WO2014079198A1 - Degradable wound-repairing material and preparation method thereof - Google Patents

Abstract

The present invention relates to a degradable wound-repairing material and the preparation method thereof, and the raw material of the degradable wound-repairing material comprises matrix components and auxiliary components, the matrix components comprise protein components and the auxiliary components comprise at least one of an antimicrobial agent and an active factor, wherein the antimicrobial agent is a synthetic antimicrobial drug, an inorganic antimicrobial agent, an organic antimicrobial agent or a natural antimicrobial agent; the active factor is at least one of the following: an epidermal growth factor, an FGF vascular endothelial growth factor, a platelet derived growth factor, a platelet activating factor, an insulin-like growth factor, a tumour necrosis factor, an interleukin, colony stimulating factor-1, various bone morphogenetic proteins or a transforming growth factor. The antimicrobial agent and the active factor are loaded on the bases of the matrix components, and thereby the present invention can also possess biological activity on the basis of satisfying biodegradability conditions to better promote wound repairing, and displays a good anti-infective performance during the application process.

Description

 Degradable wound repairing material and preparation method thereof

 [0001] The present invention relates to the field of biomedical materials and biomedical engineering technology, and in particular, to a degradable wound repairing material and a preparation method thereof.

Background technique

 [0002] Skin is one of the most important organs of the human body. It is an important barrier between the human body and the external environment, and one of the most vulnerable organs. The repair of skin wounds is also one of the oldest medical problems in humans.

 [0003] From the composition of the skin, the dermis layer of the skin is rich in collagen fibers and glycoproteins, which is rich in blood supply and has strong regeneration and repair ability. Therefore, in the design of wound repair materials, high-molecular scaffolds with good biocompatibility and excellent degradation properties provide sites for early cell attachment, which can promote their proliferation and migration more effectively. Supplemented with the necessary antibacterial ingredients and growth-promoting growth factors, it can reduce the early infection of wound repair and accelerate wound healing.

[0004] In recent years, degradable wound repair materials have been applied for skin wound repair treatment. At present, the clinically used degradable wound repair materials are mainly collagen sponges, which have good biocompatibility, are one of the important components of human tissues, and have excellent degradability and degradation. The process can guide tissue proliferation and migration. However, this type of material degrades too quickly, and it is easy to lose the mechanical support structure prematurely. Moreover, during wound repair, the wound is susceptible to infection, and such stents are also difficult to load with antibacterial agents and factors that promote wound healing. Therefore, the development of components and structures that are more similar to natural skin and diverse in morphology, and the ability to load a variety of components to prevent infection and promote wound repair has far-reaching significance.

Summary of the invention

[0005] The technical problem to be solved by the present invention is to provide a degradable wound repairing material with good biocompatibility.

[0006] A further technical problem to be solved by the present invention is to provide a method for preparing a degradable wound repairing material to obtain a biodegradable biodegradable wound repairing material.

 [0007] In order to solve the above technical problems, the present invention provides the following technical solutions: A degradable wound repairing material, the raw material comprising a matrix component, the matrix component comprising a protein component, the protein component is derived from a mammal The collagen in the skin or Achilles or the silk fibroin or silk fibroin-modified derivative.

[0008] Further, the raw material of the degradable wound repairing material further comprises an auxiliary component, wherein the auxiliary component comprises at least one of an antibacterial agent and an active factor, wherein the antibacterial agent is a synthetic antibacterial drug, an inorganic antibacterial agent, and an organic An antibacterial agent or a natural antibacterial agent; the active factor is at least one of the following active factors: epidermal growth factor, vascular endothelial growth factor, platelet-derived growth factor, platelet activating factor, insulin-like growth factor, tumor necrosis factor, leukocyte-mediated Prime, Colony stimulating factor-1, various bone morphogenetic proteins or transforming growth factors.

 [0009] Further, the matrix component further includes a polysaccharide component, the polysaccharide component comprising at least one of the following components: hyaluronic acid, chitin and chitosan derived from the arthropod shell, source Alginic acid or sodium alginate in seaweed or kelp or chondroitin sulfate extracted from organs of pigs, cattle, sheep or sharks.

[0010] Further, the mass ratio of the protein component to the sum of all other raw material components is 1/350 to 4000/1.

 [0011] In another aspect, the present invention provides a method for preparing a degradable wound repairing material according to any of the above, comprising the following steps;

Preparing a substrate material step, configuring a matrix component solution;

In the molding step, the composite is subjected to a molding process to obtain a desired material form.

[0012] Further, in the step of preparing the base material, mixing the respective matrix component solutions under stirring conditions, and performing a pore-forming process to obtain a composite with a support pore structure as a matrix material, wherein the pore-forming process is as follows At least one of: particle leaching, stirred foaming or defoaming, initial concentration adjustment or complex concentration.

 [0013] Further, a composite auxiliary component step is further performed between the preparation of the base material and the molding step, and at least one of an antibacterial agent and an active factor is loaded onto the composite having the pore structure of the stent, wherein the antibacterial agent The loading mode is any one of the following modes: grafting, physical blending, adsorption or microsphere encapsulation, and the loading method of the active factor is any one of the following modes: grafting, physical blending, adsorption Or microspheres are embedded.

 [0014] Further, the molding step adopts a flow film forming process or a film forming process or a granulation process, and after the forming step, a crosslinking step is further performed, and the crosslinked composite is washed and dried to obtain the following One of the composites of the state: a dry sponge, a wet sponge, a dry granule or a wet granule, the crosslinking step using a physical crosslinking process, a crosslinking process involving a chemical crosslinking agent, or a natural crosslinking agent Cross-linking process, wherein

The physical crosslinking process is any one of the following processes: a vacuum high temperature crosslinking process, an ultraviolet irradiation process, a Y irradiation process, a high energy electron beam irradiation, and the like;

The crosslinking agent used in the crosslinking process involving the chemical crosslinking agent is any one of the following components: an aldehyde crosslinking agent, an imide crosslinking agent or a diisocyanate crosslinking agent;

The crosslinking agent used in the crosslinking process involving natural crosslinking agents is genipin.

 [0015] Further, the molding step uses a gel process or a compound solution dilution process to obtain an injectable gel or a sprayable solution of a degradable wound repair material, respectively.

[0016] Further, the preparation method further includes a terminal sterilization step as a last step, the obtained material is sterilized to obtain a finished product, and the terminal sterilization step adopts any one of the following sterilization processes. Species: High-energy electron beam irradiation process, Y-radiation process, filtration process, gas sterilization process. [0017] After adopting the above technical solution, the present invention has at least the following beneficial effects: by using collagen derived from mammalian skin or Achilles tendon or silk fibroin derived from silk or silk fibroin modified by derivatization As a protein component, it has good biocompatibility and is more conducive to degradation and absorption.

 [0018] In addition, the loading of the active factor component can also be biologically active on the basis of satisfying the degradable property, and can better promote wound repair; and the anti-infective property of the product is improved by loading the antibacterial agent on the basis of the matrix component.

DRAWINGS

 1 is a flow chart of a method for preparing a degradable wound repairing material of the present invention.

 2 is a schematic view showing the path of a dispersion of a solution of a living factor or a suspension of a sustained release carrier onto a formed sponge block in the method for preparing a degradable wound repairing material of the present invention.

detailed description

 [0021] It should be noted that, in the case of no conflict, the embodiments in the present application and the features in the embodiments may be combined with each other. The present invention will be further described in detail below with reference to the accompanying drawings and specific embodiments.

[0022] The present invention provides a degradable wound repairing material, the raw materials of which include:

Protein-based components, including various types of collagen derived from skins such as cows, sheep, pigs, and the like, and derivatives of silk fibroin or silk fibroin derived from silk, such as: The collagen may be type I, type II, type III, type IV, type V, type VI, type VD, type VIII, type IX, type X, type XI, type ΧΠ, type X IV, type XV, type X VI , χνιπ type,

X X type and other types of collagen;

a polysaccharide component, chitin and chitosan derived from the arthropod shell, alginic acid or sodium alginate derived from seaweed or kelp, chondroitin sulfate extracted from various organs of pig, cow, sheep and shark, and transparent Acidic acid, etc.

[0023] The above protein components and polysaccharide components may be regarded as the matrix component of the degradable wound repairing material of the present invention, but in preparation, the two are still different, and the protein component is an indispensable matrix component, and more The sugar component can be selectively added according to the needs of product differentiation.

 [0024] In the actual preparation process, if necessary, the wound repairing material of the present invention may optionally be added with one or more of the following antibacterial agents:

Synthetic antibacterial drugs, such as: penicillins, cephalosporins, other beta-lactamase inhibitors, aminoglycosides, amides, glycopeptides, macrolides, tetracyclines, sulfonamides, quinolones , furans, antifungals, nitrazoles;

Inorganic antibacterial agents, such as: nano-silver, nano-titanium dioxide, zinc oxide, copper oxide, ammonium dihydrogen phosphate, lithium carbonate, etc.; organic antibacterial agents, such as: vanillin, ethyl vanillin, acyl aniline, imidazole, Thiazoles, isothiazolone derivatives, quaternary ammonium salts, biguanides, phenols, etc.; Natural antibacterial agents, such as: chitin, mustard, castor oil, wasabi, etc.

 [0025] In order to increase the prosthetic repair ability of the material, one or more of the following active factors may be optionally added to the wound repair material of the present invention: epidermal growth factor (EGF), vascular endothelial growth factor (VEGF), platelet Derived growth factor (PDGF), platelet activating factor (PAF), insulin-like growth factor (IGF), tumor necrosis factor (TNF), interleukin (IL-1, IL-3, IL-4, IL-6, etc.) , colony stimulating factor-1, various bone morphogenetic proteins (BMPs) and other transforming growth factors (TGF).

 [0026] It should be noted that the core of the innovation of the present invention is that the above-mentioned raw materials are prepared into a porous scaffold material according to the method to be described below, and as the raw materials used, in addition to protein components, other raw materials are required. The amount of the component may be in any ratio, and in the case where other raw material components are added in addition to the protein component, the mass ratio of the protein component to the sum of all other raw material components is 1/350 to 4000/1.

 As shown in FIG. 1, the preparation method of the degradable wound repairing material of the present invention includes the main steps as will be described in detail below.

 [0028] -, preparation of the base material steps:

 1. Preparation of solution: Select one or more of the above raw materials to prepare a solution. The solution concentration (mass fraction concentration) and solvent are as follows: Collagen: 0.01%~10%, acid (mass percentage is 0.05) Aqueous solution or pure water of %~10% acetic acid or hydrochloric acid; silk fibroin: 0.02%~10%, pure water; polysaccharide: 0.001%~15%, acid (0.001%~10% acetic acid or hydrochloric acid, etc.) or pure water.

 [0029] 2, compounding: using the protein component as a first component, the second component or the third component or other component solution of the complex is sequentially added to the solution of the first component under stirring In the method of direct pouring or gradual addition, stirring is continued for a certain period of time after the addition, until a uniform composite liquid is formed, and the solute components in the composite liquid, the protein component and the other components as the first component The mass ratio of the sum is 1/350 to 4000/1.

[0030] 3. Control hole structure: Particle leaching method, stirring foaming or defoaming method, initial concentration adjustment method or composite liquid concentration method may be selected as the pore forming process. If particle leaching is used, porogens (sodium, potassium and ammonium salts such as NaCl, KN0 ₃ , NH ₄ C1) are added, and the pore structure can be controlled by the content of the porogen. The mass ratio of the composite liquid prepared in step 2 is 1/100~20/100; the pore structure of the support can also be controlled by stirring or defoaming, for example, the step poly 2 solution compound is stirred and stirred at a certain stirring speed. One can select the rotation speed, then immediately mold injection, and then the composite can be defoamed and then injection molded, wherein the defoaming vacuum is 2~200Pa, the defoaming time is 1~30 minutes; the initial concentration adjustment rule is based on the solution The initial concentration is controlled according to the concentration of the solution prepared in the above step 1; the composite liquid concentration method is to concentrate the composite liquid prepared in the step 2 by centrifugation or filtration to control the pore structure. . It is possible to select any one of the above several boring processes, or two, three or four at the same time. [0031] Second, the composite auxiliary component steps:

 When necessary, the composite may also be selectively compounded with an antibacterial agent or an active factor, or both may be selected as an auxiliary component, as follows:

 A, the composite of antibacterial agents in the complex:

 1. For soluble antibacterial drugs, select a suitable solvent (see Table 1) to dissolve, the concentration of which can be determined according to the dosage of the drug; for ionic or granular antibacterial agents, choose a suitable dispersion medium for dispersion; And an ionic or granular antibacterial agent, which can be optionally embedded in a sustained-release carrier (polymer microspheres, microcapsules, particles, etc.), and then prepared into a dispersion;

 Table 1. Solvents or dispersants selected for various antibacterial agents

 2. The solution or dispersion of the antibacterial agent prepared in the step 1 is added to the complex prepared in the first step under stirring. In the resulting complex, the dose of the antibacterial agent should take into account the pharmacodynamic and pharmacokinetic parameters, as well as the difference between the concentration-dependent drug and the time-dependent drug.

 [0032] B, complexation of active factors in the complex:

 Method 1 : Graft blending

 1. Various active factors may also be selected by first-dissolving various solvents listed in Table 1. Alternatively, the active factors may be embedded in a sustained-release carrier (polymer microspheres, microcapsules, particles, etc.), sustained-release carrier. The preparation method includes a microemulsion method, a spray drying method, etc., and the base material of the prepared sustained-release carrier can be selected from the polymer materials shown in Table 2;

 Table 2 Base material of sustained release carrier

2. preparing an active factor or a sustained release carrier into a solution or dispersion; 3. Under stirring, (2) is added dropwise to the solution of the composite prepared in the first step, and in the dry weight of the formed composite, the mass ratio of the active factor to the dry weight of the composite is 10 ng/g~ 100mg/g.

Method 2: Physical adsorption

The solution of the active factor prepared in the step 2 of the method 1 or the dispersion of the sustained-release carrier is added dropwise to the matrix material obtained in the first step in a row-by-row manner, and can be fully adsorbed by means of a negative pressure, specific The dropping path is as shown by the arrow in Fig. 2, and is dropped back and forth from the first line.

[0034] Step three, the molding step:

The molding process of the wound repairing material of the present invention may be selected from, but not limited to, one of the following processes: flow film formation, film formation, granulation, gelation, and dilution of a composite solution. And with freeze drying, air conditioning at room temperature.

[0035] Specifically, the molding of the composite obtained in the second step may select one of the following routes:

Route 1, rogue - freezing - freeze drying

The process parameters of the route: the temperature during freezing is -5~-150 °C; the temperature during freeze-drying is -65~45 °C, and the pressure is 0.1~Pa.

 [0036] Route 2, rogue - freezing - freeze drying - pressing into a film

The process parameters of the route: the temperature during freezing is -5~-150 °C; the temperature during freeze-drying is -65~45 °C, the pressure is 0.1~200Pa; the pressure when pressed into film is 5Pa~5000MPa, time It is 0.5~168h.

[0037] Route 3, rogue - air drying at room temperature - pressed into a film

The process parameters of the route: the temperature at room temperature is 5~45 °C; the pressure when pressed into film is 5Pa~5000MPa.

[0038] Route 4, injection molding - freezing - freeze drying

The process parameters of the route: the temperature during freezing is -5~-150 °C; the temperature during freeze-drying is -65~45 °C, and the pressure is 0.1~Pa.

 [0039] Route 5, composite solution - defoaming - filtration - granulation

When filtering, the pore size of the filter is lm~10mm; when the granulation is carried out, the flow rate of the pump head is 0.05~200mL/min, and the flow of the pump head can be selected from air, nitrogen or other inert gas, and the pumped composite droplets are filled with hydrocarbons. For a compound (such as hexane) collection tank, the temperature of the collection tank is maintained at -40 to -2 °C.

 [0040] Route 6, composite solution - defoaming - filtration - granulation - freeze drying

When filtering, the pore size of the filter is lm~10mm; when the granulation is carried out, the flow rate of the pump head is 0.05~200mL/min, and the flow of the pump head can be selected from air, nitrogen or other inert gas, and the pumped composite droplets are filled with organic A collection tank of a mixture of solvent and water (such as alcohol and water), the temperature of the collection tank is maintained at -40 to -2 ° C; the temperature during freeze-drying is -65 to 45 ° C, and the pressure is 0.1 to 200 Pa. [0041] Route 7, composite solution gel

The composite solution obtained in the second step is allowed to stand in an environment of 1 to 8 ° C, or diluted in a certain ratio, and then allowed to stand in an environment of 1 to 8 ° C to obtain a composite gel.

[0042] Route 8, dilution of the composite solution

The composite solution obtained in the second step is diluted in a certain ratio to obtain a composite sprayable solution.

[0043] Step four, cross-linking steps:

The crosslinking method of the wound repairing material of the invention may be selected but not limited to: physical crosslinking method such as vacuum high temperature crosslinking, ultraviolet irradiation, Y irradiation, high energy electron beam irradiation, or various chemical crosslinking agents (aldehyde Cross-linking agent: Glyoxal, glutaraldehyde, formaldehyde, etc., imine cross-linking agent: carbodiimide, etc., diisocyanate cross-linking agent: diisocyanate, carbamate derivative, etc. A cross-linking method involving natural cross-linking agents such as Genipin.

 [0044] The sample of the dried composite obtained by the route 1-6 in the third step may be crosslinked by one or more crosslinking methods. The crosslinking method includes: physical crosslinking method such as vacuum high temperature crosslinking, ultraviolet irradiation, Y irradiation, high energy electron beam irradiation, or various chemical crosslinking agents (aldehyde crosslinking agent: glyoxal, glutaraldehyde, Formaldehyde, etc., imine cross-linking agent: carbodiimide, etc., diisocyanate cross-linking agent: diisocyanate, carbamate derivative, etc.) and natural cross-linking agent [eg Genipin (Genipin) ) etc.] Participate in the cross-linking method. The specific crosslinking conditions are as follows:

In the method of vacuum high-temperature crosslinking, the crosslinking temperature is 50 to 200 ° C, the crosslinking time is 2 to 120 hours, the vacuum pressure during crosslinking is 0.1 to 200 Pa, and the irradiation time is 5 minutes in the ultraviolet irradiation crosslinking. ~ 48 hours, the irradiation intensity is 10~100mW/cm ² ; the dose of Y irradiation is l~40KGy; the dose of high energy electron beam irradiation is l~50KGy.

[0045] In the chemical crosslinking method in which the crosslinking agent is involved, the dried composite film sample is immersed in a solution of various concentrations of various crosslinking agents for a certain period of time. The mass fraction of the carbodiimide solution is 1 μg/mL~100 mg/mL, the crosslinking time is 5 minutes to 72 hours; the mass fraction concentration of the glyoxal, glutaraldehyde, and formaldehyde solution is 0.01 to 10%, The crosslinking time is 1 to 168 hours; the mass fraction concentration of the diisocyanate crosslinking agent solution is 0.1 to 8%, the crosslinking time is 30 minutes to 84 hours; the mass fraction concentration of the Genipin solution is 0.05~ 8%, the crosslinking time is 2 to 168 hours.

[0046] The final morphology of the wound repair material obtained by the cross-linking of the cross-linking agent after the cross-linking agent can be prepared into one of the following states: dry sponge, wet sponge, injectable coagulation Glue, dry or wet granules, sprayable solution.

[0047] Step 5, sterilization treatment:

The sterilization method of the wound repairing material of the present invention may be one of the following methods: terminal sterilization process, for example: high energy electron beam irradiation, Y radiation, filtration, gas sterilization (ethylene oxide, formaldehyde vapor, ethanol, etc.) or Other terminal sterilization processes; process aseptic processes can also be employed. [0048] The aseptic mode of the composite in various states obtained in the fourth step can be selected by using both the process aseptic process and the terminal sterilization. The aseptic process of the process, that is, the entire preparation process is carried out in a clean room of a suitable degree, and aseptic control is carried out. Terminal sterilization refers to the terminal sterilization process after the composite produced in the clean workshop, such as: high energy electron beam irradiation, Y radiation, filtration, gas sterilization (ethylene oxide, formaldehyde vapor, ethanol, etc.) or Other sterilization methods are sterilized.

[0049] wherein, the dry sponge, the wet sponge, the dry particles or the wet particles, and the gel state composite may be sterilized by one of high energy electron beam or Y radiation, and the sterilization dose is 5 to 35 KGy. The dry sponge and the dry granules may be sterilized by a gas sterilization method (ethylene oxide, formaldehyde vapor, ethanol, etc.), and the sterilization parameters are determined according to the state of the product and the purpose to be achieved. The gel-state and solution-state complexes may be sterilized by a filter sterilization method, and the pores of the filter membrane should be less than or equal to 0.22 μm.

[0050] The specific details of the preparation process of the present invention are described in detail below by means of several examples.

Embodiment 1

A 1% collagen solution was prepared by using 3% aqueous acetic acid as a solvent, and a 0.001% chitosan solution was prepared using 0.001% aqueous acetic acid as a solvent. Then, 4 parts by mass of the type I collagen solution was taken and stirred at 500 rpm, and 1 part by mass of chitosan solution was added at a rate of 2 ml/min. After the completion of the addition, stirring was continued to form a uniform composite liquid. The mass ratio of collagen to chitosan in the obtained composite solution was 4000:1.

 [0052] The VEGF-loaded PBS solution was added to the composite solution and stirred and uniformly mixed so that the mass ratio of the active factor to the dry weight of the composite in the resulting composite was 1/100. The resulting composite solution was defoamed under vacuum of 20 Pa for 20 min.

[0053] Weigh the defoamed composite liquid in a mold and then freeze at -150 ° C for 5 min, and then in a freeze dryer (temperature is -65 ° C after 45 ° C, pressure is O.lPa Freeze in the middle.

[0054] A carbodiimide solution having a concentration of 100 g/L was prepared, and the film obtained above was immersed therein and immersed for 30 hours. Thereafter, after washing with water for 5 times, a wet spongy material was obtained. It was sterilized by irradiation in Y-rays at a sterilization dose of 25 KGy.

Example 2

A 10% type III collagen solution was prepared by using 0.05% aqueous acetic acid as a solvent, and a 0.02% silk fibroin solution was prepared using a pure aqueous solution as a solvent. Then, 6 parts by mass of the type III collagen solution was taken, and while stirring at 100 rpm, 1 part by mass of the silk fibroin solution was added at a rate of 0.5 ml/min, and after the completion of the addition, stirring was continued to form a uniform composite liquid. The mass ratio of collagen to silk fibroin in the resulting composite solution was 3000:1.

 [0056] A volume of 5 parts by volume of a pure aqueous dispersion of gentamicin-loaded PLGA microspheres (2 g/L) was added to the composite solution, and the mixture was stirred and homogenized. The resulting composite solution was defoamed for 10 min under a vacuum of 10 Pa.

[0057] The defoamed composite liquid was weighed in a mold and then air-dried at room temperature (30 ° C), and the air-dried film was continuously pressed at a static pressure of 5 MPa for 168 hours to obtain a pressed film. [0058] The sterility of the resulting film is controlled by using a "process aseptic process."

Embodiment 3

A 0.5% V-type collagen solution was prepared by using 10% aqueous acetic acid as a solvent, and a 1% chondroitin sulfate solution was prepared by using 0.5% aqueous acetic acid as a solvent. Then, 4 parts by mass of the collagen solution was taken and stirred at 800 rpm, and 1 part by mass of chondroitin sulfate solution was added at a rate of 0.02 mL/min. After the addition was completed, stirring was continued to form a uniform composite liquid. The mass ratio of collagen to chondroitin sulfate in the obtained composite liquid was 2:1.

 [0060] In the above composite liquid, NaCl was added to make the mass ratio of the mass to the composite liquid 1/100, and after stirring uniformly, it was defoamed under a vacuum of 10 Pa for 5 minutes.

 [0061] Weigh the defoamed composite liquid in a mold and then freeze it in a -70 ° C refrigerator for 3 h, and then freeze in a freeze dryer (temperature first -45 ° C, 30 ° C, pressure 200 Pa) The dried, lyophilized sponge membrane was washed with water for 8 times to obtain a wet sponge-like material.

 [0062] It was crosslinked and sterilized by irradiation in Y rays at a dose of 15 KGy.

Example 4

A 0.1% VD-type collagen solution was prepared by using 5% aqueous acetic acid as a solvent, and a 1% hyaluronic acid solution was prepared using water as a solvent. Then, 4 parts by mass of the collagen solution was taken and stirred at 200 rpm, and 1 part by mass of hyaluronic acid solution was added at a rate of 5 mL/min. After the completion of the addition, stirring was continued to form a uniform composite liquid. The mass ratio of collagen to hyaluronic acid in the obtained composite liquid was 2:5.

[0064] The pure water dispersion of the PCL microspheres containing riiBMP-2 is added to the composite liquid to be uniformly mixed, so that the mass ratio of the active factor to the dry weight of the composite in the resulting composite is 1/5*10 ⁴ . The resulting composite solution was defoamed under vacuum at 12 Pa for 20 min.

 [0065] Weigh the defoamed composite liquid into a fully enclosed mold, and then freeze it in a refrigerator at -5 ° C for 24 h, then in a freeze dryer (temperature is -40 ° C after 45 ° C, pressure is 5 Pa ) lyophilized to obtain a sponge-like film.

 The film was crosslinked by a high temperature vacuum, the crosslinking temperature was 200 ° C, the crosslinking time was 2 h, and the vacuum pressure at the time of crosslinking was 0.1 Pa.

[0067] A solution of 0.01% glyoxal was prepared, and the film obtained above was immersed therein and immersed for 48 hours. Thereafter, after washing 5 times, a wet spongy material was obtained. Then, it was frozen at -150 ° C for 4 hours, and then lyophilized in a freeze dryer (temperature of -40 ° 45, 45 ° 〇, pressure: 5 Pa) to obtain a sponge-like film.

 [0068] Sterilization was carried out using ethylene gas sterilization using a gas sterilization method at a temperature of 60, a humidity of 50%, and a concentration of 600 mg/mL.

Example 5 A 0.2% type II collagen solution was prepared by using 0.5% aqueous hydrochloric acid as a solvent, and a 2% chitin solution was prepared by using 10% aqueous acetic acid as a solvent. Then, 2 parts by mass of the collagen solution was taken, and while stirring at 1000 rpm, 5 parts of a mass of chitin solution was added at a rate of 2 mL/min, and after the completion of the addition, stirring was continued to form a uniform composite liquid. The mass ratio of collagen to chitin in the obtained composite liquid was 2:50.

[0070] The resulting composite liquid was defoamed under vacuum of 20 Pa for 20 min.

 [0071] The composite liquid after defoaming is filtered (the pore diameter of the sieve is 1 mm), and then granulated, the granulation conditions: the flow rate of the pump head is 0.05 mL/min, the air flow of the pump head circulation selects air, and the pumping compound The droplets were placed in a hexane-containing collection tank, and the temperature of the collection tank was maintained at -40 °C. A wet gel particle composite is obtained after granulation.

[0072] It was sterilized by irradiation in Y rays at a sterilization dose of 25 KGy.

Example 6

A 0.01% type I collagen solution was prepared by using 3% aqueous hydrochloric acid as a solvent, and a 1.5% chondroitin sulfate solution was prepared by using 0.5% aqueous hydrochloric acid as a solvent. Then, 2 parts by mass of the collagen solution was taken and stirred at 450 rpm, and 5 parts by mass of chondroitin sulfate solution was added at a rate of 0.01 ml/min. After the completion of the addition, stirring was continued to form a uniform composite liquid. The mass ratio of collagen to chondroitin sulfate in the obtained composite solution was 1:350.

[0074] The pure aqueous dispersion of the PAF-loaded hyaluronic acid microspheres was added to the composite liquid to be stirred and mixed uniformly, so that the mass ratio of the active factor to the dry weight of the composite in the resulting composite was 1/10 ⁵ . The resulting composite solution was defoamed under lOPa vacuum for 20 min.

[0075] The composite liquid after defoaming is filtered (the pore size of the sieve is 1 μ ηι), and then granulated, the granulation conditions: the flow rate of the pump head is 200 mL/min, the gas flow of the pump head is selected, the helium gas is pumped out. The composite droplets were transferred to a collection tank containing octane, and the temperature of the collection tank was maintained at -2 °C. After granulation, the wet gel particles were frozen in a refrigerator at -65 °C for 3 h, and then lyophilized (the temperature at lyophilization was -30 ° C after 20 ° C, the pressure was lOPa). Particle complex.

[0076] The high energy electron beam sterilization was carried out at a sterilization dose of 50 KGy.

Example 7

Prepare 10% X IV collagen solution with 0.5% acetic acid aqueous solution as solvent, 0.12% alginic acid solution with 0.2% acetic acid aqueous solution as solvent, and 0.2% chondroitin sulfate solution with 2% acetic acid aqueous solution as solvent. . Then, 20 g of the collagen solution was taken and stirred at 500 rpm, and 1 part by mass of alginic acid solution was added at a rate of 2 ml/min. Then, while stirring at 800 rpm, 1 part by mass of chondroitin sulfate solution was added at a rate of 0.002 ml/min. Stirring was continued after the addition was completed to form a uniform composite liquid. The mass ratio of the obtained composite liquid was: collagen: alginic acid: chondroitin sulfate = 1000: 3: 5.

 [0078] The resulting composite liquid was defoamed under vacuum of 20 Pa for 20 min.

[0079] Weigh the defoamed composite liquid in a mold and then freeze it in a refrigerator at -50 ° C for 3 h, then in a freeze dryer (The temperature is -45 ° C after -45 ° C, pressure is 10 Pa) and lyophilized. The lyophilized sponge-like film was continuously pressed at a static pressure of 500 MPa for 10 hours to obtain a pressed film.

[0080] The crosslinking was then carried out by ultraviolet irradiation for 48 hours, and the irradiation intensity was 1000 mW/cm ² .

A carbodiimide solution having a concentration of 100 g/L was prepared, and the film obtained above was immersed therein and immersed for 30 hours. Thereafter, after washing with water for 5 times, a wet spongy material was obtained. It was sterilized by irradiation in Y-rays at a sterilization dose of 25 KGy.

Example 8

Prepare 0.6% type I collagen solution with 0.3% acetic acid aqueous solution as solvent, prepare 1% chitosan solution with 3% acetic acid aqueous solution as solvent, prepare 1% chondroitin sulfate solution with aqueous solution as solvent, take water as The solvent was formulated with a 1% hyaluronic acid solution. Then, 4 parts by mass of the collagen solution was taken, and while stirring at 500 rpm, 1 part of a mass chitosan solution was added at a rate of 2 mL/min. Then, while stirring at 800 rpm, 1 part by mass of chondroitin sulfate solution was added at a rate of 0.1 mL/min. Next, while stirring at 600 rpm, 1 part by mass of a hyaluronic acid solution was added at a rate of 1 ml/min. Stirring was continued after the addition was completed to form a uniform composite liquid. The mass ratio of the obtained composite liquid was, collagen: chitosan: chondroitin sulfate: hyaluronic acid = 12: 5: 5: 5.

 [0083] A pure aqueous dispersion (20 g/L) of 2 parts by mass of nano-titanium-loaded chitin microspheres was added to the composite liquid, and the mixture was stirred and uniformly mixed. The resulting composite solution was degassed under vacuum at 20 Pa for 20 min.

 [0084] Weigh the defoamed composite liquid in a mold and then freeze it in a refrigerator at -50 ° C for 3 h, and then in a freeze dryer (temperature is -45 ° C, 45 ° C, pressure 10 Pa) Freeze dried. The lyophilized sponge-like film was continuously pressed at a static pressure of 500 MPa for 10 hours to obtain a pressed film.

 A glutaraldehyde solution having a concentration of 0.5% was prepared, and the film obtained above was immersed therein and immersed for 168 hours. After washing with water for 6 times, a wet spongy material was obtained, which was then frozen in a refrigerator at -50 ° C for 3 h, and then lyophilized in a freeze dryer (temperature: -45 ° C, 45 ° C, pressure: 10 Pa). .

 [0086] This is sterilized by high energy electron beam irradiation at a dose of lKGy.

 [0087]

Example 9

A 10% silk fibroin solution was prepared by using water as a solvent, and a 1% chitosan solution was prepared by using 1% acetic acid aqueous solution as a solvent. Then, 2 parts of the silk fibroin solution was taken and stirred at 400 rpm, 5 parts of chitosan solution was added at a rate of 3 ml/min, and stirring was continued to form a uniform composite liquid. The mass ratio of collagen to chitosan in the obtained composite liquid was 4:1.

[0088] 4 ml of an aqueous dispersion of penicillin-containing alginate microspheres (40 g/L) was added to the composite solution and stirred and mixed uniformly. The resulting composite solution was defoamed under a vacuum of 20 Pa for 20 min. [0089] Weigh the defoamed composite liquid in a mold and then freeze it in a refrigerator at -50 ° C for 3 h, and then in a freeze dryer (temperature is -45 ° C, 45 ° C, pressure 10 Pa) Freeze dried. The lyophilized sponge-like film was continuously pressed at a static pressure of 500 MPa for 10 hours to obtain a pressed film.

 [0090] A carbodiimide solution having a concentration of 100 g/mL was prepared, and the film obtained above was immersed therein and immersed for 30 hours. After washing with water for 5 times, a wet spongy material was obtained.

 [0091] It was sterilized by irradiation in Y rays at a sterilization dose of 5 KGy.

 Example 10

A 0.01% type I collagen solution was prepared by using 0.5% aqueous acetic acid as a solvent.

 A pure aqueous dispersion of vanillin-loaded PHA microspheres (50 mg/ml) was added to the collagen solution and stirred and mixed uniformly. The resulting composite solution was degassed under vacuum at 20 Pa for 20 min.

 [0094] The prepared composite liquid was allowed to stand in a refrigerator at 4 ° C for 72 hours to obtain a composite gel.

 [0095] This was sterilized by a filter sterilization method, and the pores of the filtration membrane were 0.22 μm to obtain a final gel solution.

 [0096] While the embodiments of the present invention have been shown and described, it will be understood by those skilled in the art The scope of the invention is defined by the appended claims and their equivalents.

Claims

WO 2014/079198 Right and j request PCT/CN2013/074697 A degradable wound repairing material, characterized in that the raw material comprises a matrix component, the matrix component comprising a protein component, which is derived from collagen or derived from a mammalian skin or Achilles Silk fibroin or silk fibroin graft modified derivative.  2. The degradable wound repairing material according to claim 1, wherein the raw material of the degradable wound repairing material further comprises an auxiliary component, wherein the auxiliary component comprises at least one of an antibacterial agent and an active factor, wherein The antibacterial agent is a synthetic antibacterial agent, an inorganic antibacterial agent, an organic antibacterial agent or a natural antibacterial agent; the active factor is at least one of the following active factors: epidermal growth factor, vascular endothelial growth factor, platelet-derived growth factor, Platelet activating factor, insulin-like growth factor, tumor necrosis factor, interleukin, colony stimulating factor-1, various bone morphogenetic proteins or transforming growth factors. The degradable wound repairing material according to claim 1 or 2, wherein the matrix component further comprises a polysaccharide component, and the polysaccharide component comprises at least one of the following components: hyaluronic acid , chitin and chitosan derived from the arthropod shell, alginic acid or sodium alginate derived from seaweed or kelp or chondroitin sulfate extracted from organs of pig, cow, sheep or shark.  The degradable wound repairing material according to claim 3, wherein a mass ratio of the protein component to the sum of all other raw material components is 1/350 to 4000/1.  A method for preparing a degradable wound repairing material according to any one of claims 1 to 4, which comprises the following steps; Preparing a substrate material step, configuring a matrix component solution; In the molding step, the composite is subjected to a molding process to obtain a desired material form. The method for preparing a degradable wound repairing material according to claim 5, wherein in the step of preparing the base material, each base component solution is mixed under stirring conditions, and a belt as a base material is prepared by a pore-forming process. The composite of the pore structure of the stent, the pore-forming process being at least one of the following methods: particle leaching, stirring foaming or defoaming, initial concentration adjustment or complex concentration.  7. The method of preparing a degradable wound repairing material according to claim 5, wherein a composite auxiliary component step is further performed between the preparation of the base material and the molding step, and the antibacterial agent is loaded onto the composite with the support pore structure. And at least one of the active factors, wherein the antibacterial agent is loaded in any one of the following ways: grafting, physical blending, adsorption or microsphere entrapment, and the loading method of the active factor is Any of the following: grafting, physical blending, adsorption or microsphere entrapment. The method for preparing a degradable wound repairing material according to claim 5, wherein the molding step is performed by a flow film forming process or a film forming process or a granulation process, and further crosslinking is performed after the forming step. Step, after cross-linking The composite is washed and dried to obtain one of the following composites: a dry sponge, a wet sponge, a dry granule or a wet granule, and the cross-linking step employs a physical cross-linking process and cross-linking with a chemical cross-linking agent. a process or a cross-linking process involving natural cross-linking agents, wherein The physical crosslinking process is any one of the following processes: a vacuum high temperature crosslinking process, an ultraviolet irradiation process, a Y irradiation process, a high energy electron beam irradiation, and the like; The crosslinking agent used in the crosslinking process involving the chemical crosslinking agent is any one of the following components: an aldehyde crosslinking agent, an imide crosslinking agent or a diisocyanate crosslinking agent; The crosslinking agent used in the crosslinking process involving natural crosslinking agents is genipin.  9. The method for preparing a degradable wound repairing material according to claim 5, wherein the forming step is performed by a gel process or a compound solution dilution process to obtain an injectable gel or a sprayable solution for degradable wound repair. material.  The method for preparing a degradable wound repairing material according to claim 5 or 8 or 9, wherein the preparation method further comprises the step of sterilizing the obtained material as a final step of sterilizing the obtained material. Processing, obtaining the finished product, the terminal sterilization step adopts any one of the following sterilization processes: high energy electron beam irradiation process, Y radiation process, filtration process, gas sterilization process.