A regenerative wound repair membrane with antibacterial barrier function and a preparation method thereof

Abstract

The application discloses a wound repairing membrane with regeneration and bacteriostatic barrier function and a preparation method thereof, which comprises the following steps of preparing collagen with different molecular weights, preparing silk fibroin microspheres, preparing a film-forming solution and mixing to obtain the membrane; collagen with different molecular weights is obtained from bovine Achilles tendon and pig skin tissues through an acid enzyme method, and is compounded in a certain proportion; silk fibroin microsphere particles are used as microporous agents, and are prepared together with a cellulose film-forming solvent, so that beneficial substances can be fixed on the surface of a wound, and an isolation barrier membrane is formed in 30s to 60s at room temperature. The membrane has good tension, flexibility and compliance, has little foreign body sensation on the skin, has a broad-spectrum bacteriostatic agent in the composition, and greatly reduces the risk of delayed wound healing caused by wound infection.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of biomedical materials, in particular to a regenerative wound repair membrane with antibacterial barrier function and a preparation method thereof. BACKGROUND

[0002] Wound repair has always been a research hotspot in the medical field. In life, skin, as the largest and outermost organ of the human body, is extremely vulnerable to various physical injuries, such as abrasions, burns, scalds, chronic skin ulcers, etc. According to statistics, about 1.1 million people in the United States suffer from acute / chronic trauma every year, and about 300,000 people are hospitalized. Given the larger population base and more serious aging in China, this statistical number should be much higher than the statistical data in the United States.

[0003] Skin has a certain self-repair mechanism, and wound repair (or other organ tissues) is a complex dynamic process of self-repair through the interaction between various repair cells, growth factors and extracellular matrix after injury. In the current clinical consensus, it can be summarized into three stages: ① hemostasis and inflammatory reaction stage; ② cell proliferation and differentiation stage; ③ tissue reconstruction or scar formation stage.

[0004] At the same time, wound repair is affected by many factors (including local factors and systemic factors) ① foreign body. Foreign body is the most important local influencing factor for wound healing. Its bacteria, tissue toxicity and stimulation can all aggravate the inflammatory response and delay wound healing. ② infection. Bacterial toxins, proteolytic enzymes and cytotoxic effects in infected wounds can prolong the inflammatory response of the wound, thereby delaying wound healing. ③ oxygen. Early transient hypoxia after trauma can induce the initiation of the wound healing process, induce macrophages, keratinocytes and fibroblasts to synthesize and release cytokines and growth factors. However, the wound healing process requires oxygen to maintain, and long-term hypoxia can increase the chances of wound infection, hinder the release of various cytokines and growth factors of cells, etc., leading to delayed wound healing.

[0005] There are some collagen protein dressings on the market, which use recombinant collagen and some conventional dressing additives, such as sodium lactate, sodium hyaluronate, etc. They have no obvious pertinence in promoting wound repair and skin healing, and lack obvious internal connection in the above mechanism. Collagen itself has excellent cell activation performance and biocompatibility, and is abundant in animal tissues, so it is an excellent tissue repair raw material. However, it is difficult for the human body to effectively absorb collagen with a large molecular weight, so the molecular weight control is important when using collagen as a substrate.

[0006] Chinese patent application (publication number CN 116113448A) discloses a collagen-based gel / freeze-dried sponge for repairing skin wounds, but the pH value of the collagen matrix is generally 2-5. The slightly acidic repair environment still has great controversy on cell repair of wound tissue. Li Zihong et al. (publication number CN 109675085A) use I, III and V collagen proteins mixed with purified water in different proportions to prepare sponge, fibrous membrane and powder repair dressings in different forms. The use of collagen sponge and powder in such forms is inconvenient, the doctor's operation is complex, and the patient also has no aesthetic appearance. At the same time, the function of this invention is single, and it cannot solve the subsequent problems caused by infection during wound repair.

[0007] With the progress of society, people's demand for beauty is gradually increasing, and the current level and quality of wound repair cannot meet the patient's requirements, and the contradiction between the two is becoming increasingly acute. Therefore, further development of new treatment products is still a more urgent problem. SUMMARY

[0008] The purpose of the present application is to provide a regenerative wound repair membrane with antibacterial barrier function and a preparation method thereof to overcome the shortcomings of the prior art.

[0009] To achieve the above-mentioned purpose, the present application provides the following technical solutions:

[0010] The present application discloses a preparation method of a regenerative wound repair membrane with antibacterial barrier function, which specifically comprises the following steps:

[0011] S1, take bovine Achilles tendon tissue or pig skin tissue, and perform dehydration pretreatment; then perform enzymatic treatment for different lengths of time, and finally perform salting-out and dialysis treatment to obtain high molecular collagen protein with a molecular weight of 100KD-200KD and low molecular collagen protein with a molecular weight of 2KD-8KD;

[0012] S2, take silk fibroin and perform emulsification, freezing and ion water washing to obtain microsphere particles with a particle size of 5-100 microns;

[0013] S3, dissolve ethyl cellulose, glyceryl triethyl, and antibacterial agent according to a mass ratio of 50-40:1:0.005-0.01 to prepare a 5%-10% ethyl cellulose solution;

[0014] S4, mix the high molecular collagen protein and the low molecular collagen protein according to a mass ratio of 1:0.1-5 to obtain mixed collagen protein; then uniformly mix the mixed collagen protein, the microsphere particles and the ethyl cellulose solution according to a mass ratio of 1:2-10:50-80 to obtain the regenerative wound repair membrane with antibacterial barrier function.

[0015] As preferred, the specific operation of the dehydration pretreatment in step S1 is as follows: taking the Achilles tendon tissue or pigskin tissue, soaking in ethanol or sodium chloride for 16-20 hours at room temperature.

[0016] As preferred, the specific operation of the enzymatic treatment in step S1 is as follows: taking the Achilles tendon tissue or pigskin tissue, and performing enzymatic treatment with pepsin at a mass ratio of 1:0.05-0.1 at room temperature for 24-96 hours.

[0017] As preferred, the salting-out in step S1 specifically includes the following operation: adding sodium chloride to the enzymatically treated tissue protein, standing at room temperature for 2-4 hours, and then filtering, repeating 4-5 times; the dialysis specifically includes the following operation: placing the salting-out tissue protein into a dialysis bag, and changing the water every 6-12 hours, lasting for 3-5 days.

[0018] As preferred, the specific operation of the emulsification in step S2 is as follows: taking the silk fibroin, and emulsifying with polyethylene glycol for 1-3 hours.

[0019] As preferred, the freezing temperature in step S2 is-40℃ to-10℃.

[0020] As preferred, the bacteriostatic agent is one or a combination of the two of polyvidone iodine and benzalkonium chloride.

[0021] The application also discloses a regenerated and bacteriostatic barrier functional wound repair membrane prepared by the above method.

[0022] The application has the following advantages:

[0023] The application uses Achilles tendon and pigskin tissue to obtain collagen proteins with different molecular weights by acid enzyme method, and the collagen proteins are compounded in a certain proportion, combined with silk fibroin microsphere particles as a micropore agent, and then prepared with a cellulose film-forming solvent, so that beneficial substances can be fixed on the surface of a wound, and an isolation barrier film is formed in 30-60 seconds at room temperature. The film has good tension and flexibility, and has little foreign body sensation on the skin. The film contains a broad-spectrum bacteriostatic agent, which greatly reduces the risk of delayed wound healing caused by wound infection.

[0024] The collagen proteins with different molecular weights are main substrates, which can be degraded in a gradient within a certain time, small molecular collagen is easily absorbed by the human epidermis, and the collagen protein molecules can lock the moisture of the wound, maintain a slightly humid environment, and provide a good repair environment for cells. The collagen protein has excellent biocompatibility, and can be converted into various amino acids after degradation, which transports nutrient substances for cell proliferation and differentiation.

[0025] The silk fibroin microsphere particle makes the isolation film not a completely closed barrier film, but has a certain micron-level pore structure, maintains oxygen transport with the outside world, and guarantees the required oxygen for cell growth;

[0026] The ethyl cellulose solution as a film-forming solvent is transparent after being sprayed, forms a film quickly, can effectively isolate the wound surface from the external environment, and the bacteriostatic agent component can cooperate with the silk fibroin to resist bacteria, reduce the risk of wound infection, and accelerate tissue healing.

[0027] The features and advantages of the present application will be described in detail by combining the embodiments with the drawings. BRIEF DESCRIPTION OF DRAWINGS

[0028] Figure 1 is an HE staining diagram in the wound repair process of the rat back skin fascia;

[0029] Figure 2 is an electron microscope diagram of the wound repair film; DETAILED DESCRIPTION

[0030] In order to make the purpose, technical scheme and advantages of the present application more clear and obvious, the present application will be further described in detail by combining the drawings and examples. However, it should be understood that the specific examples described here are only used to explain the present application, and are not used to limit the scope of the present application. In addition, in the following description, the description of the known structures and technologies is omitted to avoid unnecessary confusion of the concept of the present application.

[0031] Example 1:

[0032] S1 Preparation of different molecular weight collagens: take pig skin tissue, soak in ethanol at room temperature for 16h for dehydration, then enzymolysis with pepsin at room temperature for 24h according to the mass ratio of 1:0.1; add sodium chloride for salting-out, stand at room temperature for 4h, then filter, repeat 4 times; then put the salting-out tissue protein into a dialysis bag for dialysis, change water every 6 hours for 3 days; obtain collagen with a molecular weight of 120KD~180KD; after enzymolysis for 96h, the subsequent steps are carried out in the same way, and collagen with a molecular weight of 4KD~6KD is obtained;

[0033] S2 Preparation of silk fibroin microspheres: commercially available silk fibroin powder is emulsified with polyethylene glycol for 2h, placed in a-20℃ refrigerator for freezing, and after the end, washed with deionized water for 3 times to obtain silk fibroin microspheres with an average particle size of 50 microns;

[0034] S3 Preparation of film-forming solution: dissolve ethyl cellulose, glyceryl triacetate and povidone-iodine according to the mass ratio of 50:1:0.005 to prepare a 5% ethyl cellulose solution;

[0035] S4 Mixing: high and low molecular weight collagen were mixed in a mass ratio of 1:0.1, and then mixed with silk fibroin particles and film-forming solution in a mass ratio of 1:2:50 using a homogenizer;

[0036] The solution was bottled and sterilized by irradiation to obtain a regenerated and antibacterial barrier function wound repair membrane. It was sprayed on fresh pig skin and formed a film in 40 seconds. After peeling off, scanning electron microscopy, mechanical testing, and air permeability testing were performed;

[0037] The tensile strength was 2.75 Mpa and the elongation at break was 11.4% by universal testing machine.

[0038] Air permeability test: CaCl2 was added to the cup, the film was packaged at the cup opening, and the sample assembly was placed upright in a stable environment at 37°C and RH of 80%. The weight was measured every 1h, and the formula APR=(24m / s t) 100% calculated air permeability, m is the difference between two weightings of the same sample, s is the sample area, and t is the test time. The average air permeability was 33.5%.

[0039] The antibacterial test used E. coli and S. aureus as direct inoculation, with 3 parallel samples per experimental sample, and the antibacterial rate reached 85%.

[0040] Enzymatic degradation was performed in vitro using collagenase (20 units / mL), with an average degradation time of 556±6.79 min.

[0041] Example 2:

[0042] S1 Preparation of collagen proteins with different molecular weights: The bovine Achilles tendon tissue was soaked in ethanol for 18h at room temperature for dehydration, then enzymolysis was performed at room temperature for 48h with a mass ratio of tissue to pepsin of 1:0.08. Sodium chloride was added for salting-out, and after standing at room temperature for 3h, filtration was performed, repeated 4 times. Then it was loaded into a dialysis bag, and the water was changed every 8 hours for 4 days to obtain collagen proteins with a molecular weight of 100KD~180KD. The remaining parameters were unchanged, and after enzymolysis for 72h, the subsequent steps were performed in the same way to obtain collagen proteins with a molecular weight of 4KD~8KD;

[0043] S2 Preparation of silk fibroin microspheres: commercially available silk fibroin powder was emulsified with polyethylene glycol for 1h, frozen in a-10°C refrigerator for 8h, and washed with deionized water for 3 times to obtain silk fibroin microspheres with an average particle size of 100 microns;

[0044] S3 Preparation of film-forming solution: ethyl cellulose, glyceryl triethylhexanoate, and povidone-iodine were dissolved in a mass ratio of 50:1:0.008 to prepare a 10% ethyl cellulose solution;

[0045] S4 Mixing: high and low molecular weight collagen were mixed in a mass ratio of 1:1, and then mixed with silk fibroin microspheres and film-forming solution in a mass ratio of 1:5:80 using a homogenizer;

[0046] The solution was bottled and sterilized by irradiation to obtain a regenerated and antibacterial barrier function wound repair membrane. It was sprayed on fresh pig skin and formed a film in 48 seconds. After peeling off, scanning electron microscopy, mechanical testing, and air permeability testing were performed;

[0047] The tensile strength was 1.99 Mpa and the elongation at break was 12.7% as tested by a universal testing machine.

[0048] Air permeability test: The cup was filled with anhydrous CaCl2, the film was packaged at the cup opening, and the sample assembly was placed upright in a stable environment at 37°C and RH of 80%. The weight was measured every 1h. The average air permeability was 37.5%.

[0049] The antibacterial test used E. coli and S. aureus as direct inoculation, with 3 parallel samples per experimental sample, and the antibacterial rate reached 91%.

[0050] Enzymatic degradation was performed in vitro using collagenase (20 units / mL), and the average degradation time was 428 ± 4.17 min.

[0051] Example 3:

[0052] S1 Preparation of collagen with different molecular weights: The pig skin tissue was soaked in sodium chloride at room temperature for 20h to dehydrate, then the tissue was digested with pepsin at room temperature for 24h at a mass ratio of 1:0.1. Sodium chloride was added for salting-out, and the mixture was filtered after standing at room temperature for 2h. The process was repeated 5 times, and the collagen with a molecular weight of 120KD~160KD was obtained by dialysis for 5 days with water changed every 10 hours. The remaining parameters were unchanged, and the collagen with a molecular weight of 4KD~8KD was obtained by following the same procedure after 72h of enzymatic digestion.

[0053] S2 Preparation of silk fibroin microspheres: Commercially available silk fibroin powder was emulsified with polyethylene glycol for 3h and frozen at -40°C. After washing with deionized water for 3 times, silk fibroin microspheres with an average particle size of 5 microns were obtained.

[0054] S3 Preparation of film-forming solution: Ethyl cellulose, glyceryl triethylhexanoate, and benzalkonium chloride were dissolved in a mass ratio of 50:1:0.01 to prepare a 6% ethyl cellulose solution.

[0055] S4 Mixing: high and low molecular weight collagen were mixed in a mass ratio of 1:2, and then mixed with silk fibroin particles and film-forming solution in a mass ratio of 1:4:60 using a homogenizer;

[0056] The solution is bottled, and after irradiation sterilization, a regenerated and bacteriostatic wound repair film is obtained. It is sprayed on fresh pig skin and forms a film within 45 seconds; after peeling, electron microscopy scanning, mechanical testing, and air permeability testing are performed;

[0057] The tensile strength is 2.26 Mpa and the elongation at break is 11.8% by universal testing machine testing;

[0058] Air permeability test: the cup is filled with anhydrous CaCl2, the film is packaged at the cup opening, and the sample assembly is placed vertically in a stable environment at 37°C and RH of 80%. The weight is measured every 1h. The average air permeability is 36.9%.

[0059] The bacteriostatic test uses E. coli and S. aureus as direct inoculation, and each experimental sample is 3 parallel samples, with a bacteriostatic rate of 94%;

[0060] Enzymatic degradation experiment in vitro using collagenase (20 units / mL) with an average degradation time of 494±5.82min.

[0061] Example 4:

[0062] S1 Preparation of collagen with different molecular weights: The bovine Achilles tendon tissue is soaked in sodium chloride for 16h at room temperature for dehydration, and then enzymolysis is carried out at room temperature for 36h with a mass ratio of tissue to pepsin of 1:0.05. Sodium chloride is added for salting-out, and after standing at room temperature for 3h, filtration is repeated 4 times, and then the dialysis bag is placed for dialysis, and the water is changed every 12 hours for 5 days to obtain collagen with a molecular weight of 120KD~160KD. The remaining parameters remain unchanged, and after enzymolysis for 96h, the subsequent steps are carried out in the same way to obtain collagen with a molecular weight of 4KD~8KD;

[0063] S2 Preparation of silk fibroin microspheres: commercially available silk fibroin powder is emulsified with polyethylene glycol for 2h, and then frozen in a-25℃ refrigerator. After washing with deionized water for 3 times, silk fibroin microspheres with an average particle size of 35 microns are obtained;

[0064] S3 Preparation of film-forming solution: ethyl cellulose, glyceryl triethylhexanoate, and benzalkonium chloride are dissolved according to a mass ratio of 50:1:0.008 to prepare a 5% ethyl cellulose solution;

[0065] S4 Mixing: high and low molecular weight collagens are mixed according to a mass ratio of 1:5, and then the mixture is mixed with silk fibroin microspheres and film-forming solution according to a mass ratio of 1:10:80 using a homogenizer.

[0066] The solution is bottled, and after irradiation sterilization, a regenerative and antibacterial barrier wound repair membrane is obtained. It is sprayed on fresh pigskin and forms a film within 40 seconds. After peeling off, scanning electron microscopy, mechanical testing, and air permeability testing are performed.

[0067] The tensile strength is 1.54 Mpa and the elongation at break is 13.1% by universal testing machine.

[0068] Air permeability test: The cup is filled with anhydrous CaCl2, the film is packaged at the cup opening, and the sample assembly is placed upright in a stable environment at 37°C and RH of 80%. The weight is measured every 1h. The average air permeability is 38.2%.

[0069] The antibacterial test uses E. coli and S. aureus as direct inoculation, with 3 parallel samples per experimental sample, and the antibacterial rate reaches 92%.

[0070] Enzymatic degradation experiment in vitro using collagenase (20 units / mL) with an average degradation time of 407 ± 7.19 min.

[0071] The mechanical properties, air permeability, antibacterial rate, and in vitro degradation experiment of the above different groups are summarized in Table 1

[0072]

[0073] Table 1

[0074] In the four groups of examples, the mechanical properties, air permeability, and antibacterial rate of the regenerative and antibacterial barrier wound repair membrane can meet the clinical use requirements. According to the severity of the wound, the ratio of different collagen molecular weights can be adjusted to control the degradation time of the repair membrane. For example, if the wound is more serious, the product needs to have high antibacterial properties to kill bacteria deeply and provide oxygen supply and other conditions.

[0075] Comparative Example 1:

[0076] In Example 1, the collagen proteins are all large molecular weight and are not mixed with small molecular weight proteins.

[0077] S1 Preparation of collagen proteins with different molecular weights: The pigskin tissue is soaked in ethanol at room temperature for 16h to dehydrate, then enzymolysis with pepsin at room temperature for 24h at a mass ratio of 1:0.1. Sodium chloride is added for salting-out, and the mixture is left to stand at room temperature for 4h, then filtered and repeated 4 times. The salting-out tissue protein is then placed in a dialysis bag and dialyzed, with water changed every 6 hours for 3 days. Collagen proteins with a molecular weight of 120KD-180KD are obtained.

[0078] S2 Preparation of silk fibroin microspheres: Commercially available silk fibroin powder was emulsified with polyethylene glycol for 2 hours, then frozen at -20°C. After freezing, the microspheres were rinsed three times with deionized water to obtain silk fibroin microspheres with an average particle size of 50µm.

[0079] S3 film-forming solution preparation: Ethyl cellulose, triethyl glycerol, and povidone-iodine are dissolved in a mass ratio of 50:1:0.005 to prepare a 5% ethyl cellulose solution;

[0080] S4 Mixing: Mix high molecular weight collagen with silk fibroin microspheres and film-forming solution at a mass ratio of 1:2:50 using a homogenizer until homogenized.

[0081] The solution was bottled and sterilized by irradiation to obtain a regenerative wound repair membrane with antibacterial barrier function. When sprayed onto fresh pigskin, it formed a film within 50 seconds; after peeling, electron microscopy, mechanical testing, and air permeability testing were performed.

[0082] According to the universal testing machine, its tensile strength is 3.62 MPa and its elongation at break is 10.21%.

[0083] Air permeability test: Anhydrous CaCl2 was placed inside the cup, the membrane was sealed at the mouth of the cup, and the sample assembly was placed upright in a stable environment of 37°C and 80% RH. The weight was measured every 1 hour. The average air permeability was 34.6%.

[0084] The antibacterial test used two bacteria, Escherichia coli and Staphylococcus aureus, for direct inoculation. Each experimental sample consisted of 3 parallel samples, and the antibacterial rate reached 86%.

[0085] Enzymatic hydrolysis experiments were conducted in vitro using collagenase (20 units / mL), and the average degradation time was 672±6.86 min.

[0086] Compared to Example 1, the mechanical properties showed a slight increase, but in the in vitro degradation test by collagenase, the degradation time of collagen was significantly prolonged. See also... Figure 1 Figure A shows the wound staining results of Example 1, and Figure B shows the wound staining results of Comparative Example 1. Figure A shows that after spraying the sample of Example 1 onto the rat's back wound, a new stratum corneum was formed within one week, fibroblasts were dispersed, capillaries were abundant, the epidermis of the skin tissue was intact, and a new basal layer covered with a scab was present on the wound. From the attached... Figure 1 As shown in Figure B, after spraying the rat's back with the sample from Comparative Example 1, collagen accumulation formed on the skin surface. A small number of capillaries and a large number of fibroblasts were visible in the skin tissue, accompanied by a large number of inflammatory cell infiltrations. A large number of vacuoles of different shapes and sizes were visible in the tissue, and the scarring on the body surface was aggravated.

[0087] Comparative Example 2:

[0088] In the steps of Example 2, the silk fibroin microsphere component was removed.

[0089] S1 Preparation of collagen with different molecular weights: Bovine Achilles tendon tissue was dehydrated by soaking in ethanol at room temperature for 18 hours, then enzymatically hydrolyzed at room temperature for 48 hours at a tissue-to-pepsin mass ratio of 1:0.08; sodium chloride was added for salting out, and the mixture was allowed to stand at room temperature for 3 hours before filtration. This process was repeated 4 times. The tissue was then placed in a dialysis bag, and the water was changed every 8 hours for 4 days to obtain collagen with a molecular weight of 100KD~180KD. With other parameters unchanged, enzymatic hydrolysis was performed for 72 hours, and subsequent steps were carried out in the same way to obtain collagen with a molecular weight of 4KD~8KD.

[0090] S2 film-forming solution preparation: Ethyl cellulose, triethyl glycerol, and povidone-iodine are dissolved in a mass ratio of 50:1:0.008 to prepare a 10% ethyl cellulose solution.

[0091] S3 Mixing: Mix high and low molecular weight collagen at a mass ratio of 1:1, then mix it with the film-forming solution at a mass ratio of 1:80 using a homogenizer until homogeneous.

[0092] The solution was bottled and sterilized by irradiation to obtain a regenerative wound repair membrane with antibacterial barrier function. It was sprayed onto fresh pigskin, forming a film within 40 seconds; after peeling, electron microscopy, mechanical testing, and air permeability testing were performed.

[0093] According to the universal testing machine, its tensile strength is 3.47 MPa and its elongation at break is 11.38%.

[0094] Air permeability test: Anhydrous CaCl2 was placed inside the cup, the membrane was sealed at the mouth of the cup, and the sample assembly was placed upright in a stable environment at 37℃ and 80% RH. The weight was measured every 1 hour. The average air permeability was 12.6%.

[0095] The antibacterial test used two bacteria, Escherichia coli and Staphylococcus aureus, for direct inoculation. Each experimental sample consisted of 3 parallel samples, and the antibacterial rate reached 76%.

[0096] Enzymatic hydrolysis experiments were conducted in vitro using collagenase (20 units / mL), and the average degradation time was 467±4.49 min.

[0097] like Figure 2 As shown, C is an electron micrograph of the sample in Example 2, and D is an electron micrograph of the sample in Comparative Example 2. Compared with the sample in Example 2, the sample lacks silk fibroin microspheres. After film formation, the structure is more compact, and the air permeability is reduced by more than 50% compared with other examples. The skin is in a hypoxic state for a longer period of time, so cell respiration is hindered, growth is slow, and wound healing is delayed.

[0098] The above description is only the preferred embodiment of the present application, and is not intended to limit the present application. Any modification, equivalent replacement or improvement made within the spirit and principle of the present application should be included in the protection scope of the present application.

Claims

1. A method for preparing a wound repair membrane that is regenerated and has a bacteriostatic barrier function, characterized by, Specifically comprising the following steps: S1, taking the bovine Achilles tendon tissue or pig skin tissue, and performing dehydration pretreatment; then performing enzymatic treatment for different lengths of time, and finally performing salting-out and dialysis treatment to obtain high-molecular collagen with a molecular weight of 100KD-200KD and low-molecular collagen with a molecular weight of 2KD-8KD; S2, taking silk fibroin to perform emulsification, freezing and ion water washing to obtain microspheres with a particle size of 5-100 microns; S3, dissolving ethyl cellulose, glyceryl triethylhexanoate and bacteriostatic agent according to a mass ratio of 50-40:1:0.005-0.01 to prepare a 5%-10% ethyl cellulose solution; S4, mixing the high-molecular collagen and the low-molecular collagen according to a mass ratio of 1:0.1-5 to obtain mixed collagen; and then uniformly mixing the mixed collagen, the microspheres and the ethyl cellulose solution according to a mass ratio of 1:2-10:50-80 to obtain the regenerated wound repair membrane with a bacteriostatic barrier function.

2. The method for preparing a regenerable wound repair membrane with antibacterial barrier function as described in claim 1, characterized in that, The specific operation of the dehydration pretreatment in step S1 is as follows: taking the bovine Achilles tendon tissue or pig skin tissue, and immersing it in ethanol or sodium chloride at room temperature for 16-20h.

3. The method for preparing a regenerable wound repair membrane with antibacterial barrier function as described in claim 1, characterized in that, The specific operation of the enzymatic treatment in step S1 is as follows: taking the bovine Achilles tendon tissue or pig skin tissue and pepsin according to a mass ratio of 1:0.05-0.1, and performing enzymatic treatment at room temperature for 24-96h.

4. The method for preparing a regenerable wound repair membrane with antibacterial barrier function as described in claim 1, characterized in that, The salting-out in step S1 specifically comprises the following operations: taking the tissue protein after enzymatic treatment, adding sodium chloride, and standing at room temperature for 2-4h before filtering, and repeating 4-5 times; and the dialysis specifically comprises the following operations: placing the tissue protein after salting-out into a dialysis bag, and changing the water every 6-12h for 3-5 days.

5. The method for preparing a regenerable wound repair membrane with antibacterial barrier function as described in claim 1, characterized in that, The specific operation of the emulsification in step S2 is as follows: taking silk fibroin, and emulsifying it with polyethylene glycol for 1-3h.

6. The method for preparing a regenerable wound repair membrane with antibacterial barrier function as described in claim 1, characterized in that, The freezing temperature in step S2 is-40℃--10℃.

7. The method for preparing a regenerable wound repair membrane with antibacterial barrier function as described in claim 1, characterized in that, The bacteriostatic agent is one or a combination of the other of polyvidone iodine and benzalkonium chloride.

8. A wound healing membrane which is regenerated and has a bacteriostatic barrier function, characterized by, The wound repair membrane is prepared by the preparation method of the regenerated wound repair membrane with a bacteriostatic barrier function according to any one of claims 1-7.

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