Polymer bioprotein natural silk fibroin repair liquid for covering deep III-III-degree wound surface and preparation method of polymer bioprotein natural silk fibroin repair liquid

Abstract

The invention belongs to the field of biomedical materials. The method is applied to biological medicine and traditional Chinese medicine. The polymer bioprotein liquid coating is prepared by taking natural silk fibroin with natural human protein as a base material and adding 2-roots alcohol solute and edible plant light oil, is used for repairing deep III-III-degree wounds, and is used for covering and sealing the wounds, easing pain, resisting bacteria and diminishing inflammation. Due to the action of high-content glycine, alanine and serine in the natural silk fibroin, the cells are activated quickly, the wound surface is repaired quickly, and the healing is comprehensive. Neonatal skin and skin appendages (hair, hair follicles, sebaceous glands and sweat glands) are well recovered, and the wound appearance is comfortable. The problems that in the prior art, survival is difficult and the skin repairing effect is poor due to exclusiveness and incompatibility when the wound is repaired through homogeneous or heterogeneous skin transplantation or other skin substitutes are solved. The purpose of solving the medical problem is achieved.

Description

Technical Field

[0001] The present invention belongs to the field of biomedical materials and is applied to biomedicine and traditional Chinese medicine. Specifically, it provides a substrate and a preparation method of a polymer bio-protein tussah fibroin repair solution for covering deep degree III - III wounds. Background Art

[0002] The repair of deep wounds is an important medical and social issue. Because deep wounds are a special type of wounds with relatively high incidence and disability rates, and the fundamental problem is the wound repair problem. If the wound surface cannot be covered and sealed in a timely and effective manner, it will surely cause disorders of the patient's internal circulation, infection, and multiple organ dysfunction. Non-physiological wound healing will also occur, resulting in the formation of keloids and exacerbating the degree of disability. Therefore, it has always been the goal of the biological field and the medical community to strive to find a polymer bio-protein pharmaceutical substrate that can meet the clinical needs for repairing deep degree III - III wounds to replace the need for transplanting autologous skin, allogeneic (species) skin, plant polymers, or synthetic polymers for wound matrix repair.

[0003] The skin is the largest organ of the human body, which is composed of the epidermis, dermis, subcutaneous tissue, etc. Skin appendages have important functions and roles in protecting life and health. When skin wounds caused by various reasons such as physical, chemical, and biological factors involve the dermis in daily life, it is generally very difficult to restore the original shape and function of the skin.

[0004] Currently, the main method for treating dermal wounds is to transplant split-thickness autologous skin grafts. The prominent problems are that if large-area wounds occur, autologous skin grafts cannot meet the needs of the wound surface, and the shortage of skin will threaten the patient's life; secondly, it may cause secondary or multiple pains to the patient; thirdly, although it is autologous skin grafts, due to the split-thickness skin grafts, it is very difficult to anastomose with the cells of the wound cross-section, resulting in scar hyperplasia in the recipient area and affecting the appearance and functional use.

[0005] Another method for treating skin wounds is to cover them with skin substitutes. Currently, there are two types of skin substitutes on the market, namely "wound dressings" and "artificial skin". One type is temporary, such as regenerated fibroin membranes, chitosan membranes, polyurethane membranes, split-thickness pigskins, etc.; the other type is permanent, such as defatted cellular allogeneic dermis, defatted cellular pigskin, collagen, chitosan porous materials, etc. Temporary skin substitutes can generally prevent a large amount of body fluid loss and bacterial invasion to a certain extent. After the wound heals, they need to be removed or fall off naturally, and they are generally only suitable for the repair of shallow epidermal defect wounds. Permanent skin substitutes can only cover the wound surface to a certain extent. Before the emergence of the present invention, many research institutions (researchers) at home and abroad have been committed to the research of permanent skin substitutes or wound repair coatings and made some progress. For example, according to the technical solution of the Chinese patent "Cross-linked Porcine Acellular Dermal Sheet" with the publication number of CN 1266716, white pigs are selected, slaughtered, washed and dehaired, and the full-thickness skin pieces are peeled off. After removing fat, skin pieces about 0.1 - 0.4 mm thick are taken, immersed in a composite digestive solution containing 0.25% trypsin at 4°C for 16 - 24 hours, the epidermis is removed, and after punching holes in the dermis reticulum, it is cross-linked with glutaraldehyde or a formaldehyde-ethanol mixed solution to obtain xenogeneic skin for wound repair. However, due to the immune rejection reaction of allografts, its application and curative effect have been greatly restricted.

[0006] According to the technology adopted in the Chinese patent "Medical Collagen Sponge and Its Preparation Method" with the publication number of CN 1416907, the animal tissue rich in collagen is removed of impurities and crushed, washed, degreased, dissolved, treated with protease, precipitated, dissolved in acetic acid, freeze-dried, and then fixed to obtain a medical collagen sponge for wound repair and hemostasis.

[0007] According to the invention patent "A Porous Silk Fibroin Membrane and Its Preparation Method" with the announcement number of CN1118518C, the technology adopted is to degum, dissolve, dialyze, and concentrate silk, add an additive containing 10 - 60% of R(OH)n, 10 - 60% of RCH(NH 2 )COOH, and the rest is a cross-linking agent, and then processed into a porous silk fibroin membrane by freeze-drying. When using the porous silk fibroin membrane, it is still necessary to cover it with autologous skin or allogeneic or xenogeneic skin, otherwise the porous silk fibroin membrane is prone to dry up due to water shortage.

[0008] According to the skin substitute described in the Chinese patent "A Skin Substitute and Its Preparation Method" with the publication number of CN 100342924C, the skin substitute is used as a scaffold for dermal regeneration to cover the wound surface, creating an environment for the repair of dermal morphology and function. After blood vessels grow into the lower layer of the skin substitute, the upper and middle layers are removed, and the patient's surface is transplanted and covered, so as to achieve the purpose of deep wound repair.

[0009] In summary, for the "artificial skin" or skin substitutes currently used for repairing dermal defects in deep degree III - III burn wounds, the problems that still need to be solved are as follows: (1) Usually, when transplanting such skin substitutes, it is necessary to transplant autologous or allogeneic (species) skin for coverage at the same time. Otherwise, it is easy to dry out due to water shortage. And the skin donor sites are seriously insufficient for large - area trauma at one time; (2) Using defatted cellular allogeneic (species) dermis to repair wounds has great rejection and insecurity; (3) Specific types of collagen can be used to prepare permanent skin substitutes, which also cover the wounds. And the purification process of specific types of collagen is rather cumbersome. Although the manufacturing cost of the substitutes can be reduced, the effect is still not satisfactory; (4) As mentioned above, due to the fact that therapeutic drugs for nutritional moisturizing, analgesia, antibacterial, anti - inflammatory, and anti - swelling, as well as human proteins needed to supplement the wounds, cannot quickly penetrate into the wounds for degradation, absorption, promoting granulation and moisturizing the skin, the treatment is affected. As a result, autologous skin and skin appendages (hair, hair follicles, sebaceous glands, sweat glands) cannot grow, causing functional defects in the damaged part of the wound surface.

[0010] It must be admitted that all the above - mentioned methods have made arduous and positive efforts and contributions of love to repair traumatic skin. Summary of the Invention

[0011] Aiming at the deficiencies existing in the prior art, the present invention provides a high - molecular biological protein tussah silk fibroin repair solution for covering deep degree III - III burn wounds and a preparation method thereof. This method uses a biological material with a natural human protein structure - tussah silk fibroin. This material with a protein structure is non - toxic and harmless (Guangdong Provincial Microbial Analysis and Testing Center, Report No.: 2019SP7391R01), has good biological activity, and a relatively high content of glycine, alanine, and serine (Jinan Medical Device Quality Supervision and Inspection Center of the State Food and Drug Administration, Report No.: S2020011512). When used to cover and seal the wound surface, it can quickly penetrate into the wound muscle, timely supplement the nutrients urgently needed for the trauma, as well as therapeutic drugs such as analgesia, antibacterial, anti - inflammatory, and anti - swelling, enabling the collagen and cells in the dermis to have the correct orientation. It activates the growth of injured muscle cells, repairs the wound matrix, promotes wound healing, realizes the repair of deep degree III - III dermal defect wounds and the regeneration of skin appendages, and makes the wound appearance comfortable.

[0012] To achieve the above - mentioned purpose, the materials used in the present invention have the following characteristics: 1. The high - molecular biological protein tussah silk fibroin material is derived from the cocoons of tussah silkworms in the Tussah Science and Technology Research Center Farm in Tongdao Dong Autonomous County. The functions of tussah silk fibroin protein in the present invention are: supplying energy to the wound surface, and various peptides decomposed can participate in the body's immune regulation, promote the absorption of minerals, and manufacture meat, blood, skin, etc. See Table (1) before the structural information of the tussah silkworm cocoon silk protein in the present invention.

[0013] 2. The added 2 - borneol solute, common name: borneol, chemical formula C10 H 18 O. Ancient medical pharmacopoeias such as *Newly Revised Materia Medica*, *Compendium of Materia Medica*, *Annotations on the Classic of Materia Medica*, and *Compendium of Medical Sciences* have recorded its good curative effects. Through modern means of detection, 2-camphol has components such as "humulene, β-elemene, caryophyllene, oleanolic acid, asiatic acid", dipterocarpol, hydroxydarenonell, dryobalanone, erythrodiol, etc. These components are the characteristics of 2-camphol solute in this technology, and they lay a solid foundation for the unique effect of being quickly and evenly absorbed through mucous membranes and subcutaneous tissues after being fused with the macromolecular biological protein fibroin in light oil. Its miraculous functions such as analgesic and antiseptic, antibacterial and anti-inflammatory, clearing heat and promoting granulation have remarkable effects when applied to the preparation method of the present invention.

[0014] 3. Using edible plant light oil that is both a solute and a solvent endows the active ingredients in the dosage form with enhanced, solubilizing, and cosolubilizing effects, and can quickly and conveniently penetrate the nutrients required for wounds and the drugs for treating and repairing wounds into muscle cells, playing a role in lubricating and nourishing the skin, maintaining the original correct orientation of collagen and cell tissues in the dermis, promoting the uniform and rapid degradation and absorption of subcutaneous tissues, and improving the quality of skin repair. When applied to the preparation method of the present invention, edible plant light oil also has the following characteristics: (1) Edible plant light oil has extremely strong moisturizing properties, which can prevent the skin from drying and make it soft and supple; (2) Edible plant light oil has antibacterial and antiviral properties, which can prevent skin infections; its anti-inflammatory properties help relieve irritation and itching caused by specific skin conditions.

[0015] (3) The protective barrier formed by edible plant light oil on the skin can prevent ultraviolet damage, detoxify the skin, and promote skin regeneration.

[0016] (4) The rich vitamins and minerals in edible plant light oil can promote wound growth. In particular, the healing properties of vitamin K help the wound recover its blood coagulation function faster, improve blood circulation, and promote the healing and health of new skin and skin appendages (hair, hair follicles, sebaceous glands, sweat glands).

[0017] 4. The preparation method of a macromolecular biological protein fibroin repair solution for covering deep degree III - III wounds described in the above technical solution is characterized by: (1) After the tussah cocoons are washed, soaked, and cleaned, they are washed three times with distilled water. Then they are fished out and placed in a nylon mesh tray in a 1m×2m stainless steel enclosure, and naturally dried to extract fibroin for standby.

[0018] (2) Preparation of an edible plant light oil composite solution with a total concentration of 99.8 - 98.00%, where the ratio of each solute is: 1.0 - 6% borneol: 98.80 - 92.00% edible plant light oil. Then stir. Stirring time: 10 - 15 minutes. Name of stirring equipment: overhead stirrer; Model: AS20; Display type: LED digital tube; Rotation speed range (RPM): 30 - 2000; Maximum viscosity (mPAS): 10000.

[0019] (3) Weigh the silk fibroin according to the ratio and immerse it in the above composite light oil solution. The temperature of the light oil solution is 5°C - 18°C, and the immersion time is more than 5 minutes. Then stir with the same type of equipment as described above. Stirring time: 10 - 40 minutes.

[0020] (4) According to the preparation method of the tussah silk fibroin repair liquid described in claim 6, it is characterized in that: before step (3), step (2) needs to be repeated for stirring 1 - 3 times. To quickly dissolve the borneol solute into the plant light oil.

[0021] Compared with the prior art, the high - molecular biological protein tussah silk fibroin repair liquid of the present invention has the following remarkable advantages: 1. The high - molecular biological protein tussah silk fibroin repair liquid has created a new type of high - molecular active biological protein tussah silk fibroin composite coating and preparation method; 2. The high - molecular biological protein tussah silk fibroin repair liquid can meet the substrate characteristics of biological proteins required for modern wound surface coating repair; 3. The high - molecular biological protein tussah silk fibroin repair liquid can be quickly and conveniently used for wounds, with analgesic, antibacterial, anti - inflammatory effects, enabling the collagen and cell tissues in the dermis to maintain the original correct orientation. During the repair process of the dermis morphology and function, the tussah silk protein is gradually degraded and absorbed; 4. The high - molecular biological protein tussah silk fibroin repair liquid can shield the wound surface from contact with air, prevent the invasion of bacteria and fluid loss, and create a good aseptic environment for the repair of dermis morphology and function; 5. The high - molecular biological protein tussah silk fibroin repair liquid can activate the skin, strengthen the muscles, enhance immunity, activate and induce the growth of new blood vessels, and induce the regeneration of dermal cell tissues. Repair the wound matrix and avoid the formation of keloids; 6. The high - molecular biological protein tussah silk fibroin repair liquid can repair the skin of deep degree III - III wounds, induce the growth of new skin and skin appendages (hair, hair follicles, sebaceous glands, sweat glands) on the wound surface. And achieve the best function and appearance comfort during treatment; 7. The high - molecular biological protein tussah silk fibroin repair liquid avoids the influence of viruses, antigens and the spread of diseases that may be carried by materials from vertebrate animals; 8. The high-molecular biological protein fibroin repair solution does not require autologous skin transplantation or coverage with allogeneic (species) skin during use; 9. The high-molecular biological protein fibroin is easy to purify, can avoid complex and cumbersome processing techniques, is easy to achieve large-scale production, has a low manufacturing cost, and has broad application prospects.

[0022] 10. The high-molecular biological protein fibroin repair solution is convenient to use and carry; in case of sudden accidents, self-help or mutual rescue can be carried out. Description of the Drawings Figure 1 It is a diagram of layer-by-layer incision of the subcutaneous tissue and deep fascia of experimental animals and removal of the dermis; Figure 2 It is a diagram of the wound surface covered with hair shaved off with a shaver on the 45th day; Figure 3 It is a color image diagram of the microscopic photo of the interference group section magnified 40 times for observation and analysis; Figure 4 It is a color image diagram of the microscopic photo of the interference group section magnified 200 times for observation and analysis; Figure 5 It is a color image diagram of the microscopic photo of the interference group section magnified 400 times for clear observation, analysis and description; Figure 6 It is a color image diagram of the microscopic photo of the control group section magnified 40 times for comparison with the interference group; Figure 7 It is a color image diagram of the microscopic photo of the control group section magnified 200 times for comparison with the interference group; Specific Embodiments The present invention will be further described below in combination with experiments.

[0024] Experiment (1) 1. Sample name: Tussah cocoon silk powder 2. Experimental item: Detection of tussah silk protein 3. Equipment: Mass spectrometer: Thermo Orbitrap Fusion Lumos; Database search software: ProteomeDiscoverer 4. Test materials: 50 mM NH4HCO3; cocktail protease inhibitor; 0.45 mg / ml BSA standard solution; Coomassie Brilliant Blue standard solution; DTT solution: 50 mM in 50 mM NH4HCO3 (prepared freshly); IAA solution: 60 mM in 50 mM NH4HCO3 (prepared freshly); Trypsin solution: 1 μg / μl in 10 mM acetic acid (prepared freshly); 0.1% FA-ACN; 0.1% FA; Millpore 4 ml 3 kDa cut-off ultrafiltration tube, 96-well plate, pH test paper.

[0025] 5. Experimental Procedures Protein Extraction and Proteolysis Prevention: (1) Take 10 mg of silkworm powder and dissolve it in 0.5 ml of 9 M lithium thiocyanate solution, vortex for 2 h at room temperature; (2) Centrifuge at 12000 g for 1 min at 4 °C and take the supernatant; (3) Add the sample to a 4 ml ultrafiltration tube with a 3 kDa cut-off, centrifuge at 7000 g for 15 min, discard the filtrate, add 500 μl of 50 mM NH4HCO3 to the sample, centrifuge at 7000 g for 15 min, discard the filtrate, then add 500 μl of 50 mM NH4HCO3 to the sample again, repeat 4 times, collect the sample, and make up the volume to 300 μL.

[0026] (4) Measure the protein concentration with Coomassie Brilliant Blue. First, add 200 μl of Coomassie Brilliant Blue to 9 wells in a 96-well plate, the standard solution concentration is 0.45 mg / ml, add 0, 1, 2, 4, 8, 16 μl to the standard curve respectively, add 2 μl, 8 μl, 16 μl to the sample, measure the absorbance at 595 nm within 5 min, and calculate the sample protein concentration according to the formula.

[0027] (5) Take 50 μg of protein according to the protein concentration, add 50 mM NH4HCO3 to adjust the pH of the supernatant to 7 - 8.

[0028] (6) Add 50 nM DTT / 50 mM NH4HCO3 to a final DTT concentration of 5 mM, place at 56 °C for 1 h (1 M DTT = 0.154 g / ml).

[0029] (7) Add 60 mM IAA / 50 mM NH4HCO3 to a final IAA concentration of 10 nM, place in the dark for 40 min (600 mM IAA = 0.111 g / ml) (8) Add trypsin to the sample at a mass ratio of sample:Trypsin = 25:1 (trypsin is dissolved in 10 mM acetic acid to 1 μg / μl), adjust the pH value to 7 - 8.5, and digest at 37 °C for 20 h. (9) Add 4 times the amount of 0.1% FA - acetonitrile to the digested solution, shake for 5 min, centrifuge at 20000 g for 10 min, and collect the supernatant.

[0030] (10) Add 50 μl of 0.1% FA to the precipitate, shake for 5 min, then add 200 μl of 0.1% FA - acetonitrile, shake for 5 min, centrifuge at 20000 g for 10 min, and collect the supernatant. Repeat twice.

[0031] (11) Combine the supernatants from the three times and spin dry at room temperature with a centrifugal concentrator.

[0032] (12) Reconstitute to 50 μL of 10% ACN - water (containing 0.1% FA), centrifuge at 20,000 g for 10 min, and take 40 μL to a dedicated proteomics liquid vial. Perform LC - MS / MS detection.

[0033] Chromatograph: Ultimate3000RSLCnano Systems (Dionex, Thermo Fisher FisherScientific) Chromatographic column: PepMapTM Trap Column (75 μm×20 mm, Thermo Fisher Scientifie) PepMapTM RSLCC18Snail Column (75 μm×500 mm, Thermo Fisher Scientifie) Mobile phase A: 0.1% formic acid; B: acetonitrile (containing 0.1% formic acid) Table 1 Gradient elution conditions Mass spectrometer: Thermo Orbitrap Fusion Lumos Scanning mode: positive ion Scanning range: 300 - 1700 m / z Resolution: 120,000 Select 20 strongest high - energy fragmentations Database: human database Search library parameters: Precursor tolerance: ±10 ppm Fragment ion tolerance: ±0.6 ppm Enzyme: Trypsin Maximum missed cleavage: 2 Fixed modification: Carbamylation (C) Variable modifications: Oxidation (M); Acetyl (N) Search library software: Proteome Discoverer2.2 5. Detection results: (See Appendix 1 for the information of the silk protein of Antheraea yamamai from Qiannanling) Experiment (2) 1. Sample name: A high - molecular biological protein fibroin repair solution for covering deep third - degree wounds 2. Experimental item: Deep third - degree wound repair 3. Materials (1) Preparation of the test substance: Prepare 150 ml of Repair Solution A and 500 ml of Repair Solution B for standby. The ratio of each solute solution is: silk protein 0.04 - 0.20%: 2 - butanol 1.0 - 6%: plant light oil 98.96 - 93.80%.

[0034] (2) Experimental animals: 4 healthy adult New Zealand rabbits, half male and half female, with a body weight range of 2500 - 3000 g. They are from Yelo Rabbit Farm in Tongdao Dong Autonomous County, Hunan Province. Breeding environment: Temperature range (°C): 16 - 24, relative humidity (%): 40 - 70. The feed is produced in Shandong, with a certificate SCXK(Xiang) 2006—0001.

[0035] 4. Methods (1) Experimental method: The experimental animals were respectively labeled as 1, 2, 3, and 4. Without anesthesia, they were manually fixed in the lateral position and placed in a 40×60 cm stainless steel tray on the animal experiment table.

[0036] (2) Surgical procedure: The New Zealand rabbits were fixed in the lateral position with their limbs abducted on the operating table. Use a shaver to remove the hair larger than the size of the skin trauma to be repaired in the experiment. Then, draw lines for disinfection, cover with sterile towels, and use conventional medical instruments to cut through the skin, subcutaneous tissue, deep fascia, and remove the skin deeply layer by layer along the drawn lines ( Figure 1 ); After hemostasis with gauze, apply the repair liquid type A to the wound (uniformly drip with a dropper; adopt the methods of full dressing, half dressing, and exposure (the same surgical procedure is adopted, and the experiment is carried out 3 times in summer, autumn, and spring, with 4 rabbits in each group each time, a total of 12 rabbits), and all are raised under the same conditions for dynamic observation.

[0037] (3) Control and intervention skin pathological sections: Using the same method of non-anesthetizing and manually fixing the experimental animals, take the surgical interference and control samples of the same rabbits. After a series of tissue cell fixation, dehydration, wax infiltration, section staining and other technological operations of the samples, observe the color images of the specimen sections under an Olpympus optical microscope.

[0038] 5. Observation and analysis (1) Dynamic observation: Through dynamic repeated observation of the cell cortex test of rabbits in summer, spring, and autumn and the cell cortex repair situation of the interference group, on the 5th day after applying the repair liquid type A, the wound repair effect was good, without fluid accumulation or exudation. Continuously apply the type B, and the wound repair was good on the 10th and 15th days. On the 20th day, some wounds healed, and hair follicles and hairs could be seen. On the 45th day, the wound was covered with hair, and use a shaver to shave off the hair on the repaired wound ( Figure 2 ), (2) Static analysis Through color image analysis, the infiltration of inflammatory cells in the local tissue of the repaired wound in the interference group ( Figure 1 -HE-40; Figure 2 -HE-200; Figure 3 -HE-400) and the control group ( Figure 1 -HE-40; Figure 2Compared with the control group (-HE-200), under the microscope, a section of epidermis with normal structure can be seen, and local epidermal defects can be seen. No abnormal pathological changes are found in the dermis layer, and no abnormalities are found in the dermis layer and skin appendages (hair, hair follicles, sebum, sweat glands, etc.). The cell structure of the skin cell tissue repaired in the interference group is the same as that of the control group.

[0039] 6. Conclusion Through dynamic observation of the experimental effect and static color image analysis: Using Covered with polymer bio-protein tussah fibroin The external use of Repair Liquid A and Auxiliary Repair Liquid B for traumatic skin has the following functions and effects in animal experiments conducted in spring, summer, autumn and pathological sections: easy to use, rapid penetration, excellent performance, inhibition of bleeding; moistening the wound and preventing drying; having anti-inflammatory properties and resisting wound infection; adsorbing exudate and preventing the accumulation of exudate under the coating; maintaining the adhesion of silk fibroin to the wound cell tissue, supplying energy to the wound surface, and various decomposed peptides participating in the body's immune regulation, promoting the absorption of minerals and silk fibroin, and creating flesh, blood, skin, etc. for the growth of traumatic tissue cells, achieving the purpose of healing the dermal defect of deep degree III-III wounds, and having obvious characteristics of complete cell tissue repair. This new type of wound coating has strong application suitability and broad prospects.

Claims

1. A high-molecular biological protein tussah fibroin repair solution for covering deep third-degree wounds belongs to the field of biomedical materials and is applied to the biomedical and traditional Chinese medicine industries. It uses tussah cocoons, which are high-molecular biological active materials containing natural human proteins, as the base material, and adds 2-borneol solute and edible plant light oil in proportion to form a coating for repairing deep wound matrices. The structural information of the Nanjin tussah cocoon silk protein is shown in Table (1): Table of Nanjin tussah silk protein information 2. A preparation method of a high-molecular biological protein tussah fibroin repair solution for covering deep third-degree wounds. Solution A is composed of 0.002 - 0.020 / ml silk protein, 0.010 - 0.060 / ml of 2-borneol solute and 1 ml of edible plant light oil; Solution B is composed of 0.010 - 0.060 / ml of 2-borneol solute and 1 ml of edible plant light oil. Both solutions are used for covering and repairing deep third-degree wounds.

3. According to the high-molecular biological protein tussah fibroin repair solution described in claim 1, the tussah fibroin protein base material is derived from tussah cocoons in the breeding farm of the Nanjin Tussah Science and Technology Research Center in Tongdao Dong Autonomous County.

4. According to the high-molecular biological protein tussah fibroin repair solution described in claim 1, the 2-borneol solute is effectively fused with the high-molecular biological tussah fibroin protein and has characteristics and functions such as analgesia, antibacterial, anti-inflammatory, and anti-inflammatory.

5. According to the high-molecular biological protein tussah fibroin repair solution described in claim 1, the edible plant light oil (both a solute and a solution) is effectively fused and linked with the tussah fibroin protein and 2-borneol solute, and has the special functions of quickly covering, sealing, and penetrating the wound surface.

6. According to the preparation method of the high-molecular biological protein tussah fibroin repair solution described in claim 2, Its characteristics are shown as follows: (1) After the tussah cocoons are washed, soaked, and cleaned, they are washed three times with distilled water. Then they are fished out and placed in a nylon mesh tray in a 1m×2m stainless steel enclosure and naturally dried to extract fibroin protein for standby.

7. (2) Preparation of an edible plant light oil composite solution with a total concentration of 99.8 - 98.00%. The proportion of each solute is: 1.0 - 6% 2-borneol : 98.80 - 92.00% edible plant light oil. Then mechanical stirring is carried out. Stirring time: 10 - 15 minutes. Name of stirring equipment: overhead stirrer; Model: AS20; Display type: LED digital tube; Rotation speed range (RPM): 30 - 2000; Maximum viscosity (mPAS): 10000.

8. (3) Immerse the fibroin protein in the above composite light oil solution. The temperature of the light oil solution is 5 - 18°C, and the immersion time is more than 5 minutes. Then stir with the same type of equipment as described above. Stirring time: 10 - 40 minutes.

9. (4) According to the preparation method of the tussah fibroin protein repair solution described in claim 2, It is characterized in that: Before step (3), step (2) needs to be repeated for stirring 1 - 3 times to quickly dissolve the 2-borneol solute into the edible plant light oil.

10. The tussah silk fibroin repair liquid according to claim 2 is in the state of a light yellow or dark yellow liquid.

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