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Method for preparing high-purity chondroitin sulfate A from rabbit ear cartilage

A chondroitin sulfate, high-purity technology, applied in the field of preparation of high-purity chondroitin sulfate, can solve the problems of chondroitin sulfate application limitations, difficult to obtain raw materials, preparation of chondroitin sulfate, etc., to make up for insufficient market supply, low cost, Product safety and reliability

Active Publication Date: 2014-05-14
OCEAN UNIV OF CHINA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In recent years, swine flu, porcine foot-and-mouth disease, porcine blue-ear disease and other diseases have occurred from time to time, which is very likely to limit the application of chondroitin sulfate derived from it
[0004] In terms of CSA preparation, a Chinese patent application (publication number CN102898541A) involves a method for extracting chondroitin sulfate from fish bones. Although chondroitin sulfate derived from fish is less polluted than terrestrial animals, its disaccharide The ratio of C-4 sulfate group and C-6 sulfate group in the repeating unit is quite different from that of medicinal chondroitin sulfate A
Chinese patent application (publication number CN102850466A) discloses a method for preparing chondroitin sulfate from yak bone. Due to the obvious regional distribution of yak, raw materials are difficult to obtain
Chinese patent application (publication number CN102690372A) involves a method of producing chondroitin sulfate from chicken skeletons. Also due to the frequent prevalence of bird flu, chicken-derived products also have certain safety hazards
With the development of the rabbit breeding industry, a large number of rabbit ears are eaten by people as various common foods. The rabbit ear cartilage contains a large amount of chondroitin sulfate, which is an excellent raw material of chondroitin sulfate. According to domestic and foreign patent searches, it is currently There is no report on the preparation of chondroitin sulfate from rabbit ears

Method used

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  • Method for preparing high-purity chondroitin sulfate A from rabbit ear cartilage
  • Method for preparing high-purity chondroitin sulfate A from rabbit ear cartilage
  • Method for preparing high-purity chondroitin sulfate A from rabbit ear cartilage

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preparation example Construction

[0028] The preparation method of the high-purity rabbit ear cartilage chondroitin sulfate A provided by the present invention comprises the following steps:

[0029] (1) Degreasing: After the death of the rabbit, the fresh rabbit ears are peeled and fleshed to obtain the rabbit ear cartilage, soaked in an organic solvent, stirred intermittently, taken out and dried, and pulverized to obtain defatted cartilage powder.

[0030] (2) Enzymolysis: Soak defatted cartilage powder in Tris-HCl buffer solution and boil to cool down to 37°C, add trypsin, stir at 37°C for enzymolysis, then boil to inactivate the enzyme.

[0031] (3) Ethanol precipitation: Centrifuge the enzymatic hydrolysis solution obtained in (2), add ethanol to the supernatant to precipitate, take the precipitate after centrifugation, add distilled water to redissolve, then dialyze against distilled water, and obtain the crude chondroitin product after the dialyzed liquid is concentrated and dried under reduced pressure...

Embodiment 1

[0037] Take 3 pairs of fresh rabbit ears, peel and remove the flesh to obtain 35g of rabbit ear cartilage, add 350mL of acetone to soak, stir intermittently for 4h (every 1h, stir for 10 minutes), discard the acetone soaking solution, add 350mL of chloroform / methanol (chloroform and Methanol (volume ratio 4:1) was soaked for 4 hours, the organic solvent was removed, dried and pulverized, and dried to obtain 10.1 g of defatted cartilage powder. Soak defatted cartilage powder in 200mL, 0.1mol / L, Tris-HCl buffer solution with pH=8.0, boil for 2h, cool down to 37°C, add 1.5g trypsin, stir at 37°C for 22h, boil at 100°C for 5min Enzyme inactivation. Centrifuge the enzymatic solution, take the supernatant and add 600mL of absolute ethanol, centrifuge to collect the precipitate, add distilled water to redissolve the precipitate, use a dialysis bag with a cut-off molecular weight of 3500Da for dialysis desalination, concentrate the liquid in the dialysis bag under reduced pressure, and ...

Embodiment 2

[0039] Take 5 pairs of fresh rabbit ears, peel and remove the flesh to obtain 57g of rabbit ear cartilage, add 850mL of acetone to soak, stir intermittently for 8h (every 1h, stir for 5 minutes), remove the acetone, dry and pulverize to obtain 19.4g of defatted cartilage powder. Soak defatted cartilage powder in 390mL, 0.1mol / L, pH=8.0, Tris-HCl buffer solution and boil for 3 hours, then add 5.0g trypsin after cooling down to 37°C, stir at 37°C for 21h, boil at 100°C for 5min to make the enzyme Inactivate. Centrifuge the enzymatic solution, take the supernatant and add 780mL of ethanol with a concentration of 95wt%, centrifuge to collect the precipitate, add distilled water to redissolve the precipitate, and dialyze with a dialysis bag with a cut-off molecular weight of 7000Da until the conductivity of the liquid outside the dialysis bag does not change, and the inside of the dialysis bag The solution was concentrated under reduced pressure and dried to obtain 2.37 g of crude ...

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Abstract

The invention relates to a technology for preparing high-purity chondroitin sulfate from animal organ, and particularly relates to a method for preparing high-purity chondroitin sulfate A from rabbit ear cartilage. The method comprises the following steps: skinning and fleshing the rabbit ear to obtain fresh rabbit ear cartilage; performing degreasing, trypsin enzymolysis, ethanol precipitation and anion exchange resin purification to obtain high-purity chondroitin sulfate A with molecular weight of 16-59kDa. The method provided by the invention is used for preparing chondroitin sulfate A from rabbit ear cartilage, has the characteristics of abundant sources of raw material, high product yield and high purity, and can meet the need for high-purity chondroitin sulfate A in the medical market.

Description

technical field [0001] The invention relates to a technology for preparing high-purity chondroitin sulfate by using animal organs, in particular a method for preparing high-purity chondroitin sulfate A by using rabbit ear cartilage. Background technique [0002] Chondroitin sulfate (CS) is a sulfated polysaccharide that exists widely in various animal tissues and organs in the form of proteoglycan, and has various physiological functions. Chondroitin sulfate in biological tissues mainly includes chondroitin sulfate A (CSA), chondroitin sulfate B (CSB) and chondroitin sulfate C (CSC), among which: CSA is composed of glucuronic acid (GlcA) and β-1 , 3-glucosidic bonds and 4-sulfate-acetylgalactosamine (GalNAc4S) are connected alternately by β-1,4-glycosidic bonds. Chondroitin sulfate A has good anti-inflammatory, hypolipidemic and other physiological activities, and is widely used in clinical practice. my country is a major producer and exporter of chondroitin sulfate, and c...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C08B37/08
Inventor 于广利张玉倩祝贺郎银芝赵小亮
Owner OCEAN UNIV OF CHINA
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