Silk fibroin material and extraction method thereof

Abstract

The invention relates to an extraction method of a silk fibroin material, which comprises the following steps: degumming silkworm cocoons / raw silk, dissolving, dialyzing, directionally stimulating, pre-cooling at ultralow temperature, and freeze-drying to obtain the silk fibroin material. In the silk fibroin extraction process, an ordered physical stimulation process is introduced, so that a silk fibroin raw material which has an ordered crystal nucleus region and is easy to form an ordered crystal region through process induction is finally obtained; in the extraction process of the silk fibroin, oriented short physical stimulation is added, so that the secondary structure of the purified silk fibroin has ordered transition state sites which are converted from alpha type to beta type.

Description

Technical Field

[0001] The invention relates to the technical field of biomaterials, and in particular to a silk fibroin material and an extraction method thereof. Background Art

[0002] Silk fibroin is a biomaterial with excellent biocompatibility and biosafety. It is composed of 18 amino acids and has a five-level structure. Through the regulation of the five-level structure, it is widely used in medicine, daily chemicals, food, agricultural chemicals, etc. In recent years, by regulating the secondary structure of silk fibroin, controlling the transformation of silk fibroin from an α-type amorphous structure to a β-folded structure, the physical properties of silk fibroin solutions have been regulated and applied to different scenarios in the medical field.

[0003] The prior art mostly studies and applies the secondary structure of silk fibroin, that is, by regulating the degree of the β-folding structure of silk fibroin to obtain silk fibroin with suitable physical properties, but does not regulate the tertiary structure of silk fibroin, that is, controlling the orderly arrangement of the crystalline regions of silk fibroin. Based on this, the present invention further optimizes it. Summary of the invention

[0004] In view of the defects of the prior art, the purpose of the present invention is to provide a silk fibroin material and an extraction method thereof to solve the problems raised in the above background technology.

[0005] The present invention solves the technical problem by adopting the following technical solution: The present invention provides a method for extracting silk fibroin material, comprising the following steps:

[0006] The silk cocoons / raw silk are degummed, dissolved, dialyzed, and directionally stimulated, and then pre-cooled and freeze-dried to obtain silk fibroin material.

[0007] Preferably, the ultra-low temperature precooling method is: precooling under liquid nitrogen at -80°C for 20-30 minutes.

[0008] Preferably, the directional stimulation is: S1: shear treatment: a high-speed shear treatment of 5-10 min is used to induce the silk fibroin to produce transition state sites distributed along the tangential direction;

[0009] S2: Ultrasonic treatment: 500W ultrasonic frequency was used for 10 min to induce different transition state sites in silk fibroin;

[0010] S3: Shaking on a shaking table: To simulate daily shaking, the shaking program of the shaking table is random shaking, which induces the generation of disordered transition state sites.

[0011] Preferably, the specific operation steps of the degumming are: placing the cocoons / raw silk in sufficient water for 5 hours at a treatment temperature of 90°C, then cooling to 50°C, and then adding alkaline protease of 10% of the total amount of the cocoons / raw silk until the glue content of the cocoons / raw silk is less than 3%, and the degumming technology is completed;

[0012] The specific operation method of the dissolution treatment is: dissolve the degummed silk in a 9-12 mol / L lithium bromide solution for 24 hours.

[0013] Preferably, the dialyzed solution is stirred on a plate by low-frequency magnetic force to produce silk fibroin with planar ordered transition state sites, forming a film-forming silk fibroin solution / gel for skin barrier repair / care;

[0014] The solution thickness is less than 30 mm, magnetic stirring is used, the stirring speed is 50 to 500 rpm, and the stirring time is 5 to 60 minutes.

[0015] Preferably, a rod-shaped ultrasound source is placed in the dialyzed solution to perform ultrasonic vibration stimulation to form a planar orderly transitional state site of silk fibroin with tentacles. During wound repair, the damaged part of the tentacles is embedded in the damaged part, and the planar body forms a silk fibroin membrane.

[0016] Ultrasonic vibration for 5 to 60 minutes, vibration frequency 20k to 100kHz.

[0017] The invention also provides a silk fibroin material obtained by a method for extracting the silk fibroin material.

[0018] The disordered silk fibroin prepared by shaking on a shaker is used to prepare a silk fibroin gel with excellent viscoelasticity and easy injection, which is suitable for preparing an injection filling material for pre-filled packaging;

[0019] A spherical ultrasound source is placed in the solution for ultrasonic vibration stimulation to form tetrahedral silk fibroin. The silk fibroin material is used to obtain a silk fibroin gel with excellent viscoelasticity and mechanical support properties, which is suitable for various types of injectable filling materials.

[0020] Compared with the prior art, the present invention has the following beneficial effects:

[0021] It is difficult to regulate the tertiary structure of the silk fibroin extracted by conventional methods in the present invention. In order to achieve the orderly arrangement of the silk fibroin crystal region, the present invention introduces an orderly physical stimulation process during the silk fibroin extraction process, and finally obtains a silk fibroin raw material with an ordered "crystal core region" that is easy to be induced to form an ordered crystal region through a process; during the extraction of the silk fibroin, a directional short-term physical stimulation is added, so that the secondary structure of the purified silk fibroin has an orderly transition state site that transforms from α type to β type, so that the silk fibroin retains the soluble characteristics, and the silk fibroin can easily induce β folding from the transition state site during the subsequent processing, thereby making it easier to produce ordered crystals. DETAILED DESCRIPTION

[0022] The following is a clear and complete description of the technical solutions in the embodiments of the present invention in conjunction with specific embodiments. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0023] A method for extracting silk fibroin material of this embodiment comprises the following steps:

[0024] The silk cocoons / raw silk are degummed, dissolved, dialyzed, and directionally stimulated, and then pre-cooled and freeze-dried to obtain silk fibroin material.

[0025] The ultra-low temperature precooling method is: precooling under liquid nitrogen at -80°C for 20-30 minutes;

[0026] Silk cocoons / raw silk are made from mulberry silk produced by feeding and feeding in a modern sericulture environment using GMP technology, ensuring that the cocoons have better consistency and traceability.

[0027] Directed stimulation is:

[0028] Shear: Short-term continuous shear force in the same local direction can induce silk fibroin to produce transitional state sites distributed along the tangential direction;

[0029] Ultrasound: Different transition state sites are induced according to the ultrasound frequency and the location of the ultrasound source;

[0030] Shaking on a shaker: Shaking on a shaker simulates daily shaking, providing a way to stimulate the chaotic state and induce the generation of disordered transition state sites.

[0031] The silk fibroin with planar ordered transition state sites produced by low-frequency magnetic stirring on a plate forms a film-forming silk fibroin solution / gel for skin barrier repair / care;

[0032] The dialyzed solution was placed in a flat dish with a thickness of less than 30 mm. The solution was stirred magnetically at a speed of 50 to 500 rpm for 5 to 60 min.

[0033] The application method of the silk fibroin material of this embodiment also includes the following:

[0034] A rod-shaped ultrasound source is placed in a solution to perform ultrasonic vibration stimulation, thereby forming a planar orderly transitional site of silk fibroin with tentacles. During wound repair, the tentacles can pass through the injured part and embed into the injured part, and the planar body can form a silk fibroin membrane.

[0035] The dialyzed solution is placed in a beaker or other container, and a rod-shaped ultrasonic source is placed vertically in the solution. Ultrasonic vibration is performed for 5 to 60 minutes at a vibration frequency of 20k to 100kHz.

[0036] The application method of the silk fibroin material of this embodiment also includes the following:

[0037] The disordered silk fibroin prepared by shaking on a shaker can produce a silk fibroin gel with excellent viscoelasticity and easy injection, which is suitable for preparing an injection filling material for pre-filled packaging;

[0038] Oscillation frequency: 100~1000rpm / min, oscillation time: 1~120min.

[0039] In this embodiment, a spherical ultrasound source is placed in a solution to perform ultrasonic vibration stimulation, thereby forming a tetrahedral-shaped silk fibroin. This silk fibroin material can be used to obtain a silk fibroin gel with excellent viscoelasticity and excellent mechanical support properties, and is suitable for various injection filling materials.

[0040] The dialyzed solution is placed in a beaker or other container, a spherical ultrasonic source is placed under the liquid surface, and ultrasonic vibration is applied for 5 to 60 minutes at a vibration frequency of 20k to 100kHz.

[0041] The silk fibroin material of this embodiment is obtained by a method for extracting a silk fibroin material.

[0042] The soluble silk fibroin prepared by the process of the present invention has stronger adaptability in different application scenarios.

[0043] (1) For example, silk fibroin with planar ordered transition state sites produced by low-frequency magnetic stirring on a plate can easily form a silk fibroin solution / gel with excellent film-forming properties, which can be well used for skin barrier repair / care.

[0044] Reference parameters: Place the dialyzed solution in a plate, the solution thickness is less than 30 mm (preferably less than 10 mm), magnetic stirring, stirring speed 50-500 rpm (preferably 50-150 rpm), stirring time 5-60 min (preferably 15-20 min)

[0045] For example, placing a rod-shaped ultrasound source in a solution and conducting ultrasonic vibration stimulation can form a planar, ordered transition state site of silk fibroin with tentacles. In wound repair, the tentacles can pass through the damaged area and embed into the damaged area. The planar body can form a good silk fibroin membrane, which can play a regenerative repair role while blocking external environmental interference, thereby better promoting damage repair.

[0046] Place the dialyzed solution in a beaker or other container, place a rod-shaped ultrasonic source vertically in the solution, contact the solution as much as possible, and vibrate ultrasonically for 5 to 60 minutes (preferably 10 to 15 minutes) at a vibration frequency of 20k to 100kHz (preferably 30k to 40kHz).

[0047] Placing a spherical ultrasound source in a solution and conducting ultrasonic vibration stimulation can form tetrahedral silk fibroin. Using this silk fibroin material, a silk fibroin gel with excellent viscoelasticity can be obtained, which has excellent mechanical support properties and is suitable for various types of injectable filling materials.

[0048] Place the dialyzed solution in a beaker or other container, place a spherical ultrasound source below the liquid surface (preferably half the sphere is submerged in the liquid surface or the entire sphere is just submerged in the liquid surface), and vibrate ultrasonically for 5 to 60 minutes (preferably 10 to 15 minutes) at a vibration frequency of 20k to 100kHz (preferably 30k to 40kHz).

[0049] Multiple ultrasound sources are added to the solution in an orderly manner for synchronous ultrasound stimulation; or magnetic materials are added to the solution for magnetic vibration; and then the solution is quickly frozen by liquid nitrogen or the like. The resulting silk fibroin raw material can form a snowflake-like silk fibroin structure, which can be injected into the subcutaneous layer of the skin on a large scale in a solution state. After the water evaporates, a dense silk fibroin network structure can be formed, thereby forming a new skin barrier, which can protect the skin from external stimuli such as ultraviolet rays, bacteria, etc. for a long time, and can reduce the loss of water in the deep skin.

[0050] Place the dialyzed solution in a beaker or other container, arrange multiple ultrasound sources in a regular polygonal position inside the solution or outside the container (preferably inside the solution to prevent damage to the container due to resonance), and perform ultrasonic vibration for 5 to 60 minutes (preferably 10 to 15 minutes) at a vibration frequency of 20k to 100kHz (preferably 30k to 40kHz).

[0051] The disordered silk fibroin prepared by shaking on a shaker can be used to prepare a silk fibroin gel with excellent viscoelasticity and easy injection, which is suitable for preparing an injection filling material for pre-filled packaging.

[0052] Oscillation frequency: 100-1000 rpm / min (preferably 100-300 rpm / min), oscillation time: 1-120 min (preferably 30-60 min).

[0053] Example 1.

[0054] A method for extracting silk fibroin material of this embodiment comprises the following steps:

[0055] The silk cocoons / raw silk are degummed, dissolved, dialyzed, and directionally stimulated, and then pre-cooled and freeze-dried to obtain silk fibroin material.

[0056] The ultra-low temperature precooling method is: precooling under liquid nitrogen at -80°C for 30 minutes;

[0057] Silk cocoons / raw silk are made from mulberry silk produced by feeding and feeding in a modern sericulture environment using GMP technology, ensuring that the cocoons have better consistency and traceability.

[0058] Directed stimulation is:

[0059] Shear: Short-term continuous shear force in the same local direction can induce silk fibroin to produce transitional state sites distributed along the tangential direction;

[0060] Ultrasound: Different transition state sites are induced according to the ultrasound frequency and the location of the ultrasound source;

[0061] Shaking on a shaker: Shaking on a shaker simulates daily shaking, providing a way to stimulate the chaotic state and induce the generation of disordered transition state sites.

[0062] The silk fibroin with planar ordered transition state sites produced by low-frequency magnetic stirring on a plate forms a film-forming silk fibroin solution / gel for skin barrier repair / care;

[0063] The dialyzed solution was placed in a petri dish with a thickness of less than 30 mm. The solution was stirred magnetically at a speed of 50 rpm for 5 min.

[0064] The application method of the silk fibroin material of this embodiment also includes the following:

[0065] A rod-shaped ultrasound source is placed in a solution to perform ultrasonic vibration stimulation, thereby forming a planar orderly transitional site of silk fibroin with tentacles. During wound repair, the tentacles can pass through the injured part and embed into the injured part, and the planar body can form a silk fibroin membrane.

[0066] The dialyzed solution is placed in a beaker or other container, and a rod-shaped ultrasonic source is placed vertically in the solution. Ultrasonic vibration is performed for 5 minutes at a vibration frequency of 20 kHz.

[0067] The application method of the silk fibroin material of this embodiment also includes the following:

[0068] The disordered silk fibroin prepared by shaking on a shaker can produce a silk fibroin gel with excellent viscoelasticity and easy injection, which is suitable for preparing an injection filling material for pre-filled packaging;

[0069] Oscillation frequency: 100 pm / min, oscillation time: 20 min.

[0070] In this embodiment, a spherical ultrasound source is placed in a solution to perform ultrasonic vibration stimulation, thereby forming a tetrahedral-shaped silk fibroin. This silk fibroin material can be used to obtain a silk fibroin gel with excellent viscoelasticity and excellent mechanical support properties, and is suitable for various injection filling materials.

[0071] Place the dialyzed solution in a beaker or other container, place a spherical ultrasound source under the liquid surface, and perform ultrasonic vibration for 5 minutes at a vibration frequency of 20 kHz.

[0072] The silk fibroin material of this embodiment is obtained by a method for extracting a silk fibroin material.

[0073] Example 2.

[0074] A method for extracting silk fibroin material of this embodiment comprises the following steps:

[0075] The silk cocoons / raw silk are degummed, dissolved, dialyzed, and directionally stimulated, and then pre-cooled and freeze-dried to obtain silk fibroin material.

[0076] The ultra-low temperature precooling method is: precooling under liquid nitrogen at -80°C for 30 minutes;

[0077] Silk cocoons / raw silk are made from mulberry silk produced by feeding and feeding in a modern sericulture environment using GMP technology, ensuring that the cocoons have better consistency and traceability.

[0078] Directed stimulation is:

[0079] Shear: Short-term continuous shear force in the same local direction can induce silk fibroin to produce transitional state sites distributed along the tangential direction;

[0080] Ultrasound: Different transition state sites are induced according to the ultrasound frequency and the location of the ultrasound source;

[0081] Shaking on a shaker: Shaking on a shaker simulates daily shaking, providing a way to stimulate the chaotic state and induce the generation of disordered transition state sites.

[0082] The silk fibroin with planar ordered transition state sites produced by low-frequency magnetic stirring on a plate forms a film-forming silk fibroin solution / gel for skin barrier repair / care;

[0083] The dialyzed solution was placed in a petri dish with a thickness of less than 30 mm. The solution was stirred magnetically at a speed of 500 rpm for 60 min.

[0084] The application method of the silk fibroin material of this embodiment also includes the following:

[0085] A rod-shaped ultrasound source is placed in a solution to perform ultrasonic vibration stimulation, thereby forming a planar orderly transitional site of silk fibroin with tentacles. During wound repair, the tentacles can pass through the injured part and embed into the injured part, and the planar body can form a silk fibroin membrane.

[0086] The dialyzed solution is placed in a beaker or other container, and a rod-shaped ultrasonic source is placed vertically in the solution. Ultrasonic vibration is performed for 60 minutes at a vibration frequency of 100 kHz.

[0087] The application method of the silk fibroin material of this embodiment also includes the following:

[0088] The disordered silk fibroin prepared by shaking on a shaker can produce a silk fibroin gel with excellent viscoelasticity and easy injection, which is suitable for preparing an injection filling material for pre-filled packaging;

[0089] Oscillation frequency: 1000rpm / min, oscillation time: 120min.

[0090] In this embodiment, a spherical ultrasound source is placed in a solution to perform ultrasonic vibration stimulation, thereby forming a tetrahedral-shaped silk fibroin. This silk fibroin material can be used to obtain a silk fibroin gel with excellent viscoelasticity and excellent mechanical support properties, and is suitable for various injection filling materials.

[0091] The dialyzed solution is placed in a beaker or other container, a spherical ultrasonic source is placed under the liquid surface, and ultrasonic vibration is applied for 60 minutes at a vibration frequency of 100 kHz.

[0092] The silk fibroin material of this embodiment is obtained by a method for extracting a silk fibroin material.

[0093] Example 3.

[0094] A method for extracting silk fibroin material of this embodiment comprises the following steps:

[0095] The silk cocoons / raw silk are degummed, dissolved, dialyzed, and directionally stimulated, and then pre-cooled and freeze-dried to obtain silk fibroin material.

[0096] The ultra-low temperature precooling method is: precooling under liquid nitrogen at -80°C for 25 minutes;

[0097] Silk cocoons / raw silk are made from mulberry silk produced by feeding and feeding in a modern sericulture environment using GMP technology, ensuring that the cocoons have better consistency and traceability.

[0098] Directed stimulation is:

[0099] Shear: Short-term continuous shear force in the same local direction can induce silk fibroin to produce transitional state sites distributed along the tangential direction;

[0100] Ultrasound: Different transition state sites are induced according to the ultrasound frequency and the location of the ultrasound source;

[0101] Shaking on a shaker: Shaking on a shaker simulates daily shaking, providing a way to stimulate the chaotic state and induce the generation of disordered transition state sites.

[0102] The silk fibroin with planar ordered transition state sites produced by low-frequency magnetic stirring on a plate forms a film-forming silk fibroin solution / gel for skin barrier repair / care;

[0103] The dialyzed solution was placed in a flat dish with a thickness of less than 30 mm. The solution was stirred magnetically at a speed of 100 rpm for 25 min.

[0104] The application method of the silk fibroin material of this embodiment also includes the following:

[0105] A rod-shaped ultrasound source is placed in a solution to perform ultrasonic vibration stimulation, thereby forming a planar orderly transitional site of silk fibroin with tentacles. During wound repair, the tentacles can pass through the injured part and embed into the injured part, and the planar body can form a silk fibroin membrane.

[0106] The dialyzed solution was placed in a beaker or other container, and a rod-shaped ultrasonic source was placed vertically in the solution. Ultrasonic vibration was performed for 55 minutes at a vibration frequency of 60 kHz.

[0107] The application method of the silk fibroin material of this embodiment also includes the following:

[0108] The disordered silk fibroin prepared by shaking on a shaker can produce a silk fibroin gel with excellent viscoelasticity and easy injection, which is suitable for preparing an injection filling material for pre-filled packaging;

[0109] Oscillation frequency: 500 rpm / min, oscillation time: 20 min.

[0110] In this embodiment, a spherical ultrasound source is placed in a solution to perform ultrasonic vibration stimulation, thereby forming a tetrahedral-shaped silk fibroin. This silk fibroin material can be used to obtain a silk fibroin gel with excellent viscoelasticity and excellent mechanical support properties, and is suitable for various injection filling materials.

[0111] Place the dialyzed solution in a beaker or other container, place a spherical ultrasonic source under the liquid surface, and perform ultrasonic vibration for 20 minutes at a vibration frequency of 30 kHz.

[0112] The silk fibroin material of this embodiment is obtained by a method for extracting a silk fibroin material.

[0113] Example 4.

[0114] A method for extracting silk fibroin material of this embodiment comprises the following steps:

[0115] The silk cocoons / raw silk are degummed, dissolved, dialyzed, and directionally stimulated, and then pre-cooled and freeze-dried to obtain silk fibroin material.

[0116] The ultra-low temperature precooling method is: precooling under liquid nitrogen cooling at -80°C for 22 minutes;

[0117] Silk cocoons / raw silk are made from mulberry silk produced by feeding and feeding in a modern sericulture environment using GMP technology, ensuring that the cocoons have better consistency and traceability.

[0118] Directed stimulation is:

[0119] Shear: Short-term continuous shear force in the same local direction can induce silk fibroin to produce transitional state sites distributed along the tangential direction;

[0120] Ultrasound: Different transition state sites are induced according to the ultrasound frequency and the location of the ultrasound source;

[0121] Shaking on a shaker: Shaking on a shaker simulates daily shaking, providing a way to stimulate the chaotic state and induce the generation of disordered transition state sites.

[0122] The silk fibroin with planar ordered transition state sites produced by low-frequency magnetic stirring on a plate forms a film-forming silk fibroin solution / gel for skin barrier repair / care;

[0123] The dialyzed solution was placed in a petri dish with a thickness of less than 30 mm. The solution was stirred magnetically at a speed of 20 rpm for 55 min.

[0124] The application method of the silk fibroin material of this embodiment also includes the following:

[0125] A rod-shaped ultrasound source is placed in a solution to perform ultrasonic vibration stimulation, thereby forming a planar orderly transitional site of silk fibroin with tentacles. During wound repair, the tentacles can pass through the injured part and embed into the injured part, and the planar body can form a silk fibroin membrane.

[0126] The dialyzed solution is placed in a beaker or other container, and a rod-shaped ultrasonic source is placed vertically in the solution. Ultrasonic vibration is performed for 10 minutes at a vibration frequency of 30 kHz.

[0127] The application method of the silk fibroin material of this embodiment also includes the following:

[0128] The disordered silk fibroin prepared by shaking on a shaker can produce a silk fibroin gel with excellent viscoelasticity and easy injection, which is suitable for preparing an injection filling material for pre-filled packaging;

[0129] Oscillation frequency: 200 rpm / min, oscillation time: 10 min.

[0130] In this embodiment, a spherical ultrasound source is placed in a solution to perform ultrasonic vibration stimulation, thereby forming a tetrahedral-shaped silk fibroin. This silk fibroin material can be used to obtain a silk fibroin gel with excellent viscoelasticity and excellent mechanical support properties, and is suitable for various injection filling materials.

[0131] Place the dialyzed solution in a beaker or other container, place a spherical ultrasonic source under the liquid surface, and perform ultrasonic vibration for 10 minutes at a vibration frequency of 30 kHz.

[0132] Example 5.

[0133] A method for extracting silk fibroin material of this embodiment comprises the following steps:

[0134] The silk cocoons / raw silk are degummed, dissolved, dialyzed, and directionally stimulated, and then pre-cooled and freeze-dried to obtain silk fibroin material.

[0135] The ultra-low temperature precooling method is: precooling under liquid nitrogen at -80°C for 25 minutes;

[0136] Silk cocoons / raw silk are made from mulberry silk produced by feeding and feeding in a modern sericulture environment using GMP technology, ensuring that the cocoons have better consistency and traceability.

[0137] Directed stimulation is:

[0138] Shear: Short-term continuous shear force in the same local direction can induce silk fibroin to produce transitional state sites distributed along the tangential direction;

[0139] Ultrasound: Different transition state sites are induced according to the ultrasound frequency and the location of the ultrasound source;

[0140] Shaking on a shaker: Shaking on a shaker simulates daily shaking, providing a way to stimulate the chaotic state and induce the generation of disordered transition state sites.

[0141] The silk fibroin with planar ordered transition state sites produced by low-frequency magnetic stirring on a plate forms a film-forming silk fibroin solution / gel for skin barrier repair / care;

[0142] The dialyzed solution was placed in a petri dish with a thickness of less than 30 mm. The solution was stirred magnetically at a speed of 200 rpm for 50 min.

[0143] The application method of the silk fibroin material of this embodiment also includes the following:

[0144] A rod-shaped ultrasound source is placed in a solution to perform ultrasonic vibration stimulation, thereby forming a planar orderly transitional site of silk fibroin with tentacles. During wound repair, the tentacles can pass through the injured part and embed into the injured part, and the planar body can form a silk fibroin membrane.

[0145] The dialyzed solution is placed in a beaker or other container, and a rod-shaped ultrasonic source is placed vertically in the solution. Ultrasonic vibration is performed for 50 minutes at a vibration frequency of 70 kHz.

[0146] The application method of the silk fibroin material of this embodiment also includes the following:

[0147] The disordered silk fibroin prepared by shaking on a shaker can produce a silk fibroin gel with excellent viscoelasticity and easy injection, which is suitable for preparing an injection filling material for pre-filled packaging;

[0148] Oscillation frequency: 300rpm / min, oscillation time: 70min.

[0149] In this embodiment, a spherical ultrasound source is placed in a solution to perform ultrasonic vibration stimulation, thereby forming a tetrahedral-shaped silk fibroin. This silk fibroin material can be used to obtain a silk fibroin gel with excellent viscoelasticity and excellent mechanical support properties, and is suitable for various injection filling materials.

[0150] Place the dialyzed solution in a beaker or other container, place a spherical ultrasound source under the liquid surface, and perform ultrasonic vibration for 20 minutes at a vibration frequency of 80 kHz.

[0151] The silk fibroin material of this embodiment is obtained by a method for extracting a silk fibroin material.

[0152] The products of Examples 1-5 were tested for performance with reference to the existing CN109276764A patent, and the compression modulus of the products was tested. The test results are as follows:

[0153] Compression modulus (MPa) Example 1 11.5 Example 2 11.7 Example 3 11.9 Example 4 11.6 Example 5 11.7

[0154] The products of Examples 1-5 have excellent compression modulus. During the extraction process of silk fibroin, directional and short-term physical stimulation is added, so that the secondary structure of the purified silk fibroin has an orderly transition state site from α type to β type, so that the silk fibroin retains its soluble properties and can easily induce β folding from the transition state site during subsequent processing, thereby making it easier to produce ordered crystals. The silk fibroin obtained by the method of the present invention can effectively support the mechanical properties and better serve as a filling material.

[0155] It will be apparent to those skilled in the art that the present invention is not limited to the details of the exemplary embodiments described above and that the present invention can be implemented in other specific forms without departing from the spirit or essential features of the present invention. Therefore, the embodiments should be considered exemplary and non-restrictive in all respects, and the scope of the present invention is defined by the appended claims rather than the above description, and it is intended that all changes falling within the meaning and scope of the equivalent elements of the claims be included in the present invention.

[0156] In addition, it should be understood that although the present specification is described according to implementation modes, not every implementation mode contains only one independent technical solution. This description of the specification is only for the sake of clarity. Those skilled in the art should regard the specification as a whole. The technical solutions in each embodiment may also be appropriately combined to form other implementation modes that can be understood by those skilled in the art.

Claims

1. A method for extracting silk fibroin material, characterized in that: The following steps are involved: The silk cocoons / raw silk are degummed, dissolved, dialyzed, and directionally stimulated, and then pre-cooled and freeze-dried to obtain silk fibroin material.

2. The method for extracting silk fibroin material according to claim 1, characterized in that: The ultra-low temperature precooling method is: precooling under liquid nitrogen at -80°C for 20-30 minutes.

3. The method for extracting silk fibroin material according to claim 2, characterized in that: Directed stimulation is: S1: Shear treatment: 5-10 min of high-speed shear treatment is used to induce silk fibroin to produce transition state sites distributed along the tangential direction; S2: Ultrasonic treatment: 500W ultrasonic frequency was used for 10 min to induce different transition state sites in silk fibroin; S3: Shaking on a shaking table: To simulate daily shaking, the shaking program of the shaking table is random shaking, which induces the generation of disordered transition state sites.

4. The method for extracting silk fibroin material according to claim 3, characterized in that: The specific operation steps of the degumming are: The cocoons / raw silk are placed in sufficient water for 5 hours at a temperature of 90°C, then cooled to 50°C, and alkaline protease is added at 10% of the total amount of the cocoons / raw silk until the glue content of the cocoons / raw silk is less than 3%, and the degumming technology is completed; The specific operation method of the dissolution treatment is: dissolve the degummed silk in a 9-12 mol / L lithium bromide solution for 24 hours.

5. The method for extracting silk fibroin material according to claim 1, characterized in that: The dialyzed solution is stirred on a plate by low-frequency magnetic force to produce silk fibroin with planar ordered transition state sites, forming a film-forming silk fibroin solution / gel for skin barrier repair / care; The solution thickness is less than 30 mm, magnetic stirring is used, the stirring speed is 50 to 500 rpm, and the stirring time is 5 to 60 minutes.

6. The method for extracting silk fibroin material according to claim 1, characterized in that: A rod-shaped ultrasound source is placed in the dialyzed solution to perform ultrasonic vibration stimulation, thereby forming a planar orderly transitional site of silk fibroin with tentacles. During wound repair, the damaged part of the tentacles is embedded in the damaged part, and the planar main body forms a silk fibroin membrane. Ultrasonic vibration for 5 to 60 minutes, vibration frequency 20k to 100kHz.

7. The silk fibroin material obtained by the extraction method of the silk fibroin material according to any one of claims 1 to 6.