Fish skin-derived medical collagen membrane with compact outer layer and loose inner layer and preparation method of medical collagen membrane

A collagen and fish skin technology, applied in the field of medical materials, can solve the problems of cytotoxicity, inability to effectively block growth, and inhibit cell adhesion and proliferation in cross-linked collagen membranes.

Pending Publication Date: 2021-09-10
OCEAN UNIV OF CHINA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] 1) The in vivo degradation rate is too fast to maintain structural integrity during tissue regeneration;
[0006] 2) The partially cross-linked collagen membrane has cytotoxicity and inhibits cell adhesion and proliferation;
[0007] 3) There is a large difference in mechanical properties between collagen membranes;
[0008] 4) May transmit animal diseases to humans;
[0009] 5) It cannot effectively block the fibrous connective tissue cells and epithelial cells of the surrounding soft tissue and guide the growth of osteoblasts

Method used

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  • Fish skin-derived medical collagen membrane with compact outer layer and loose inner layer and preparation method of medical collagen membrane
  • Fish skin-derived medical collagen membrane with compact outer layer and loose inner layer and preparation method of medical collagen membrane
  • Fish skin-derived medical collagen membrane with compact outer layer and loose inner layer and preparation method of medical collagen membrane

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Example 1: Preparation of non-denatured collagen derived from fish skin

[0045]Weigh 100g of fish skin, add 20 times the volume of 0.1mol / L NaOH solution and magnetically stir for 24 hours to remove non-collagen components and impurities, and eliminate the influence of endogenous proteases on the collagen structure. During this period, 0.1mol / L NaOH solution Replace 2 times. After the fish skin is swollen with alkali, the fish scales are removed with a knife and cut into small square pieces of about 5×5 mm. Wash the fish skin repeatedly with pre-cooled deionized water until neutral, then add 0.5mol / L acetic acid solution and 0.5% (w:w) pepsin to carry out enzymatic extraction at a material-liquid ratio of 1:60, and stir with medium speed magnetic force for 48h . After the enzymolysis extract was refrigerated and centrifuged at 9000r / min for 30min, the supernatant was collected and salted out with finely ground NaCl. The final actual concentration of NaCl was controll...

Embodiment 2

[0048] Example 2: Preparation of a medical graded collagen membrane with a dense outer layer and a loose inner layer from fish skin

[0049] The non-denatured collagen derived from fish skin obtained according to Example 1 was dissolved in 0.05M acetic acid to prepare 2.5%, 1.5%, and 0.5% collagen suspensions, stood at 4°C for 48 hours, and then stirred until homogenized. After the dissolution is complete, put the 2.5% collagen solution into a stainless steel dish, spread a thin layer, and stand at a low temperature to obtain a glue block. Then, according to the gradient, 1.5% collagen solution was covered on the lower layer of 2.5% collagen gel block, and the uppermost layer was 0.5% collagen solution, which was pre-cooled at 40°C for 12 hours and then freeze-dried. 60mM EDC / NHS (nEDC / nNHS=2.5:1, pH 5.5), soaked in 50mM MES in 40% ethanol solution for 4h, with 0.1M Na 2 HPO 4 The solution was washed for 4 hours, and then washed repeatedly with deionized water. After freeze...

Embodiment 3

[0050] Example 3: Preparation of a medical gradient collagen film with a dense outer layer and a loose inner layer from fish skin

[0051] The non-denatured collagen derived from fish skin obtained according to Example 1 was dissolved in 0.05M acetic acid to prepare 3%, 2%, and 1% collagen suspensions, left standing at 4°C for 48 hours, and then stirred until homogenized. After the dissolution is complete, put the 3% collagen solution into a stainless steel dish, spread a thin layer, and stand at a low temperature to obtain a glue block. Then, according to the gradient, 2% collagen solution was covered on the lower layer of 3% collagen gel block, and the uppermost layer was 1% collagen solution, which was pre-cooled at 40°C for 12 hours and then freeze-dried. 60mM EDC / NHS (nEDC / nNHS=2.5:1, pH 5.5), soaked in 40% ethanol solution containing 50mM MES for 4h, with 0.1M Na 2 HPO 4 The solution was washed for 4 hours, and then washed repeatedly with deionized water. After freeze...

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Abstract

The invention provides a medical collagen membrane with a compact outer layer and a loose inner layer in structure. Fish skin pepsin-solubilized collagen is dissolved in an acid solution to prepare collagen solutions with different concentrations, then laying is performed according to the concentrations from high to low, and pressing is performed to prepare a collagen membrane, wherein the fish skin pepsin-solubilized collagen is prepared by treating fish skin with alkali liquor, and then carrying out enzymolysis with pepsin under a weak acid condition; an enzymatic hydrolysate is centrifuged, a supernate is collected, and the supernate is salted out; precipitates are collected, the precipitates are redissolved with a weak acid solution, and then dialysis treatment is performed; and a dialysate is freeze-dried to prepare the fish skin pepsin-solubilized collagen. The collagen membrane prepared in the invention has three layers of different structures, the aperture of the inner layer is relatively large to promote the growth of osteoblasts, the small aperture of the outer layer can prevent soft tissue cells from entering and allow stem cells and nutrient substances to pass through, and a certain transition layer exists between the inner layer structure and the outer layer structure to facilitate tissue integration.

Description

technical field [0001] The invention belongs to the technical field of medical materials, and in particular relates to a medical collagen film derived from fish skin and a preparation method thereof, that is, a medical collagen film with a dense outer layer and a loose inner layer in structure and a preparation method thereof. Background technique [0002] The amino acids of collagen are arranged in (Gly-X-Y)n period, and the molecular structure is highly repetitive, so its antigenicity is low and immune rejection is less. Collagen shows good biocompatibility and has a harmonious interaction mechanism with the surrounding environment of cells, which can better promote the repair of human defective tissue and participate in the reconstruction process of new tissue. It is an ideal medical biomaterial. [0003] Terrestrial mammals such as pigs and cattle are currently the main source of raw materials for extracting collagen, but there are problems with the spread of diseases su...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/78C12P21/06C07K1/14C07K1/36C07K1/30C07K1/34A61L31/12A61L31/04A61L31/14
CPCC07K14/78C12P21/06A61L31/044A61L31/14A61L31/148
Inventor 侯虎宋雪樊燕王静凤
Owner OCEAN UNIV OF CHINA
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