Multifunctional silk fibroin hydrogels that do not require dialysis, their preparation methods and applications

Abstract

This invention discloses a method for preparing a non-dialysis-required multifunctional silk fibroin hydrogel, specifically comprising the following steps: Step 1, preparing silk fibroin fibers; Step 2, preparing a silk fibroin solution based on the product obtained in Step 1; Step 3, preparing a silk fibroin hydrogel based on the product obtained in Step 2. This invention also discloses the non-dialysis-required multifunctional silk fibroin hydrogel and its applications. This invention solves the problems of poor mechanical properties and environmental responsiveness of hydrogels prepared using SF4 (silicon fibroin).

Description

Technical Field

[0001] This invention belongs to the field of biomedical materials technology, and relates to a multifunctional silk fibroin hydrogel that does not require dialysis. This invention also relates to a method for preparing and applying the multifunctional silk fibroin hydrogel that does not require dialysis. Background Technology

[0002] Hydrogels made from silk fibroin (SF) hold great promise for wound management due to their excellent biocompatibility, natural renewability, and chemical modifiability. However, the cumbersome dialysis steps in conventional preparation processes inevitably lead to thermodynamic instability and prolonged gelation time in SF solutions. These drawbacks result in insufficient mechanical properties (stretchability and adhesion) and a lack of environmental responsiveness. Furthermore, SF lacks intrinsic antibacterial and anti-inflammatory activity.

[0003] To address the aforementioned drawbacks, existing methods utilize anionic surfactants such as sodium dodecyl sulfate, ultrasonic treatment, and temperature or pH adjustment to accelerate the gelation process of silk fibroin hydrogels. Furthermore, combining them with other materials, such as polyvinyl alcohol, and chemical modification enhances the adhesiveness of the silk fibroin hydrogels. In the biomedical and tissue engineering fields, 3D bioprinting into composite scaffolds or using electrospinning / wet spinning techniques to fabricate silk fibroin fiber pads can also shorten gelation time and enhance mechanical strength to meet diverse biomedical requirements.

[0004] Currently, Chinese patent (application number: 202010053074.7, application date: 2020.1.17, publication number: CN111228563A) discloses a method for preparing a composite medical adhesive of silk fibroin and tannic acid. This method discloses a composite adhesive of silk fibroin and tannic acid with good adhesion. The adhesive is obtained by mixing a silk fibroin solution (after dissolving and dialyzing lithium bromide) with a tannic acid solution, stirring evenly, allowing it to stand, freeze-drying the resulting precipitate, and then grinding it into powder. Chinese patent (application number: 202011358899.6, application date: 2020.11.27, publication number: CN113769151A) discloses a silk fibroin antibacterial dressing with silver loaded on one side, its preparation method, and its application. This dressing utilizes a calcium chloride and formic acid system to dissolve silk fibroin, and achieves good antibacterial properties by adding silver ions. Chinese Patent (Application No.: 202310053807.0, Application Date: 2023.2.3, Publication No.: CN116655945A) discloses a photocurable silk fibroin hydrogel and its preparation method. The method involves mixing a pure silk fibroin solution with water-soluble fullerene and horseradish peroxidase, then introducing oxygen into the mixture and sealing it. The mixture is then irradiated with near-ultraviolet light for 20-30 minutes to obtain the photocurable silk fibroin hydrogel. Chinese Patent (Application No.: 202310404123.0, Application Date: 2023.4.10, Publication No.: CN116271206A) discloses a gallium porphyrin complex / silk fibroin antibacterial dressing and its preparation method. In this method, after degumming, dissolving, and dialysis of silkworm cocoons, a concentrated silk fibroin solution is obtained by reverse dialysis using polyethylene glycol. This solution is then crosslinked with a novel metal antibacterial material, gallium porphyrin, and thoroughly frozen in a -80 ℃ freezer. After freeze-drying, the solution is finally sterilized under high temperature and high pressure in a high-pressure steam sterilizer to obtain the final gallium porphyrin complex / silk fibroin antibacterial dressing.

[0005] The gel materials in the above patents all require dialysis of the dissolved silk fibroin solution during preparation. This not only consumes a large amount of deionized water but also consumes a significant amount of time and effort. Furthermore, the metal ions in the dialysis water are difficult to recover. Moreover, the tensile and adhesive properties of the hydrogels prepared through this complex dialysis process still rely on the addition of toxic cross-linking agents to maintain them. In addition, the subsequent hydrogel preparation process requires several or more operations such as freezing and freeze-drying, complicating the production process for enterprises and making it difficult to build a complete production line. This limits silk fibroin hydrogel dressings to the laboratory stage. Summary of the Invention

[0006] The purpose of this invention is to provide a method for preparing multifunctional silk fibroin hydrogels that do not require dialysis, thereby solving the problems of poor mechanical properties and environmental responsiveness of hydrogels prepared by SF.

[0007] A second objective of this invention is to provide a multifunctional silk fibroin hydrogel that does not require dialysis.

[0008] A third objective of this invention is to provide the application of a dialysis-free, multifunctional silk fibroin hydrogel in wound dressings.

[0009] The first technical solution adopted in this invention is a method for preparing a multifunctional silk fibroin hydrogel that does not require dialysis, specifically including the following steps: Step 1: Prepare silk fibroin fibers; Step 2: Prepare a silk fibroin solution based on the product obtained in Step 1; Step 3: Prepare silk fibroin hydrogel based on the product obtained in Step 2.

[0010] The first technical solution of this invention is further characterized by: The specific process of step 1 is as follows: Add Na2CO3 to deionized water and stir to mix evenly to obtain Na2CO3 solution. Then heat the Na2CO3 solution to boiling in an electric heating mantle, add silkworm cocoons that have been pupated, and start timing from the time the solution boils again after the silkworm cocoons are added. Boil for 30 min-60 min and repeat the above process 1-4 times. Wash with deionized water and dry in an oven to obtain silk fibroin fiber.

[0011] In step 1, the ratio of the depupaed silkworm cocoons to the sodium carbonate solution is 1 g:400 mL - 1 g:25 mL.

[0012] In step 1, the oven temperature is 60-70 ℃.

[0013] The specific process of step 2 is as follows: Add anhydrous CaCl2, anhydrous ethanol and deionized water to the round-bottomed sesame cake, add a rotor and stir thoroughly to mix evenly, then add the silk fibroin fiber obtained in step 1, place it in a water bath at 60-80 ℃ and heat to dissolve, transfer the dissolved silk fibroin solution to a centrifuge tube, centrifuge for 5-15 minutes, take the supernatant and transfer it to a clean centrifuge tube for storage, and obtain the silk fibroin solution.

[0014] In step 2, the molar ratio of anhydrous CaCl2, anhydrous ethanol, and deionized water is 1:2:8.

[0015] In step 2, the heating and dissolving time is 60 min-90 min.

[0016] The specific process of step 3 is as follows: Add tannic acid to deionized water to prepare a tannic acid solution. Mix the tannic acid solution with the silk fibroin solution prepared in step 2 at a mass ratio of 1:1-3 and stir evenly with a glass rod to obtain silk fibroin hydrogel.

[0017] The second technical solution adopted in this invention is a dialysis-free multifunctional silk fibroin hydrogel, which is prepared by the above-mentioned method for preparing dialysis-free multifunctional silk fibroin hydrogel. The third technical solution adopted in this invention is the application of a non-dialysis multifunctional silk fibroin hydrogel in wound dressings.

[0018] The beneficial effects of this invention are that, using silk fibroin hydrogel as a substrate, its three-dimensional network structure, low density, high specific surface area, and large porosity provide more binding sites for tannic acid. This fully utilizes the advantages of Ca... 2+ Its functions include: breaking the hydrogen bonds between silk fibroin molecular chains during dissolution, thus allowing the silk fibroin fibers to dissolve; inhibiting the formation of β-coagulation nuclei in the silk fibroin solution during storage, thus prolonging its storage time; and enhancing the cross-linking between silk fibroin and tannic acid through the chelating effect of metal ions during gelation, resulting in excellent mechanical strength of the hydrogel. Attached Figure Description

[0019] Figure 1 This is a SEM image of the silk fiber surface obtained in Example 3 of the preparation method of the non-dialysis multifunctional silk fibroin hydrogel of the present invention. Figure 2 Example 3 shows the structural diagram of the hydrogel prepared by the preparation method of the non-dialysis multifunctional silk fibroin hydrogel of the present invention. Figure 3 This is a physical image of the silk fibroin hydrogel obtained in Example 3 of the preparation method of the non-dialysis multifunctional silk fibroin hydrogel of the present invention. Figure 4 This is a stretched image of the silk fibroin hydrogel obtained in Example 3 of the preparation method of the non-dialysis multifunctional silk fibroin hydrogel of the present invention. Figure 5 This is a picture of the silk fibroin hydrogel obtained in Example 3 of the preparation method of the non-dialysis multifunctional silk fibroin hydrogel of the present invention, which adheres to and lifts a 500 g weight. Figure 6 This is an example of the preparation method of the non-dialysis multifunctional silk fibroin hydrogel of the present invention. The silk fibroin hydrogel obtained in Example 3 is adhered to different objects. Figure 7 This is a physical image of the silk fibroin hydrogel obtained from Example 3 of the preparation method of the non-dialysis multifunctional silk fibroin hydrogel of the present invention after being peeled off from human skin. Figure 8 This is a physical image of the pigskin overlap shearing of the silk fibroin hydrogel obtained in Example 3 of the preparation method of the non-dialysis multifunctional silk fibroin hydrogel of the present invention. Figure 9The preparation method of the non-dialysis multifunctional silk fibroin hydrogel of the present invention is shown in Examples 1-4, which are the shear tensile stress-strain diagrams of the pigskin overlap of the silk fibroin hydrogel obtained. Figure 10 These are antibacterial images of the silk fibroin hydrogels obtained in Examples 1-4 of the preparation method of the non-dialysis multifunctional silk fibroin hydrogel of the present invention. Figure 11 These are SEM images of the internal surface of the silk fibroin hydrogel obtained in Examples 1-4 of the preparation method of the non-dialysis multifunctional silk fibroin hydrogel of the present invention after quenching. Figure 12 These are the infrared spectra of the silk fibroin hydrogels obtained in Examples 2-4 of the preparation method of the multifunctional silk fibroin hydrogel that does not require dialysis according to the present invention. Detailed Implementation

[0020] The following detailed description is provided in conjunction with specific implementation methods.

[0021] The preparation method of the non-dialysis multifunctional silk fibroin hydrogel of the present invention specifically includes the following steps: Step 1: Weigh 1.06 g of Na2CO3 and add it to 500 mL of deionized water. Stir and mix well to obtain 500 mL of 0.02 M Na2CO3 solution. Then, heat it in an electric heating mantle until it boils. Add 1.25-20 g of depupaed silkworm cocoons, so that the ratio of depupaed silkworm cocoons to sodium carbonate solution is 1 g:400 mL-1 g:25 mL. Start timing from the time the solution boils again after the silkworm cocoons are added. Boil for 30 min-60 min. Repeat the above process 1-4 times. Wash with deionized water 3 times. Place in an oven at 60-70 ℃ for 24-12 hours to obtain silk fibroin fiber.

[0022] Step 2: Prepare the dissolving solution. Add 11.1 g of anhydrous CaCl2, 9.2 g of anhydrous ethanol and 14.4 g of deionized water to 100 mL of round-bottom sesame cake. The molar ratio of the resulting ternary system is 1:2:8. After thoroughly mixing with a rotor, add 3-5 g of the silk fibroin fiber obtained in Step 1. Place the mixture in a 60-80 ℃ water bath and heat to dissolve for 60-90 min. Transfer the dissolved silk fibroin solution to a 50 mL centrifuge tube and centrifuge at 5800 rpm for 5-15 min. Transfer the supernatant to a clean centrifuge tube and store at 4 ℃ to obtain the silk fibroin solution.

[0023] Step 3: Add 5.26~25g of tannic acid to 100mL of deionized water to prepare 100mL of tannic acid solution with a concentration of 5%~20%. Mix the tannic acid solution with the silk fibroin solution prepared in step 2 at a mass ratio of 1:1-1:3 and stir rapidly with a glass rod to obtain silk fibroin hydrogel.

[0024] Example 1 Step 1: Weigh 1.06 g of Na2CO3 and add it to 500 mL of deionized water. Stir and mix well to obtain 500 mL of 0.02 M Na2CO3 solution. Heat the solution in an electric heating mantle until boiling, then add 1.25 g of depupaed silkworm cocoons to make the ratio of depupaed silkworm cocoons to sodium carbonate solution 1 g: 400 mL. Start timing from the time the solution boils again after the silkworm cocoons are added. Boil for 30 min, wash three times with deionized water, and place in a 60 ℃ oven for 24 hours to obtain silk fibroin fiber.

[0025] Step 2: Prepare the dissolving solution. Add 11.1 g of anhydrous CaCl2, 9.2 g of anhydrous ethanol and 14.4 g of deionized water to 100 mL of round-bottom sesame cake. The molar ratio of the resulting ternary system is 1:2:8. After thoroughly mixing with a rotor, add 3 g of the silk fibroin fiber obtained in Step 1. Place the mixture in a 60 ℃ water bath and heat to dissolve for 60 min. Transfer the dissolved silk fibroin solution to a 50 mL centrifuge tube and centrifuge at 5800 rpm for 5 min. Transfer the supernatant to a clean centrifuge tube and store at 4 ℃ to obtain the silk fibroin solution.

[0026] Step 3: Add 5.26 g of tannic acid to 100 mL of deionized water to prepare 100 mL of 5% tannic acid solution. Mix the tannic acid solution with the silk fibroin solution prepared in Step 2 at a mass ratio of 1:1 and stir rapidly with a glass rod to obtain silk fibroin hydrogel.

[0027] Example 2 Step 1: Weigh 1.06 g of Na2CO3 and add it to 500 mL of deionized water. Stir and mix well to obtain 500 mL of 0.02 M Na2CO3 solution. Heat the solution in an electric heating mantle until boiling, then add 5 g of depupaed silkworm cocoons, so that the ratio of depupaed silkworm cocoons to sodium carbonate solution is 1 g: 100 mL. Start timing from the time the solution boils again after the silkworm cocoons are added. Boil for 30 min, wash 3 times with deionized water, and place in a 65 ℃ oven for 18 hours to obtain silk fibroin fiber.

[0028] Step 2: Prepare the dissolving solution. Add 11.1 g of anhydrous CaCl2, 9.2 g of anhydrous ethanol and 14.4 g of deionized water to 100 mL of round-bottom sesame cake. The molar ratio of the resulting ternary system is 1:2:8. After thoroughly mixing with a rotor, add 4 g of the silk fibroin fiber obtained in Step 1. Place the mixture in a 70 ℃ water bath and heat to dissolve for 70 min. Transfer the dissolved silk fibroin solution to a 50 mL centrifuge tube and centrifuge at 5800 rpm for 10 min. Transfer the supernatant to a clean centrifuge tube and store at 4 ℃ to obtain the silk fibroin solution.

[0029] Step 3: Add 17.65 g of tannic acid to 100 mL of deionized water to prepare 100 mL of 15% tannic acid solution. Mix the tannic acid solution with the silk fibroin solution prepared in Step 2 at a mass ratio of 1:2 and stir rapidly with a glass rod to obtain silk fibroin hydrogel.

[0030] Example 3 Step 1: Weigh 1.06 g of Na₂CO₃ and add it to 500 mL of deionized water. Stir and mix thoroughly to prepare 500 mL of 0.02 M Na₂CO₃ solution. Heat the solution in an electric heating mantle until boiling. Add 20 g of depupaed silkworm cocoons, making the ratio of depupaed silkworm cocoons to sodium carbonate solution 1 g: 25 mL. Start timing from when the solution boils again after adding the silkworm cocoons. Boil for 5 minutes, then rinse three times with tap water. Before rinsing, weigh 1.06 g of Na₂CO₃ again and add it to 500 mL of deionized water. Stir and mix thoroughly, then heat and boil in an electric heating mantle. Squeeze out the water from the rinsed silkworm silk and place it in the boiling solution. Boil for 10 minutes from when the solution boils again. Rinse three times with tap water. Before rinsing, weigh 1.06 g of Na₂CO₃ again and add it to 500 mL of deionized water. Mix the silk fibers thoroughly with deionized water for 15 minutes, then heat in an electric heating mantle until boiling. Squeeze out the water from the washed silk and place it in the boiling solution. Boil for 15 minutes after the solution boils again. Finally, wash the silk fibers three times with deionized water, squeeze out the water, and place it in a 70 ℃ oven for 12 hours to obtain silk fibroin fibers.

[0031] Step 2: Prepare the dissolving solution. Add 11.1 g of anhydrous CaCl2, 9.2 g of anhydrous ethanol and 14.4 g of deionized water to 100 mL of round-bottom cake. The molar ratio of the resulting ternary system is 1:2:8. After stirring thoroughly with a rotor, add 4 g of the silk fibroin fiber obtained in Step 1. Place the mixture in a 70 ℃ water bath and heat to dissolve for 90 min. Transfer the dissolved silk fibroin solution to a 50 mL centrifuge tube and centrifuge at 5800 rpm for 10 min. Transfer the supernatant to a clean centrifuge tube and store at 4 ℃ to obtain the silk fibroin solution.

[0032] Step 3: Add 11.11g of tannic acid to 100mL of deionized water to prepare 100mL of tannic acid solution with a concentration of 10%. Mix the tannic acid solution with the silk fibroin solution prepared in step 2 at a mass ratio of 1:2 and stir rapidly with a glass rod to obtain silk fibroin hydrogel. Remove the hydrogel from the beaker, rinse it with deionized water, and it can be stored at 4℃ or frozen for long-term storage.

[0033] Figure 1 This is a SEM image of the silk fibroin fiber surface obtained in Example 3 of the present invention. It can be seen that the surface of the silk fibroin fiber obtained by boiling three times for 10 minutes each time is clean and smooth, with almost no sericin, wax or other impurities remaining, and the fiber is free of burrs and has not been damaged.

[0034] Figure 2 This is a schematic diagram of the internal cross-linking of the silk fibroin hydrogel obtained in Example 3 of the present invention, which is mainly composed of intermolecular hydrogen bonds and metal ion coordination bonds forming a cross-linking network.

[0035] Figure 3 The image shows the actual silk fibroin hydrogel obtained in Example 3 of this invention. It can be seen that its appearance structure is relatively complete and its elasticity is good.

[0036] Figure 4 This is a stretched image of the silk fibroin hydrogel obtained in Example 3 of the present invention. It can be seen that the hydrogel's elongation can reach 10,000%, demonstrating excellent stretchability.

[0037] Figure 5 This is a picture of the silk fibroin hydrogel obtained in Example 3 of the present invention, which adheres to and lifts a 500 g weight. Figure 5 The attached diagrams on the left and right show different adhesion positions of the hydrogel on the weight. It can be seen that the hydrogel adhering between the acrylic plate and the 500g weight can easily stick the weight, proving that it has excellent adhesion properties. Figure 5 The top of the attached image on the left shows uniform adhesion force. Figure 5 The attached diagram on the right shows a small adhesion area on the side, resulting in uneven stress, yet stable adhesion is still achieved.

[0038] Figure 6 These are actual images of the silk fibroin hydrogel obtained in Example 3 of this invention adhering to different objects. It can be seen that the hydrogel exhibits good adhesion performance to different objects and materials.

[0039] Figure 7 This is a photograph of the silk fibroin hydrogel obtained in Example 3 of the present invention being removed from human skin. It can be seen that although the hydrogel exhibits excellent adhesion to human skin, no residue is left during the removal process, and there are no adverse effects on the skin due to excessive adhesion.

[0040] Figure 8 This is a physical image of the pigskin overlap and shearing of the silk fibroin hydrogel obtained in Example 3 of the present invention. The wet pigskin is used to simulate human skin after sweating. After longitudinal stretching, the hydrogel is still firmly adhered to the surface of the pigskin without falling off, which can ensure that the hydrogel can still adhere to the human skin during large-scale exercise.

[0041] Figure 9 These are the shear-tensile stress-strain diagrams of the silk fibroin hydrogel obtained in Examples 1-4 of this invention. It can be seen that the obtained hydrogel can withstand stresses greater than 100 kPa even when adhered to the pigskin surface, and can still maintain nearly 1,000% strain performance, allowing it to adhere to the skin surface even during vigorous exercise and sweating.

[0042] Figure 10 These are actual images of the antibacterial silk fibroin hydrogels obtained in Examples 1-4 of this invention. It can be seen that when the concentration of tannic acid increases to 10%, the resulting hydrogel exhibits excellent bactericidal ability against both Escherichia coli and Staphylococcus aureus, effectively preventing bacterial infection of wounds and thus promoting wound healing.

[0043] Figure 11 These are SEM images of the internal surface of the silk fibroin hydrogels obtained in Examples 1-4 of this invention after quenching. The images show that hydrogels with different TA (tannic acid) concentrations all possess abundant porous structures, which facilitates gas exchange between the wound and the external environment, preventing inflammation and hindering wound healing due to lack of air permeability.

[0044] Figure 12 These are the infrared spectra of silk fibroin hydrogels obtained in Examples 2-4 of this invention. SF at 3435 cm⁻¹ -1 The single peaks appearing in the left and right spectral regions are typical characteristic absorption peaks of the NH stretching vibration of proteins. (1628 cm⁻¹) -1 The resulting absorption peak is due to the C=O stretching vibration, a characteristic absorption peak of amide I (random coil). (1550 cm⁻¹) -1 The resulting absorption peak is due to the NH bending vibration, a characteristic absorption peak of amide II (β-sheet). 1255 cm⁻¹ -1 The resulting absorption peaks are due to NH bending and CH stretching vibrations, characteristic absorption peaks of amide III (random coil). 952 cm⁻¹ -1 The resulting absorption peak is the characteristic absorption peak of the -OH group in the carboxyl group.

[0045] For TA, at 1710, 1611, 1535, and 1447 cm 1Numerous absorption bands were identified, which were attributed to the stretching vibrations of the C=O group in carboxylic esters, the stretching of aromatic C=C groups, and the C=O group stretching. In-plane bending of the OH group.

[0046] In contrast, TA / SF hydrogels have a range of 1741–1605 cm⁻¹. 1 The characteristic peak bands shown are due to the formation of a typical β-sheet structure and a small number of α-helical conformations. Furthermore, the 3435 cm⁻¹ of SF... -1 And TA's 3416 cm -1 The peak at that location (attributed to) NH2 and (OH stretching vibrations), which migrate to 3490 cm⁻¹ after gelation. -1 At approximately 1033 and 1628 cm 1 The absorption band at that point corresponds to the C of TA. The OH stretching vibration and the C=O stretching vibration of amide I in SF migrate to 1023 and 1648 cm⁻¹, respectively, in TA / SF gel. -1 The shifts in these absorption peaks are likely due to molecular complexation between TA and SF during gelation. The results indicate that the introduction of TA not only transforms the conformation of SF from random coil to β-sheet, but also promotes rapid gelation of SF through hydrogen bonding interactions, demonstrating the successful preparation of the hydrogel.

[0047] Example 4 Step 1: Weigh 1.06 g of Na2CO3 and add it to 500 mL of deionized water. Stir and mix well to obtain 500 mL of 0.02 M Na2CO3 solution. Heat the solution in an electric heating mantle until boiling, then add 1.25 g of depupaed silkworm cocoons to make the ratio of depupaed silkworm cocoons to sodium carbonate solution 1 g: 400 mL. Start timing from the time the solution boils again after the silkworm cocoons are added. Boil for 30 min, wash three times with deionized water, and place in a 60 ℃ oven for 24 hours to obtain silk fibroin fiber.

[0048] Step 2: Prepare the dissolving solution. Add 11.1 g of anhydrous CaCl2, 9.2 g of anhydrous ethanol and 14.4 g of deionized water to 100 mL of round-bottom sesame cake. The molar ratio of the resulting ternary system is 1:2:8. After thoroughly mixing with a rotor, add 3 g of the silk fibroin fiber obtained in Step 1. Place the mixture in a 60 ℃ water bath and heat to dissolve for 60 min. Transfer the dissolved silk fibroin solution to a 50 mL centrifuge tube and centrifuge at 5800 rpm for 5 min. Transfer the supernatant to a clean centrifuge tube and store at 4 ℃ to obtain the silk fibroin solution.

[0049] Step 3: Add 25g of tannic acid to 100 mL of deionized water to prepare 100 mL of 20% tannic acid solution. Mix the tannic acid solution with the silk fibroin solution prepared in Step 2 at a mass ratio of 1:1 and stir rapidly with a glass rod to obtain silk fibroin hydrogel.

[0050] Example 5 Compared to Example 1, in step 2, the temperature in the water bath is 65°C.

[0051] Example 6 Compared with Example 1, in step 1, the boiling time is 40 minutes.

[0052] This invention employs a three-pronged approach to calcium utilization: ① In the dissolution stage, calcium ions disrupt the hydrogen bonds between silk fibroin fiber molecular chains, and with the synergistic effect of ethanol, rapidly dissolve the silk fibroin fibers to obtain a silk fibroin solution; ② In the storage stage, the presence of calcium ions hinders the formation of β-coagulation nuclei in the silk fibroin solution, thereby inhibiting the coagulation of silk fibroin into antiparallel β-sheets; ③ In the gelation stage, the retained calcium ions fully utilize their metal ion chelating effect in the gelation system, greatly enhancing the stretchability of the hydrogel. This invention solves the problems of existing technologies where silk fibroin dissolution requires dialysis, the metal ions after dialysis are difficult to recover, toxic cross-linking agents are needed during gelation, and the resulting gel has poor mechanical properties, poor adhesion, poor antibacterial properties, and is difficult to industrialize. The silk degumming process, through the method of "large number of silkworm cocoons and multiple degumming", can not only remove impurities but also reduce the waste of electricity and water resources. The silk fibroin fiber dissolution stage eliminates the energy and water consumption caused by dialysis. The high concentration of calcium ions and water precipitated after hydrogel synthesis can be recycled, which greatly reduces production costs and is conducive to the formation of industrialization (water-saving advantage).

Claims

1. A method for preparing a non-dialysis-required multifunctional silk fibroin hydrogel, characterized in that: Specifically, the steps include the following: Step 1: Prepare silk fibroin fibers; Step 2: Prepare a silk fibroin solution based on the product obtained in Step 1; Step 3: Prepare silk fibroin hydrogel based on the product obtained in Step 2.

2. The method for preparing the non-dialysis multifunctional silk fibroin hydrogel according to claim 1, characterized in that: The specific process of step 1 is as follows: Add Na2CO3 to deionized water and stir to mix evenly to obtain Na2CO3 solution. Then heat the Na2CO3 solution to boiling in an electric heating mantle, add silkworm cocoons that have been pupated, and start timing from the time the solution boils again after the silkworm cocoons are added. Boil for 30 min-60 min and repeat the above process 1-4 times. Wash with deionized water and dry in an oven to obtain silk fibroin fiber.

3. The method for preparing the non-dialysis multifunctional silk fibroin hydrogel according to claim 2, characterized in that: In step 1, the ratio of the depupaed silkworm cocoon to the sodium carbonate solution is 1 g:400 mL - 1 g:25 mL.

4. The method for preparing the non-dialysis multifunctional silk fibroin hydrogel according to claim 2, characterized in that: In step 1, the temperature of the oven is 60-70 ℃.

5. The method for preparing the non-dialysis multifunctional silk fibroin hydrogel according to claim 2, characterized in that: The specific process of step 2 is as follows: add anhydrous CaCl2, anhydrous ethanol and deionized water to the round-bottomed sesame cake, add a rotor and stir thoroughly to mix evenly, add the silk fibroin fiber obtained in step 1, put it in a water bath at 60-80 ℃ and heat to dissolve, transfer the dissolved silk fibroin solution to a centrifuge tube, centrifuge for 5-15 minutes, take the supernatant and transfer it to a clean centrifuge tube for storage to obtain the silk fibroin solution.

6. The method for preparing the non-dialysis multifunctional silk fibroin hydrogel according to claim 5, characterized in that: In step 2, the molar ratio of anhydrous CaCl2, anhydrous ethanol, and deionized water is 1:2:

8.

7. The method for preparing the non-dialysis multifunctional silk fibroin hydrogel according to claim 5, characterized in that: In step 2, the heating and dissolving time is 60 min-90 min.

8. The method for preparing the non-dialysis multifunctional silk fibroin hydrogel according to claim 5, characterized in that: The specific process of step 3 is as follows: Add tannic acid to deionized water to prepare a tannic acid solution. Mix the tannic acid solution with the silk fibroin solution prepared in step 2 at a mass ratio of 1:1-3 and stir evenly with a glass rod to obtain silk fibroin hydrogel.

9. A non-dialysis-required multifunctional silk fibroin hydrogel, prepared by the method for preparing a non-dialysis-required multifunctional silk fibroin hydrogel as described in any one of claims 1 to 8.

10. Application of non-dialysis multifunctional silk fibroin hydrogel in wound dressings.

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