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Preparation method of active collagen with triple-helix structure

A triple-helix and collagen technology, applied in the field of protein engineering, can solve the problems of destroying the triple-helix structure, low extraction efficiency, unable to realize the triple-helix structure of active collagen, etc., and achieves the effects of improving extraction efficiency, improving purity and low immunogenicity.

Active Publication Date: 2015-01-07
无锡贝迪生物工程股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The methods disclosed in the prior art often destroy the triple helix structure of collagen during the extraction and preparation of collagen, or the extraction efficiency is low, and it is impossible to preserve the triple helix structure of active collagen while improving the extraction efficiency and purity

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] (1) Take fresh cowhide, cut and remove the grease layer and fur layer, remove impurities, wash, break the cowhide into small particles, add 0.03mol / L sodium hydroxide aqueous solution according to the ratio of solid to liquid 1:30, and Soak at 6-8°C for 2 hours, filter, and set aside;

[0027] (2) Add anhydrous ether or acetone whose mass is 8 times the mass of cowhide to the cowhide after the above pretreatment, reflux at 35°C for 8 hours, then rinse with distilled water until there is no peculiar smell, and set aside;

[0028] (3) Immerse the above cowhide in a mixture of 0.6mol / L glacial acetic acid and 700mg / L pepsin, and keep stirring for about 25 hours;

[0029] (4) Use progressive ultrasonic treatment on the above mixture, first use 100W ultrasonic treatment for 30 minutes, then use 300W ultrasonic treatment for 30 minutes, and finally use 400W ultrasonic treatment for 1 hour;

[0030] (5) Add 2% H 2 o 2 Solution, mix and stand for 4 hours, adjust the pH to 5....

Embodiment 2

[0035] (1) Take fresh beef Achilles tendon, cut and remove the grease layer and fur layer, remove impurities, wash, break the beef Achilles tendon into small particles, and add 0.01mol / L of hydroxide according to the ratio of solid to liquid 1:40 Sodium aqueous solution, soak at 6-8°C for 1 hour, filter, and set aside;

[0036] (2) Add anhydrous ether or acetone with a mass 6 times the mass of the Achilles tendon to the Achilles tendon after the above pretreatment, reflux at 40°C for 5 hours, then rinse with distilled water until there is no peculiar smell, and set aside;

[0037] (3) Immerse the above bovine Achilles tendon in a mixture of 0.7mol / L glacial acetic acid and 600mg / L pepsin, and keep stirring for about 30 hours;

[0038] (4) Use progressive ultrasonic treatment on the above mixture, first use 100W ultrasonic treatment for 30 minutes, then use 200W ultrasonic treatment for 30 minutes, and finally use 300W ultrasonic treatment for 1 hour;

[0039] (5) Add 3% H 2 o ...

Embodiment 3

[0044] (1) Take fresh pigskin, cut and remove the grease layer and fur layer, remove impurities, wash, break the pigskin into small particles, and add 0.02mol / L sodium hydroxide aqueous solution according to the ratio of solid to liquid 1:35 , soaked at 6-8°C for 1.5 hours, filtered, and set aside;

[0045] (2) Add anhydrous ether or acetone whose mass is 7 times that of the pigskin to the above pretreated pigskin, reflux at 37°C for 6 hours, then rinse with distilled water until there is no peculiar smell, and set aside;

[0046] (3) Immerse the above pigskin in a mixture of 0.65mol / L glacial acetic acid and 650mg / L pepsin, and keep stirring for about 28 hours;

[0047] (4) Use progressive ultrasonic treatment on the above mixture, first use 150W ultrasonic treatment for 30 minutes, then use 250W ultrasonic treatment for 30 minutes, and finally use 350W ultrasonic treatment for 1 hour;

[0048] (5) Add 2.5% H2O2 solution, mix and stand for 3 hours, adjust the pH to 5.0, and ...

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Abstract

The invention relates to a preparation method of collagen; fresh pigskin is used as a raw material; the raw material is treated by pretreatment, fat removal, and acid-enzyme treatment; ultrasonic treatment is employed to further improve extraction efficiency; and finally purification is performed to prepare a collagen product with high purity. The preparation method of collagen has high extraction efficiency, and the product purity is up to above 98%. The product is active collagen with a maintained triple-helix structure, has low immunogenicity and high biocompatibility, is biodegradable, and is applicable to medical purposes.

Description

technical field [0001] The invention relates to a preparation method of active collagen with a triple helix structure, which belongs to the field of protein engineering. The product is active collagen maintaining a triple helix structure, has low immunogenicity, high biocompatibility, is biodegradable, and is suitable for use in medical use. Background technique [0002] Collagen is a group of hard proteins with the most abundant content and widest distribution in vertebrates. 25%-33% of the total protein content of the animal body. Several types of collagen are often contained in the same tissue, and a certain type is often the main one. Type Ⅰ collagen is distributed in all parts of the body, mainly in the skin, tendons and ligaments, and has a strong tensile capacity. Type II collagen mainly exists in the hyaline cartilage and vitreous body, and has a strong ability to resist pressure. Type III collagen is widely distributed in tissues with high extensibility, such as...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/78
Inventor 任伟业
Owner 无锡贝迪生物工程股份有限公司
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