Mussel mucoprotein hydrogel dressing with self-repairing function as well as preparation method and application of mussel mucoprotein hydrogel dressing

Abstract

The invention discloses a mussel mucoprotein hydrogel dressing with a self-repairing function and a preparation method of the mussel mucoprotein hydrogel dressing. Comprising a gel system, and the gel system comprises a composite hydrogel matrix and a functionalized nano-modification component; the composite hydrogel matrix is of a network structure formed by light-enzyme synergistic crosslinking of mussel mucin, methacrylated gelatin and phenylboronic acid modified gelatin, and the weight part ratio of the mussel mucin to the methacrylated gelatin to the phenylboronic acid modified gelatin is (2-5): (15-20): 5; the functionalized nano modification component comprises mesoporous silica nanoparticles loaded with small molecule polypeptide and graphene oxide, and the graphene oxide accounts for 0.1-0.5 part by weight in the gel system; the mesoporous silica nanoparticles account for 1-3 parts by weight in the gel system, and the polypeptide loading rate is 30-50%. According to the application, the defects of the traditional wound repair material in adhesion stability, antibacterial property and anti-inflammatory property are overcome.

Description

Technical Field

[0001] The present invention relates to the technical field of biological adjuvants, and particularly relates to a mussel adhesive protein hydrogel dressing with self-healing function, a preparation method thereof and an application thereof. Background Art

[0002] Hydrogel is a polymer macromolecular material with a three-dimensional network structure, which is formed by physical or chemical bonding in an aqueous solution. It has hydrophilic groups and can swell in water or body fluid without dissolving. This unique structure endows hydrogel with many excellent properties, making it show great potential in the application of wound dressings. Hydrogel has good biocompatibility, which means that when it comes into contact with human tissues, it is not easy to cause adverse reactions such as inflammatory reactions.

[0003] Since the 1960s, research scholars have injected hydrogel into the body of experimental subjects for treating osteoarticular injuries, etc., which has verified this property. Its biodegradability is also very excellent. High-performance hydrogels with biodegradability can be prepared by modification or compounding means, and they have great application potential in implantation and drug delivery, and can effectively solve safety problems such as rejection of intervention materials and difficult metabolism of drug delivery systems. Hydrogel also has high water absorption and water retention properties, and its water content can be as high as 99%. The medical dressing made of hydrogel can absorb the wound exudate, while maintaining a moist environment, avoiding adhesion to the wound and causing secondary trauma, and has a large liquid absorption capacity, so there is no need to replace it frequently. In addition, the surface of the hydrogel is smooth and has high elasticity. When used as a dressing, it can closely fit the wound without adhesion, reducing the contact with bacteria.

[0004] Although hydrogel dressings have significant advantages, there are still many deficiencies in the prior art. In terms of breathability, the existing hydrogel dressings have limited breathability, which may affect gas exchange at the wound site and is not conducive to wound healing. For example, after some traditional hydrogel dressings cover the wound, the wound area will be in a relatively closed state, resulting in the skin being unable to breathe normally, which may cause problems such as skin maceration. Poor environmental adaptability is also a prominent problem. Some hydrogel dressings are difficult to maintain stable performance in special environments such as extreme temperatures. For example, in a low-temperature environment, some hydrogel dressings may become brittle and lose elasticity, and cannot fit the wound well. In terms of antibacterial properties, some hydrogel dressings may cause bacteria to develop drug resistance after long-term use. For some hydrogel dressings added with antibacterial agents, as the use time increases, bacteria may become adapted to the antibacterial components therein, reducing the antibacterial effect. Moreover, there are limitations in the drug selection of current hydrogel dressings, and it is difficult to achieve effective loading and release for some special drugs or treatment requirements. When absorbing the wound exudate, the existing hydrogel wound dressings will adhere to the skin and swell, resulting in wound stretching and expansion, which not only causes pain to the patient, but also increases the risk of bacterial infection due to wound area expansion.

[0005] Mussels can firmly attach to various surfaces in the humid and complex marine environment, thanks to the mussel adhesive protein they secrete. Incorporating mussel adhesive protein into hydrogels can significantly enhance the adhesion between the hydrogels and wound tissues. Compared with ordinary hydrogel dressings, the hydrogels added with mussel adhesive protein can fit more closely to the wounds. Even when the patient is in a mobile state, they are not easily displaced or shed, ensuring the continuity and effectiveness of wound care and reducing the risk of bacterial invasion caused by dressing loosening. Mussel adhesive protein contains multiple dopamine groups, which can promote cell adhesion, spreading, and proliferation. When used as a hydrogel component, it can provide a good growth microenvironment for cells at the wound site. Research shows that under the action of the hydrogel containing mussel adhesive protein, the proliferation rate of cells closely related to wound healing, such as fibroblasts, significantly accelerates, which helps to accelerate the wound healing process, shorten the healing cycle, and reduce the possibility of scar formation. Mussel adhesive protein has certain antibacterial properties and can effectively inhibit the growth of common wound pathogens. This antibacterial effect does not simply rely on killing bacteria, but rather interferes with the bacterial adhesion mechanism to prevent bacteria from colonizing on the wound surface, thereby reducing the probability of wound infection and creating a clean environment for wound healing. Therefore, incorporating mussel adhesive protein into hydrogel dressings can well solve the problem that current hydrogel dressings lack antibacterial properties. However, after the mussel adhesive protein is prepared into a hydrogel, its adhesion will also be affected in a humid environment; in addition, since the hydrogel dressing needs to have a certain mechanical strength to support the wound and at the same time maintain sufficient flexibility to adapt to the movement and deformation of the skin, but the addition of mussel adhesive protein will affect the mechanical properties of the hydrogel. Therefore, the main problem to be solved currently is to make the adhesion of the hydrogel added with mussel adhesive protein persistent and maintain good mechanical properties. Summary of the Invention

[0006] In order to overcome the deficiencies of the prior art, the purpose of this application is to provide a mussel adhesive protein hydrogel dressing with self-healing function to solve the defects existing in the adhesion stability, antibacterial, and anti-inflammatory properties of traditional wound repair materials.

[0007] To solve the above problems, the technical solutions adopted in this application are as follows: The embodiments of this application provide a mussel adhesive protein hydrogel dressing with self-healing function, including a gel system, and the gel system includes a composite hydrogel matrix and a functionalized nano-modified component; wherein The composite hydrogel matrix is a network structure formed by photo-enzyme co-crosslinking of mussel adhesive protein, methacrylated gelatin, and phenylboronic acid-modified gelatin, and the weight ratio of mussel adhesive protein, methacrylated gelatin, and phenylboronic acid-modified gelatin is (2 - 5):(15 - 20):5; The functionalized nano-modified component includes mesoporous silica nanoparticles loaded with small molecule polypeptides and graphene oxide, wherein the graphene oxide is 0.1-0.5 parts by weight in the gel system; the mesoporous silica nanoparticles are 1-3 parts by weight in the gel system, and the polypeptide loading rate is 30-50%.

[0008] As a further preferred solution, the gel system described in the embodiments of the present application further includes an auxiliary component, and the auxiliary component includes a humectant, a temperature-responsive polymer and deionized water, wherein the humectant is 5-10 parts by weight, the temperature-responsive polymer is 2-4 parts by weight, and the deionized water is 30-80 parts by weight; the addition amount of the auxiliary component in the gel system is 8-12 parts by weight.

[0009] As a further preferred solution, the small molecule polypeptides described in the embodiments of the present application are one or more mixtures of GD peptide, EGF, and VEGF; the humectant is one or more mixtures of glycerol, sorbitol, hyaluronic acid, trehalose, polyethylene glycol, and aloe polysaccharide; the temperature-responsive polymer is one or more mixtures of poly(N-isopropylacrylamide), poly[di(ethylene glycol) methacrylate], polyvinylcaprolactam, and poly(ethylene oxide)-poly(propylene oxide) block copolymer.

[0010] As a further preferred solution, the functional modification component described in the embodiments of the present application further includes ε-polylysine-PLGA microspheres, and a hydrophobic network structure fiber membrane is formed on the surface of the gel system by the ε-polylysine-PLGA microspheres.

[0011] As a further preferred solution, the gel system described in the embodiments of the present application further includes a pH-responsive fluorescent probe, and the pH-responsive fluorescent probe is dispersed in the hydrogel in the form of microcapsules.

[0012] The embodiments of the present application also provide a preparation method of a mussel adhesive protein hydrogel dressing with a self-healing function, including: Preparing a mixed hydrogel prepolymer solution: Dissolve methacrylated gelatin and phenylboronic acid-modified gelatin in PBS respectively, mix them in proportion, add a photoinitiator and disperse evenly to obtain a hydrogel prepolymer solution; Mesoporous silica loaded with polypeptides: Mix a small molecule polypeptide solution with nano-mesoporous silica, and vacuum impregnate to adsorb the small molecule polypeptide into the pores of the mesoporous silica, and freeze-dry after treatment to obtain mesoporous silica nanoparticles loaded with small molecule polypeptides; Functional modification of the mixed hydrogel: Add the above-mentioned mesoporous silica nanoparticles loaded with polypeptides into the hydrogel prepolymer solution, and perform ultrasonic treatment and dispersion homogenization in a cold water area to obtain a functionally modified hydrogel prepolymer solution; Preparation of mussel adhesive protein premix: The mussel adhesive protein solution is obtained by dissolving mussel adhesive protein in PBS. Graphene oxide is uniformly dispersed in the mussel adhesive protein solution to form a stable suspension, and then catalytic enzyme is added and dispersed evenly to obtain the mussel adhesive protein premix; Photo-enzyme co-reaction: The above functionalized hydrogel prepolymer solution and the mussel adhesive protein premix are mixed in proportion, and photo-polymerized with blue light to form a primary network, and then incubated to complete the enzyme-catalyzed reaction to obtain a composite hydrogel with a network structure.

[0013] As a further preferred solution, the preparation method of the mussel adhesive protein hydrogel dressing with self-healing function described in the embodiment of the present application further includes spraying ε-polylysine-PLGA microspheres on the surface of the composite hydrogel to form a surface hydrophobic network layer of ε-polylysine. The specific steps are as follows: Preparation of ε-polylysine-PLGA microspheres: First, dissolve PLGA in ethyl acetate to obtain an oil phase; then dissolve ε-polylysine in acetic acid aqueous solution to obtain an aqueous phase; slowly drop the aqueous phase into the oil phase and homogenize in an ice bath to form a primary emulsion; add the primary emulsion to a PBS solution containing polyvinyl alcohol, homogenize to obtain an emulsion, stir the emulsion and volatilize ethyl acetate, collect the microspheres by centrifugation, wash with deionized water, freeze-dry and sieve, and retain microspheres with a particle size of 1.5-2.5 μm; Spraying microspheres to form a surface hydrophobic network layer: Disperse the above microspheres in an ethanol solution containing Tween 80, transfer them into a high-pressure airless spray gun, and spray the microspheres onto the surface of the composite hydrogel; perform hot air annealing to melt the surface of the microspheres and embed them in the dressing surface layer; Among them, the weight ratio of ε-polylysine to PLGA is 1: (3-5).

[0014] As a further preferred solution, the preparation method of the mussel adhesive protein hydrogel dressing with self-healing function described in the embodiment of the present application further includes the step of adding auxiliary components: Dissolve the humectant in deionized water to obtain a humectant solution; then add the temperature-responsive polymer to pure water at 0-5°C and stir magnetically to form a uniform sol; then raise the temperature of the sol to 20-25°C and add the humectant solution drop by drop; obtain an auxiliary composition; Add the auxiliary composition to the functionalized hydrogel prepolymer solution and ultrasonically treat it in an ice bath for 10-20 minutes until it is evenly dispersed.

[0015] As a further preferred solution, in the preparation method described in the embodiments of the present application, when preparing the mussel adhesive protein premixed solution, graphene oxide is uniformly dispersed in the mussel adhesive protein solution by ultrasonic-centrifugation method to form a stable suspension. The ultrasonic power is 280-350W, and the time is 25-35 minutes; the addition amount of the catalytic enzyme is 100-300U / g calculated according to the mass of the mussel adhesive protein.

[0016] As a further preferred solution, the photoinitiator described in the embodiments of the present application is LAP or / and Na-TPO (sodium phenyl-2,4,6-trimethylbenzoylphosphonate), and the catalytic enzyme is one or a mixture of two or more of laccase, tyrosinase, transglutaminase, and lysozyme.

[0017] Compared with the prior art, the beneficial effects of the present invention are as follows: 1. The mussel adhesive protein hydrogel dressing with self-healing function described in the present application combines mussel adhesive protein, methacrylated gelatin and phenylboronic acid modified gelatin, and forms a network structure by photo-enzyme co-crosslinking, endowing the hydrogel dressing with unique self-healing ability; this self-healing property can automatically restore its integrity when the dressing is damaged, thus maintaining the continuity and effectiveness of wound coverage, reducing the frequency of dressing changes, and being beneficial to the wound healing process. By adjusting the ratio of mussel adhesive protein, methacrylated gelatin and phenylboronic acid modified gelatin, both the stability and strength of the gel system are ensured, and good biocompatibility and adhesiveness are also ensured. It helps to achieve a tight fit between the dressing and the wound tissue, and promotes cell proliferation and wound healing.

[0018] 2. The mussel adhesive protein hydrogel dressing with self-healing function described in the present application is added with functionalized nano-modified components, including mesoporous silica nanoparticles loaded with small molecule polypeptides and graphene oxide. The mesoporous silica nanoparticles loaded with small molecule polypeptides not only provide additional structural support, but also promote wound repair and regeneration through the loaded small molecule polypeptides, ensuring the effective release of their biological activity, and helping to accelerate the wound healing process. The trace addition of graphene oxide significantly enhances the conductivity and mechanical strength of the hydrogel, while maintaining good biocompatibility, which helps to promote the micro-current activity at the wound site and can further promote cell growth and tissue repair.

[0019] 3. In the mussel adhesive protein hydrogel dressing with self-healing function described in the present application, phenylboronic acid-modified gelatin is used to replace part of the chemical cross-linking agent with phenylboronic acid-catechol dynamic covalent bonds, endowing the hydrogel with self-healing ability; graphene oxide enhances the adhesion performance of mussel adhesive protein through π-π stacking. And in a further solution, the surface of the composite hydrogel is sprayed with ε-polylysine / PLGA microspheres to form an antibacterial outer layer with hydrophobic function, which can effectively block the invasion of bacteria.

[0020] The present invention will be further described in detail below in conjunction with the accompanying drawings and specific embodiments. Description of the Drawings

[0021] In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the accompanying drawings required for describing the embodiments will be briefly introduced below. Obviously, the accompanying drawings in the following description are only some embodiments of the present invention. For those of ordinary skill in the art, without creative efforts, other accompanying drawings can also be obtained based on these drawings.

[0022] Figure 1 SEM image of the mussel adhesive protein hydrogel dressing with self-healing function described in Embodiment 1 of the present application.

[0023] Figure 2 SEM image of the mussel adhesive protein hydrogel dressing with self-healing function described in Embodiment 2 of the present application.

[0024] Figure 3 SEM image of the mussel adhesive protein hydrogel dressing with self-healing function described in Embodiment 3 of the present application.

[0025] Figure 4 Appearance morphology diagram of the mussel adhesive protein hydrogel dressing with self-healing function described in the embodiments of the present application. Specific Embodiments

[0026] To make the objectives, technical solutions, and advantages of the embodiments of the present application clearer, the technical solutions in the embodiments of the present application will be clearly and completely described below in conjunction with the accompanying drawings in the embodiments of the present application. Obviously, the described embodiments are some, but not all, of the embodiments of the present application. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present application without creative efforts fall within the scope of protection of the present application.

[0027] The term "including" and other equivalent descriptive methods involved in the specification and claims of the present application are all intended to cover non-exclusive inclusion, that is, including both the content clearly described in the specification and claims and the steps or units that are inherent in the product, method, or structure but not described in the specification and claims.

[0028] An embodiment of the present application provides a mussel adhesive protein hydrogel dressing with a self-healing function, which includes a gel system. The gel system includes a composite hydrogel matrix and a functionalized nano-modified component. The composite hydrogel matrix is a network structure formed by photo-enzyme co-crosslinking of mussel adhesive protein, methacrylated gelatin, and phenylboronic acid-modified gelatin. The weight ratio of mussel adhesive protein, methacrylated gelatin, and phenylboronic acid-modified gelatin is (2-5):(15-20):5. The functionalized nano-modified component includes mesoporous silica nanoparticles loaded with small molecule polypeptides and graphene oxide, where the graphene oxide is 0.1-0.5 parts by weight in the gel system; the mesoporous silica nanoparticles are 1-3 parts by weight in the gel system, and the polypeptide loading rate is 30-50%. In the composite hydrogel matrix described in the embodiment of the present application, methacrylated gelatin (GelMA) forms a stable covalent network through photoinitiated polymerization, while the borate bond of phenylboronic acid-modified gelatin (Gel-PBA) has dynamic reversibility. The two cooperate to construct a dual-network structure with both rigidity and flexibility; the borate bond reversibly breaks and re-binds when damaged, realizing the dynamic repair of the internal structure of the material, prolonging the service life of the dressing. The phenylboronic acid group can reversibly bind to substances such as glucose in wound exudate, endowing the gel with pH responsiveness and optimizing the moist environment; mussel adhesive protein (MAP) is rich in catechol groups, which can tightly adhere to the tissue surface through hydrogen bonds, π-π stacking, etc., and at the same time participate in the formation of the dynamic network, improving the overall mechanical strength. Further, in the embodiment of the present application, the purpose of synergistic effect is achieved by adding functionalized nano-components; the high specific surface area mesoporous structure of MSNs can achieve efficient loading of small molecule polypeptides (such as antimicrobial peptides or growth factors) and slowly release them in the wound microenvironment (such as weak acidity or enzyme-triggered). The released polypeptides can inhibit the formation of bacterial biofilms (such as antimicrobial peptides) or activate cell migration / proliferation signal pathways (such as RGD peptides), accelerating wound repair; graphene oxide (GO) has a nanosheet structure, and the nanosheet structure can achieve broad-spectrum antibacterial by physically cutting the bacterial membrane and oxidative stress (generating reactive oxygen species); its conductivity can also promote intercellular electrical signal transmission and accelerate epidermal regeneration; graphene oxide is embedded in the gel network as a nano-filler, and the tensile strength (about 1.5-3 times) and toughness are improved through hydrogen bonds and π-π interactions. In the above scheme, the weight ratio of mussel adhesive protein, methacrylated gelatin (GelMA), and phenylboronic acid-modified gelatin has a significant impact on the performance of the hydrogel dressing.For example, mussel adhesive protein has excellent adhesion and biocompatibility, can closely adhere to wounds, reduce the risk of infection, and promote wound healing. However, an excessive proportion will lead to a decline in the mechanical properties of the gel. When forming a network structure, as the weight ratio of GelMA increases, the mechanical properties such as the tensile strength and toughness of the hydrogel will improve. An appropriate proportion of GelMA helps to achieve a good cross-linking effect, thereby improving the overall performance of the hydrogel. An appropriate amount of phenylboronic acid-modified gelatin can significantly improve the self-healing ability of the hydrogel. In order to achieve the balance and optimization of adhesion, biocompatibility, mechanical properties, and self-healing function, in the embodiments of the present application, the weight ratio of mussel adhesive protein, methacrylated gelatin, and phenylboronic acid-modified gelatin is (2-5):(15-20):5. In a preferred embodiment, the weight ratio of the mussel adhesive protein, methacrylated gelatin, and phenylboronic acid-modified gelatin is 4:20:5. Graphene oxide can enhance the adhesion performance of mussel adhesive protein through π-π stacking. Graphene oxide has oxygen-containing functional groups and can also act as a physical cross-linking agent to bind to -COOH in the gel system through hydrogen bonds, thereby increasing the cross-linking density and intermolecular force of the hydrogel, increasing the tensile strength and elongation at break of the hydrogel, and improving its overall mechanical properties. Therefore, when preparing the hydrogel dressing, appropriately increasing the content of GO can enhance its mechanical stability and make it more suitable for application scenarios that need to withstand a certain amount of external force or pressure. In addition, the addition of GO can also significantly improve the thermal stability of the hydrogel dressing and improve the elastic recovery ability of the hydrogel dressing. Graphene oxide (GO) is both a reinforcing agent (mechanical, electrical conductivity) and a functional carrier (antibacterial, photothermal response) in the dynamic covalent cross-linked hydrogel system. A slight change in its mass ratio will significantly affect the key performance of the dressing. When the mass ratio of GO is too low, the rigid skeleton effect of GO weakens, the storage modulus of the hydrogel decreases, and the tensile strength of the dressing also decreases accordingly. At the same time, the insufficient participation of the carboxyl groups on the surface of GO in the phenylboronic acid dynamic covalent bond will also affect the network self-healing efficiency. However, if the dosage ratio of graphene oxide (GO) is too high, there will also be many negative effects. For example, excessive filling of GO results in too small pore size of the hydrogel, which will hinder oxygen / nutrient penetration and inhibit cell migration. The exposure probability of the active edges (oxygen-containing functional groups) of GO increases, resulting in a decrease in the survival rate of fibroblasts. Therefore, in the embodiments of the present application, the weight portion of graphene oxide in the gel system needs to be controlled at 0.1-0.5 parts; in some preferred embodiments, the graphene oxide in the gel system is 0.2-0.4 parts by weight.

[0029] Further, the gel system described in the embodiments of the present application further includes auxiliary components, and the auxiliary components include a humectant, a temperature-responsive polymer, and deionized water. Among them, the humectant is 5-10 parts by weight, the temperature-responsive polymer is 2-4 parts, and deionized water is 50-80 parts; the addition amount of the auxiliary components in the gel system is 8-12 parts by weight. The small molecule polypeptide is one or more mixtures of GD peptide, EGF, and VEGF. The humectant is one or more mixtures of glycerol, sorbitol, hyaluronic acid, trehalose, polyethylene glycol, and aloe polysaccharide. In some embodiments, glycerol is added to the hydrogel dressing. Glycerol can form a hydrogen bond network with water molecules through hydroxyl groups to increase the water absorption rate; at the same time, it reduces the glass transition temperature of the hydrogel and improves the flexibility of the hydrogel dressing. However, too high a dosage will cause a sticky feeling and interfere with the oxidative cross-linking efficiency of mussel adhesive protein. Therefore, in the gel system of the embodiments of the present application. Sorbitol has low viscosity characteristics, can maintain high porosity, is suitable for highly exuding wounds, and also has the effects of scavenging free radicals and prolonging the activity of polypeptides. In a preferred embodiment, sorbitol is compounded with glycerol (volume ratio of 3:2), which can effectively balance moisture retention and air permeability. Hyaluronic acid has super water retention capacity, can activate CD44 receptors to promote fibroblast migration, and it can dynamically bind to the phenylboronic acid of GelPB in the gel system to enhance network stability. However, too much dosage will cause a decrease in the gel cross-linking density. Therefore, in the gel system. In the embodiments of the present application, the humectant in the auxiliary components may also include trehalose, polyethylene glycol, aloe polysaccharide, etc. The role of trehalose is equivalent to a biomolecule protectant, which stabilizes the polypeptide conformation through the "water displacement" mechanism; polyethylene glycol is mainly used for film-forming and osmotic pressure regulation, forms a semi-permeable membrane on the wound surface, and inhibits bacterial adhesion; aloe polysaccharide has dual anti-inflammatory and moisturizing effects, and can effectively inhibit the growth of G+ bacteria and reduce the erythema area. In the embodiments of the present application, a temperature-responsive polymer is also added to the auxiliary components. The temperature-responsive polymer is a type of intelligent polymer material that can respond to changes in the external temperature. Such polymers have a lower critical solution temperature (LCST) or cloud point (CT) in aqueous solution. When the temperature is lower than the LCST, the polymer chain is hydrophilic, dissolves in water and is in an extended state, and the volume expands; when the temperature is higher than the LCST, the polymer chain is hydrophobic, phase separation occurs, the chain is in a coiled state, and the volume shrinks. The temperature-responsive polymer may specifically be selected from one or more mixtures of poly(N-isopropylacrylamide), poly[bis(ethylene glycol) methacrylate], polyvinylcaprolactam, and poly(ethylene oxide)-poly(propylene oxide) block copolymer. In the embodiments of the present application, by precisely controlling the addition amounts and ratios of the humectant, temperature-responsive polymer, deionized water, and small molecule polypeptide, fine regulation and optimization of the performance of the hydrogel dressing can be achieved. Not only can the comprehensive performance of the hydrogel dressing be improved, but also the needs of different wound types and healing stages can be met.

[0030] Furthermore, on the basis of the above solution, the functional modification component described in the embodiments of the present application further includes ε-polylysine-PLGA microspheres. The ε-polylysine-PLGA microspheres are sprayed on the surface of the gel system to form a fiber membrane with a hydrophobic network structure, that is, an antibacterial outer layer is formed. Combining the above solution, the mussel adhesive protein hydrogel dressing described in the embodiments of the present application can form a biomimetic structure, specifically including a hydrophobic network layer formed by ε-polylysine-PLGA microspheres as the outer antibacterial layer for blocking the invasion of bacteria; mesoporous silica nanoparticles loaded with small molecule polypeptides are assembled in the network structure formed by the photo-enzyme co-crosslinking of mussel adhesive protein, methacrylated gelatin and phenylboronic acid modified gelatin to form a shrinkage pore support layer in the middle to ensure the stability of the pores under mechanical stress; the π-π stacking interaction and dopamine oxidative crosslinking of mussel adhesive protein and graphene oxide form a strong adhesion interface to form a high adhesion matrix layer.

[0031] As a further preferred solution, on the basis of the above embodiment solution, in some embodiments, a pH-responsive fluorescent probe is further added to the gel system, and the pH-responsive fluorescent probe is dispersed in the hydrogel in the form of microcapsules. Specifically, the pH-responsive fluorescent probe is a microencapsulated bromocresol green derivative, the shell material is calcium alginate, and the wall thickness is 0.5-1 μm. When pH>7, the fluorescence emission peak redshifts from 515 nm to 620 nm.

[0032] The embodiments of the present application also provide a preparation method of a mussel adhesive protein hydrogel dressing with self-healing function, including: Preparing a mixed hydrogel prepolymer solution: Dissolve methacrylated gelatin and phenylboronic acid modified gelatin with PBS respectively. After dissolution, mix the methacrylated gelatin and phenylboronic acid modified gelatin in proportion, add a photoinitiator and disperse evenly to obtain a hydrogel prepolymer solution; Mesoporous silica loaded with polypeptides: Mix the small molecule polypeptide solution and nano mesoporous silica, and vacuum impregnate to adsorb the small molecule polypeptides into the pores of the mesoporous silica. After completion, freeze-dry to obtain mesoporous silica nanoparticles loaded with small molecule polypeptides; Functional modification of the mixed hydrogel: Add the above-mentioned mesoporous silica nanoparticles loaded with polypeptides to the hydrogel prepolymer solution, and perform ultrasonic treatment and dispersion homogenization in a cold water bath to obtain a functionally modified hydrogel prepolymer solution; Preparing a mussel adhesive protein premixed solution: The mussel adhesive protein solution obtained by dissolving mussel adhesive protein in PBS, uniformly disperse graphene oxide in the mussel adhesive protein solution to form a stable suspension, and then add a catalytic enzyme and disperse evenly to obtain a mussel adhesive protein premixed solution; Photo-enzyme co-reaction: The above functionalized hydrogel prepolymer solution and mucin premix solution are mixed in proportion, irradiated with blue light for photopolymerization to form a primary network, and then incubated to complete the enzyme-catalyzed reaction to obtain a composite hydrogel with a network structure.

[0033] In a specific embodiment of the present application, when preparing the mixed hydrogel prepolymer solution, methacrylated gelatin and phenylboronic acid-modified gelatin are dissolved separately in PBS with a pH of 7.0 - 7.5. The dissolution process is carried out in a water bath at 30 - 40 °C to avoid gelatin deformation, and the viscosity is controlled at 500 - 800 mPa·s; the dosage of the photoinitiator is 0.1 wt% - 0.3 wt% of the weight of the gel system. Preferably, after obtaining the hydrogel prepolymer solution, bubbles are removed by vacuum centrifugation, and the centrifugation conditions are 1800 - 2500 rpm for 8 - 15 minutes to avoid structural defects after curing. In some preferred embodiments, when loading polypeptides on mesoporous silica, small molecule polypeptides and nano-mesoporous silica are mixed in a weight ratio of (0.5 - 1.0):2. The pore size of the mesoporous silica is 4 - 6 nm, and the specific surface area is 800 - 1000 m² / g; the vacuum impregnation conditions are: -0.1 MPa, oscillating adsorption at 20 - 27.5 °C for 10 - 15 hours; freeze-drying adopts a combination of pre-freezing and freeze-drying steps. The pre-freezing conditions are -80 °C for 20 - 25 hours, and the vacuum freeze-drying conditions are 0.1 Pa for 20 - 25 hours, with the residual moisture ≤ 2%. In a specific embodiment of the present application, when functionalizing the mixed hydrogel, probe sonication is used, and the conditions are 180 - 250 W for 25 - 35 minutes. The temperature of the cold water area is controlled ≤ 25 °C, and the particle size distribution CV ≤ 15%. In the preparation method described in the embodiments of the present application, when preparing the mussel mucin premix solution, graphene oxide is uniformly dispersed in the mussel mucin solution by ultrasonic-centrifugation to form a stable suspension. The size of the graphene oxide sheet layer is 200 - 500 nm. The ultrasonic treatment is carried out in a water bath at a temperature ≤ 25 °C to prevent protein denaturation caused by overheating. The ultrasonic power is 280 - 350 W, and the time is 25 - 35 minutes; the addition amount of the catalytic enzyme is 50 - 100 U / mL. The enzyme solution is prepared and used immediately and stored at 4 °C for ≤ 6 hours to avoid inactivation. In the embodiments of the present application, during the photo-enzyme co-reaction, blue light with a wavelength of 405 nm and an intensity of 10 mW / cm² is irradiated for 50 - 70 seconds. The incubation conditions are a constant temperature and humidity incubator at 37 °C, and the reaction time is 1.5 - 2.5 hours, with the humidity ≥ 90% to prevent dehydration.

[0034] Furthermore, on the basis of the above scheme, in some embodiments, the preparation method of the mussel mucin hydrogel dressing with self-healing function described in the embodiments of the present application further includes spraying ε-polylysine-PLGA microspheres on the surface of the composite hydrogel to form a surface hydrophobic network layer of ε-polylysine. The specific steps are as follows: Preparation of ε-polylysine-PLGA microspheres: First, dissolve PLGA in ethyl acetate to obtain the oil phase; then dissolve ε-polylysine in acetic acid aqueous solution to get the water phase; slowly drip the water phase into the oil phase, homogenize in an ice bath to form a primary emulsion; add the primary emulsion to a PBS solution containing polyvinyl alcohol, homogenize to obtain an emulsion, stir the emulsion and evaporate ethyl acetate, collect the microspheres after centrifugation, wash with deionized water, sieve after freeze-drying, and retain microspheres with a particle size of 1.5 - 2.5 μm; in this step, when PLGA is dissolved in ethyl acetate, magnetic stirring is used to promote dissolution, and the conditions of magnetic stirring are: 500 - 800 rpm, 25 - 30 °C, 1.5 - 2.5 hours, and the concentration of PLGA after dissolution is 10 - 15% w / v; when ε-polylysine is dissolved in acetic acid aqueous solution, ultrasonic assistance is used for dissolution, and the ultrasonic conditions are: frequency 35 - 45 kHz, power 100 - 150 W, time 10 - 20 minutes, and the concentration of ε-polylysine after dissolution is 2 - 4% w / v; the process of forming the primary emulsion is carried out in a water bath with a temperature of ≤10 °C, the dropping rate of the water phase is 0.3 - 0.8 mL / min, and a homogenizer is used to accelerate dispersion during the dropping process, and the rotation speed of the homogenizer is 10000 - 12000 rpm; Spray the microspheres to form a surface hydrophobic network layer: Disperse the above microspheres in an ethanol solution containing Tween 80, transfer them into a high-pressure airless spray gun, and spray the microspheres onto the surface of the composite hydrogel; perform hot air annealing to melt the surface of the microspheres and embed them in the dressing surface layer; Among them, the weight ratio of ε-polylysine to PLGA is 1: (3 - 5).

[0035] As a further preferred scheme, the preparation method of the mussel adhesive protein hydrogel dressing with self-healing function described in the embodiment of the present application further includes the step of adding auxiliary components: Dissolve the moisturizer in deionized water to obtain a moisturizing solution; then add the temperature-responsive polymer to pure water at 0 - 5 °C, and stir magnetically to form a uniform sol; raise the temperature of the sol to 20 - 25 °C, and dropwise add the moisturizing solution; obtain an auxiliary composition; Add the auxiliary composition to the functionalized modified hydrogel prepolymer solution, and perform ultrasonic treatment in an ice bath for 10 - 20 minutes until evenly dispersed.

[0036] Further, in some embodiments, the photoinitiator used is LAP or / and Na-TPO (sodium phenyl-2,4,6-trimethylbenzoylphosphinate). Among them, LAP matches 405 nm blue light, which can avoid ultraviolet damage to bioactive components, has high initiation efficiency, and its degradation products are lactic acid and pyruvic acid, with low toxicity. Na-TPO has deep curing ability and is suitable for thick hydrogels or tissue repair scaffolds; it has long-wavelength adaptability and can respond to a wide-spectrum light source of 365 - 420 nm. Its radical activity is high, and its antioxidant interference ability is better than that of LAP. Preferably, in some embodiments, a mixture of LAP and Na-TPO (mixed in a weight ratio of 1:1) is used as the photoinitiator to combine rapid surface curing and deep penetration. In the embodiments of the present application, the catalytic enzyme is one or more of laccase, tyrosinase, transglutaminase, and lysozyme. Among them, laccase has specificity for dopa oxidation, efficiently catalyzes the oxidation of dopa (DOPA) in mussel adhesive protein to dopaquinone, forms a covalent crosslinking network, and promotes the construction of a dynamic crosslinking and self-repair network of mussel adhesive protein; moreover, laccase also has an antibacterial auxiliary effect, and its oxidation products inhibit the formation of bacterial biofilms. Tyrosinase catalyzes the hydroxylation and crosslinking of tyrosine / dopa, is suitable for collagen, silk fibroin, etc., and is active and stable at 25 - 37 °C, avoiding high-temperature damage to cells; it can regulate melanin production and assist in the aesthetic repair of wounds. Transglutaminase enhances the mechanical strength of the gelatin / collagen network through ε-(γ-glutamyl) lysine bonds. Lysozyme can lyse the cell walls of Gram-positive bacteria to achieve efficient antibacterial. When combined with a photoinitiator, it can achieve synchronous antibacterial and gel formation. Preferably, in some embodiments, the catalytic enzyme is a composite enzyme of laccase and lysozyme, and the weight ratio of the two is 2:1; it can simultaneously achieve the crosslinking of mussel adhesive protein and long-term antibacterial. The addition amount of the catalytic enzyme is calculated as 100 - 300 U / g based on the mass of mussel adhesive protein.

[0037] Example 1 This example provides a mussel adhesive protein hydrogel dressing with self-repair function, including a gel system. The gel system includes a composite hydrogel matrix, a functionalized nano-modified component, and an auxiliary component; wherein the composite hydrogel matrix is a network structure formed by the photo-enzyme co-crosslinking of mussel adhesive protein, methacrylated gelatin, and phenylboronic acid-modified gelatin; the functionalized nano-modified component includes mesoporous silica nanoparticles loaded with small molecule polypeptides and graphene oxide. In this example, the specific composition and dosage are as shown in Table 1 below.

[0038] Table 1: Composition and dosage table of the examples The preparation method of the mussel adhesive protein hydrogel dressing with self-repair function is as follows: Preparation of mixed hydrogel prepolymer solution: Add 20 g of GelMA to 100 mL of PBS (pH 7.4, 37 °C), and stir in a water bath until dissolved (rotation speed 200 rpm, 30 min); dissolve 5 g of phenylboronic acid-modified gelatin in 50 mL of PBS (pH 7.4), and after stirring and dissolving in a 50 °C water bath, mix it with the methacrylated gelatin solution; add 0.15 g of LAP and 0.15 g of Na-TPO, stir for 10 min until completely dissolved, and remove air bubbles by vacuum centrifugation (2000 rpm, 10 min) to obtain a clear prepolymer solution; Mesoporous silica loaded with polypeptide: Dissolve 0.5 g of small molecule polypeptide (RGD polypeptide) in 10 mL of deionized water, add 2 g of mesoporous silica nanoparticles (particle size 50 - 100 nm), and vacuum impregnate to adsorb the small molecule polypeptide into the pores of mesoporous silica (vacuum impregnation conditions -0.1 MPa, 25 °C, oscillate for 12 h), after treatment, pre-freeze at -80 °C for 24 h, and freeze-dry at 0.1 Pa for 24 h to obtain mesoporous silica nanoparticles loaded with small molecule polypeptide; Preparation of auxiliary composition: Dissolve 5 g of glycerol and 3 g of sorbitol in 30 g of deionized water to obtain a moisturizing solution; add 2 g of PNIPAM to 10 mL of ice water (0 - 5 °C), stir magnetically for 30 min to form a sol, heat the sol to 25 °C, and gradually add the moisturizing solution, mix well and then ultrasonicate in an ice bath for 15 min (power 200 W); obtain the auxiliary composition; Functional modification of mixed hydrogel: Add the above-mentioned mesoporous silica nanoparticles loaded with polypeptide to the hydrogel prepolymer solution, ultrasonically disperse and homogenize in a cold water bath (≤25 °C) (ultrasonic power 250 W, time 20 min), then add 10 g of the above-mentioned auxiliary composition, and stir evenly; obtain the functionalized modified hydrogel prepolymer solution; Preparation of mussel adhesive protein premixed solution: 4 g of freeze-dried mussel adhesive protein is ultrasonically dispersed (300 W, 30 min, ≤25 °C) in PBS to obtain a mussel adhesive protein gel with a concentration of 10 mg / mL, add 0.2 g of GO, and remove aggregates by ultrasonic centrifugation (3000 rpm, 10 min) to form a stable suspension, then add 400 U of laccase (freshly prepared enzyme solution, stored at 4 °C for ≤6 h) and mix well to obtain the mussel adhesive protein premixed solution; Photo-enzyme synergistic reaction: Mix the above-mentioned functionalized modified hydrogel prepolymer solution and mussel adhesive protein premixed solution, irradiate with blue light (405 nm, 10 mW / cm², 60 s) to form a primary network, transfer to a 37 °C humid box (humidity 95%), and incubate with the enzyme for 2 h to obtain a composite hydrogel with a network structure; Preparation of ε-polylysine-PLGA microspheres: Dissolve 4 g of PLGA in 40 mL of ethyl acetate (stirring speed 500 rpm, 25 °C, 2 h) to prepare the oil phase; dissolve 1 g of ε-polylysine in 10 mL of 1% acetic acid water (ultrasonic for 10 min, power 100 W) to prepare the aqueous phase; drop the aqueous phase into the oil phase (0.5 mL / min, ≤10 °C), homogenize (10000 rpm, 10 min) to form an emulsion, pour the emulsion into 500 mL of PBS containing 2% PVA, homogenize (5000 rpm, 20 min), stir to evaporate the solvent (4 h), and obtain microspheres after centrifugation (8000 rpm, 15 min), washing with water, freeze-drying, and sieving (2 μm); Spray the microspheres to form a surface hydrophobic network layer: Disperse the microspheres in 50% ethanol (containing 0.5% Tween 80, concentration 10 mg / mL), and evenly spray them onto the gel surface with a high-pressure spray gun (0.3 MPa) to a thickness of 30 μm, followed by hot air annealing (50 °C, 30 min). The microspheres melt and embed in the surface layer to form a surface hydrophobic network layer.

[0039] Figure 1 This is the SEM image of the mussel adhesive protein hydrogel dressing with self-healing function described in this example. From Figure 1 it can be seen that the mussel adhesive protein hydrogel dressing presents a porous network structure. These pores are of different sizes and are interconnected to form a complex three-dimensional network. This porous structure provides a large specific surface area for the hydrogel dressing, which is beneficial for material exchange with surrounding tissues. For example, it can better absorb wound exudate, and at the same time, it is also convenient for nutrients and drugs. The interconnected pores and network structure provide conditions for the self-healing of the hydrogel; when the hydrogel is damaged, the molecular chains inside it can interact through the pores, rewind and crosslink to achieve self-healing. The mussel adhesive protein itself has special adhesion properties and can play a better role in this structure to help the hydrogel re-establish connections at the damaged site.

[0040] Example 2 This example provides a mussel adhesive protein hydrogel dressing with self-healing function, including a gel system. The gel system includes a composite hydrogel matrix, a functionalized nano-modified component, and an auxiliary component; among them, the composite hydrogel matrix is a network structure formed by the photocatalytic-enzymatic co-crosslinking of mussel adhesive protein, methacrylated gelatin, and phenylboronic acid-modified gelatin; the functionalized nano-modified component includes mesoporous silica nanoparticles loaded with small molecule polypeptides and graphene oxide. The specific composition and dosage are shown in Table 2 below.

[0041] Table 2: Composition and dosage table of Example 2 The preparation method of the mussel adhesive protein hydrogel dressing with self-healing function is as follows: Preparation of the mixed hydrogel prepolymer solution: Add 15 g of GelMA to 120 mL of PBS (pH 7.2, 37 °C), and stir in a water bath until dissolved (rotation speed 200 rpm, 30 min); add 5 g of phenylboronic acid-modified gelatin to 50 mL of PBS (pH 7.4), soak in water and swell, then disperse by magnetic stirring in a 50 °C water bath (300 rpm), gradually add deionized water until it becomes transparent, and then mix with the methacrylated gelatin solution; add 0.2 g of Na-TPO, stir for 15 min until completely dissolved to obtain the prepolymer solution. Ultrasonically disperse 0.1 g of fluorescent microcapsules (power 180 W, 10 min) into the prepolymer solution, and remove bubbles under vacuum (1800 rpm, 8 min); Mesoporous silica loaded with polypeptide: Dissolve 1.5 g of small molecule polypeptide (EGF) in 15 mL of deionized water, add 3 g of mesoporous silica nanoparticles (particle size 50 - 100 nm), and perform vacuum impregnation to adsorb the small molecule polypeptide into the pores of the mesoporous silica (vacuum impregnation conditions: -0.1 MPa, 20 °C, oscillate for 15 h). After treatment, pre-freeze at -80 °C for 20 h, and freeze-dry at 0.1 Pa for 20 h to obtain mesoporous silica nanoparticles loaded with small molecule polypeptide; Preparation of the auxiliary composition: Dissolve 4 g of hyaluronic acid and 4 g of trehalose in 20 g of deionized water to obtain a moisturizing gel; add 3 g of polyvinylcaprolactam to 10 mL of ice water (0 - 5 °C), stir magnetically for 40 min to form a sol, heat the sol to 20 °C, add the moisturizing gel to the sol, stir magnetically and mix evenly, and then perform ice bath ultrasonic treatment for 10 min (power 200 W); obtain the auxiliary composition; Functional modification of the mixed hydrogel: Add the above-mentioned mesoporous silica nanoparticles loaded with polypeptide to the hydrogel prepolymer solution, perform ultrasonic treatment in a cold water bath (≤25 °C) to disperse and homogenize (ultrasonic power 250 W, time 20 min), then add 12 g of the above-mentioned auxiliary composition, and stir evenly; obtain the functionally modified hydrogel prepolymer solution; Preparation of the mussel adhesive protein premixed solution: Disperse 2 g of freeze-dried mussel adhesive protein by ultrasonic treatment (300 W, 30 min, ≤25 °C) in PBS to obtain a mussel adhesive protein gel with a concentration of 5 mg / mL. Add 0.2 g of GO, perform ultrasonic - centrifugation (3000 rpm, 10 min) to remove aggregates, form a stable suspension, and then add 400 U of laccase (freshly prepared enzyme solution, stored at 4 °C for ≤6 h) and mix evenly to obtain the mussel adhesive protein premixed solution; Photo - enzyme co - reaction: Mix the above-mentioned functionally modified hydrogel prepolymer solution and the mussel adhesive protein premixed solution, irradiate with blue light (405 nm, 10 mW / cm², 60 s) to form a primary network, transfer to a 37 °C humid box (humidity 95%), and incubate with the enzyme for 2 h to obtain a composite hydrogel with a network structure.

[0042] Figure 2SEM image of the mussel adhesive protein hydrogel dressing with self - healing function described in Example 2 Figure 2 It presents a three - dimensional porous structure with interconnected pores. The white area is the hydrogel skeleton (including the cross - linked network of mussel adhesive protein and polymer), and the black area is the pore. The porous network allows the molecular chains (including the adhesion groups of mussel adhesive protein) at the damaged site to contact and recombine with each other through the pores, and realizes the repair by means of the adhesiveness of the mucin and the dynamic cross - linking of the polymer chains.

[0043] Example 3 This example provides a mussel adhesive protein hydrogel dressing with self - healing function, which includes a gel system. The gel system includes a composite hydrogel matrix, a functionalized nano - modification component and an auxiliary component. Among them, the composite hydrogel matrix is a network structure formed by the photocatalytic - enzymatic co - crosslinking of mussel adhesive protein, methacrylated gelatin and phenylboronic acid - modified gelatin. The functionalized nano - modification component includes mesoporous silica nanoparticles loaded with small - molecule polypeptides and graphene oxide. The specific composition and dosage are shown in Table 3 below.

[0044] Table 3: Composition and dosage table of Example 3 The preparation method of the mussel adhesive protein hydrogel dressing with self - healing function is as follows: Prepare a mixed hydrogel prepolymer solution: Dissolve 18 g of GelMA and 5 g of phenylboronic acid - modified gelatin in 100 mL of PBS (pH 7.4, 40 °C), stir in a water bath until dissolved (rotation speed 200 rpm, 30 min); add 0.2 g of LAP, stir until clear, and remove bubbles under vacuum (2500 rpm, 15 min) to obtain a clear prepolymer solution; Mesoporous silica loaded with polypeptides: Dissolve 0.3 g of small - molecule polypeptide (LL - 37 peptide) in 10 mL of deionized water, add 1 g of mesoporous silica nanoparticles, and perform vacuum impregnation to adsorb the small - molecule polypeptide into the pores of the mesoporous silica (vacuum impregnation conditions: - 0.1 MPa, 27.5 °C, oscillate for 10 h). After treatment, pre - freeze at - 80 °C for 24 h and lyophilize at 0.1 Pa for 24 h to obtain mesoporous silica nanoparticles loaded with small - molecule polypeptides; Functional modification of the mixed hydrogel: Add the above - mentioned mesoporous silica nanoparticles loaded with polypeptides to the hydrogel prepolymer solution, and perform ultrasonic treatment and dispersion homogenization in a cold water bath (≤25 °C) (ultrasonic power 250 W, time 20 min) to obtain a functionally modified hydrogel prepolymer solution; Preparation of mussel adhesive protein premixed solution: 5 g of freeze-dried mussel adhesive protein was ultrasonically dispersed (300 W, 30 min, ≤25 °C) in 20 mL of PBS to obtain a mussel adhesive protein solution. 0.2 g of GO was added, and ultrasonic centrifugation (3000 rpm, 10 min) was performed to remove aggregates, forming a stable suspension. Then, 400 U of laccase (freshly prepared enzyme solution, stored at 4 °C for ≤6 h) was added and mixed evenly to obtain a mussel adhesive protein premixed solution; Photo-enzyme co-reaction: The above functionalized hydrogel prepolymer solution was mixed with the mussel adhesive protein premixed solution, and irradiated with blue light (405 nm, 10 mW / cm², 60 s) to form a primary network. It was transferred to a 37 °C humid box (humidity 95%) and incubated with the enzyme for 2 h to obtain a composite hydrogel with a network structure; Preparation of ε-polylysine-PLGA microspheres: 3 g of PLGA was dissolved in 30 mL of ethyl acetate (stirring speed 800 rpm, 30 °C, 1.5 h) to prepare an oil phase; 1 g of ε-polylysine was dissolved in 5 mL of 1% acetic acid water (ultrasonic for 20 min, power 150 W) to prepare an aqueous phase; the aqueous phase was dropped into the oil phase (0.8 mL / min, ≤5 °C), and homogenized (6000 rpm, 15 min) to form an emulsion. The emulsion was poured into 600 mL of PBS containing 3% PVA, homogenized (6000 rpm, 15 min), stirred to evaporate the solvent (4 h), and after centrifugation (10000 rpm, 10 min), washing, freeze-drying, and sieving (2.5 μm), microspheres were obtained; Spraying microspheres to form a surface hydrophobic network layer: The microspheres were dispersed in 60% ethanol (containing 1% Tween 80, concentration 15 mg / mL), and evenly sprayed onto the gel surface with a high-pressure spray gun (0.3 MPa) to a thickness of 50 μm, followed by hot air annealing (55 °C, 25 min). The microspheres were melted and embedded in the surface layer to form a surface hydrophobic network layer.

[0045] Figure 3 This is the SEM image of the mussel adhesive protein hydrogel dressing with self-healing function described in Example 3. Similarly Figure 3 The network structure of the three-dimensional porous structure can also be clearly seen.

[0046] Performance testing The mussel adhesive protein hydrogel dressing with self-healing function obtained in Example 1 was subjected to performance testing. The specific test items and methods are as follows: 1. Tensile property and compressive property testing After the mussel adhesive protein hydrogel dressing with self-healing function was dried in an oven at 50 °C to remove moisture, it was cut into strips with a length of 50 mm, a width of 10 mm, and a thickness of 3 mm. A Stema S6 series electronic universal testing machine was used for tensile strength testing and compressive strength testing. The test strain rate was set to 50 mm / min, and each test was performed 5 times, and the average value was calculated.

[0047] 2. Adhesion Strength (Peel) Test After the mussel adhesive protein hydrogel dressing with self-healing function is dried in an oven at 50 °C to remove moisture, it is cut into strips with a length of 25 mm, a width of 20 mm, and a thickness of 3 mm. The adhesion performance is tested on a glass plate and a polytetrafluoroethylene plate respectively on an electronic universal testing machine at a loading rate of 50 mm / min and a force of 100 N.

[0048] 3. Self-Healing Performance Test After the mussel adhesive protein hydrogel dressing with self-healing function is dried in an oven at 50 °C to remove moisture, it is cut into strips with a length of 25 mm, a width of 20 mm, and a thickness of 10 mm. After the strip is cut into two pieces with a blade, they are immediately brought into contact without external force, and the self-healing process of the hydrogel is observed with a microscope, and the healing time after complete healing without visible cracks is recorded.

[0049] 4. Antibacterial Activity Test Using Gram-positive Staphylococcus aureus (CICC 10145) and Gram-negative Escherichia coli AB93154 as test objects, the surface antibacterial activity of the mussel adhesive protein hydrogel is tested. The specific operation is to place the composite hydrogel into a 36-well plate, and add 25 µL of a bacterial dispersion with a concentration of 1×10 7 CFU / mL dropwise onto the surface of the mussel adhesive protein hydrogel. After incubating at 37 °C for 2 h, the surface of the hydrogel is rinsed with sterile physiological saline to redisperse the bacteria; 100 µL of the test strain suspension is pipetted onto an LB solid medium, and quickly spread evenly with a sterile spreading rod, and cultured at 37 °C for 24 h, and the number of viable bacteria is recorded.

[0050] The performance test results of Examples 1-3 are shown in Table 4.

[0051] Table 4: Performance Test of the Hydrogel Dressings Described in Examples 1-3 The above embodiments are only the preferred embodiments of the present invention and cannot be used to limit the scope of protection of the present invention. Any non-substantial changes and substitutions made by those skilled in the art based on the present invention fall within the scope of protection required by the present invention.

Claims

1. A mussel adhesive protein hydrogel dressing with self-healing function, characterized in that, It includes a gel system, and the gel system includes a composite hydrogel matrix and a functionalized nano-modified component; wherein the composite hydrogel matrix is a network structure formed by the co-crosslinking of mussel adhesive protein, methacrylated gelatin, and phenylboronic acid-modified gelatin through photo-enzyme synergistic crosslinking. The weight ratio of mussel adhesive protein, methacrylated gelatin, and phenylboronic acid-modified gelatin is (2-5):(15-20):5; the functionalized nano-modified component includes mesoporous silica nanoparticles loaded with small molecule polypeptides and graphene oxide, wherein graphene oxide is 0.1-0.5 parts by weight in the gel system; mesoporous silica nanoparticles are 1-3 parts by weight in the gel system, and the polypeptide loading rate is 30-50%.

2. The mussel adhesive protein hydrogel dressing with self-healing function according to claim 1, characterized in that, The gel system further includes an auxiliary component, and the auxiliary component includes a humectant, a temperature-responsive polymer, and deionized water. Among them, the humectant is 5-10 parts by weight, the temperature-responsive polymer is 2-4 parts by weight, and deionized water is 30-80 parts by weight; the addition amount of the auxiliary component in the gel system is 8-12 parts by weight.

3. The mussel adhesive protein hydrogel dressing with self-healing function according to claim 2, characterized in that, The small molecule polypeptide is one or a mixture of two or more of GD peptide, EGF, and VEGF; the humectant is one or a mixture of two or more of glycerol, sorbitol, hyaluronic acid, trehalose, polyethylene glycol, and aloe polysaccharide; the temperature-responsive polymer is one or a mixture of two or more of poly(N-isopropylacrylamide), poly[di(ethylene glycol) methacrylate], polyvinylcaprolactam, and poly(ethylene oxide)-poly(propylene oxide) block copolymer.

4. The mussel adhesive protein hydrogel dressing with self-healing function according to claim 1, characterized in that, The functional modification component further includes ε-polylysine-PLGA microspheres, and the ε-polylysine-PLGA microspheres form a fibrous membrane with a hydrophobic network structure on the surface of the gel system.

5. The mussel adhesive protein hydrogel dressing with self-healing function according to any one of claims 1-4, characterized in that, The gel system further includes a pH-responsive fluorescent probe, and the pH-responsive fluorescent probe is dispersed in the hydrogel in the form of microcapsules.

6. A preparation method of a mussel adhesive protein hydrogel dressing with a self-healing function as described in any one of claims 1-5, characterized in that, It includes: Preparing a mixed hydrogel prepolymer solution: Dissolve methacrylated gelatin and phenylboronic acid-modified gelatin in PBS respectively, mix them in proportion, add a photoinitiator and disperse evenly to obtain a hydrogel prepolymer solution; Mesoporous silica loading polypeptide: Mix the small molecule polypeptide solution with nano-mesoporous silica, vacuum impregnate to adsorb the small molecule polypeptide into the pores of mesoporous silica, and freeze-dry after treatment to obtain mesoporous silica nanoparticles loaded with small molecule polypeptides; Functional modification of the mixed hydrogel: Add the above-mentioned mesoporous silica nanoparticles loaded with polypeptides to the hydrogel prepolymer solution, and perform ultrasonic treatment and dispersion homogenization in a cold water bath to obtain a functionally modified hydrogel prepolymer solution; Preparing a mussel adhesive protein premixed solution: The mussel adhesive protein solution obtained by dissolving mussel adhesive protein in PBS, uniformly disperse graphene oxide in the mussel adhesive protein solution to form a stable suspension, and then add a catalytic enzyme and disperse evenly to obtain a mussel adhesive protein premixed solution; Photo-enzyme synergistic reaction: Mix the above-mentioned functionally modified hydrogel prepolymer solution and the mussel adhesive protein premixed solution in proportion, irradiate with blue light for photopolymerization to form a primary network, and then incubate to complete the enzyme-catalyzed reaction to obtain a composite hydrogel with a network structure.

7. The preparation method of the mussel adhesive protein hydrogel dressing with self-healing function according to claim 6, characterized in that, It also includes spraying ε-polylysine-PLGA microspheres on the surface of the composite hydrogel to form a surface hydrophobic network layer of ε-polylysine. The specific steps are as follows: Prepare ε-polylysine-PLGA microspheres: First, dissolve PLGA in ethyl acetate; then dissolve ε-polylysine in acetic acid aqueous solution to obtain an aqueous phase; slowly drip the aqueous phase into the oil phase and homogenize in an ice bath to form a primary emulsion; add the primary emulsion to a PBS solution containing polyvinyl alcohol, homogenize to obtain an emulsion, stir the emulsion and evaporate ethyl acetate, collect the microspheres after centrifugation, wash with deionized water, freeze-dry and sieve, and retain microspheres with a particle size of 1.5-2.5 μm; Spray microspheres to form a surface hydrophobic network layer: Disperse the above microspheres in an ethanol solution containing Tween 80, transfer them into a high-pressure airless spray gun, and spray the microspheres onto the surface of the composite hydrogel; perform hot air annealing to melt the surface of the microspheres and embed them in the dressing surface layer; Among them, the weight ratio of ε-polylysine to PLGA is 1:(3-5).

8. The preparation method of the mussel adhesive protein hydrogel dressing with self-healing function according to claim 6, characterized in that, It also includes the step of adding auxiliary components: Dissolve the humectant in deionized water to obtain a humectant solution; then add the temperature-responsive polymer to pure water at 0-5°C and magnetically stir to form a uniform sol; then raise the temperature of the sol to 20-25°C and gradually add the humectant solution; obtain an auxiliary composition; Add the auxiliary composition to the functionalized modified hydrogel prepolymer solution and perform ultrasonic treatment in an ice bath for 10-20 minutes until evenly dispersed.

9. The preparation method of the mussel adhesive protein hydrogel dressing with self-healing function according to claim 8, characterized in that, When preparing the mussel adhesive protein premixed solution, use the ultrasonic-centrifugation method to uniformly disperse graphene oxide in the mussel adhesive protein solution to form a stable suspension. The ultrasonic power is 280-350 W and the time is 25-35 minutes; the addition amount of the catalytic enzyme is calculated as 100-300 U / g based on the mass of the mussel adhesive protein.

10. The preparation method of the mussel adhesive protein hydrogel dressing with self-healing function according to claim 8, characterized in that, The photoinitiator is LAP or / and Na-TPO, and the catalytic enzyme is one or a mixture of two or more of laccase, tyrosinase, transglutaminase, and lysozyme.

Images