Silk fibroin gel material, preparation method and application

Abstract

This invention relates to a method for preparing silk fibroin gel materials and their applications. Its advantages lie in the fact that this invention utilizes a combined process of "structure regulator adjustment + heat treatment + cold treatment + mechanical dispersion" to promote the formation of a stable β-sheet structure in silk fibroin, constructing a robust gel network. The resulting silk fibroin gel microparticles remain stable under complex process conditions and when compounded with various raw materials, effectively solving the problem of insufficient stability in existing silk fibroin gels. Furthermore, no toxic chemical cross-linking agents are used in the preparation process of this invention; the selected structure regulators are all non-toxic and biocompatible substances, and silk fibroin itself possesses excellent biocompatibility. Therefore, the prepared silk fibroin gel microparticles are non-toxic and non-irritating, and can be safely applied in biomedical and other fields with high safety requirements.

Description

Technical Field

[0001] This invention relates to the field of biomedical materials technology, and specifically to a method for preparing silk fibroin gel microparticles, a method for preparing silk fibroin gel dressings, silk fibroin gel materials, and their applications. Background Technology

[0002] In recent years, the field of biomedical materials has continued to develop rapidly, and clinical practice has placed higher demands on the biosafety, compatibility, and functionality of medical materials used in wound care and soft tissue repair. Natural biomaterials, with their excellent biocompatibility, biodegradability, and low immunogenicity, have gradually become a core research direction in wound repair, tissue engineering, and other fields. Among them, silk fibroin, a natural high-molecular-weight protein extracted from silk, is rich in hydrophilic amino acids such as glycine and alanine. It can provide an ideal biomimetic microenvironment for cell adhesion and proliferation, and can also form a stable three-dimensional network gel by regulating its molecular conformation, demonstrating outstanding application potential in medical scenarios such as wound dressings, soft tissue filling, and drug delivery systems.

[0003] Despite the promising application prospects of silk fibroin materials, their large-scale preparation and clinical translation are still limited by several technical bottlenecks. Current preparation processes for silk fibroin gels are quite demanding. During processing under extreme pH conditions, drastic temperature fluctuations, or high-intensity mechanical stirring, or when used in combination with other functional components, the system is prone to protein aggregation, sedimentation, and damage to the gel network structure. This directly leads to unstable material performance and limited applicability, severely hindering its industrial production and clinical application.

[0004] Existing technologies have also failed to effectively address the aforementioned shortcomings. For example, Chinese invention patent (CN202210758330.1) discloses a silk fibroin hydrogel dressing and its preparation method. This method involves loading a silk fibroin protein solution onto a carrier after cyclone shearing and then achieving in-situ gelation through heat treatment. However, the gel dressing obtained by this technology has limited application scenarios, and after conventional irradiation sterilization, the gel disintegrates, resulting in an uneven mixture of liquid and gel, which significantly reduces the user experience and clinical application effectiveness.

[0005] To improve the stability of silk fibroin gel microparticles, existing technologies generally employ chemical cross-linking modification strategies, adding cross-linking agents such as glutaraldehyde and genipin to enhance the stability of the gel structure. However, these chemical cross-linking agents are prone to leaving toxic and harmful residues, which can significantly reduce the biocompatibility of the material, posing potential biosafety risks and failing to meet the safety requirements for high-end medical materials.

[0006] In summary, existing silk fibroin gel materials generally suffer from problems such as insufficient stability, complex preparation processes, and easy introduction of toxic components, making it difficult to simultaneously meet the comprehensive needs of industrial production, multi-scenario compounding, and clinical biosafety. Summary of the Invention

[0007] The purpose of this invention is to address the shortcomings of existing technologies by providing a method for preparing silk fibroin gel microparticles, a method for preparing silk fibroin gel dressings, silk fibroin gel materials and their applications, in order to solve problems such as poor stability, complex preparation processes, poor biocompatibility, and the presence of toxic and harmful substances in related technologies.

[0008] To achieve the above objectives, the technical solution adopted by the present invention is as follows: In a first aspect, a method for preparing silk fibroin gel microparticles is provided, comprising: A structure modifier is added to a silk fibroin solution and mixed to obtain a mixed solution. The mixed solution is subjected to heat treatment and cold treatment in sequence to obtain the treated mixed solution; The treated mixture was mechanically dispersed to obtain silk fibroin gel microparticles.

[0009] In some of these embodiments, the structure modifier is at least one selected from polyvinyl alcohol, polyethylene glycol, glycerin, propylene glycol, butanediol, pentanediol, hexanediol, and sorbitol.

[0010] In some of these embodiments, the solution concentration of the structure modifier is 20% to 65%.

[0011] In some of these embodiments, the mass ratio of the structure modifier to the silk fibroin solution is 1 to 1.5.

[0012] In some of these embodiments, the mixing rate is 100-600 rpm, the mixing time is 1-12 h, and the mixing temperature is 24-60°C.

[0013] In some of these embodiments, the heat treatment temperature is 80~121°C and the heat treatment time is 0.3~8 h.

[0014] In some of these embodiments, the cold treatment temperature is -20 to 10°C and the cold treatment time is 1 to 24 hours.

[0015] In some of these embodiments, the heat treatment and cold treatment are performed 1 to 3 times.

[0016] In some of these embodiments, the mechanical dispersion treatment is at least one of ultrasonic dispersion treatment, high-pressure homogenization treatment, and high-speed shearing treatment.

[0017] In some of these embodiments, the ultrasonic dispersion power is 100-500 W and the ultrasonic dispersion time is 10-60 min.

[0018] In some of these embodiments, the high-pressure homogenization pressure is 50-150 MPa, and the number of high-pressure homogenization cycles is 2-10.

[0019] In some of these embodiments, the high-speed shearing process involves a high-speed shearing rotation speed of 1000-15000 rpm and a high-speed shearing time of 10-60 min.

[0020] In some of these embodiments, it also includes: Regenerated silk fibroin is dissolved in water to obtain a regenerated silk fibroin solution; The regenerated silk fibroin solution was centrifuged and filtered to obtain a silk fibroin solution.

[0021] In some of these embodiments, the molecular weight of the regenerated silk fibroin is 2 to 10 W, and the concentration of the regenerated silk fibroin solution is 1% to 20%.

[0022] In some of these embodiments, the centrifugation speed is 3000~8000 rpm and the centrifugation time is 5~10 min.

[0023] In some of these embodiments, the filter mesh size is 100 to 1000 mesh during the filtration process.

[0024] In some of these embodiments, the particle size of the silk fibroin gel microparticles is 100 nm to 50 μm.

[0025] Secondly, a method for preparing a silk fibroin gel dressing is provided, comprising: The silk fibroin gel microparticles prepared by the preparation method described in the first aspect are added to the dressing solution and mixed and homogenized in sequence to obtain a mixed solution. The mixed solution is sterilized to obtain a silk fibroin gel dressing.

[0026] In some embodiments, the dressing solution includes water and a dressing material, wherein the dressing material is at least one selected from trehalose, sodium hyaluronate, agar, gelatin, collagen, recombinant collagen, chitosan, xanthan gum, carbomer, carboxymethyl cellulose, hydroxyethyl cellulose, fruitwood fiber, and regenerated cellulose fiber.

[0027] In some of these embodiments, the homogenization rate is 1000-5000 rpm and the homogenization time is 10-60 min.

[0028] In some of these embodiments, the sterilization process is at least one of moist heat sterilization and irradiation sterilization.

[0029] Thirdly, a silk fibroin gel material is provided, comprising: Silk fibroin gel microparticles prepared by the preparation method described in the first aspect; or Silk fibroin gel dressing prepared by the preparation method described in the second aspect.

[0030] Fourthly, an application of the silk fibroin gel material as described in the third aspect is provided, the application including at least one of the following: Application of coverings for non-chronic wounds; Applications in the care of non-chronic wounds; Wrinkle filling application; Application for filling skin tissue defects.

[0031] The present invention adopts the above technical solution and has the following technical effects compared with the prior art: 1) Excellent stability: This invention uses a combination of "structure regulator + heat treatment + cold treatment + mechanical dispersion" to promote the formation of a stable β-sheet structure of silk fibroin and build a strong gel network. The prepared silk fibroin gel microparticles can remain stable under complex process conditions and when compounded with a variety of raw materials, effectively solving the problem of insufficient stability of silk fibroin gel in the prior art. 2) High biocompatibility: No toxic chemical cross-linking agents are used in the preparation process of this invention. The selected structure modifiers are all non-toxic and biocompatible substances. Silk fibroin itself has excellent biocompatibility. Therefore, the prepared silk fibroin gel microparticles are non-toxic and non-irritating, and can be safely applied in fields with high safety requirements such as biomedicine. 3) The preparation process is simple and controllable: The preparation method of the present invention has clear steps, and each process parameter (such as solution concentration, structure regulator ratio, temperature, time, etc.) can be precisely controlled, which is convenient for industrial production; at the same time, mechanical dispersion treatment can flexibly control the particle size to meet the personalized needs of different application scenarios. Attached Figure Description

[0032] Figure 1 This is a schematic diagram of silk fibroin gel microparticles according to Embodiment 2 of the present invention; Figure 2 This is a schematic diagram of the particle size distribution of silk fibroin gel microparticles according to Embodiment 2 of the present invention; Figure 3 This is a schematic diagram of the silk fibroin gel dressing according to Embodiment 6 of the present invention; Figure 4 These are comparative diagrams of Embodiments 3 to 5 and Comparative Example 1 according to the present invention. Detailed Implementation

[0033] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0034] It should be noted that, unless otherwise specified, the embodiments and features described in the present invention can be combined with each other.

[0035] The present invention will be further described below with reference to the accompanying drawings and specific embodiments, but this is not intended to limit the scope of the invention.

[0036] Example 1 This embodiment relates to the preparation method of silk fibroin gel microparticles, the preparation method of silk fibroin gel dressing, silk fibroin gel materials and their applications.

[0037] An illustrative embodiment of the present invention provides a method for preparing silk fibroin gel microparticles, comprising: Mixing step (step S3): The structure regulator is added to the silk fibroin solution and mixed to obtain a mixed solution; The hot and cold treatment step (step S4) involves sequentially performing heat treatment and cold treatment on the mixed solution to obtain the treated mixed solution. Mechanical dispersion step (step S5): The treated mixed solution is mechanically dispersed to obtain silk fibroin gel microparticles.

[0038] The silk fibroin gel microparticles of the present invention have a particle size of 100 nm to 50 μm.

[0039] In the mixing step, the structure modifier is at least one of polyvinyl alcohol, polyethylene glycol, glycerin, propylene glycol, butanediol, pentanediol, hexanediol, and sorbitol.

[0040] It should be noted that polyethylene glycol is any one or more combinations of polyethylene glycol 200 to polyethylene glycol 4000.

[0041] It should be noted that, in this invention, the structure modifier is added to the silk fibroin solution in solution form.

[0042] In the mixing step, the solution concentration of the structure modifier is 20% to 65%.

[0043] In the mixing step, the mass ratio of the structure regulator to the silk fibroin solution is 1 to 1.5.

[0044] It should be noted that the mass ratio of the structure modifier to the silk fibroin solution is the mass ratio of the structure modifier solution to the silk fibroin solution.

[0045] In the mixing step, the mixing rate is 100~600 rpm, the mixing time is 1~12h, and the mixing temperature is 24~60℃.

[0046] In the heat treatment step, the heat treatment temperature is 80~121℃ and the heat treatment time is 0.3~8 h.

[0047] In the hot and cold treatment steps, the cold treatment temperature is -20~10℃ and the cold treatment time is 1~24 h.

[0048] In the hot and cold treatment steps, the number of times heat treatment and cold treatment are performed is 1 to 3.

[0049] It is understandable that one heat treatment and one cold treatment constitute a heat-cold treatment process, while multiple heat treatments and cold treatments refer to multiple heat-cold treatment processes. Taking two processes as an example, it requires heat treatment, cold treatment, heat treatment, and cold treatment in sequence.

[0050] It should be noted that the purpose of alternating heat treatment and cold treatment in this invention is to precisely control molecular conformation and crystallization behavior, thereby achieving targeted optimization of material properties. Specifically: 1) High temperature disrupts the intramolecular hydrogen bonds of silk fibroin, causing the random coil / α-helix structure (Silk I) to transform into β-sheet (Silk II). The β-sheet forms a rigid crystalline domain through intermolecular hydrogen bonds, which improves the thermal stability of the material. 2) Low temperature inhibits molecular chain movement and retains part of the Silk I structure. Its hydrophilic region, which is rich in amino acids, can capture water molecules to form an ordered hydration layer.

[0051] In the mechanical dispersion step, the mechanical dispersion treatment is at least one of ultrasonic dispersion treatment, high pressure homogenization treatment, and high-speed shearing treatment.

[0052] In the ultrasonic dispersion process, the ultrasonic dispersion power is 100~500 W and the ultrasonic dispersion time is 10~60 min.

[0053] In the high-pressure homogenization process, the high-pressure homogenization pressure is 50~150 MPa, and the number of high-pressure homogenization cycles is 2~10.

[0054] In the high-speed shearing process, the high-speed shearing speed is 1000~15000 rpm and the high-speed shearing time is 10~60 min.

[0055] Furthermore, prior to the mixing step, the following steps are also included: Dissolution step (step S1): Dissolve the regenerated silk fibroin in water to obtain a regenerated silk fibroin solution; Centrifugation and filtration step (step S2): The regenerated silk fibroin solution is centrifuged and filtered to obtain a silk fibroin solution.

[0056] In the dissolution step, the molecular weight of the regenerated silk fibroin is 2-10 W, and the concentration of the regenerated silk fibroin solution is 1%-20%.

[0057] In the centrifugal filtration step, the centrifugal speed is 3000~8000 rpm and the centrifugation time is 5~10 min.

[0058] In the centrifugal filtration step, the filter mesh size is 100~1000 mesh.

[0059] It should be noted that in the centrifugal filtration step, centrifugation and filtration can be performed simultaneously, or centrifugation can be performed first, followed by filtration.

[0060] It should be noted that the key innovation of this invention lies in steps S3 to S5, namely, the combination of introducing a structure modifier, heat treatment-cold treatment, and mechanical dispersion treatment.

[0061] Furthermore, the silk fibroin gel microparticles obtained based on the above preparation method can be used to prepare silk fibroin gel dressings. The preparation method for silk fibroin gel dressings is as follows: Mixing step (step S6): Silk fibroin gel microparticles are added to the dressing solution and mixed and homogenized sequentially to obtain a mixed solution; Sterilization step (step S8): The mixed solution is sterilized to obtain silk fibroin gel dressing.

[0062] In the mixing step, the dressing solution includes water and dressing material, wherein the dressing material is at least one of trehalose, sodium hyaluronate, agar, gelatin, collagen, recombinant collagen, chitosan, xanthan gum, carbomer, carboxymethyl cellulose, hydroxyethyl cellulose, fruit wood fiber, and regenerated cellulose fiber.

[0063] In the mixing step, the homogenization rate is 1000~5000 rpm and the homogenization time is 10~60 min.

[0064] In the sterilization step, the sterilization treatment is at least one of moist heat sterilization and irradiation sterilization.

[0065] It should be noted that the sterilization process includes the following steps: Filling step (step S7): Fill the mixed solution into a sealed container.

[0066] The silk fibroin gel microparticles and silk fibroin gel dressings described above can be applied to the following scenarios: Application of coverings for non-chronic wounds; Applications in the care of non-chronic wounds; Wrinkle filling application; Application for filling skin tissue defects.

[0067] It should be noted that non-chronic wounds refer to wounds caused by laser surgery, chemical peels, minimally invasive cosmetic procedures, etc.

[0068] It should be noted that wrinkle filling and skin tissue defect filling refer to filling in areas such as the face and lips.

[0069] It should be noted that the skin tissue defect filling application is a volume defect filling application.

[0070] The preparation method of the silk fibroin gel microparticles and dressings of the present invention and their applications have the following technical effects: 1) Excellent stability: This invention uses a combination of "structure regulator + heat treatment + cold treatment + mechanical dispersion" to promote the formation of a stable β-sheet structure of silk fibroin and build a strong gel network. The prepared silk fibroin gel particles can exist stably under complex process conditions and when compounded with a variety of raw materials, effectively solving the problem of insufficient stability of silk fibroin gel in the prior art. 2) High biocompatibility: No toxic chemical cross-linking agents are used in the preparation process of this invention. The selected structure modifiers are all non-toxic and biocompatible substances. Silk fibroin itself has excellent biocompatibility. Therefore, the prepared silk fibroin gel microparticles are non-toxic and non-irritating, and can be safely applied in fields with high safety requirements such as biomedicine. 3) The preparation process is simple and controllable: The preparation method of the present invention has clear steps, and each process parameter (such as solution concentration, structure regulator ratio, temperature, time, etc.) can be precisely controlled, which is convenient for industrial production; at the same time, mechanical dispersion treatment can flexibly control the particle size to meet the personalized needs of different application scenarios.

[0071] Example 2 This embodiment relates to a specific implementation of the silk fibroin gel microparticles of the present invention.

[0072] In this embodiment, the preparation method of silk fibroin gel microparticles is as follows: S1. Dissolve regenerated silk fibroin with a molecular weight of 9w in water to obtain a 5% silk fibroin solution. Centrifuge at 4000 rpm for 8 min and filter through a 500-mesh sieve. Then add a 20% polyethylene glycol 400 solution. The mass ratio of silk fibroin solution to polyethylene glycol solution is 1.2. Start stirring and stir at 500 rpm and 50℃ for 8 h to obtain a mixture.

[0073] S2. Place the mixture prepared in S1 at 100℃ for 1 hour. (i.e., heat treatment) S3. Place the mixture obtained in S2 at -15℃ for 2 hours. (i.e., cold treatment) S4. Repeat steps S2 and S3 three times.

[0074] S5. The system obtained in S4 is sheared at 5000 rpm for 20 min to obtain silk fibroin gel microparticles (e.g. Figure 1 (As shown).

[0075] The particle size distribution of the silk fibroin gel microparticles prepared in this embodiment was detected, and the results are as follows: Figure 2 As shown in the table below.

[0076] Example 3 This embodiment relates to a specific implementation of the silk fibroin gel dressing of the present invention.

[0077] In this embodiment, the preparation method of the silk fibroin gel dressing is as follows: S1. Dissolve regenerated silk fibroin with a molecular weight of 6w in water to obtain a silk fibroin solution with a concentration of 8%, centrifuge at 5000 rpm for 6 min, and filter through an 800-mesh sieve; then add a 30% glycerol solution, with a mass ratio of silk fibroin solution to glycerol solution of 1.5, start stirring, and stir at 200 rpm and 45℃ for 12 h.

[0078] S2. The mixture prepared in S1 is placed in an environment of 80℃ for 4 hours (i.e., heat treatment), and then placed in an environment of 2℃ for 12 hours (i.e., cold treatment). This cycle is repeated twice.

[0079] S3. The system obtained in S2 is sheared at 3000 rpm for 30 min, and then homogenized twice in a high-pressure homogenizer at a pressure of 100 MPa to obtain silk fibroin gel microparticles.

[0080] S4. Mix the silk fibroin gel microparticles prepared in S3 with a 0.2% hydroxyethyl cellulose solution and homogenize at 2000 rpm for 40 min to obtain a mixture.

[0081] S5. Fill the mixture prepared in S4 into an aluminum foil bag containing nonwoven fabric and seal it to obtain silk fibroin gel dressing.

[0082] Example 4 This embodiment relates to a specific implementation of the silk fibroin gel dressing of the present invention.

[0083] In this embodiment, the preparation method of the silk fibroin gel dressing is as follows: S1. Dissolve regenerated silk fibroin with a molecular weight of 6.5w in water to obtain a silk fibroin solution with a concentration of 8%, centrifuge at 5000 rpm for 6 min, and filter through an 800-mesh sieve; then add a 30% glycerol solution, with a mass ratio of silk fibroin solution to glycerol solution of 1.5, start stirring, and stir at 200 rpm and 45℃ for 12 h.

[0084] S2. The mixture prepared in S1 is placed in an environment of 80℃ for 4 hours (i.e., heat treatment), and then placed in an environment of 2℃ for 12 hours (i.e., cold treatment). This cycle is repeated twice.

[0085] S3. The system obtained in S2 is sheared at 3000 rpm for 30 min, and then homogenized twice in a high-pressure homogenizer at a pressure of 100 MPa to obtain silk fibroin gel microparticles.

[0086] S4. Mix the silk fibroin gel microparticles prepared in S3 with a 0.2% hydroxyethyl cellulose solution and homogenize at 2000 rpm for 40 min to obtain a mixture.

[0087] S5. Fill the mixture prepared in S4 into an aluminum foil bag containing nonwoven fabric, then sterilize it by 15 kGy irradiation, and finally seal it to obtain the silk fibroin gel dressing.

[0088] Example 5 This embodiment relates to a specific implementation of the silk fibroin gel dressing of the present invention.

[0089] In this embodiment, the preparation method of the silk fibroin gel dressing is as follows: S1. Dissolve regenerated silk fibroin with a molecular weight of 6.8w in water to obtain a silk fibroin solution with a concentration of 8%, centrifuge at 5000 rpm for 6 min, and filter through an 800-mesh sieve; then add a 30% glycerol solution, with a mass ratio of silk fibroin solution to glycerol solution of 1.5, start stirring, and stir at 200 rpm and 45℃ for 12 h.

[0090] S2. The mixture prepared in S1 is placed in an environment of 80℃ for 4 hours (i.e., heat treatment), and then placed in an environment of 2℃ for 12 hours (i.e., cold treatment). This cycle is repeated twice.

[0091] S3. The system obtained in S2 is sheared at 3000 rpm for 30 min, and then homogenized twice in a high-pressure homogenizer at a pressure of 100 MPa to obtain silk fibroin gel microparticles.

[0092] S4. Mix the silk fibroin gel microparticles prepared in S3 with a 0.2% hydroxyethyl cellulose solution and homogenize at 2000 rpm for 40 min to obtain a mixture.

[0093] S5. Fill the mixture prepared in S4 into an aluminum foil bag containing nonwoven fabric, then sterilize it by irradiation at 25 kGy, and finally seal it to obtain the silk fibroin gel dressing.

[0094] Example 6 This embodiment relates to a specific implementation of the silk fibroin gel dressing of the present invention.

[0095] In this embodiment, the preparation method of the silk fibroin gel dressing includes: S1. Dissolve regenerated silk fibroin with a molecular weight of 3w in water to obtain a silk fibroin solution with a concentration of 15%. Centrifuge at 8000 rpm for 5 min and filter through a 200-mesh sieve. Then add a polyethylene glycol 600 solution with a concentration of 55%. The mass ratio of silk fibroin solution to polyethylene glycol 600 solution is 1.3. Start stirring and stir at 100 rpm and 35℃ for 5 h.

[0096] S2. The mixture prepared in step S1 is placed in an environment of 121℃ for 0.5 h (i.e., heat treatment), and then placed in an environment of 8℃ for 24 h (i.e., cold treatment). This cycle is repeated once.

[0097] S3. The system obtained in step S2 is sheared at 10,000 rpm for 10 min, and then placed in an ultrasonic disperser for 20 min at 300 W to obtain silk fibroin gel microparticles.

[0098] S4. Mix the silk fibroin gel microparticles prepared in S3 with 0.5% carbomer solution and 2% trehalose solution, and homogenize at 5000 rpm for 10 min to obtain a mixture (in gel state).

[0099] S5. Fill the mixture prepared in S4 into a pre-filled syringe, and sterilize by moist heat to obtain the silk fibroin gel dressing (e.g. Figure 3 (As shown).

[0100] It should be noted that, in this embodiment, the silk fibroin gel dressing is in gel form.

[0101] Comparative Example 1 The comparative example is a silk fibroin dressing, which is prepared as follows: S1. Dissolve regenerated silk fibroin with a molecular weight of 5.5w in water to obtain a silk fibroin solution with a concentration of 8%, centrifuge at 5000 rpm for 5 min, and filter through an 800-mesh sieve; then add a glycerol solution with a concentration of 30%, with a mass ratio of silk fibroin solution to glycerol solution of 1.5, start stirring, and stir at 200 rpm and 45℃ for 0.5 h.

[0102] S2. Mix the mixture prepared in S1 with a 0.2% hydroxyethyl cellulose solution and homogenize at 2000 rpm for 40 min to obtain the mixture.

[0103] S3. Fill the mixture prepared in S2 into an aluminum foil bag containing non-woven fabric, seal it, and sterilize it by irradiation to obtain the control sample.

[0104] Test Example 1 This test example describes the physicochemical properties of Examples 3 to 5 and Comparative Example 1. Test parameters include appearance, pH, and viscosity.

[0105] In addition, for Examples 4 and 5, the silk fibroin gel dressing was placed in an environment of 50°C and 75% humidity (high temperature and high humidity environment) for 90 days to test its stability.

[0106] Among them, appearance indicators are determined by visual observation; pH indicators are determined by pH meter; and viscosity indicators are determined by rotational viscometer.

[0107] The test results are shown in the table below.

[0108] Compared with Comparative Example 1, Examples 4 and 5 still had high viscosity after irradiation sterilization, and maintained stable physicochemical properties after 90 days of high temperature and high humidity accelerated test, indicating that the silk fibroin gel dressing of the present invention has good stability.

[0109] Test Example 2 This test example is a physicochemical performance test of Example 6. The test indicators include appearance, pH, and viscosity.

[0110] In addition, the stability of the silk fibroin gel dressing was tested for 90 days in an environment of 50°C and 75% humidity (high temperature and high humidity environment).

[0111] Among them, appearance indicators are determined by visual observation; pH indicators are determined by pH meter; and viscosity indicators are determined by rotational viscometer.

[0112] The test results are shown in the table below.

[0113] The silk fibroin gel dressing (gel-like) prepared in Example 6 maintained stable physicochemical properties after 90 days of high temperature and high humidity accelerated testing, indicating that the silk fibroin gel dressing of the present invention has good stability.

[0114] The above description is merely a preferred embodiment of the present invention and does not limit the implementation and protection scope of the present invention. Those skilled in the art should realize that any equivalent substitutions and obvious changes made based on the description and illustrations of the present invention should be included within the protection scope of the present invention.

Claims

1. A method for preparing silk fibroin gel microparticles, characterized in that, include: A structure modifier is added to a silk fibroin solution and mixed to obtain a mixed solution. The mixed solution is subjected to heat treatment and cold treatment in sequence to obtain the treated mixed solution; The treated mixture was mechanically dispersed to obtain silk fibroin gel microparticles.

2. The preparation method according to claim 1, characterized in that, The structure modifier is at least one of polyvinyl alcohol, polyethylene glycol, glycerin, propylene glycol, butanediol, pentanediol, hexanediol, and sorbitol; and / or The solution concentration of the structure modifier is 20%–65%; and / or The mass ratio of the structure modifier to the silk fibroin solution is 1~1.5; and / or In the mixing process, the mixing rate is 100–600 rpm, the mixing time is 1–12 h, and the mixing temperature is 24–60 °C; and / or In the heat treatment, the heat treatment temperature is 80~121℃, and the heat treatment time is 0.3~8 h; and / or In the cold treatment, the cold treatment temperature is -20~10℃, and the cold treatment time is 1~24 h; and / or The number of heat treatments and cold treatments is 1 to 3; and / or Mechanical dispersion treatment includes at least one of ultrasonic dispersion treatment, high-pressure homogenization treatment, and high-speed shearing treatment.

3. The preparation method according to claim 2, characterized in that, In ultrasonic dispersion, the ultrasonic dispersion power is 100~500 W, and the ultrasonic dispersion time is 10~60 min; and / or In high-pressure homogenization, the homogenization pressure is 50~150 MPa, and the number of homogenization cycles is 2~10; and / or In the high-speed shearing process, the high-speed shearing speed is 1000~15000 rpm, and the high-speed shearing time is 10~60 min.

4. The preparation method according to any one of claims 1 to 3, characterized in that, Also includes: Regenerated silk fibroin is dissolved in water to obtain a regenerated silk fibroin solution; The regenerated silk fibroin solution was centrifuged and filtered to obtain a silk fibroin solution.

5. The preparation method according to claim 4, characterized in that, The molecular weight of regenerated silk fibroin is 2-10 W, and the concentration of the regenerated silk fibroin solution is 1%-20%; and / or In centrifugation, the centrifugation speed is 3000~8000 rpm, and the centrifugation time is 5~10 min; and / or In the filtration process, the filter mesh size is 100~1000 mesh.

6. The preparation method according to any one of claims 1 to 3, characterized in that, The particle size of the silk fibroin gel microparticles ranges from 100 nm to 50 μm.

7. A method for preparing a silk fibroin gel dressing, characterized in that, include: The silk fibroin gel microparticles prepared by any one of the preparation methods described in claims 1 to 6 are added to the dressing solution and mixed and homogenized sequentially to obtain a mixed solution. The mixed solution is sterilized to obtain a silk fibroin gel dressing.

8. The preparation method according to claim 7, characterized in that, The dressing solution comprises water and dressing material, wherein the dressing material is at least one of the following: trehalose, sodium hyaluronate, agar, gelatin, collagen, recombinant collagen, chitosan, xanthan gum, carbomer, carboxymethyl cellulose, hydroxyethyl cellulose, fruit fiber, and regenerated cellulose fiber; and / or In the homogenization process, the homogenization rate is 1000–5000 rpm, and the homogenization time is 10–60 min; and / or The sterilization process includes at least one of moist heat sterilization and irradiation sterilization.

9. A silk fibroin gel material, characterized in that, include: Silk fibroin gel microparticles prepared by any one of the preparation methods described in claims 1 to 6; or Silk fibroin gel dressing prepared by any one of the preparation methods described in claims 7 to 8.

10. An application of the silk fibroin gel material as described in claim 9, characterized in that, The application includes at least one of the following: Application of coverings for non-chronic wounds; Applications in the care of non-chronic wounds; Wrinkle filling application; Application for filling skin tissue defects.

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