A strong wet-skin tissue adhesive hydrogel and its preparation method and application

Abstract

The application provides a strong wet tissue adhesion hydrogel and a preparation method and application thereof, a long-chain modified silk fibroin with higher purity is prepared by shortening the degumming time of a sodium carbonate solution to raw rubber, prolonging the reaction time with glycidyl methacrylate, and dialysis with a dialysis bag with a molecular weight of not less than 50 kDa; the long-chain modified silk fibroin is compounded with polyphenol materials, especially tannic acid, and the prepared hydrogel has a strong wet tissue adhesion effect, the shear stress reaches 62.5 N, the anti-burst capacity reaches 844 mmHg, the Young's modulus reaches 160 kpa, the degradation speed is slower, the adhesion time is longer, a strong hemostatic effect can be realized on internal organs such as liver and heart, and tissues such as blood vessels, a good plugging effect can be realized on stomach perforation, intestinal perforation, pneumothorax and the like, the hydrogel can also replace sutures to realize the closure of skin wounds, scar formation caused by the increase of local stress of sutures is avoided, and thus the clinical requirements can be better met.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of biomedical materials, and particularly relates to a natural-based material for wet surface tissue adhesive glue field, and especially relates to a strong wet surface tissue adhesive hydrogel and a preparation method and application thereof. BACKGROUND

[0002] In the field of wet surface tissue glue, many high-performance wet surface tissue glues based on artificial materials such as polymers and macromolecules have been studied, but artificial synthetic materials have problems such as low biocompatibility, slow degradation, and potential toxicity of degradation products. Natural biological materials do not have the above-mentioned drawbacks of artificial materials, but natural materials generally have poor cohesion, and therefore cannot become an ideal wet surface tissue glue.

[0003] Silk fibroin, as a natural biological material, can greatly enhance its cohesion by forming beta-sheet molecular chains, so as to design a wet surface tissue glue with strong adhesion capacity by combining the advantages of artificial materials and natural materials. Polyphenol materials such as tannic acid are also substances existing in nature, which are rich in hydroxyl groups and can form a large number of hydrogen bonds with silk fibroin and tissue to form a tight adhesion between silk fibroin and wet surface tissue.

[0004] CN111228563A provides a medical adhesive composed of silk fibroin and tannic acid, but the prepared hydrogel has weak adhesion performance and mechanical properties, and the degradation speed is fast, the wet surface tissue adhesion effect is poor, and the adhesion effect is not long-lasting.

[0005] CN114621464A provides a method for preparing a hydrogel by modifying silk fibroin with glycidyl methacrylate to prepare SilMA, and then condensing SilMA with thiolated hyaluronic acid, but the large molecular silk fibroin is destroyed due to long degumming time during the modification process, and the obtained mixture is a mixture of short-chain SilMA and long-chain SilMA after dialysis with a low molecular weight dialysis membrane after the reaction, and the mechanical properties are weak. The preparation process is complex, and the adhesion performance and mechanical properties of the prepared hydrogel are weak, and it is difficult to realize effective adhesion of wet surface tissue.

[0006] Therefore, it is urgent to find a hydrogel with long-acting and high-strength adhesion effect on wet surface tissue, so as to realize strong hemostasis of internal organs, blood vessels and other tissues, and replace sutures to realize closure of skin wounds and the like. SUMMARY

[0007] To solve the above problems, the application provides a strong wet tissue adhesion hydrogel, which is prepared by shortening the degumming time of a sodium carbonate solution on raw rubber, prolonging the reaction time with glycidyl methacrylate, and dialysis with a dialysis bag with a molecular weight of not less than 50 kDa, so that long-chain modified silk fibroin with higher purity is prepared; the long-chain modified silk fibroin is further compounded with polyphenol materials, especially tannic acid, to prepare a hydrogel with strong wet tissue adhesion effect, a shear stress of more than 70 N, a blasting resistance of 900 mmHg, a Young's modulus of 110 kpa, a slower degradation speed, and a longer adhesion time, so that a strong hemostatic effect on internal organs such as liver and heart, blood vessels and other tissues can be achieved, and the hydrogel can also replace sutures to achieve the closure of skin wounds, avoid the formation of scars caused by the increase of local stress caused by sutures, and better meet the needs of clinical application.

[0008] In one aspect, the application provides a preparation method of long-chain SilMA, which comprises the following steps:

[0009] (1) preparing modified silk fibroin;

[0010] (2) dialysis with a dialysis bag with a molecular weight of not less than 50 kDa to obtain long-chain SilMA.

[0011] Further, the dialysis bag has a molecular weight of 50 kDa, and the dialysis time is 4 days.

[0012] The preparation process of modified silk fibroin (SilMA) generally comprises four steps of degumming, lithium bromide dissolution, modification by reaction with glycidyl methacrylate, and dialysis. However, in the preparation of modified silk fibroin (SilMA) by the prior art, a dialysis bag with a low molecular weight (the molecular weight is all below 12 kDa) is usually used for dialysis in order to improve the yield, and after dialysis, a small amount of impurities is removed, and a mixture of long-chain SilMA and short-chain SilMA is left.

[0013] In the preparation of modified silk fibroin (SilMA) by the application, a dialysis bag with a molecular weight of not less than 50 kDa is used for dialysis, and after dialysis, not only impurities are removed, but also short-chain SilMA is removed, and only high-purity long-chain SilMA is left. The prepared long-chain SilMA has a significant improvement in performance compared with SilMA obtained by dialysis with a dialysis bag with a small molecular weight, has a higher Young's modulus, has stronger adhesion, and has a slower degradation speed.

[0014] Further, the silk fibroin in step (1) is obtained by degumming raw silk with a sodium carbonate solution for 20-40 minutes, and the sodium carbonate solution has a concentration of 0.01-0.05 M.

[0015] The degumming rate of raw silk increases with the increase of the mass fraction of sodium carbonate solution and the degumming time, but when the concentration of sodium carbonate solution is greater than 0.1M and the degumming time is greater than 1h, the surface of the silk fibroin fiber obtained by degumming is easy to appear split microstructure, and the breaking strength of the fiber is also significantly reduced.

[0016] The present application has been proved by a large number of experiments that when the concentration of sodium carbonate solution is greater than 0.05M, a large amount of long-chain silk fibroin protein in the silk fibroin obtained by degumming is decomposed into short-chain silk fibroin protein, and the Young's modulus and adhesion performance of the obtained silk fibroin are significantly reduced; at the same time, when the degumming time is greater than 1h, a large amount of long-chain silk fibroin protein in the silk fibroin obtained by degumming is also decomposed into short-chain silk fibroin protein, and the Young's modulus and adhesion performance of the obtained silk fibroin are significantly reduced. Therefore, in order to obtain more long-chain silk fibroin and improve the mechanical properties of silk fibroin, the concentration of sodium carbonate solution for raw silk degumming needs to be controlled at 0.01-0.05M, and the degumming time needs to be controlled at 20-40 minutes.

[0017] Further, the preparation of modified silk fibroin in step (1) is: dissolving silk fibroin with lithium bromide, the dissolving time is 0.5-3h, and then glycidyl methacrylate is reacted with silk fibroin, the reaction time is 3-6h.

[0018] The process for preparing modified silk fibroin includes two steps of lithium bromide dissolution and modification reaction with glycidyl methacrylate; wherein, the lithium bromide dissolution time and the glycidyl methacrylate reaction time will directly affect the yield and mechanical strength of long-chain SilMA.

[0019] The complete dissolution time of silk fibroin in lithium bromide is only 5-10min, but complete dissolution does not mean that high-performance long-chain SilMA can be obtained, because the molecular chain of long-chain silk fibroin is very long, and a sufficient number of groups must be exposed to fully react with glycidyl methacrylate to achieve modification. If the dissolution time in lithium bromide is too short, the number of groups in the molecular chain of long-chain silk fibroin for modification is too small, which will directly affect the mechanical strength of SilMA obtained in the subsequent modification reaction; however, the dissolution time of silk fibroin in lithium bromide cannot be too long, because too long dissolution time will cause the molecular chain of long-chain silk fibroin to be continuously decomposed and shortened, thereby significantly reducing the content of long-chain SilMA in SilMA. Therefore, the dissolution time of silk fibroin in lithium bromide needs to be controlled at 0.5-3h.

[0020] The reaction time of glycidyl methacrylate with silk fibroin after being dissolved by lithium bromide also directly affects the performance of SilMA. Since the molecular chain of long-chain silk fibroin is long, the reaction of the amino group with glycidyl methacrylate also needs a longer time. However, if the reaction time is too long, the performance of the obtained SilMA will be significantly reduced. The lithium bromide remaining in the reaction system for a long time will lead to the decomposition of the silk fibroin molecular chain, thereby reducing the molecular weight of the silk fibroin and weakening the mechanical properties of the silk fibroin. Therefore, the reaction time of glycidyl methacrylate with silk fibroin needs to be controlled within 3-6 hours.

[0021] Further, the dissolution time of silk fibroin by lithium bromide is 1 hour, and the reaction time of glycidyl methacrylate with silk fibroin is 4 hours.

[0022] In some ways, the reaction temperature of glycidyl methacrylate with silk fibroin is 60°C.

[0023] In another aspect, the present application provides a method for preparing a strong wet tissue tissue adhesive hydrogel, which comprises the following steps:

[0024] (a) preparing long-chain SilMA by the method described above;

[0025] (b) adding a photoinitiator to the long-chain SilMA solution and mixing with a polyphenol solution to prepare a wet tissue adhesive hydrogel by photocuring.

[0026] The photoinitiator is LAP, and the final concentration of LAP in the solution is 0.1%-0.5%.

[0027] The photocuring is irradiated by 405 nm ultraviolet light for 5-60 seconds, which can realize the preparation of a hydrogel glue with strong adhesion to wet tissues, and the strong cohesive force of the glue is provided by photocrosslinking.

[0028] Further, the polyphenol solution in step (b) is one or more selected from the group consisting of tannic acid, dopamine, catechol, procyanidin, pentagalloyl glucose, and resveratrol solution.

[0029] Studies have shown that the long-chain SilMA prepared by the present application can be combined with a polyphenol solution to prepare a hydrogel with long-acting and high-strength adhesion effect to wet tissues.

[0030] The existing hydrogel will swell immediately after entering the body, and the adhesion will also decrease. The hydrogel provided by the application will not swell immediately after entering the body, but will undergo a conformational transition (β-sheet content increases) in the body (37°C) to increase the mechanical properties of the glue matrix, which is beneficial to the stability and adhesion of the tissue, and thus exhibits time-dependent tissue adhesion enhancement characteristics, providing long-term high-strength tissue adhesion effect for wet surfaces. The polyphenol material can form a large number of hydrogen bonds with the long-chain SilMA and the tissue, thereby forming a tight adhesion between the long-chain SilMA and the wet tissue.

[0031] Further, the polyphenol solution in step (b) is a tannic acid solution.

[0032] Tannic acid is a small molecular polyphenol with many hydroxyl groups, and the hydroxyl groups are relatively close, so that more hydrogen bonds can be formed between the long-chain SilMA and the tissue. In addition to the hydrogen bonds between tannic acid and long-chain SilMA and the tissue, some hydrophobic bonds are also formed, which can form a pocket-like structure to enhance the stability and mechanical strength of the hydrogen bonds in the aqueous environment. Therefore, by compounding long-chain SilMA and tannic acid, a wet surface adhesion hydrogel with better mechanical properties, stronger adhesion and slower degradation rate can be prepared.

[0033] The photocuring refers to irradiating with 405 nm ultraviolet light for 5-60 seconds to obtain a silk fibroin hydrogel glue that strongly adheres to wet tissue.

[0034] Further, the long-chain SilMA solution in step (b) is a solution prepared by redissolving long-chain SilMA to a mass fraction of 5-40%, and the mass fraction of the polyphenol solution is 5-50%; the mass ratio of the long-chain SilMA solution to the polyphenol solution is 20:1-1:1.

[0035] The 40% long-chain SilMA solution has the strongest mechanical properties and the best adhesion, but is not easy to operate. Reducing the concentration appropriately can improve the convenience of operation.

[0036] In some modes, due to the limitation of solubility, the polyphenol solution cannot be prepared to a too high concentration, and the mass fraction is generally 5-50%.

[0037] In some modes, too high concentration of the polyphenol solution will also affect the quality of the prepared hydrogel, and it is necessary to try to reduce the influence of the polyphenol solution concentration on the hydrogel.

[0038] In some modes, the polyphenol solution is a tannic acid solution, and after mixing the tannic acid solution with long-chain SilMA, the final concentration of tannic acid is 3%.

[0039] In another aspect, the application provides a use of long-chain SilMA for preparing a strong wet tissue adhesion hydrogel, wherein the long-chain SilMA is prepared by the above method.

[0040] In still another aspect, the present application provides a use of the strong wet surface tissue adhesive hydrogel for internal organs and blood vessels hemostasis.

[0041] In still another aspect, the present application provides a use of the strong wet surface tissue adhesive hydrogel for replacing sutures to achieve skin wound closure.

[0042] The strong wet surface tissue adhesive hydrogel, the preparation method and the application thereof provided by the present application have the following beneficial effects:

[0043] 1. The preparation method of SilMA is improved: by shortening the degumming time of sodium carbonate solution to raw rubber, prolonging the reaction time with glycidyl methacrylate, and using a dialysis bag with a molecular weight of not less than 50 kDa for dialysis, a long-chain SilMA with higher purity and better mechanical properties is prepared.

[0044] 2. By mixing long-chain SilMA and polyphenol materials, a strong wet surface tissue adhesive hydrogel with strong wet surface tissue adhesion effect and high mechanical properties is prepared.

[0045] 3. Preferably, by mixing long-chain SilMA and tannic acid, a strong wet surface tissue adhesive hydrogel with strong wet surface tissue adhesion effect, shear stress reaching 62.5 N, anti-burst capacity reaching 844 mmHg, and Young's modulus reaching 160 kpa is prepared.

[0046] 4. The prepared strong wet surface tissue adhesive hydrogel has a slower degradation rate and a longer adhesion time, can achieve strong hemostasis effect on internal organs such as liver and heart, blood vessels and other tissues, and can replace sutures to achieve skin wound closure, avoid scar formation caused by local stress increase of sutures, and thus better meet the needs of clinical application. BRIEF DESCRIPTION OF DRAWINGS

[0047] Figure 1 A schematic diagram of the adhesion mechanism between long-chain SilMA-polyphenol (tannic acid) prepared in Example 2 and tissues;

[0048] Figure 2 A shear stress comparison chart of the hydrogels prepared by long-chain SilMA respectively compounded with tannic acid, hyaluronic acid, chondroitin sulfate in Example 7, or single SilMA;

[0049] Figure 3 An anti-burst capacity comparison chart of the hydrogels prepared by long-chain SilMA respectively compounded with tannic acid, hyaluronic acid, chondroitin sulfate in Example 7, or single SilMA;

[0050] Figure 4 A comparison chart of Young's modulus detection results in Example 8;

[0051] Figure 5 Shear stress comparison chart for hydrogels of groups 8, 9, 12 in Example 8;

[0052] Figure 6 Burst resistance comparison chart for Example 8;

[0053] Figure 7 Picture of liver hemostasis in Example 7, left side is continuous bleeding state before glue use, right side is immediate hemostasis state after use. DETAILED DESCRIPTION

[0054] The application will be further described below in conjunction with the drawings and examples, it should be noted that the following examples are intended to facilitate the understanding of the application, and do not have any limiting effect on it. The advantages and features of the application will be more apparent according to the following description and claims. It should be noted that the drawings are very simplified and all use non-precise proportions, only to facilitate, clear and assist the purpose of explaining the examples of the application.

[0055] Example 1 Preparation of long-chain SilMA provided by the application

[0056] The preparation method of long-chain SilMA provided by this example is as follows:

[0057] (1) 50g raw silk is degummed with 2 liters of 0.01M-0.05M (preferably 0.025M in this example) concentration of sodium carbonate at 100℃ for 20-40 minutes (preferably 40 minutes in this example) to obtain 38g of silk fibroin, which is dried in an oven at 65℃;

[0058] (2) 30g of silk fibroin is dissolved in 150ml of 9.3M lithium bromide solution, the dissolution time is 0.5-3h (preferably 1h in this example), 150ml of silk fibroin solution is obtained, which is reacted with 30ml of glycidyl methacrylate at 60℃, so that glycidyl methacrylate reacts with the amino group of silk to join, the reaction time is 4 hours, 150ml of product is obtained, which is dialyzed in a 50kDa dialysis bag (purchased from Yuan Ye Bio, model 31mm / 1m) at 4℃ for 4 days, and freeze-dried to obtain 26g of long-chain SilMA sponge.

[0059] The prepared long-chain SilMA sponge can be redissolved into a long-chain SilMA solution, and a photoinitiator LAP (the final concentration of LAP in the solution is 0.1%-0.5%, preferably 0.25%) is added, and the long-chain SilMA hydrogel is prepared by light curing, or it can be compounded with other materials (such as polyphenol material) to prepare hydrogel.

[0060] Example 2 Preparation of strong wet surface adhesive hydrogel provided by the application

[0061] The preparation method of the strong wet surface adhesive hydrogel provided by the embodiment is as follows:

[0062] (a) The long-chain SilMA is prepared by the method provided in Embodiment 1; 1.2 g of the long-chain SilMA is redissolved into a hydrogel solution with a mass fraction of 5%-40% (preferably 30% in the embodiment) by using deionized water, and 2.8 ml of a photoinitiator LAP is added into the solution, and the final concentration of the LAP in the solution is 0.1%-0.5% (preferably 0.25% in the embodiment);

[0063] (b) The polyphenol material such as tannic acid is prepared into a tannic acid solution with a mass fraction of 5%-50% (preferably 50% in the embodiment).

[0064] (c) The long-chain SilMA solution and the tannic acid solution are mixed on the interface between the wet surface tissue in a ratio of 20:1-1:1 (preferably 10:1 in the embodiment), and the mixture is irradiated by 405 nm ultraviolet light for 5-60 seconds (preferably 30 seconds in the embodiment) to obtain the hydrogel which is strongly adhered to the wet surface tissue, and the adhesion mechanism between the long-chain SilMA-polyphenol (tannic acid) and the tissue is as shown in Figure 1 .

[0065] Embodiment 3: Effect of different degumming times on preparation of long-chain SilMA hydrogel

[0066] In the embodiment, the long-chain SilMA is prepared by the method provided in Embodiment 1, and the fibroin is obtained by degumming the long-chain SilMA at 100°C for 10, 20, 30, 40, 50 and 60 minutes respectively by using 0.02M sodium carbonate, and the fibroin is dried in an oven at 65°C; and then the long-chain SilMA sponge is prepared by a modification reaction, and the long-chain SilMA solution with a mass fraction of 30% is redissolved by using deionized water, and 0.25% of the photoinitiator LAP is added into the solution to obtain the long-chain SilMA hydrogel by light curing, and the Young's modulus, the shear stress, the burst pressure resistance and the degradation time of the long-chain SilMA hydrogel prepared by different degumming times are detected, wherein the detection method of the Young's modulus is Compression test, the detection method of the shear stress is Lap Shear test, the detection method of the burst pressure resistance is Burst Pressure test, and the detection method of the degradation time is rat subcutaneous implantation experiment, and the detection results are shown in Table 1.

[0067] Table 1: Effect of different degumming times on preparation of long-chain SilMA hydrogel

[0068] Degumming time (min) Young's modulus (kPa) Shear stress (N) Resistance to bursting (mmHg) Degradation time (days) 10 76 11.3 155 32 20 104 16.9 183 30 30 116 17.7 191 29 40 121 18.5 202 28 50 102 16.1 174 25 60 94 15.4 161 20

[0069] The stronger the mechanical property of the long-chain SilMA hydrogel is, the better the adhesion property is, and the mechanical property and adhesion of the wet surface adhesion hydrogel prepared by compounding the long-chain SilMA hydrogel with the polyphenol material are also stronger, and the degradation time is also slower.

[0070] As shown in Table 1, the length of the degumming time has a great influence on the mechanical property of the long-chain SilMA hydrogel. When the degumming time is within 20-40 min, the Young's modulus, shear stress and burst resistance of the prepared long-chain SilMA hydrogel are maintained at a high level, and when the degumming time is further prolonged, the mechanical property begins to decline, and the degradation time also begins to shorten. The reason may be that when the degumming time is too long, a lot of long-chain silk fibroin in the obtained silk fibroin is decomposed into short-chain silk fibroin, and the Young's modulus and adhesion of the obtained silk fibroin are obviously decreased. Therefore, in order to obtain more long-chain silk fibroin and improve the mechanical property of the silk fibroin, the concentration of the sodium carbonate solution used for degumming of raw silk needs to be controlled at 0.01-0.05 M, and the degumming time needs to be controlled at 20-40 min. The most preferred concentration of the sodium carbonate solution is 0.025 M, and the degumming time is 40 min, at which the Young's modulus reaches 12 kPa, the shear stress reaches 18.5 N, the burst resistance reaches 202 mmHg, and the degradation time reaches 28 days.

[0071] Example 4 Influence of lithium bromide dissolution time on preparation of wet surface adhesion hydrogel

[0072] In this example, the long-chain SilMA was prepared according to the method provided in Example 1, wherein the dissolution time of the lithium bromide in dissolving the silk fibroin was 0.5, 1, 2 and 3 h, respectively, and then the long-chain SilMA sponge was prepared by a modification reaction. The long-chain SilMA solution with a mass fraction of 30% was prepared by re-dissolving the long-chain SilMA sponge in deionized water, and 0.25% of the photoinitiator LAP was added to the solution to obtain the long-chain SilMA hydrogel by light curing. The Young's modulus, shear stress, burst resistance and degradation time of the long-chain SilMA hydrogel prepared by different lithium bromide dissolution times were detected, and the detection methods of the Young's modulus, shear stress, burst resistance and degradation time were shown in Example 3. The detection results are shown in Table 2.

[0073] Table 2, Influence of lithium bromide dissolution time on preparation of long-chain SilMA hydrogel

[0074]

[0075] As can be seen from Table 2, the lithium bromide dissolution time also directly affects the mechanical properties of the long-chain SilMA hydrogel. When the lithium bromide dissolution time is 0.5 h, the mechanical strength of the obtained SilMA is affected due to the short dissolution time, which results in too few groups in the long-chain silk fibroin molecular chain being exposed for modification, thereby directly affecting the mechanical strength of the SilMA obtained after the subsequent modification reaction. When the lithium bromide dissolution time is more than 1 h, the long-chain silk fibroin molecular chain is continuously decomposed and shortened due to the long time, thereby significantly reducing the content of the long-chain SilMA in the SilMA and continuously reducing the mechanical properties. Therefore, the dissolution time of the lithium bromide for dissolving the silk fibroin is most preferably 1 h.

[0076] Example 5 Influence of glycidyl methacrylate reaction time on preparation of wet surface-adhesive hydrogel

[0077] In this example, long-chain SilMA sponges were prepared according to the method provided in Example 1, in which the glycidyl methacrylate reaction time was 1, 2, 3, 4, 5, 6, and 7 h, respectively, and then the long-chain SilMA sponges were subjected to a modification reaction to obtain long-chain SilMA sponges. The long-chain SilMA sponges were re-dissolved in deionized water to obtain a long-chain SilMA solution with a mass fraction of 30%, and 0.25% of a photoinitiator LAP was added to the solution to obtain long-chain SilMA hydrogels by light curing. The Young's modulus, shear stress, burst resistance, and degradation time of the long-chain SilMA hydrogels prepared at different glycidyl methacrylate reaction times were detected, and the detection methods of the Young's modulus, shear stress, burst resistance, and degradation time were as shown in Example 3. The detection results are shown in Table 3.

[0078] Table 3 Influence of glycidyl methacrylate reaction time on preparation of long-chain SilMA hydrogel

[0079]

[0080] As can be seen from Table 3, the glycidyl methacrylate reaction time also directly affects the mechanical properties of the long-chain SilMA hydrogel. Since the long-chain silk fibroin molecular chain is long, a longer reaction time is required for the reaction between the amino group and the glycidyl methacrylate. Therefore, when the reaction time is less than 3 h, the mechanical strength of the obtained SilMA is affected due to the short reaction time, which results in part of the long-chain silk fibroin not being reacted. When the reaction time is more than 6 h, the performance of the obtained SilMA is significantly reduced due to the continued splitting of the silk fibroin by the lithium bromide in the reaction system, which makes the obtained SilMA mainly in the form of small molecular weight, thereby reducing the mechanical properties of the SilMA. Therefore, the glycidyl methacrylate reaction time needs to be controlled in the range of 3-6 h, and most preferably 4 h.

[0081] Example 6 Influence of dialysis bag molecular weight on preparation of long-chain SilMA hydrogel / wet surface-adhesive hydrogel

[0082] This embodiment is prepared according to the method provided in Example 1. Long-chain SilMA sponges are prepared by dialysis at 4°C for 4 days using dialysis bags with molecular weights of 12, 18, 20, 30, 40, 50, 60, and 70 kDa, respectively, and then lyophilized. The long-chain SilMA solution with a mass fraction of 30% is re-dissolved in deionized water, and 0.25% of a photoinitiator LAP is added to the solution to obtain a long-chain SilMA hydrogel by light curing, or a tannic acid solution is added to the solution, and then a photoinitiator LAP is added to obtain a wet-surface-adhesion hydrogel by light curing. The Young's modulus, shear stress, anti-burst capacity, and degradation time of the long-chain SilMA hydrogels prepared by dialysis using dialysis bags with different molecular weights are detected, respectively. The detection methods of the Young's modulus, shear stress, anti-burst capacity, and degradation time are shown in Example 3. The detection results are shown in Table 4.

[0083] Table 4, Effect of molecular weight of dialysis bag on preparation of long-chain SilMA hydrogel

[0084]

[0085] As can be seen from Table 4, the stronger the mechanical properties of the long-chain SilMA hydrogel, the better the adhesion performance, and the slower the degradation time. The mechanical properties and adhesion of the wet-surface-adhesion hydrogel prepared by compounding the long-chain SilMA hydrogel with tannic acid are also stronger, and the degradation time is also slower.

[0086] At the same time, as can be seen from Table 4, the molecular weight of the dialysis bag also directly affects the mechanical properties of the prepared SilMA. When the molecular weight of the dialysis bag is greater than 50 kDa, the impurities and short-chain SilMA in the dialysis bag can be removed together, and only high-purity long-chain SilMA is left. The long-chain SilMA prepared by dialysis has significantly improved performance compared to the SilMA obtained by dialysis using a dialysis bag with a small molecular weight, has a higher Young's modulus, stronger adhesion, and slower degradation rate.

[0087] When the molecular weight of the dialysis bag continues to increase to 60 and 70 kDa, the mechanical properties of the obtained long-chain SilMA increase very slowly compared to 50 kDa, but the long-chain SilMA obtained after dialysis becomes more and more difficult to dissolve, which brings inconvenience to subsequent operations. Therefore, the dialysis bag with a molecular weight of 50-60 kDa is the most preferred.

[0088] Example 7 Effect of compounding long-chain SilMA with different materials on preparation of wet-surface-adhesion hydrogel

[0089] This example uses long chain SilMA prepared in Example 1 to complex with polyphenol materials or polysaccharides, respectively, including catechol (TP), dopamine (DOPA), tannic acid (TA), procyanidine, pentagalloylglucose, resveratrol, and polysaccharides including hyaluronic acid (HA), chondroitin sulfate (CS) to prepare wet face adhesive hydrogel, as follows:

[0090] 1. Long chain SilMA with catechol

[0091] LAP (0.25%) was added to 30% SilMA and mixed evenly, then mixed with 30% catechol at a ratio of 10:1, and SilMA was obtained after 405 nm UV light for 30 seconds 30 -TP3.

[0092] 2. Long chain SilMA with dopamine

[0093] LAP (0.25%) was added to 30% SilMA and mixed evenly, then mixed with 30% dopamine at a ratio of 10:1, and SilMA was obtained after 405 nm UV light for 30 seconds 30 -DOPA3.

[0094] 3. Long chain SilMA with tannic acid

[0095] LAP (0.25%) was added to 30% SilMA and mixed evenly, then mixed with 30% tannic acid at a ratio of 10:1, and SilMA was obtained after 405 nm UV light for 30 seconds 30 -TA3.

[0096] 4. Long chain SilMA with procyanidine

[0097] LAP (0.25%) was added to 30% SilMA and mixed evenly, then mixed with 30% procyanidine at a ratio of 10:1, and SilMA was obtained after 405 nm UV light for 30 seconds 30 -Procyanidine3.

[0098] 5. Long chain SilMA with pentagalloylglucose

[0099] LAP (0.25%) was added to 30% SilMA and mixed evenly, then mixed with 30% pentagalloylglucose at a ratio of 10:1, and SilMA was obtained after 405 nm UV light for 30 seconds 30 -Pentagalloylglucose3.

[0100] 6. Long chain SilMA with Resveratrol

[0101] LAP (0.25%) was added to 30% SilMA and mixed evenly, then mixed with 30% Resveratrol at a ratio of 10:1, and SilMA was obtained after 405 nm ultraviolet light irradiation for 30 seconds 30 - Resveratrol3.

[0102] 7. Long chain SilMA with Hyaluronic acid

[0103] LAP (0.25%) was added to 30% SilMA and mixed evenly, then mixed with 30% Hyaluronic acid at a ratio of 10:1, and SilMA was obtained after 405 nm ultraviolet light irradiation for 30 seconds 30 - HA3.

[0104] 8. Long chain SilMA with Chondroitin sulfate

[0105] LAP (0.25%) was added to 30% SilMA and mixed evenly, then mixed with 30% Chondroitin sulfate at a ratio of 10:1, and SilMA was obtained after 405 nm ultraviolet light irradiation for 30 seconds 30 - CS3.

[0106] 9. Long chain SilMA alone

[0107] LAP (0.25%) was added to 30% SilMA and mixed evenly, and SilMA was obtained after 405 nm ultraviolet light irradiation for 30 seconds 30 .

[0108] The 9 groups of wet surface-adhesive hydrogels prepared were respectively detected for Young's modulus, shear stress, anti-burst capacity, degradation time, and hemostatic effect on internal organs; the detection methods for Young's modulus, shear stress, anti-burst capacity, and degradation time were as shown in Example 3; the detection method for the hemostatic effect on internal organs was that SilMA and tannic acid were directly sprayed onto the surface of a continuously bleeding organ and ultraviolet irradiation was simultaneously performed to observe the bleeding stop condition; the detection results were as shown in Table 5. The comparison chart of shear stress of the hydrogels prepared by long chain SilMA respectively compounded with tannic acid, hyaluronic acid, and chondroitin sulfate, or long chain SilMA alone was as shown in Figure 2 , Figure 3 .

[0109] Table 5, Influence of long chain SilMA compounded with different materials on preparation of wet surface-adhesive hydrogel

[0110]

[0111] From Table 5 and Figure 2 , Figure 3It can be seen that compared with polysaccharides (hyaluronic acid, chondroitin sulfate), long-chain SilMA composite polyphenol materials can significantly improve the mechanical properties and adhesion of hydrogels, and have better effect when used for internal organ hemostasis. Figure 7 For liver hemostasis, the left side is the continuous bleeding state before the glue is used, and the right side is the immediate hemostasis state after use); and when long-chain SilMA is compounded with polysaccharides, the mechanical properties are not much different from those of SilMA alone, and it is difficult to achieve good hemostasis effect.

[0112] At the same time, compared with different polyphenol materials, long-chain SilMA composite tannic acid and procyanidine has a better effect than other polyphenol materials, especially when compounded with tannic acid, it can make the mechanical properties and adhesion of hydrogels reach a higher level, and the degradation rate is slower, which can achieve the effect of immediate hemostasis after application.

[0113] In addition, the long-chain SilMA composite tannic acid hydrogel is also successfully used for skin wound closure in this embodiment. LAP (0.25%) is added to 30% SilMA and mixed uniformly, and then mixed with 30% tannic acid at a ratio of 10:1, applied to a 60um silk film, and irradiated with 405nm ultraviolet light for 30 seconds to obtain SF / SilMA 30 The hydrogel silk film can replace sutures to achieve closure of skin wounds, and can completely avoid sutures, avoiding the formation of scars caused by increased local stress of sutures.

[0114] Example 8 Effect of different contents of long-chain SilMA and tannic acid on the mechanical properties of hydrogels

[0115] In this embodiment, long-chain SilMA prepared in Example 1 is used to prepare SilMA solutions with different mass percentages, and is compounded with tannic acid with different mass percentages to prepare different wet surface adhesive hydrogels, as follows:

[0116] 1. SilMA5-TA 2.5 (The hydrogel contains 5% long-chain SilMA and 2.5% tannic acid)

[0117] LAP (0.25%) is added to 5% SilMA and mixed uniformly, and then mixed with 50% tannic acid at a ratio of 20:1, and irradiated with 405nm ultraviolet light for 30 seconds to obtain SilMA5-TA 2.5 .

[0118] 2. SilMA 15 -TA 2.5 (The hydrogel contains 15% long-chain SilMA and 2.5% tannic acid)

[0119] LAP (0.25%) was added to 15% SilMA and mixed well, then mixed with 50% tannic acid in a ratio of 20:1 to obtain SilMA after 405 nm UV light for 30 seconds 15 -TA 2.5 .

[0120] 3. SilMA 30 -TA 2.5 (30% long chain SilMA and 2.5% tannic acid in hydrogel)

[0121] LAP (0.25%) was added to 30% SilMA and mixed well, then mixed with 50% tannic acid in a ratio of 20:1 to obtain SilMA after 405 nm UV light for 30 seconds 30 -TA 2.5 .

[0122] 4. SilMA 40 -TA 2.5 (40% long chain SilMA and 2.5% tannic acid in hydrogel)

[0123] LAP (0.25%) was added to 40% SilMA and mixed well, then mixed with 50% tannic acid in a ratio of 20:1 to obtain SilMA after 405 nm UV light for 10 seconds 40 -TA 2.5 .

[0124] 5. SilMA 30 -TA 0.25 (30% long chain SilMA and 0.25% tannic acid in hydrogel)

[0125] LAP (0.25%) was added to 30% SilMA and mixed well, then mixed with 5% tannic acid in a ratio of 20:1 to obtain SilMA after 405 nm UV light for 30 seconds 30 -TA 0.25 .

[0126] 6. SilMA 30 -TA 1.5 (30% long chain SilMA and 1.5% tannic acid in hydrogel)

[0127] LAP (0.25%) was added to 30% SilMA and mixed well, then mixed with 30% tannic acid in a ratio of 20:1 to obtain SilMA after 405 nm UV light for 20 seconds 30 -TA 1.5 .

[0128] 7. SilMA 30TA5 (30% long chain SilMA and 5% tannic acid in hydrogel)

[0129] LAP (0.25%) was added to 30% SilMA and mixed well, then mixed with 50% tannic acid at a ratio of 10:1. SilMA was obtained after 405 nm UV light for 45 seconds 30 TA5.

[0130] 8. SilMA 30 TA6 (30% long chain SilMA and 6% tannic acid in hydrogel)

[0131] LAP (0.25%) was added to 30% SilMA and mixed well, then mixed with 30% tannic acid at a ratio of 5:1. SilMA was obtained after 405 nm UV light for 30 seconds 30 TA6.

[0132] 9. SilMA 30 TA3 (30% long chain SilMA and 3% tannic acid in hydrogel)

[0133] LAP (0.25%) was added to 30% SilMA and mixed well, then mixed with 15% tannic acid at a ratio of 5:1. SilMA was obtained after 405 nm UV light for 30 seconds 30 TA3.

[0134] 10. Long chain SilMA alone 10

[0135] LAP (0.25%) was added to 10% SilMA and mixed well. SilMA was obtained after 405 nm UV light for 30 seconds 30 .

[0136] 11. Long chain SilMA alone 20

[0137] LAP (0.25%) was added to 20% SilMA and mixed well. SilMA was obtained after 405 nm UV light for 30 seconds 30 .

[0138] 12. Long chain SilMA alone 30

[0139] LAP (0.25%) was added to 30% SilMA and mixed well. SilMA was obtained after 405 nm UV light for 30 seconds 30 .

[0140] The prepared 12 groups of wet surface adhering hydrogels were respectively detected for Young's modulus, shear stress, anti-bursting capacity, degradation time, hemostatic effect on internal organs, and effect of replacing suture to achieve skin wound closure. The detection methods of Young's modulus, shear stress, anti-bursting capacity, and degradation time are shown in Example 3. The detection method of hemostatic effect on internal organs is that the SilMA and tannic acid are directly sprayed onto the surface of the continuously bleeding organs and simultaneously subjected to ultraviolet irradiation to form a gel to observe the bleeding stop condition. The detection method of effect of replacing suture to achieve skin wound closure is that the long-chain SilMA and tannic acid are mixed and then applied to a 60 um silk film, and the hydrogel is obtained after the silk film is subjected to 405 nm ultraviolet light irradiation for 30 seconds, and then applied to a skin wound to detect whether the skin wound can be immediately closed. The detection results are shown in Table 6. The comparison of the Young's modulus detection results of the 10%, 20%, and 30% long-chain SilMA hydrogels of the 10th, 11th, and 12th groups is shown in Figure 4 Fig. 8. The comparison of the shear stress of the hydrogels of the 8th, 9th, and 12th groups is shown in Figure 5 Fig. 9. The comparison of the anti-bursting capacity is shown in Figure 6 Fig. 10.

[0141] Table 6, Effect of different contents of long-chain SilMA and tannic acid on the preparation of wet surface adhering hydrogels

[0142]

[0143] It can be seen from Table 6, Figure 4 that the mass percentage content of the long-chain SilMA solution directly affects the mechanical properties of the hydrogel, and the mass percentage content of the long-chain SilMA solution needs to be controlled to be at least 30%. Of course, the mass percentage content of the long-chain SilMA solution can continue to increase to 40%, and the gel mechanical properties of the 40% long-chain SilMA solution are stronger, and the adhesion is definitely better. However, since it is too viscous to be operated, the concentration is reduced to 30% by optimization and selection.

[0144] It can be seen from Table 6, Figure 5 and Figure 6 that the mass percentage contents of the long-chain SilMA solution and the tannic acid solution must be controlled in a suitable range to obtain better adhesion performance, better mechanical properties, slow degradation speed, and good hemostatic effect and effect of achieving skin wound closure. This is mainly because the long-chain SilMA solution is not easy to operate when the concentration is too high, and the optimal selection is 30%. Too much tannic acid solution will dilute the long-chain SilMA solution, thereby affecting the mechanical properties and adhesion performance. Too low tannic acid solution content will also affect the formation of hydrogen bonds. Therefore, the preferred strong wet surface adhering hydrogel is SilMA 30 -TA 2.5～6 , and the most preferred is SilMA 30 -TA3.

[0145] The application is not limited to the details of the foregoing description. While the application has been described in connection with specific embodiments thereof, it will be understood that it is capable of further modifications and this application is intended to cover any variations, uses, or adaptations of the application following, in general, the principles of the application and including such departures from the present disclosure as come within known or customary practice within the art to which the application pertains.

Claims

1. A method for preparing a strong wet surface tissue adhesion hydrogel, characterized in that, Includes the following steps: (a) Preparation of long-chain SilMA; (b) A photoinitiator was added to a long-chain SilMA solution and mixed with a polyphenol solution, and then photocured to prepare a wet-surface adhesive hydrogel. The long-chain SilMA solution is prepared by redissolving long-chain SilMA to a mass fraction of 5-40%, and the polyphenol solution has a mass fraction of 5-50%; the mass ratio of the long-chain SilMA solution to the polyphenol solution is 20:1 to 1:

1. The method for preparing long-chain SilMA in step (a) includes the following steps: (1) Preparation of modified silk fibroin; (2) Dialysis with a dialysis bag with a molecular weight of not less than 50 kDa yielded long-chain SilMA.

2. The preparation method according to claim 1, characterized in that, The dialysis bag used in step (2) has a molecular weight of 50 kDa and a dialysis time of 4 days.

3. The preparation method according to claim 2, characterized in that, The silk fibroin in step (1) is obtained by degumming raw silk with sodium carbonate solution for 20-40 minutes, wherein the concentration of sodium carbonate solution is 0.01-0.05M.

4. The preparation method according to claim 3, characterized in that, The preparation of modified silk fibroin in step (1) is as follows: dissolve silk fibroin with lithium bromide for 0.5 to 3 hours, and then react glycidyl methacrylate with silk fibroin for 3 to 6 hours.

5. The preparation method according to claim 4, characterized in that, The dissolution time of the silk fibroin with lithium bromide is 1 hour, and the reaction time of the silk fibroin with glycidyl methacrylate is 4 hours.

6. The preparation method according to claim 1, characterized in that, The polyphenol solution in step (b) is selected from one or more of tannic acid, dopamine, catechol, proanthocyanidins, pentagalloglucoside, or resveratrol solution.

7. The preparation method according to claim 6, characterized in that, The polyphenol solution in step (b) is a tannic acid solution.

8. A strong wet surface tissue adhesion hydrogel, characterized in that, Prepared using the method described in any one of claims 1 to 7.

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