Freeze-dried repair dressing containing type I collagen

Abstract

The invention belongs to the field of dressings, and particularly relates to a freeze-dried repair dressing containing I-type collagen, which comprises pigskin-derived I-type collagen, chitosan, glycosaminoglycan and a cross-linking agent, the glycosaminoglycan comprises but is not limited to hyaluronic acid, chondroitin sulfate, keratin sulfate and the like, and the cross-linking agent is sodium mannuronate methylsilanol and tea polyphenol. The freeze-dried repair dressing is prepared from the type I collagen, the chitosan and the glycosaminoglycan through secondary crosslinking, has the characteristics of resisting bacteria and reducing active oxygen, and is a multifunctional skin repair dressing with biological activity and good biocompatibility.

Description

Technical Field

[0001] The invention belongs to the field of dressings, and in particular relates to a freeze-dried repair dressing containing type I collagen. Background Art

[0002] Skin is the largest organ in the human body. Its damage repair is not only related to the patient's appearance, but also to functional recovery and health. Skin repair includes wound repair and depression repair. Wound repair can accelerate wound healing and reduce scar formation; depression can make the skin plumper, improve static wrinkles and tissue contours, and revitalize the skin.

[0003] Wound repair, especially for the increase in the elderly population and the increase in diabetes cases, the incidence of chronic wounds is increasing every year. Non-healing wounds cause pain and discomfort to patients, reduce their quality of life, and are also prone to serious bacterial infections. When the wound is in the inflammatory stage, delaying its normal physiological healing, the wound becomes a chronic or non-healing wound. Infection-related complications of chronic wounds may eventually lead to amputation or even death. In the medical field, the research and development of wound repair materials has always been the focus of attention. Wound repair excipients with excellent functions can accelerate skin wound repair and reduce the occurrence of complications. Wound repair excipients with excellent functions should be able to provide the basic components required for skin wound repair and accelerate the skin wound repair process. Collagen is a core component involved in wound healing. As the main component of the extracellular matrix, collagen provides the necessary scaffold for cell migration and proliferation, and can promote the closure of wound edges and the formation of new tissues. The mesh structure of collagen not only supports the migration of epithelial cells and the contraction of wounds, but also participates in regulating inflammatory responses and provides support for new blood vessels, thereby promoting blood supply and nutrient exchange. In addition, moderate degradation and remodeling of collagen are essential for balancing the wound repair process, reducing scar formation, and restoring the structure and function of the skin. Therefore, the biological activity and physical properties of collagen play an indispensable role in effectively promoting wound healing and tissue regeneration. Therefore, skin repair dressings containing collagen are a research hotspot. However, collagen extracted from conventional sources often faces problems such as biosafety, immunogenicity, and activity retention. CN109675085A discloses a composite collagen dressing for human wound repair, which is a mixture of two or more different types of collagen; type I collagen is derived from pig skin, cowhide, and bovine Achilles tendon.

[0004] In addition to collagen, glycosaminoglycans also play an important role in the skin repair process. Glycosaminoglycans (GAGs) are polyanionic linear polysaccharides with a repeating disaccharide unit structure, which are components of various structural and connective tissues such as skin, cartilage and cornea. Hyaluronic acid and chondroitin sulfate are two important glycosaminoglycans that provide a favorable microenvironment for wound healing by promoting angiogenesis and immunomodulation, thereby accelerating wound closure, improving healing quality and reducing complications. CN116510070A discloses a collagen hydrogel, which is formed by chemical cross-linking of collagen, hydrophobically modified chitosan and dopamine through a cross-linking agent.

[0005] In addition to providing the main components necessary for skin wound repair, wound repair dressings with excellent functions are also equally important for preventing and treating bacterial infections. Bacterial contamination during wound repair can stimulate inflammatory responses, leading to excessive production of macrophages and neutrophils, which carry matrix metalloproteinases (MMPs) and reactive oxygen species (ROS). Matrix metalloproteinases (MMPs) and ROS play a vital role in the normal wound healing process. They participate in tissue remodeling and repair, promote wound contraction and epithelial regeneration, eliminate infection and promote necessary inflammatory responses. However, overexpression of MMPs and ROS is not conducive to wound healing. MMPs are zinc-dependent proteases that can degrade collagen, elastin and structural units of the extracellular matrix (ECM). Overexpression or abnormal activation of MMPs can cause excessive proteolysis of collagen, elastin and growth factors, hindering wound healing. At the same time, overexpression or abnormal activation of MMPs can also lead to the massive production of ROS. Excessive accumulation of ROS can damage cell membranes, proteins and DNA, causing cell dysfunction or even death, thereby destroying the healing process. It can be seen that controlling bacterial infection can reduce the overexpression of matrix metalloproteinases (MMP) and reactive oxygen species (ROS) carried by immune cells, which is of great significance for promoting wound healing. In short, for wound repair, supplementing the matrix components necessary for wound repair and preventing and treating wound infection are necessary conditions for skin wound repair dressings.

[0006] The body filling injection of repair dressings is also one of the important aspects of skin repair. Collagen, as a key component to maintain the elasticity of the dermis of the skin, is responsible for maintaining the structure and function of the dermis. With the increase of age, the concentration of collagen in the skin gradually decreases, and the density of elastic fibers and the repair ability of connective tissue also decrease. This not only leads to the weakening of skin elasticity and water retention, but also reduces the volume of the dermis, especially in the facial area, which is more likely to cause wrinkles and sagging. Therefore, supplementing collagen has a significant effect on successfully repairing skin depressions, reshaping facial contours and improving overall beauty. The ideal skin injection repair dressing should be able to continue to absorb water and swell after being injected into the skin, thereby playing an excellent filling effect. This swelling effect can not only fill skin depressions instantly, but also increase the water content of the skin, making the skin moist and shiny. The ideal skin injection repair dressing can also provide long-term structural support and nutrition for the skin by promoting cell proliferation and the synthesis of extracellular matrix, further promoting skin health and repair. It can be seen that the body's injectable skin repair dressing needs to have excellent water absorption properties, provide excellent filling effect and water locking function for the skin, make the skin tender and smooth, and rejuvenate the skin. Summary of the invention

[0007] In order to solve the above technical problems, the present invention provides a freeze-dried repair dressing, which is prepared by using type I collagen, chitosan, glycosaminoglycan and secondary cross-linking, and has the characteristics of antibacterial and reducing active oxygen. It is a multifunctional, bioactive and biocompatible skin repair dressing.

[0008] The present invention adopts the following technical solution:

[0009] A freeze-dried repair dressing containing type I collagen comprises the following components in percentage by mass: 25-40% type I collagen, 10-20% chitosan, 10-20% glycosaminoglycan and 25-40% cross-linking agent.

[0010] In some embodiments, the freeze-dried repair dressing is prepared by secondary cross-linking and freeze drying. Using the two micro-cross-linking technology and freeze drying technology, it can be applied to the wound site after re-dissolving with water to promote skin wound repair. At the same time, it can be injected through the water light needle device to replenish collagen for the skin, repair skin depressions, and reshape the skin shape.

[0011] In some embodiments, the crosslinker comprises a silanol crosslinker and tea polyphenols; preferably, the silanol crosslinker is sodium methyl silanol mannuronate. The first crosslinking with a silanol crosslinker can improve the water absorption of the hydrogel and increase the ability of the repair dressing to absorb secretions from the skin lesions; after crosslinking with tea polyphenols, the crosslinking morphology of collagen and glycosaminoglycans is further changed, further improving its water-locking function. Tea polyphenols is a polyphenolic substance. Due to the formation of hydrogen bonds and electrostatic interactions, it can be used as a crosslinker to promote the formation of a mesh structure of collagen and glycosaminoglycans, further increasing the water-locking performance of the repair dressing of this patent. At the same time, tea polyphenols can promote the oxidative balance of the wound site, reduce the negative effects of reactive oxygen species (ROS), and protect cells from oxidative damage. Tea polyphenols also have anti-inflammatory effects, can reduce wound inflammatory reactions, and regulate the activity of immune cells, thereby creating a more favorable healing environment for the wound.

[0012] In some embodiments, the mass percentages of the silanol cross-linking agent and tea polyphenols in the freeze-dried repair dressing are 0.5-2% and 24.5-39.5%, respectively.

[0013] In some embodiments, the type I collagen is derived from animals, such as pigs, cattle, fish, etc., preferably pig skin type I collagen. Pig skin-derived type I collagen is highly similar to collagen in human skin and has the advantage of low risk of immune rejection.

[0014] In some embodiments, the pig skin type I collagen is extracted by defatting, high-speed homogenization, protease treatment, high-speed centrifugation, etc. By optimizing the extraction process of pig skin-derived type I collagen, the high purity and biological activity of pig skin-derived type I collagen are ensured.

[0015] In some embodiments, the glycosaminoglycan includes but is not limited to hyaluronic acid, chondroitin sulfate, keratan sulfate, etc. Preferably, the glycosaminoglycan is sodium hyaluronate and chondroitin sulfate, and the mass ratio of the two is 1:2 to 2:1.

[0016] The present invention also provides a method for preparing the freeze-dried repair dressing containing type I collagen, comprising the following steps:

[0017] (1) adding chitosan and type I collagen to an acetic acid solution, stirring at high speed to make them uniform, adding a silanol crosslinking agent solution, and stirring to complete the first crosslinking;

[0018] (2) After the first cross-linking is completed, glycosaminoglycan and tea polyphenols are added, stirred for the second cross-linking, and freeze-dried to prepare the freeze-dried repair dressing containing type I collagen.

[0019] In some embodiments, the method for preparing the freeze-dried repair dressing containing type I collagen comprises the following steps:

[0020] (1) chitosan and porcine skin type I collagen were added to 100 ml of 1% acetic acid solution, stirred at 1000 rpm, 2 ml of 1% sodium mannuronate methylsilanol was added, and the mixture was stirred at 37° C. for 2 h to complete the first crosslinking;

[0021] (2) After the first cross-linking is completed, sodium hyaluronate and chondroitin sulfate are added and stirred to mix evenly. Tea polyphenols are added as a cross-linking agent for secondary cross-linking and stirred at 37°C for 2 hours; the product is filled into an injection bottle, freeze-dried, and vacuum-sealed to prepare the freeze-dried repair dressing containing type I collagen.

[0022] The mechanism of the first cross-linking is shown in the following reaction formula:

[0023]

[0024] In some embodiments, the preparation method further comprises the step of extracting type I collagen: extracting the pig skin by defatting, high-speed homogenization, protease treatment, and high-speed centrifugation.

[0025] The present invention also provides a repair auxiliary material for a water light needle device, which is obtained by re-dissolving the freeze-dried repair dressing containing type I collagen with water.

[0026] Compared with the prior art, the beneficial effects of the present invention include at least:

[0027] (1) The freeze-dried repair dressing of the present invention contains two glycosaminoglycan components, namely type I collagen, hyaluronic acid and chondroitin sulfate, which provide the necessary components for skin wound repair and accelerate wound repair.

[0028] (2) The freeze-dried repair dressing of the present invention adopts double micro-crosslinking technology and freeze-drying technology, and can form a hydrogel after being re-dissolved in water, which has high water absorption and water retention.

[0029] (3) The present invention uses tea polyphenols as the second cross-linking agent. Tea polyphenols are polyphenols. Hydrogen bonds and electrostatic interactions can promote the formation of a mesh structure between collagen and glycosaminoglycans. They can promote the oxidative balance of the wound site, reduce the negative effects of reactive oxygen species (ROS), and protect cells from oxidative damage. They also have anti-inflammatory effects and can reduce wound inflammatory responses.

[0030] (4) The freeze-dried repair dressing of the present invention effectively maintains the three-dimensional structure and biological activity of collagen, and also significantly improves the stability and convenience of the dressing. It can play an important role in various skin repair fields such as burns, chronic wounds, surgical wounds, skin hydration, wrinkle repair, and skin morphology remodeling. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 This is a scanning electron microscope photo of the freeze-dried repair dressing of Example 2. Figure 1 Figure a is a scanning electron microscope photo of the freeze-dried dressing after one cross-linking. Figure 1 Middle b is a scanning electron micrograph of the freeze-dried dressing after two cross-linking processes.

[0032] Figure 2 To compare the water-locking capacity of different freeze-dried repair dressings.

[0033] Figure 3 To compare the wound healing effects of different freeze-dried repair dressings.

[0034] Figure 4 Scoring of eye wrinkles after injection of different freeze-dried repair dressings Demasha. DETAILED DESCRIPTION

[0035] In order to make the purpose, technical scheme and beneficial effects of the present invention clearer, the present invention is further described in detail below in conjunction with the accompanying drawings and specific embodiments. Examples of the embodiments are shown in the accompanying drawings. It should be understood that the specific embodiments described in the following embodiments of the present invention are only exemplary descriptions of the specific embodiments of the present invention, and are intended to be used to explain the present invention, but do not constitute a limitation of the present invention.

[0036] The endpoints of ranges and any values ​​disclosed herein are not limited to the exact range or value, and these ranges or values ​​should be understood to include approximations to these ranges.

[0037] Example 1: Extraction and separation of type I collagen from pig skin

[0038] 5g of fresh pig skin was cut into small pieces of about 1cm×1cm after removing subcutaneous fat, and 100ml of 5% sodium carbonate solution was added for degreasing. The mixture was stirred for 1 hour and the 5% sodium carbonate solution was replaced once. The 5% sodium carbonate solution was replaced 3 times in total. The defatted pig skin was rinsed three times with distilled water. The rinsed pig skin was placed in a meat grinder and crushed, 100ml of phosphate buffer (PBS, pH 6.8) was added, and a high-speed emulsifier was used to stir at 10,000 rpm for 2 minutes, and then stirred and extracted at 4°C for 30 minutes. The supernatant was discarded by high-speed centrifugation and washed 3 times with 100ml of deionized water. The precipitated collagen was added to 50ml of 0.5 mol / L acetic acid solution (containing 2% pepsin) and placed at 25°C overnight. The supernatant was adjusted to neutral with sodium hydroxide (NaOH) solution, the supernatant was discarded by high-speed centrifugation, washed three times with 100 ml of deionized water, and the precipitate was freeze-dried to obtain 2.32 g of porcine skin-derived type I collagen extract with an extract yield of 46.4%.

[0039] Based on the characteristic that collagen is rich in hydroxyproline, a hydroxyproline kit was used to measure the hydroxyproline content and then calculate the collagen extraction rate.

[0040]

[0041] In the formula: m2 represents the mass of pig skin collagen after freeze-drying (g); m1 represents the mass of pig skin powder before acid hydrolysis (g); h1 represents the content of hydroxyproline in pig skin powder before acid hydrolysis (μg / mg); h2 represents the content of hydroxyproline in pig skin collagen after freeze-drying (μg / mg).

[0042] After calculation, the collagen extraction rate was 67.6% and the purity of the extracted collagen was 88.7%.

[0043] Example 2: Preparation of freeze-dried repair dressing of pig skin-derived type I collagen

[0044] (1) Primary cross-linking

[0045] Add 0.5g chitosan and 1g pig skin type I collagen into 100ml 1% acetic acid solution, stir evenly at 1000rpm, add 2ml 1% sodium mannuronate methylsilanol, stir at 37℃ for 2h to complete the first crosslinking.

[0046] (2) Secondary crosslinking

[0047] After the first cross-linking, 0.2 g sodium hyaluronate and 0.2 g chondroitin sulfate were added and stirred to mix well. 1 g tea polyphenols was added as a cross-linking agent for secondary cross-linking and stirred at 37°C for 2 h.

[0048] (3) Freeze drying

[0049] The product was pre-frozen at -40℃ for 5h, gradually heated to 0℃ at 2℃ / min, then gradually heated to 30℃ at 3℃ / min, and maintained at 30℃ for 4h. The vacuum degree was ensured not to exceed 50pa during the whole process. The product was filled into injection bottles, freeze-dried, and vacuum-sealed to obtain pig skin-derived type I collagen freeze-dried repair dressing.

[0050] Example 3: Preparation of freeze-dried repair dressing of pig skin-derived type I collagen

[0051] (1) Primary cross-linking

[0052] 0.4 g chitosan and 1.2 g type I collagen from pig skin were added to 100 ml 1% acetic acid solution, stirred at 1000 rpm, 3 ml 1% sodium mannuronate methylsilanol was added, and the mixture was stirred at 37°C for 2 h to complete the first cross-linking.

[0053] (2) Secondary crosslinking

[0054] After the first cross-linking, 0.3 g sodium hyaluronate and 0.3 g chondroitin sulfate were added and stirred to mix well. 1 g tea polyphenols was added as a cross-linking agent for the second cross-linking and stirred at 37°C for 2 h.

[0055] (3) Freeze drying

[0056] The product was pre-frozen at -40℃ for 5h, gradually heated to 0℃ at 2℃ / min, then gradually heated to 30℃ at 3℃ / min, and maintained at 30℃ for 4h. The vacuum degree was ensured not to exceed 50pa during the whole process. The product was filled into injection bottles, freeze-dried, and vacuum-sealed to obtain pig skin-derived type I collagen freeze-dried repair dressing.

[0057] Example 4: Preparation of pig skin-derived type I collagen freeze-dried repair dressing

[0058] (1) Primary cross-linking

[0059] 0.6 g chitosan and 1 g pig skin-derived type I collagen were added to 100 ml 1% acetic acid solution, stirred at 1000 rpm, 2 ml 1% sodium mannuronate methylsilanol was added, and the mixture was stirred at 37°C for 2 h to complete the first cross-linking.

[0060] (2) Secondary crosslinking

[0061] After the first cross-linking, 0.3 g sodium hyaluronate and 0.3 g chondroitin sulfate were added and stirred to mix well. 0.8 g tea polyphenols were added as a cross-linking agent for secondary cross-linking and stirred at 37°C for 2 h.

[0062] (3) Freeze drying

[0063] The product was pre-frozen at -40℃ for 5h, gradually heated to 0℃ at 2℃ / min, then gradually heated to 30℃ at 3℃ / min, and maintained at 30℃ for 4h. The vacuum degree was ensured not to exceed 50pa during the whole process. The product was filled into injection bottles, freeze-dried, and vacuum-sealed to obtain pig skin-derived type I collagen freeze-dried repair dressing.

[0064] Comparative Example 1: Preparation of freeze-dried repair dressing of pig skin-derived type I collagen

[0065] The preparation method is the same as that of Example 2, except that the product is not cross-linked with sodium mannuronate methylsilanol.

[0066] Add 0.5g chitosan and 1g pig skin type I collagen to 100ml 1% acetic acid solution, stir evenly at 1000rpm, stir at 37℃ for 2h, add 0.2g sodium hyaluronate and 0.2g chondroitin sulfate, stir and mix evenly. Add 1g tea polyphenol as a cross-linking agent, stir at 37℃ for 2h. Fill the product into an injection bottle, freeze-dry, and vacuum seal to obtain pig skin type I collagen freeze-dried repair dressing.

[0067] Comparative Example 2: Preparation of freeze-dried repair dressing of pig skin-derived type I collagen

[0068] The preparation method is the same as that of Example 2, except that the product is not cross-linked with sodium mannuronate methylsilanol, and the cross-linking agent tea polyphenols is replaced by 25% glutaraldehyde.

[0069] Add 0.5g chitosan and 1g pig skin type I collagen to 100ml 1% acetic acid solution, stir evenly at 1000rpm, stir at 37℃ for 2h, add 0.2g sodium hyaluronate and 0.2g chondroitin sulfate, stir and mix evenly. Add 4ml 25% glutaraldehyde as a cross-linking agent, stir at 37℃ for 2h. Fill the product into an injection bottle, freeze-dry, and vacuum seal to obtain pig skin type I collagen freeze-dried repair dressing.

[0070] Freeze-dried repair dressing performance test:

[0071] 1. Microscopic morphology of freeze-dried repair dressing

[0072] The scanning electron microscopy of the freeze-dried repair dressing prepared in Example 2 is as follows: Figure 1 As shown, Figure 1 In the figure a, the microscopic morphology of the freeze-dried dressing obtained after freeze-drying after only one cross-linking (1% sodium mannuronate methylsilanol cross-linking) is shown. Figure 1 Figure b represents the microscopic morphology of the freeze-dried dressing obtained after two cross-linking steps and freeze-drying. Figure 2 It can be seen that the gaps in the freeze-dried dressing after two cross-linkings are denser, forming a sheet-like structure, indicating that it has higher water absorption and water locking capabilities.

[0073] 2. Test of water-locking performance of dressing

[0074] The skin repair dressings of Example 2, Comparative Example 1 and Comparative Example 2 were dissolved with equal amounts of deionized water, placed in a constant temperature and humidity chamber without lids, and weighed on days 0, 1, 2, 3, 4, 5, 6, 7, 9, 12 and 15, respectively, to calculate the residual weight percentage (Q)

[0075]

[0076] Where: W iW is the weight of the initial repair dressing, in g; s The weight of the repair dressing at different time points, in g.

[0077] The results are as follows Figure 2 As shown, the weight of skin repair excipients with different prescriptions decreases with time, indicating that the water content increases with time and is not lost in a short time. The water content of the skin repair dressing of Example 2 is the slowest, indicating that Example 2 has the strongest water-locking ability. This is because Example 2 has formed a denser grid structure between the macromolecular components after two micro-crosslinkings, which can accommodate more water and help lock in water. This property can keep the skin wound site moist for a long time and is conducive to wound healing. At the same time, after water light injection, the skin can maintain moisture for a long time, making the skin tender and smooth for a long time.

[0078] 3. Evaluation of antibacterial effect

[0079] The antibacterial activity of the wound repair dressing against Staphylococcus aureus, Bacillus subtilis, and Gram-negative Escherichia coli was evaluated. After 50 μl of suspended bacteria were applied to the Luria-Bertani medium plate, an Oxford cup was placed on the surface of the medium. Example 2, Comparative Example 1, and Comparative Example 2 were reconstituted with equal amounts of water, and 0.2 ml was injected into the Oxford cup. After culturing at 37°C for 24 hours, the diameter of the inhibition zone was measured with a vernier caliper to determine the antibacterial effect of the repair dressing. The results are shown in Table 1.

[0080] Table 1 Antibacterial effect of skin repair dressings with different prescriptions

[0081]

[0082] As can be seen from Table 1, the antibacterial effects of Example 2 and Comparative Example 1 on Escherichia coli, Bacillus subtilis and Staphylococcus aureus are significantly better than those of Comparative Example 2. Tea polyphenols are used as micro-cross-linking agents in both Example 2 and Comparative Example 1, which may be the reason why the antibacterial effect is better than that of Comparative Example 2.

[0083] 4. Repair effect of damaged skin

[0084] Twelve SD female rats (180-220g) were randomly divided into 4 groups, with three rats in each group. Anesthesia was performed by intraperitoneal injection of chloral hydrate. The hair on the back of the rat was shaved with a razor and wiped with 75% ethanol solution for disinfection. Three circular wounds with a diameter of 8 mm were made on the back of each rat with a skin puncher. The sterilized skin repair excipients of Example 2, Comparative Example 1, and Comparative Example 2 and commercially available Band-Aids were applied to different experimental groups, respectively, and applied or covered on the skin defect area on the back of the rat. At the same time, a defect area without any treatment was set as a blank control group. During the experiment, the dressing was changed every 3 days and observed continuously until the 15th day. On the 3rd, 6th, 9th, 12th, and 15th days after surgery, the skin trauma of the rat was imaged and recorded, and the wound area was measured using Image J image analysis software to calculate the percentage area of ​​wound healing. The calculation formula is as follows:

[0085]

[0086] Where: A0 is the initial wound area; A t The actual area of ​​the wound at a certain point in time.

[0087] The results are as follows Figure 3 As shown, the order of the ability of skin repair excipients with different prescriptions to promote wound healing is Example 2> Comparative Example 1> Comparative Example 2> Ordinary Band-Aid. Example 2 of this patent has the best effect in promoting wound healing, mainly due to the following aspects: (1) The skin repair excipient of Example 2 contains type I collagen, hyaluronic acid, chondroitin sulfate and other extracellular matrix, which has a promoting effect on promoting wound repair; (2) The skin repair dressing of Example 2 has a strong water-locking ability after secondary cross-linking, which can keep the wound site moist for a long time, which is conducive to rapid wound healing; (3) The skin repair dressing of Example 2 contains tea polyphenols, which has astringent and bactericidal effects, which is conducive to wound healing.

[0088] 5. Evaluation of the effect of water injection

[0089] 18 healthy volunteers with obvious fine lines in the corners of the eyes were asked not to use any cosmetics, drugs or health products that may affect the results within two weeks before the start of the experiment. These volunteers were randomly divided into 3 groups (3 men and 3 women in each group), and the three groups of volunteers were injected with Demasha water light needle (9 needles) in the left and right corners of the eyes of Example 2, Comparative Example 1 or Comparative Example 2. Before use (0 day) and 2, 4, 6, 8, 12, and 24 days after use, the eye wrinkle score was measured using the VISIA skin tester (Canfield, USA), and the average value of the wrinkle score of the left and right corners of each volunteer was taken to evaluate the anti-wrinkle effect. The wrinkle score results of each group of volunteers are shown in 4. It can be seen that compared with Comparative Examples 1 and 2, after use of Example 2, the static wrinkle score of the corners of the volunteers' eyes dropped rapidly, indicating that Example 2 has a better filling and repairing effect at the same dose, which may be related to its better water-locking ability.

[0090] Finally, it should be noted that the above embodiments are only used to illustrate the technical solution of the present invention and do not constitute a limitation on the content of the present invention. Within the technical concept of the present invention, the technical solution of the present invention can be subjected to a variety of simple modifications, including the combination of various technical features in any other suitable manner, and these simple modifications and combinations should also be regarded as the contents disclosed by the present invention and belong to the protection scope of the present invention.

Claims

1. A freeze-dried repair dressing comprising type I collagen, characterized in that: The invention comprises the following components in percentage by weight: 25-40% type I collagen, 10-20% chitosan, 10-20% glycosaminoglycan and 25-40% cross-linking agent.

2. The freeze-dried repair dressing according to claim 1, characterized in that: The freeze-dried repair dressing is prepared by secondary cross-linking and freeze-drying.

3. The freeze-dried repair dressing according to claim 1, characterized in that: The cross-linking agent comprises a silanol cross-linking agent and tea polyphenols. Preferably, the silanol cross-linking agent is sodium mannuronate methylsilanol.

4. The freeze-dried repair dressing according to claim 3, characterized in that: The mass percentages of the silanol cross-linking agent and tea polyphenols in the freeze-dried repair dressing are 0.5-2% and 24.5-39.5% respectively.

5. The freeze-dried repair dressing according to claim 1, characterized in that: The type I collagen is derived from animals, preferably pig skin type I collagen.

6. The freeze-dried repair dressing according to claim 5, characterized in that: The pig skin type I collagen is extracted through defatting, high-speed homogenization, protease treatment and high-speed centrifugation steps.

7. The freeze-dried repair dressing according to claim 1, characterized in that: The glycosaminoglycan is selected from one or more of hyaluronic acid, chondroitin sulfate, and keratan sulfate; preferably, the glycosaminoglycan is sodium hyaluronate and chondroitin sulfate, and the mass ratio of the two is 1:2 to 2:

1.

8. The method for preparing the freeze-dried repair dressing according to any one of claims 1 to 7, characterized in that: The steps include: (1) adding chitosan and type I collagen to an acetic acid solution, stirring at high speed to make them uniform, adding a silanol crosslinking agent solution, and stirring to complete the first crosslinking; (2) After the first cross-linking is completed, glycosaminoglycan and tea polyphenols are added, stirred for the second cross-linking, and freeze-dried to prepare the freeze-dried repair dressing containing type I collagen.

9. The preparation method according to claim 8, characterized in that: The steps include: (1) chitosan and porcine skin type I collagen were added to 100 ml of 1% acetic acid solution, stirred at 1000 rpm, 2 ml of 1% sodium mannuronate methylsilanol was added, and the mixture was stirred at 37° C. for 2 h to complete the first crosslinking; (2) After the first cross-linking is completed, sodium hyaluronate and chondroitin sulfate are added and stirred to mix evenly. Tea polyphenols are added as a cross-linking agent for secondary cross-linking and stirred at 37°C for 2 hours; the product is filled into an injection bottle, freeze-dried, and vacuum-sealed to prepare the freeze-dried repair dressing containing type I collagen.

10. A repair auxiliary material for hyaluronic acid injection equipment, obtained by reconstituted with water the freeze-dried repair dressing according to any one of claims 1 to 7.

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