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Method for extracting water-soluble collagen

An extraction method and water-soluble technology, applied in the field of extraction of water-soluble collagen, can solve problems such as troublesome preparation of neutral biological materials

Inactive Publication Date: 2017-10-10
BEIJING HOTWIRE MEDICAL TECH DEV CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, the collagen solubility proposed by the collagen extraction method is mostly acid-soluble or alkali-soluble, which brings a lot of trouble to the preparation of neutral biomaterials suitable for human needs.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Select 0.5 kg of fresh calfskin tissue, wash with water until no foreign matter is visible to the naked eye; remove fat with scissors, wash with water, and freeze overnight at -20°C; slice with a microtome, slice thickness is about 1mm, and set aside;

[0022] Weigh 100g slices, soak in 0.05% chlorhexidine acetate solution for 0.5h, soak in purified water and wash 5 times, drain excess water for later use. Then wash and soak the sliced ​​tissue in 0.3% (W / V) sodium carbonate solution for 6 hours. Wash with purified water until the pH is 8, then soak in 0.7% saline for about 2 hours, drain off excess water, and mash the tissue at 2-8°C;

[0023] Prepare enzymatic hydrolysis solution, weigh acid protease according to the ratio of acid protease:tissue section of 1:150, the protease titer is 1:3000, dissolve it in 4000ml hydrochloric acid solution with a molar concentration of 0.02mol / L, and stir evenly. Adjust the pH of the solution to 4.

[0024] Disperse the crushed ti...

Embodiment 2

[0027] Select 0.5 kg of fresh calfskin tissue, wash with water until no foreign matter is visible to the naked eye; remove fat with scissors, wash with water, and freeze overnight at -20°C; slice with a microtome, slice thickness is about 1mm, and set aside;

[0028] Weigh 100 g slices, soak in 0.05% chlorhexidine acetate solution for 5 hours, soak in purified water and wash 5 times, drain excess water for later use. Then wash and soak the sliced ​​tissue in 0.3% (W / V) sodium carbonate solution for 6 hours. Wash with purified water until the pH is 7, then soak in 0.7% saline for about 2 hours, drain off excess water, and mash the tissue at 2-8°C;

[0029] Prepare enzymatic hydrolysis solution, weigh acid protease according to the ratio of acid protease:tissue section of 1:300, protease titer is 1:6000, dissolve it in 4000ml hydrochloric acid solution with molar concentration of 0.02mol / L, and stir evenly. Adjust the pH of the solution to 3.

[0030] Disperse the crushed tiss...

Embodiment 3

[0033] Select 0.5 kg of fresh piglet skin tissue, wash it with water until no foreign matter is visible to the naked eye; remove the fat with scissors, wash with water, and freeze overnight at -20°C; slice with a microtome, the thickness of the slice is about 1 mm, and set aside;

[0034] Weigh 100 g slices, soak in 0.05% chlorhexidine acetate solution for 1 hour, soak in purified water and wash 5 times, drain excess water for later use. Then wash and soak the sliced ​​tissue in 3.0% (W / V) sodium carbonate solution for 6 hours. Wash with purified water until the pH is 8, then soak in 0.7% saline for about 2 hours, drain off excess water, and mash the tissue at 2-8°C;

[0035] Prepare the enzymatic solution, weigh the acid protease according to the ratio of acid protease:tissue section 1:80, the protease titer is 1:1500, dissolve it in 4000ml hydrochloric acid solution with a molar concentration of 0.01mol / L, and stir evenly. Adjust the pH of the solution to 5.

[0036] Dispe...

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PUM

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Abstract

The invention discloses a method for extracting water-soluble collagen. The method comprises the following steps: firstly slicing a tissue rich in type III collagen; then cleaning tissue slices and soaking in 0.1-40%(W / V) alkali solution for 0.5-24 hours; cleaning until pH is 5-8, then soaking for about 0.5-48 hours with normal saline, draining excessive moisture, mashing the tissue at the temperature of 2-8 DEG C, then dispersing into enzymatic hydrolysate, and carrying out enzymatic hydrolysis for 1-72 hours in an environment at the temperature of 2-40 DEG C; after the enzymatic hydrolysis is finished, regulating pH value to be 7-13, and standing for 2-24 hours at the temperature of 2-40 DEG C; filtering with a filter screen of 30-150 meshes, and removing substances which are not subjected to enzymatic hydrolysis; salting out with 20-50%(W / V) salt solution, collecting solid contents, carrying out dialysis with a dialysis bag with the molecular weight cut-off of 4000-8000, and removing excessive ions and impurities, so that polymorphic collagen is obtained; and finally, dispersing a polymorphic collagen mixture into water, regulating pH value to 1-5, filtering with a filter screen of 30-150 meshes, removing precipitates, salting out filtrate, and then carrying out dialysis, so that the target collagen is obtained. By adopting the method for extracting the water-soluble collagen, the obtained collagen is an aqueous solution, and the blank that existing collagen is acid soluble or alkali soluble is filled up.

Description

technical field [0001] The invention relates to the technical field of biomedical materials, in particular to a method for extracting water-soluble collagen. Background technique [0002] There are many types of collagen, and the common types are type Ⅰ, type Ⅱ, type Ⅲ, type Ⅴ and type Ⅺ. Collagen has good biocompatibility, biodegradability and biological activity, and is widely used in food, health products, Medical biomaterials, tissue engineering, cosmetics and other fields. In the medical field, due to the advantages of low immunogenicity, fiber re-formation, high mechanical properties and biodegradability, collagen has been made into medical collagen sponges, membranous biological patches, artificial skin, cosmetic filling implants, etc. Injection, cosmetics, etc. [0003] The solubility of collagen proposed by the current collagen extraction method is mostly acid-soluble or alkali-soluble, which brings a lot of trouble to the preparation of neutral biomaterials suita...

Claims

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Application Information

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IPC IPC(8): C12P21/06C07K14/78C07K1/36C07K1/34C07K1/30
CPCC07K14/78C12P21/06
Inventor 张春红富勇
Owner BEIJING HOTWIRE MEDICAL TECH DEV CO LTD
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