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Sericin, its extraction method, as a cell antiviral immune enhancer and its application

A technology for sericin and cell culture, which is applied in the extraction of natural protein components and in the field of virology, which can solve the problems of reducing solubility, changing the natural conformation of sericin, destroying structural integrity, etc., so as to improve the yield and protect natural organisms. Chemical activity, reducing the effect of irreversible precipitation

Active Publication Date: 2022-04-22
JINAN UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The disadvantages are: 1. The proposed sericin has many impurities, which may be mixed with a large amount of silk fibroin and other components of silkworm cocoons; 2. Sericin is very unstable in heated aqueous solution and is easy to degrade, and most of the obtained sericin is Degradation products; 3. In addition, it takes a long time, because sericin is time-consuming to dissolve in hot water
[0010] Through the above analysis, the problems and defects in the prior art are: (1) the existing sericin extraction method, the obtained sericin product has the advantages of biological safety, structural integrity, biological activity and higher product yield. There are problems, and it is impossible to meet these conditions at the same time
[0011] (2) The existing technology does not provide effective technical support for the application of active sericin in anti-virus
[0012] (3) Existing immunopotentiators are mainly to enhance the body's innate immunity and acquired immunity process, and they have a long response time to virus infection, and they are not by enhancing the cell's recognition of the virus and promoting the antiviral response of the infected cell. less specific
[0014] (2) Existing extraction methods will change the natural conformation of sericin or destroy its structural integrity, which will seriously damage the biological activity of sericin products
Practice has shown that after the concentration of urea or LiBr is gradually reduced, high-concentration sericin will undergo irreversible precipitation, indicating that misfolding of the polypeptide chain of sericin leads to a decrease in solubility

Method used

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  • Sericin, its extraction method, as a cell antiviral immune enhancer and its application
  • Sericin, its extraction method, as a cell antiviral immune enhancer and its application
  • Sericin, its extraction method, as a cell antiviral immune enhancer and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0134] Such as figure 1 As shown, the sericin protein extraction (LiBr method) with complete structure and high biological safety can be used for antiviral applications

[0135] (1) Select 1 g of silk cocoons that have been broken to remove silkworm chrysalis within 24 hours after cocooning, or silkworm cocoons that meet this condition and are stored at 4°C.

[0136] (2) Silkworm cocoon pretreatment and sterilization. In a biosafety cabinet (Thermo), shred cocoons to less than 2x2 mm using sterilized forceps and scissors 2 The fragments were dispersed in a single layer on a sterile glass plate, and placed in ultraviolet light with a light intensity of 1400 Lux for 30 minutes. After turning over the silkworm cocoon fragments, the ultraviolet rays were irradiated for another 30 minutes under the same conditions.

[0137] (3) In a biosafety level two laboratory, put each gram of silkworm cocoon fragments into 55 mL of sterile lithium bromide (LiBr, Sinopharm) aqueous solution...

Embodiment 2

[0143] Structurally complete and high biosafety can be used for sericin extraction (urea method) for antiviral applications.

[0144] (1) Select 1 g of silk cocoons that have been broken to remove silkworm chrysalis within 24 hours after cocooning, or silkworm cocoons that meet this condition and are stored at 4°C.

[0145] (2) Silkworm cocoon pretreatment and sterilization. In a biological safety cabinet (Thermo), use sterilized tweezers and scissors to cut silkworm cocoons into fragments smaller than 2x2 mm2, spread them on a sterile glass plate in a single layer, and place them in ultraviolet light with a light intensity of 1400Lux Irradiate for 30 minutes. After turning over the silkworm cocoon fragments, the ultraviolet rays were irradiated for another 30 minutes under the same conditions.

[0146] (3) In a biosafety level two laboratory, put 5 grams of silkworm cocoon fragments into 100 mL of sterile urea (Sinopharm) aqueous solution with a concentration of 480 g / L (8M...

Embodiment 3

[0152] Quality control of sericin extraction process and quality detection of product sericin solution.

[0153] (1) LiBr and urea-dissolved sericin solution, dialysis / renaturation product, concentrated product and the final sericin solution obtained during the extraction process, absorb 10uL in each step and store at -20°C. After the extraction was completed, a commercial BCA protein concentration detection kit (Milipore) was purchased, and the protein concentration of the collected solution was determined according to the operation instructions of the kit.

[0154] The results of the concentration determination are attached figure 2 As shown, through the concentration / volume control, the irreversible precipitation of sericin protein in the extraction process is avoided, which not only ensures the yield, but also ensures that the concentration of the obtained product sericin solution meets the application requirements for preventing and controlling viruses (see attached fi...

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Abstract

The invention belongs to the technical fields of extraction of natural protein components and virology, and discloses a sericin protein, an extraction method, and its application as a cell antiviral immune enhancer. Uses; the dose concentration of sericin is 10-20mg / mL, and the effective final concentration is ≥1mg / mL; the extraction method of sericin includes: selecting available silkworm cocoons and storage conditions, silkworm cocoon pretreatment and sterilization, sericin dissolution, Dialysis and renaturation of sericin protein, concentration of sericin solution, removal of endotoxin in sericin solution, filtration of sericin solution, quality control inspection of sericin solution, sericin solution that meets the testing standards as a cell antiviral immune enhancer. The present invention develops and utilizes silk protein sericin for the first time, as the application of preventing and controlling virus infection. The raw material source of the invention is wide, and compared with the high synthesis and production costs of other antiviral drugs, the cost of the invention is low.

Description

technical field [0001] The invention belongs to the technical fields of extraction of natural protein components and virology, and in particular relates to a sericin protein, an extraction method, and its use as a cell antiviral immune enhancer. Background technique [0002] Currently, sericin is a natural protein present in silkworm cocoons and is the active ingredient wrapped in the outer layer of silk. In the textile industry, silkworm cocoons are spun to extract silk fibroin, which is used to make textiles, and sericin was previously usually discarded as industrial waste. In recent years, with the development of bioengineering and materials science, researchers have gradually discovered the use value, especially the discovery that it has a variety of biological activities, which makes this substance a research hotspot and turns waste into treasure become possible. Many scientific studies have shown that sericin has low immunogenicity, promotes cell adhesion and value-a...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/435C07K1/34C07K1/14A61K38/17A61P31/12A61P31/14A61P37/04A61K8/64A61Q19/00A23L33/17
CPCC07K14/43586A61K38/1767A61P37/04A61P31/12A61P31/14A61K8/64A61Q19/00A23L33/17A23V2002/00A23V2200/30A23V2200/324
Inventor 吴建国李永奎黄颖邢曦雯罗震潘攀张业顺
Owner JINAN UNIVERSITY
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