of the water-in-oil emulsion droplets to obtain concentrated water-in-oil emulsion droplets; finally, crosslinking reaction conditions are applied to cause the polymer material in the concentrated water-in-oil emulsion droplets to undergo a crosslinking reaction with the crosslinking agent to solidify the concentrated water-in-oil emulsion, and the unreacted components are removed by washing to obtain the polymer hydrogel microspheres. After removing part of the water in the oil-in-water emulsion droplets to form concentrated oil-in-water emulsion droplets, the concentrations of the polymer material and the cross-linking agent in the shrunken oil-in-water emulsion droplets can be significantly increased, so that the polymer material that cannot undergo cross-linking reaction under low concentration conditions can be smoothly cross-linked with the cross-linking agent, thereby preparing cross-linked polymer hydrogel microspheres under conditions of low cross-linking agent dosage, or low cross-linking agent and low polymer material concentration. This can solve the problem in the prior art that the degree of cross-linking agent modification of the cross-linked polymer hydrogel microspheres is too high due to excessive use of the cross-linking agent, thereby causing reduced biocompatibility of the cross-linked polymer hydrogel microspheres. It can also solve the problem in the prior art that the viscosity of the reaction system is too high due to the excessive concentration of the polymer material in the reaction solution, resulting in increased difficulty in operation and a large difference in the chemical composition of the cross-linked products of the same batch.

[0033] 2. The method of the present invention is simple to operate for concentrating the water-in-oil emulsion droplets. Concentrated water-in-oil emulsion droplets can be prepared by placing the water-in-oil emulsion droplets in a collection solution containing a concentrator and holding them for a period of time to allow the water in the water-in-oil emulsion droplets to diffuse into the collection solution, or by allowing the water-in-oil emulsion to stand in an open container for a period of time to allow the water in the water-in-oil emulsion droplets to evaporate into the air. This process does not require complex or special instruments, nor does it require cumbersome operating procedures. In addition, the operations of the water-in-oil emulsion preparation process and the cross-linking process of the concentrated water-in-oil emulsion droplets of the method of the present invention are also very simple. Therefore, the method of the present invention is easy to operate and is conducive to its promotion and application in practical applications. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 This is a schematic diagram of using low concentration reaction solution to prepare polymer hydrogel microspheres. In the figure, low C 高分子 Refers to low concentration polymer materials, low C 交联剂 Refers to low concentration crosslinker, high C 高分子 Refers to high-concentration polymer materials, high C 交联剂 refers to a high concentration cross-linker, and PBS refers to PBS buffer.

[0035] Figure 2 These are optical micrographs of the W / O emulsion droplets and concentrated W / O emulsion droplets prepared in step (2) of Example 1.

[0036] Figure 3 This is an optical microscopic image of the cross-linked HA hydrogel microspheres prepared in step (3) of Example 1 in PBS buffer.

[0037] Figure 4 This is the nuclear magnetic resonance hydrogen spectrum of the cross-linked HA hydrogel microspheres prepared in Example 1 after enzymatic hydrolysis.

[0038] Figure 5 These are optical microscopic images of the W / O emulsion droplets and concentrated W / O emulsion droplets prepared in Example 2, as well as optical microscopic images of cross-linked HA hydrogel microspheres in PBS buffer.

[0039] Figure 6 These are optical microscopic images of the W / O emulsion droplets and concentrated W / O emulsion droplets prepared in Example 3, as well as optical microscopic images of cross-linked HA hydrogel microspheres in PBS buffer.

[0040] Figure 7 These are optical micrographs of the W / O emulsion droplets prepared in Comparative Example 1 and an optical micrograph of the cross-linked product in PBS buffer.

[0041] Figure 8 These are optical micrographs of the W / O emulsion droplets prepared in Comparative Example 2 and an optical micrograph of the cross-linked product in PBS buffer.

[0042] Figure 9 This is the hydrogen nuclear magnetic resonance spectrum of the cross-linked product prepared in Comparative Example 2 after enzymatic hydrolysis.

[0043] Figures 2-3 、 Figures 5 to 8 The size data marked on the W / O emulsion droplets, concentrated W / O emulsion droplets and cross-linked HA hydrogel microspheres are unclear. In the present invention, these size data do not serve as a reference and can be ignored. Figures 2-3 、 Figures 5 to 8 The size information of the medium W / O emulsion droplets, concentrated W / O emulsion droplets, and cross-linked HA hydrogel microspheres can be estimated by referring to the ruler in the lower right corner of the figure. DETAILED DESCRIPTION

[0044] The following examples further illustrate the method for preparing polymer hydrogel microspheres using a low-concentration reaction solution provided by the present invention. It should be noted that the following examples are intended only to further illustrate the present invention and are not to be construed as limiting the scope of the present invention. Based on the above disclosure, non-essential improvements and adjustments made by persons skilled in the art to implement the present invention remain within the scope of the present invention.

[0045] In the following examples and comparative examples, the microfluidic device used is a primary microfluidic device, including an injection tube, a collecting tube and a lead-out hose. The outlet end of the injection tube is inserted into the inlet end of the collecting tube, and the outlet end of the collecting tube is connected to the lead-out hose. The injection tube and the collecting tube are coaxially arranged and fixed on a glass slide by resin glue. The injection tube is a glass tube or steel tube with a circular cross-section, and the inner diameter of the injection tube is 10 to 1000 μm and the outer diameter is 50 to 2000 μm. The collecting tube is a glass tube or steel tube with a square or circular cross-section, and the inner diameter of the collecting tube is 50 to 2000 μm and the outer diameter is 100 to 3000 μm. The cross-section of the lead-out hose is circular, and the material can be polytetrafluoroethylene (PTFE), polyethylene (PE) or silicone, etc. The inner diameter of the lead-out hose is 100 to 3000 μm and the outer diameter is 200 to 5000 μm. A flat-ended needle can be attached to the inlet of the injection tube using resin glue, and a flat-ended needle can be fixedly attached to the inlet of the collection tube using resin glue to facilitate connection to the injection pump. Each flat-ended needle can be connected to the injection pump via a tubing. The structure of the primary microfluidic device can be found in Hao Tan and Sooyoung Park, Poly(acrylic acid) hydrogel microspheres for a metal-ion sensor, ACS Sensors, 2021, 6, 3, 1039–1048.

[0046] Example 1

[0047] In this embodiment, a method for preparing polymer hydrogel microspheres using a low concentration reaction solution is described in detail, specifically taking the preparation of cross-linked hyaluronic acid (HA) hydrogel microspheres as an example. The schematic diagram of the preparation process is shown in FIG. Figure 1 As shown, the steps are as follows:

[0048] (1) Prepare internal phase solution, external phase solution and collection solution

[0049] Preparation of an internal phase solution: HA with a molecular weight of 1 to 2 MDa, a cross-linking agent 1,4-butanediol glycidyl ether (BDDE), and sodium hydroxide were added to high-purity water and stirred until the components were completely dissolved to obtain an internal phase solution; the concentration of HA in the internal phase solution was 4 wt%, the concentration of BDDE was 0.2 wt%, the concentration of NaOH was 1 wt%, and the viscosity of the internal phase solution was 56 mPa·s.

[0050] The external phase solution was prepared by dissolving the surfactant cetyl polyethylene glycol / polypropylene glycol-10 / 1 dimethylsiloxane (Abil EM90) in liquid paraffin (PO) to obtain an external phase solution; the concentration of Abil EM90 in the external phase solution was 3 wt %.

[0051] Preparation of a collection solution: dissolving the surfactant Abil EM90 in isobutyl alcohol (IBA) to obtain a collection solution; the concentration of Abil EM90 in the collection solution is 3 wt %.

[0052] (2) Preparation of concentrated W / O emulsion droplets

[0053] The internal phase solution was poured into the external phase solution and stirred at 600 rpm for 10 minutes using a magnetic stirrer to obtain a water-in-oil (W / O) emulsion. The W / O emulsion was then added dropwise to a reaction vessel containing a collection solution and stirred at 600 rpm for 30 minutes. While the W / O emulsion droplets remained in the collection solution, water from the W / O emulsion droplets continuously diffused into the collection solution, as water and IBA are miscible while HA is insoluble in IBA. This concentrated the components in the W / O emulsion droplets until the water concentration in the W / O emulsion droplets reached equilibrium with that in the collection solution, resulting in concentrated W / O emulsion droplets.

[0054] In this step, the volume ratio of the inner phase solution, the outer phase solution and the collection solution was controlled to be 1:50:200.

[0055] (3) Preparation of cross-linked HA hydrogel microspheres

[0056] The concentrated W / O emulsion droplets obtained in step (2) were allowed to settle naturally in the reaction container, and most of the collected solution in the reaction container was carefully poured out. The reaction container was placed in an oven and cross-linked at 25° C. for 24 h. After the cross-linking was completed, the cross-linked product was poured into IBA and washed three times, and then poured into PBS buffer and washed three times to obtain cross-linked HA hydrogel microspheres.

[0057] The optical micrographs of the W / O emulsion droplets and concentrated W / O emulsion droplets prepared in step (2) of this example are as follows: Figure 2 As shown in Figures (A) and (B), the optical microscopic images of the cross-linked HA hydrogel microspheres prepared in step (3) of this embodiment in PBS buffer are as follows: Figure 3 As shown. Figure 2 It can be seen that the size of concentrated W / O emulsion droplets is significantly reduced compared to that of W / O emulsion droplets. This is mainly caused by the diffusion of water in the W / O emulsion droplets into the collection solution during the concentration process. This process will significantly increase the concentration of HA, BDDE and sodium hydroxide in the concentrated W / O emulsion droplets, which is beneficial to the subsequent cross-linking reaction. Figure 3 It can be seen that after the cross-linking reaction, the obtained cross-linked HA hydrogel microspheres are stable in PBS buffer, indicating that the hydrogel microspheres are successfully cross-linked.

[0058] The cross-linked HA hydrogel microspheres prepared in this example were enzymatically hydrolyzed according to the literature Frida J. Wende, Suresh Gohil, Lars I. Nord, Anne Helander Kenne, Corine Carbohydrate Polymers 2017, 157, 1525–1530, and tested the nuclear magnetic resonance hydrogen spectrum (H-NMR). The results are as follows Figure 4 The modification degree (MoD) of BDDE in the cross-linked HA hydrogel microspheres was calculated to be 2.0% based on H NMR spectra.

[0059] Example 2

[0060] In this example, a method for preparing polymer hydrogel microspheres using a low-concentration reaction solution is described in detail. Specifically, the preparation of cross-linked HA hydrogel microspheres is described as an example. The steps are as follows:

[0061] (1) Prepare internal phase solution, external phase solution and collection solution

[0062] Preparation of an internal phase solution: HA with a molecular weight of 1-2 MDa, a crosslinker BDDE, and sodium hydroxide were added to high-purity water and stirred until the components were completely dissolved to obtain an internal phase solution; the concentration of HA in the internal phase solution was 4 wt%, the concentration of BDDE was 0.2 wt%, and the concentration of NaOH was 1 wt%, and the viscosity of the internal phase solution was 56 mPa·s.

[0063] The external phase solution was prepared by dissolving the surfactant Abil EM90 in PO to obtain an external phase solution; the concentration of Abil EM90 in the external phase solution was 3 wt %.

[0064] Preparation of a collection solution: dissolving the surfactant Abil EM90 in isobutyl alcohol (IBA) to obtain a collection solution; the concentration of Abil EM90 in the collection solution is 3 wt %.

[0065] (2) Preparation of concentrated W / O emulsion droplets

[0066] The inner phase solution is injected into the injection tube of the first-level microfluidic device through a syringe pump, and the outer phase solution is injected into the collecting tube of the first-level microfluidic device. A monodisperse W / O emulsion is formed at the outlet of the injection tube in the collecting tube. The W / O emulsion is collected under stirring conditions using a reaction container containing the collecting solution. After receiving, the W / O emulsion is maintained in the collecting solution for a period of time and stirring is continuously applied during the process. During the process of the W / O emulsion droplets being maintained in the collecting solution, since water and IBA are miscible and HA is insoluble in IBA, the water in the W / O emulsion droplets will continuously diffuse into the collecting solution, causing the various components in the W / O emulsion droplets to be concentrated until the concentration of water in the W / O emulsion droplets and the collecting solution reaches equilibrium, thereby obtaining concentrated W / O emulsion droplets.

[0067] In this step, the injection tube of the primary microfluidic device used has an inner diameter of 650 μm and an outer diameter of 1000 μm. The collection tube is a circular tube with an inner diameter of 1050 μm. The inner diameter of the outlet hose is equal to the outer diameter of the collection tube. The flow rate of the internal phase solution is controlled to be 300 μL / h, and the flow rate of the external phase solution is controlled to be 200 μL / min; the volume of the collection solution is controlled to be 100 times the volume of the internal phase solution. After receiving for 3 hours, the collection is stopped and stirring is continued for 30 minutes.

[0068] (3) Preparation of cross-linked HA hydrogel microspheres

[0069] The concentrated W / O emulsion droplets obtained in step (2) were allowed to settle naturally in the reaction container, and most of the collected solution in the reaction container was carefully poured out. The reaction container was placed in an oven and cross-linked at 25° C. for 24 h. After the cross-linking was completed, the cross-linked product was poured into IBA and washed three times, and then poured into PBS buffer and washed three times to obtain cross-linked HA hydrogel microspheres.

[0070] The optical micrographs of the W / O emulsion droplets and concentrated W / O emulsion droplets prepared in step (2) of this example are as follows: Figure 5 As shown in Figures (A) and (B), the optical microscopic images of the cross-linked HA hydrogel microspheres prepared in step (3) of this embodiment in PBS buffer are as follows: Figure 5 As shown in Figure (C). Figure 5 As shown in Figure (A), the diameter of the W / O emulsion droplets collected at the outlet of the first-stage microfluidic device is about 400 μm. The size of the W / O emulsion droplets is uniform and no satellite droplets appear. Figure 5 As shown in Figure (B), when the concentration of water in the W / O emulsion droplets and the collected solution reaches equilibrium, the diameter of the concentrated W / O emulsion droplets is about 142 μm, and the size of the concentrated W / O emulsion droplets has been significantly reduced. Based on the diameter data of the W / O emulsion droplets before and after concentration, it is estimated that the concentration of HA in the concentrated W / O emulsion droplets is above 40 wt%. Figure 5As shown in Figure (C), after the cross-linking reaction, the obtained cross-linked HA hydrogel microspheres are stable in PBS buffer, and the size of the cross-linked HA hydrogel microspheres is uniform, indicating that the hydrogel microspheres are successfully cross-linked.

[0071] Example 3

[0072] In this example, a method for preparing polymer hydrogel microspheres using a low-concentration reaction solution is described in detail. Specifically, the preparation of cross-linked HA hydrogel microspheres is described as an example. The steps are as follows:

[0073] (1) Preparation of internal phase solution, oil phase solution and continuous phase solution

[0074] Preparation of an internal phase solution: HA with a molecular weight of 1-2 MDa, a crosslinker BDDE, and sodium hydroxide were added to high-purity water and stirred until the components were completely dissolved to obtain an internal phase solution; the concentration of HA in the internal phase solution was 4 wt%, the concentration of BDDE was 0.2 wt%, and the concentration of NaOH was 1 wt%, and the viscosity of the internal phase solution was 56 mPa·s.

[0075] The external phase solution was prepared by dissolving the surfactant Abil EM90 in PO to obtain an external phase solution; the concentration of Abil EM90 in the external phase solution was 3 wt %.

[0076] (2) Preparation of concentrated W / O emulsion droplets

[0077] The internal phase solution is injected into the injection tube of the first-level microfluidic device through a syringe pump, and the external phase solution is injected into the collection tube of the first-level microfluidic device. A monodisperse W / O emulsion is formed at the outlet of the injection tube in the collection tube. An open reaction container is used to receive the W / O emulsion. During the receiving process, the W / O emulsion droplets will gradually sink. After the receiving is completed, the external phase solution in the reaction container is carefully aspirated so that the liquid level of the external phase solution in the reaction container is 1 to 2 mm higher than the W / O emulsion droplets. The solution is allowed to stand at room temperature for 2 days. During the standing process at room temperature, a fan is placed near the reaction container and turned on to increase the air flow rate at the reaction container, thereby accelerating the escape of water from the W / O emulsion droplets to obtain concentrated W / O emulsion droplets.

[0078] In this step, the primary microfluidic device used was the same as that in Example 2, and the flow rate of the inner phase solution was controlled to be 200 μL / h, and the flow rate of the outer phase solution was controlled to be 200 μL / min.

[0079] (3) Preparation of cross-linked HA hydrogel microspheres

[0080] The reaction vessel containing the concentrated W / O emulsion droplets in step (2) was placed in an oven and cross-linked at 25° C. for 24 h. After the cross-linking was completed, the cross-linked product was poured into IBA and washed three times, and then poured into PBS buffer and washed three times to obtain cross-linked HA hydrogel microspheres.

[0081] The optical micrographs of the W / O emulsion droplets and concentrated W / O emulsion droplets prepared in step (2) of this example are as follows: Figure 6 As shown in Figures (A) and (B), the optical microscopic images of the cross-linked HA hydrogel microspheres prepared in step (3) of this embodiment in PBS buffer are as follows: Figure 6 As shown in Figure (C). Figure 6 It can be seen that compared with the W / O emulsion droplets, the size of the concentrated W / O emulsion droplets has been significantly reduced, from 263 μm before concentration to 110 μm after concentration. The diameter of the concentrated W / O emulsion droplets is 0.42 times the diameter of the W / O emulsion droplets. This is mainly caused by the escape of water in the W / O emulsion droplets into the air during the concentration process. This process will significantly increase the concentrations of HA, BDDE and sodium hydroxide in the concentrated W / O emulsion droplets, which is beneficial to the subsequent cross-linking reaction; after the cross-linking reaction, the obtained cross-linked HA hydrogel microspheres are stable in PBS buffer, indicating that the hydrogel microspheres are successfully cross-linked.

[0082] As shown in Examples 1-3, emulsion droplets prepared by various methods can all be concentrated using the method of the present invention to obtain concentrated emulsion droplets, and these concentrated emulsion droplets can all be used to successfully crosslink hydrogel microspheres. Furthermore, using both the in-liquid drying method of Examples 1-2 and the evaporative drying method of Example 3, emulsion droplets can be concentrated to obtain concentrated emulsion droplets, thereby successfully crosslinking hydrogel microspheres.

[0083] Comparative Example 1

[0084] In this comparative example, it was investigated whether cross-linked HA hydrogel microspheres could be successfully prepared without concentrating the W / O emulsion droplets.

[0085] (1) Prepare internal phase solution and external phase solution

[0086] Preparation of the internal phase solution: Add HA with a molecular weight of 1 to 2 MDa, a crosslinker BDDE, and sodium hydroxide to high-purity water, and stir until all components are completely dissolved to obtain an internal phase solution; in the internal phase solution, the concentration of HA is 4 wt%, the concentration of BDDE is 0.2 wt%, and the concentration of NaOH is 1 wt%.

[0087] The external phase solution was prepared by dissolving the surfactant Abil EM90 in liquid paraffin (PO) to obtain an external phase solution; the concentration of Abil EM90 in the external phase solution was 3 wt %.

[0088] (2) Preparation of W / O emulsion droplets

[0089] The inner phase solution was poured into the outer phase solution and stirred at 600 rpm for 10 min using a magnetic stirrer to obtain a W / O emulsion. In this step, the volume ratio of the inner phase solution, the outer phase solution, and the collection solution was controlled to be 1:50:200.

[0090] (3) Cross-linking

[0091] The W / O emulsion droplets obtained in step (2) were allowed to settle naturally in the reaction vessel, and most of the external phase solution in the reaction vessel was carefully poured out. The reaction vessel was placed in an oven and cross-linked at 25° C. for 24 h. After the cross-linking was completed, the cross-linked product was poured into IBA and washed three times, and then poured into PBS buffer for washing.

[0092] The optical micrograph of the W / O emulsion droplets prepared in step (2) of this comparative example is as follows: Figure 7 As shown in FIG. (A), in step (3) of this comparative example, after cross-linking, the cross-linked product was poured into PBS buffer for washing, and it was found that the cross-linked product disappeared. Figure 7 As shown in Figure (B), no cross-linked products were observed in the PBS buffer. This indicates that the microspheres failed to cross-link successfully in this comparative example. Although this comparative example employed the same formulation as Example 1, the failure to cross-link the microspheres was primarily due to the low concentrations of HA and BDDE in the internal phase solution. This also demonstrates the effectiveness of the present method of concentrating the W / O emulsion droplets before cross-linking.

[0093] Comparative Example 2

[0094] In this comparative example, the concentrations of the components in the internal phase solution were increased to investigate whether the cross-linked HA hydrogel microspheres could be successfully prepared by using an internal phase solution with a high reactant concentration.

[0095] (1) Prepare internal phase solution and external phase solution

[0096] Preparation of the internal phase solution: Add HA with a molecular weight of 1 to 2 MDa, a crosslinker BDDE, and sodium hydroxide to high-purity water, and stir until all components are completely dissolved to obtain an internal phase solution; in the internal phase solution, the concentration of HA is 4 wt%, the concentration of BDDE is 8 wt%, and the concentration of NaOH is 1 wt%.

[0097] The external phase solution was prepared by dissolving the surfactant Abil EM90 in liquid paraffin (PO) to obtain an external phase solution; the concentration of Abil EM90 in the external phase solution was 3 wt %.

[0098] (2) Preparation of W / O emulsion droplets

[0099] The inner phase solution was poured into the outer phase solution and stirred at 600 rpm for 10 min using a magnetic stirrer to obtain a W / O emulsion. In this step, the volume ratio of the inner phase solution, the outer phase solution, and the collection solution was controlled to be 1:50:200.

[0100] (3) Cross-linking

[0101] The W / O emulsion droplets obtained in step (2) were allowed to settle naturally in the reaction vessel, and most of the external phase solution in the reaction vessel was carefully poured out. The reaction vessel was placed in an oven and cross-linked at 25° C. for 24 h. After the cross-linking was completed, the cross-linked product was poured into IBA and washed three times, and then poured into PBS buffer and washed three times.

[0102] The optical micrograph of the W / O emulsion droplets prepared in step (2) of this comparative example is as follows: Figure 8 As shown in Figure (A), the optical micrograph of the cross-linked product of step (3) of this comparative example in PBS buffer is as follows Figure 8 As shown in Figure (B). Figure 8 As shown in Figure (B), the edge of the cross-linked product in PBS buffer is blurred and difficult to identify, which indicates that the cross-linking effect of HA is not good.

[0103] The cross-linked product of this comparative example was enzymatically hydrolyzed according to the literature by Frida J. Wende, Suresh Gohil, Lars I. Nord, Anne Helander Kenne, Corine Carbohydrate Polymers 2017, 157, 1525–1530, and tested the nuclear magnetic resonance hydrogen spectrum (H-NMR). The results are as follows Figure 9 As shown. The degree of modification (MoD) of BDDE in the cross-linked product calculated based on H NMR spectroscopy was 32.5%. Combining Comparative Example 2 with Example 1, it can be seen that the concentration of HA in the internal phase solution of Comparative Example 2 was the same as that in Example 1. However, even with the addition of BDDE at a concentration 40 times that of Example 1, which increased the BDDE modification of the cross-linked product by more than 16 times, the cross-linking effect was still inferior to that of Example 1. The cross-linked product could only maintain the morphology of a gel, and no cross-linked product with good sphericity was obtained.

[0104] Example 4

[0105] In this embodiment, a method for preparing polymer hydrogel microspheres using a low-concentration reaction solution is described in detail. Specifically, the preparation of cross-linked gelatin hydrogel microspheres is described as an example. The steps are as follows:

[0106] (1) Prepare internal phase solution, external phase solution and collection solution

[0107] Preparation of an internal phase solution: Gelatin with a molecular weight of 1-2 MDa and cross-linking agent glutaraldehyde were added to high-purity water and stirred at 50°C until all components were completely dissolved to obtain an internal phase solution; the concentration of gelatin in the internal phase solution was 15 wt%, the concentration of glutaraldehyde was 0.01 wt%, and the viscosity of the internal phase solution was 61 mPa·s.

[0108] The external phase solution was prepared by dissolving the surfactant polyglycerol ricinoleate (PGPR) in soybean oil to obtain an external phase solution; the concentration of PGPR in the external phase solution was 5 wt %.

[0109] Preparation of a collecting solution: dissolving the surfactant PGPR in ethyl acetate (EA) to obtain a collecting solution; the concentration of PGPR in the collecting solution is 5 wt %.

[0110] (2) Preparation of concentrated W / O emulsion droplets

[0111] The internal phase solution was poured into the external phase solution and stirred at 600 rpm for 10 minutes using a magnetic stirrer to obtain a W / O emulsion. The W / O emulsion was then added dropwise to a reaction vessel containing a collection solution and stirred at 600 rpm for 30 minutes. While the W / O emulsion droplets were retained in the collection solution, since water and EA are miscible while HA is insoluble in EA, the water in the W / O emulsion droplets continuously diffused into the collection solution, concentrating the components in the W / O emulsion droplets until the water concentration in the W / O emulsion droplets and the collection solution reached equilibrium, resulting in concentrated W / O emulsion droplets.

[0112] In this step, the volume ratio of the inner phase solution, the outer phase solution and the collection solution was controlled to be 1:50:200.

[0113] (3) Preparation of cross-linked gelatin hydrogel microspheres

[0114] The concentrated W / O emulsion droplets obtained in step (2) were allowed to settle naturally in the reaction container, and most of the collected solution in the reaction container was carefully poured out. The reaction container was placed in an environment of 4°C for cross-linking for 10 hours. After the cross-linking was completed, the cross-linked product was poured into IBA and washed three times, and then poured into PBS buffer and washed three times to obtain cross-linked gelatin hydrogel microspheres.

[0115] The cross-linked gelatin hydrogel microspheres prepared in this example were observed using an optical microscope. The results showed that the cross-linked gelatin hydrogel microspheres were stable in the PBS buffer solution and had clear edges, indicating good cross-linking.

[0116] Example 5

[0117] In this embodiment, a method for preparing polymer hydrogel microspheres using a low-concentration reaction solution is described in detail. Specifically, the preparation of cross-linked polyvinyl alcohol (PVA) hydrogel microspheres is described as an example. The steps are as follows:

[0118] (1) Prepare internal phase solution, external phase solution and collection solution

[0119] Preparation of an internal phase solution: PVA with a molecular weight of 1-2 MDa and crosslinking agent glutaraldehyde are added to high-purity water and stirred until all components are completely dissolved to obtain an internal phase solution; the concentration of gelatin in the internal phase solution is 0.5 wt%, the concentration of glutaraldehyde is 1 wt%, and the viscosity of the internal phase solution is 36 mPa·s.

[0120] The external phase solution was prepared by dissolving the surfactant Span 80 in mineral oil (MO) to obtain an external phase solution; the concentration of Span 80 in the external phase solution was 1 wt %.

[0121] Preparation of a collecting solution: dissolving the surfactant Span 80 in IBA to obtain a collecting solution; the concentration of Span 80 in the collecting solution is 1 wt %.

[0122] (2) Preparation of concentrated W / O emulsion droplets

[0123] The internal phase solution was poured into the external phase solution and stirred at 600 rpm for 10 minutes using a magnetic stirrer to obtain a W / O emulsion. The W / O emulsion was then added dropwise to a reaction vessel containing a collection solution and stirred at 600 rpm for 30 minutes. While the W / O emulsion droplets were retained in the collection solution, water from the W / O emulsion droplets continuously diffused into the collection solution, as water and IBA are miscible while HA is insoluble in IBA. This concentrated the components in the W / O emulsion droplets until the water concentration in the W / O emulsion droplets reached equilibrium with that in the collection solution, resulting in concentrated W / O emulsion droplets.

[0124] In this step, the volume ratio of the inner phase solution, the outer phase solution and the collection solution was controlled to be 1:50:200.

[0125] (3) Preparation of cross-linked PVA hydrogel microspheres

[0126] The concentrated W / O emulsion droplets obtained in step (2) were allowed to settle naturally in the reaction container, and most of the collected solution in the reaction container was carefully poured out. The reaction container was placed in an oven and cross-linked at 70° C. for 6 h. After the cross-linking was completed, the cross-linked product was poured into IBA and washed three times, and then poured into PBS buffer and washed three times to obtain cross-linked PVA hydrogel microspheres.

[0127] The cross-linked PVA hydrogel microspheres prepared in this example were observed using an optical microscope. The results showed that the cross-linked PVA hydrogel microspheres were stable in the PBS buffer solution and had clear edges, indicating that the cross-linking was good.

Claims

1. A method for preparing polymer hydrogel microspheres using a low-concentration reaction solution, characterized in that: The following steps are involved: (1) dissolving the components including the polymer material and the cross-linking agent in water to form an inner phase solution; dissolving the oil-soluble surfactant in the oil phase to form an outer phase solution; An internal phase solution and an external phase solution are used to prepare a water-in-oil emulsion; The concentrations of the polymer material and the crosslinking agent in the internal phase solution should be such that, when the crosslinking reaction conditions are applied, the polymer material and the crosslinking agent in the water-in-oil emulsion droplets cannot undergo a crosslinking reaction; (2) removing part of the water in the water-in-oil emulsion droplets by a liquid drying method or a volatile drying method, thereby concentrating the components in the internal phase solution of the water-in-oil emulsion droplets to obtain concentrated water-in-oil emulsion droplets; the degree of concentration of the components in the internal phase solution of the water-in-oil emulsion droplets should ensure that when the crosslinking reaction conditions are applied, the polymer material and the crosslinking agent in the concentrated water-in-oil emulsion droplets can undergo a crosslinking reaction; (3) applying cross-linking reaction conditions to allow the polymer material in the concentrated oil-in-water emulsion droplets to undergo a cross-linking reaction with the cross-linking agent to solidify the concentrated oil-in-water emulsion, and washing to remove unreacted components to obtain polymer hydrogel microspheres.

2. The method for preparing polymer hydrogel microspheres using a low-concentration reaction solution according to claim 1, characterized in that: The operation of removing part of the water in the oil-in-water emulsion droplets by the liquid drying method is as follows: dissolving an oil-soluble surfactant in a concentrator to obtain a collection solution, wherein the concentrator is an organic solvent that is miscible with water but does not dissolve the polymer material in the internal phase solution; placing the oil-in-water emulsion droplets prepared in step (1) in the collection solution, and the volume of the collection solution is at least 10 times the volume of the internal phase solution used in step (1) when preparing the oil-in-water emulsion. The water in the internal phase solution in the oil-in-water emulsion droplets will diffuse into the collection solution, so that the components in the internal phase solution of the oil-in-water emulsion droplets are concentrated to obtain concentrated oil-in-water emulsion droplets.

3. The method for preparing polymer hydrogel microspheres using a low-concentration reaction solution according to claim 2, characterized in that: The water-in-oil emulsion droplets prepared in step (1) are placed in a collecting solution until the concentration of water in the water-in-oil emulsion droplets and the collecting solution reaches equilibrium, thereby obtaining concentrated water-in-oil emulsion droplets.

4. The method for preparing polymer hydrogel microspheres using a low-concentration reaction solution according to claim 2, characterized in that: The concentrator is one or more of ethyl acetate, isobutanol, ethanol, acetone, and methyl isobutyl ketone.

5. The method for preparing polymer hydrogel microspheres using a low-concentration reaction solution according to claim 1, characterized in that: The operation of removing part of the water in the oil-in-water emulsion droplets by the volatile drying method is as follows: the oil-in-water emulsion prepared in step (1) is placed in an open container, the oil-in-water emulsion droplets will naturally sink, and the liquid level of the oil phase solution in the oil-in-water emulsion is controlled to be 1 to 3 mm higher than the upper surface of the oil-in-water emulsion droplets located at the top. After standing at room temperature, the water in the oil-in-water emulsion droplets will evaporate into the air, so that the components in the internal phase solution of the oil-in-water emulsion droplets are concentrated to obtain concentrated oil-in-water emulsion droplets.

6. The method for preparing polymer hydrogel microspheres using a low-concentration reaction solution according to claim 5, characterized in that: The standing time at room temperature is controlled so that the diameter of the concentrated water-in-oil emulsion droplets obtained is (0.3 to 0.8) times the diameter of the water-in-oil emulsion droplets prepared in step (1).

7. The method for preparing polymer hydrogel microspheres using a low-concentration reaction solution according to claim 5, characterized in that: During the standing process at room temperature, the rate at which water in the water-in-oil emulsion droplets evaporates into the air is increased by promoting the air flow rate in the environment where the open container is located.

8. The method for preparing polymer hydrogel microspheres using a low-concentration reaction solution according to any one of claims 1 to 7, characterized in that: In step (1), the concentration of the crosslinking agent in the internal phase solution is 0.001 wt% to 10 wt%, and the concentration of the polymer material in the internal phase solution should be such that the viscosity of the internal phase solution does not exceed 100 mPa·s.

9. The method for preparing polymer hydrogel microspheres using a low-concentration reaction solution according to any one of claims 1 to 7, characterized in that: The internal phase solution also contains at least one of an acid-base regulator, a catalyst, and an initiator.

10. The method for preparing polymer hydrogel microspheres using a low-concentration reaction solution according to any one of claims 1 to 7, characterized in that: The polymer material includes at least one of hyaluronic acid, sodium hyaluronate, agarose, gelatin, chitosan, polyvinyl alcohol, and sodium alginate.

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