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Biological fluid film of recombined human keratinized cell growth factor-2 and its preparing process

A biological fluid, keratinocyte technology, applied in recombinant DNA technology, drug combination, pharmaceutical formulations and other directions, can solve the problems of restricting the wide use of solid dressing films, complex process, inconvenience, etc., and achieves improved industrialization value and simple extraction process. , the effect of low cost

Inactive Publication Date: 2006-11-15
ZHEJIANG ACAD OF MEDICAL SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] However, although recombinant human keratinocyte growth factor-2 is a protein factor with great therapeutic potential for wound healing, how to apply it clinically and how to make it into a certain dosage form is an urgent problem in the current medical field. The problem
[0005] The United States once made KGF-2 into a kind of injection and powder injection, but it is only a kind of drug used to treat epithelial tissue damage. When extracting this drug, the method of inclusion body is used to extract KGF-2. Its disadvantages It is low in purity and complex in process
And in dressing, although in the prior art there is the biological dressing film that is prepared as matrix with chitin derivative (chitosan) and is used on the treatment of various wounds, but the chitin biological dressing film that goes on the market is a kind of solid However, when this solid film is applied, it cannot closely contact all aspects of the wound surface, and it is easy to produce fluid accumulation and cause infection; at the same time, the size of the solid film needs to be increased or decreased according to the wound surface, which is extremely inconvenient, thus limiting the wide use of solid dressing films

Method used

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  • Biological fluid film of recombined human keratinized cell growth factor-2 and its preparing process

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Experimental program
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Effect test

Embodiment 1

[0050] Embodiment 1: Preparation of recombinant human keratinocyte growth factor-2 biological fluid membrane matrix:

[0051] Take by weighing 492 grams of chitosan dried at 80°C (final concentration is 1.6%) and put in a stirrer, add 25 kilograms of purified water and stir evenly, the stirring speed is 196r / min, slowly add 250ml acetic acid solution while stirring, the pH value For 3, the process takes about 12 hours. After the completion, add 150g of gelatin dissolved in 1500ml of water and 150ml of glycerin while it is hot, mix well, and then slowly adjust the pH value of the chitosan solution to 6.5 with 3700ml of 0.7% sodium bicarbonate solution. , the process takes 2-3 hours, then add 24.5 grams of polyvinylpyrrolidone, fully stir, filter, heat, remove air bubbles, sterilize, measure the pH value and viscosity of the chitosan solution the next day, and then potting and packaging in medical aluminum tubes. The viscosity measured with a rotary viscometer is 3570mpa.s. The...

Embodiment 2

[0062] Embodiment 2: Preparation of recombinant human keratinocyte growth factor-2 stock solution:

[0063] The expression plasmid is pLY4, and the host bacteria is Escherichia coli BL 21 (D3). According to the cDNA sequence of KGF-2 in the gene library, synthesize 20 complementary oligonucleotides, and introduce specific enzyme cutting sites EcoR I and BamH I at the 5' end and 3' end respectively, and adopt the conventional method of molecular cloning , first treated with T4 phage polynucleotide kinase at 37°C for 30 minutes. Phosphorylated oligonucleotide fragments were mixed in equimolar numbers, denatured at 94°C for 5 minutes, and immediately annealed at 65°C for 10 minutes, then added T4 ligase and reacted overnight at 16°C.

[0064] The pLY4 plasmid was digested with EcoR I and BamH I, and the large fragment was recovered. It was ligated with the synthetic rhKGF-2 cDNA fragment overnight, and directly transformed into E. coli host strain BL 21 (D3), and then screened f...

Embodiment 3

[0065] Embodiment 3: the preparation of rhKGF-2 biological fluid film

[0066] A new formulation of rhKGF-2-fluid film is prepared by matching the fluid dressing film with an appropriate concentration of recombinant human keratinocyte growth factor (rhKGF-2), and its formula is designed as follows:

[0067] Take 1 part of sterile 0.4% recombinant human keratinocyte growth factor-2 (rhKGF-2) aqueous solution, slowly add it to 19 parts of rhKGF-2 biological fluid membrane matrix, fully stir, filter, defoam, and fill in 80 ℃ baking 4h in the medical aluminum tube. The final concentration of rhKGF-2 in the fluid film was 200μg / ml.

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Abstract

The present invention is recombinant human keratinized cell growth factor-2 (rhKGF-2) biofluid membrane for treating epithelium tissue damage and its preparation process. The rhKGF-2 biofluid membrane includes purified rhKGF-2 liquid and biofluid dressing comprising chitosan, gelatin, glycerin and PVP. The purified rhKGF-2 liquid is prepared through connecting KGF-2 gene onto the expression plasmid pLY4 to constitute expression vector transforming colibacillus and clone engineering bacillus; culturing engineering bacillus and raising temperature to induce expressing rhKGF-2; and separating and purifying the expressed rhKGF-2. The biofluid dressing is prepared is prepared with chitosan as main matrix, and gelatin, glycerin and PVP as supplementary material in optimized compounding proportion.

Description

technical field [0001] The invention relates to a recombinant human keratinocyte growth factor-2 biological fluid film for treating epithelial tissue damage, in particular to a recombinant human keratinocyte growth factor-2 biological fluid film and a preparation process thereof. Background technique [0002] Epithelial cell damage manifests as skin burns, traumatic wounds, and skin ulcers of various causes, involving a wide range of areas. At present, the clinical treatment of burns and traumatic wounds basically includes three aspects: wound cleaning, dressing application and drug treatment. Among them, dressing application and drug treatment are often carried out at the same time to help the wound heal while solving the problem of covering the wound. [0003] In the prior art, the cytokine drugs for treating epithelial tissue damage mainly include human epidermal growth factor (EGF). Since EGF is a non-specific keratinocyte-promoting factor, it may leave obvious scars wh...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K38/18A61K9/70A61P17/02C12N15/09C12N15/63
Inventor 王虎根蒋玉燕岑坚沈进毕忆群蒋建国
Owner ZHEJIANG ACAD OF MEDICAL SCI
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