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Method for preparing regenerative silk fibroin and product and application thereof

A technology for regenerating silk fibroin and degumming silk, which is applied in the field of medicine

Inactive Publication Date: 2012-11-14
ARMY MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Natural silk fibroin is insoluble in water, but soluble in strong acid and strong alkali and some neutral salts (such as lithium, strontium, barium chloride, bromide, iodide, and thiocyanate and zinc chloride) solutions. It is difficult to preserve the biological activity of silk fibroin in strong acid and alkali. Neutral salts such as lithium strontium barium chloride, bromide, iodide, and thiocyanate are harmful to the human body, so they are not suitable for use. silk fibroin

Method used

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  • Method for preparing regenerative silk fibroin and product and application thereof
  • Method for preparing regenerative silk fibroin and product and application thereof
  • Method for preparing regenerative silk fibroin and product and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1 Preparation of regenerated silk fibroin

[0037] Take the silk, add a sodium dodecyl sulfate (SDS) solution with a mass fraction of 0.1%-10%, boil for 1 hour, then rinse with deionized water, and then add a mass fraction of 0.1%-10% Na 2 CO 3 solution, boiled for 1 hour, rinsed with deionized water, and then dried overnight at 65°C to obtain degummed silk. Dissolve sericin in silk by degumming, the mass fraction of SDS solution in this step is 0.25% and Na 2 CO 3 The mass fraction of the solution is 0.25% degumming effect is the best, SDS solution and Na 2 CO 3 The amount of solution added is equivalent to 100-1000 times of silk weight, and the effect is the best. After degumming, the degummed silk is observed with a scanning electron microscope, and the results are as follows: figure 1 shown.

[0038] Preparation of Calcium Alcohol Solution: Calcium Alcohol Solution Contains CaCl 2 -Ethanol-Water, CaCl 2 -methanol-water, Ca(NO 3 ) 2 4H 2 O-meth...

Embodiment 2

[0045] Example 2 Scanning electron microscope observation of regenerated silk fibroin

[0046] In order to further observe the dissolution of regenerated silk fibroin, the regenerated silk fibroin protein prepared from degummed silk and four kinds of solutions was observed by scanning electron microscope. figure 1 As shown, the regenerated silk fibroin obtained from the four solutions was observed by scanning electron microscopy as follows: figure 2 shown. Depend on figure 1 It can be seen that the degummed silk structure remains intact, but from figure 2 In A, it can be seen that Ca(NO 3 ) 2 4H 2 After the O-methanol treatment, the degummed silk was dissolved, and the silk fibroin had a small amount of silk fiber residue, and the dissolved silk fibroin was in a relatively uniform granular shape; figure 2 In B and C, it can be seen that Ca(NO 3 ) 2 4H 2 O-ethanol and CaCl 2 -Methanol-H 2 O-treated degummed silk was dissolved, but with Ca(NO 3 ) 2 4H 2 O...

Embodiment 3

[0047] Example 3 Regenerated silk fibroin SDS-PAGE electrophoresis

[0048] Silk is rich in alanine, glycine and serine amino acids, and is connected by a heavy chain of about 300 kDa and a light chain of 26 kDa through a disulfide bond. The P25 glycoprotein with a molecular weight of 30 kDa is complexed with the light chain and the heavy chain through hydrophobic interactions. The combination of these substances constitutes silk molecules with a molecular weight of about 400 kDa. The intermolecular structure of silk protein will be destroyed under heat, chemical reagents and pH conditions. After the calcium alcohol solution of the present invention is processed, the structure of degummed silk will change. In order to measure the molecular weight of the regenerated silk fibroin protein after different treatments, the mass fraction of 12% separating gel and 4% concentrated gel mass fraction are selected, and the sample volume is 20 μL Carry out molecular weight detection...

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Abstract

The invention discloses a method for preparing regenerative silk fibroin. The method includes adding a calcium alcohol solution into degumming silks, dissolving the degumming silks through water bath, centrifugally removing suspended impurities, and performing dialysis and drying to obtain the regenerative silk fibroin. The preparation method is simple, the dissolvability of the silk fibroin is good, the structure of the prepared regenerative silk fibroin is a beta-folding sheet layer, the irregular curl structure is decreased, the regenerative silk fibroin can be used as a carrier for immobilizing enzyme, and the enzyme can be immobilized on the regenerative silk fibroin, so that the enzyme can be prevented from being degraded by degeneration enzymes in an organism, and further the stability and the half-life period of the enzyme are improved.

Description

technical field [0001] The invention relates to the field of medicine, in particular to a method for preparing regenerated silk fibroin, and also to the regenerated silk fibroin prepared by the preparation method and its application. Background technique [0002] Silk is a natural macromolecular polymer, and natural mulberry silk is mainly composed of silk protein and sericin. Mulberry silk has a core-shell structure, the outer layer is coated with sericin (Sericin), and the inner layer is silk fibroin (SF for short), of which the silk fibroin content is about 70% to 80%. Silk fibroin is mainly composed of three protein components, namely 350kd H chain protein, 25kd L chain protein and 30kd P25 protein, among which the molar ratio of H chain protein:L chain protein:P25 protein is 6:6:l . The H chain protein is a hydrophobic protein, which mainly provides a crystal-like structure for silk fibers and determines the strength of the silk thread; the L chain protein is a hydrop...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K1/14C07K14/435C12N11/02
Inventor 杨霞吴玉章
Owner ARMY MEDICAL UNIV
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