Method for extracting high-purity heparan sulfate from heparan production wastes

A technology of heparan sulfate and waste, which is applied in the extraction of high-purity heparan sulfate and the field of extracting high-purity heparan sulfate. It can solve the problems of low anticoagulant activity, difficult amplification and mixing of process conditions, and solve financial and material resources. , the effect of preserving integrity and creating value

Active Publication Date: 2018-09-14
深圳市格利科生物科技有限公司
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AI Technical Summary

Problems solved by technology

The patented method obtains heparan sulfate through resin adsorption and elution, but also mixes glycosaminoglycans such as dermatan sulfate, so the purity of heparan sulfate is not high
[0009] Chinese patent ZL201410819670 discloses a method for separating heparan sulfate from animal lungs. In this method, hydrolytic enzymes are used to degrade the saline extracts of animal lungs, and then oxidizing agents and activated carbon are added to the enzymolysis solution for oxidative adsorption and decolorization. Then use acetone to fractionally precipitate the oxidized solution, and dehydrate and dry to obtain crude heparan sulfate. Then, the aqueous solution of crude heparan sulfate is sequentially separated by membrane separation and anion exchange chromatography, and heparan sulfate is mixed with dermatan sulfate and cartilage sulfate. Separation of impurities such as heparin and heparin, and then use a 5000-7000Da ultrafiltration membrane to carry out ultrafiltration and concentration of the eluate, and finally use gel filtration chromatography to desalt and freeze-dry to obtain heparan sulfate fine products, but the process of the invention is lengthy. Complex intermediate process control is required, and the process conditions are not easy to scale up, so the prospect of industrialization is worrying
However, the higher the purity of heparan sulfate, the lower the anticoagulant activity, so the purity of heparan sulfate obtained by this patent is not high
[0011] Chinese patent ZL201510394022 discloses a method for separating heparinoids from waste protein by-products of heparin. In this method, the waste protein by-products of heparin is dissolved, added with an adsorbent, eluted with alcohol precipitation, oxidized alcohol precipitation and dried to obtain a dry heparinoid product. , but this type of heparin dry product not only contains heparan sulfate, but also at least contains mucopolysaccharides such as fast heparin with high anticoagulant activity, indicating that the purity of heparan sulfate in heparin-like dry product is not high
The raw material of sulphate obtained by this invention contains dermatan sulfate and fast heparin or heparan sulfate, therefore, the purity of heparan sulfate is not high
[0016] In summary, no environmentally friendly process for extracting high-purity heparan sulfate from heparin production waste has been reported so far.

Method used

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  • Method for extracting high-purity heparan sulfate from heparan production wastes
  • Method for extracting high-purity heparan sulfate from heparan production wastes
  • Method for extracting high-purity heparan sulfate from heparan production wastes

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Experimental program
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Embodiment 1

[0045] 1) Add 1 kg of heparin high-quality production waste, add 19 kg of water, stir and dissolve, and make a solution with a concentration of 5%;

[0046] 2) Acid-adjusting centrifugation: add 1:1 HCl to adjust the pH of the solution to 1.9, add 0.1 times ethanol, stir, and centrifuge;

[0047] 3) Collect the supernatant after centrifugation, add 30% NaOH solution to adjust the pH to 7, and add 2 times more water to make the solution salinity less than 2%;

[0048] 4) Add 50ml of strong basic anion adsorption resin (Rigson resin, USA) to the above solution, stir and adsorb for 18h;

[0049] 5) Separating the remaining adsorption liquid and the resin, washing the collected resin with water, then adding 0.5 times the volume of 3% sodium chloride solution for elution, collecting the eluate after 8 hours of elution, adding 0.8 times the volume of ethanol for precipitation;

[0050] 6) Collect the precipitate, add 3 times of water to dissolve, add hydrogen peroxide to oxidize, ...

Embodiment 2

[0057] 1) Add 1 kg of heparin high-quality production waste, add 5.67 kg of water, stir and dissolve, and make a solution with a concentration of 15%;

[0058] 2) Acid-adjusting centrifugation: add sulfuric acid solution to adjust the pH of the solution to 2.0, add 0.7 times of ethanol, stir, and centrifuge;

[0059] 3) Collect the centrifuged supernatant, add 30% NaOH solution to adjust the pH to 6.0, and add 2.5 times more water to make the solution salinity less than 2%;

[0060] 4) Add 500ml of strongly basic anion adsorption resin (Germany LANXESS resin) to the above solution, stir and adsorb for 13h;

[0061] 5) Separating the remaining adsorption liquid and the resin, washing the collected resin with water, then adding 2 times the volume of 7% sodium chloride solution for elution, collecting the eluate after 2 hours of elution, adding 0.6 times the weight acetone for precipitation;

[0062] 6) Collect the precipitate, add 3 times of water to dissolve, add hydrogen per...

Embodiment 3

[0069] 1) Add 1 kg of heparin high-quality production waste, add 9 kg of water, stir and dissolve, and make a solution with a concentration of 10%;

[0070] 2) Acid-adjusting centrifugation: add trichloroacetic acid solution to adjust the pH of the solution to 2.3, add 0.35 times of ethanol, stir, and let stand for 13 hours;

[0071] 3) Collect the supernatant after standing still, add 30% NaOH solution to adjust the pH to 8, and add 2 times more water to make the solution salinity less than 2%;

[0072] 4) above-mentioned solution loads on the ion-exchange chromatography column that 500ml strong anion-exchange gel Q Sepharose is housed;

[0073] 5) Add 0.5 times the volume of 0.6M sodium chloride solution for elution, the gravity flow rate, collect the eluate, add 1 times the weight of methanol for precipitation;

[0074] 6) Collect the precipitate, add 3 times of water to dissolve, add hydrogen peroxide to oxidize, and oxidize for 48 hours.

[0075] 7) After oxidation, the...

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Abstract

The method discloses a method for extracting high-purity heparan sulfate from heparan production wastes, and aims at providing a method used for extracting purified heparan sulfate taking the heparanproduction wastes as raw material. The method has the advantages of low cost, convenient operation and easy industrial scaling. The method is characterized by comprising the steps that 1), the heparanproduction wastes are prepared into a solution; 2), acid is added to adjust the pH to 1.9-2.3, ethanol is added, and stirring, centrifugation and standing are conducted; 3), supernatant liquid is collected, aqueous alkali is added to adjust pH to 6-8, and water is added; 4), absorbent is added to the solution is step 3), and stirring is conducted until all mucopolysaccharide in the solution is absorbed by the absorbent; 5), the absorbent is collected and washed with clean water, a sodium chloride solution is added to conduct elution, elution liquid is collected after the elution is completed,and precipitant is added for precipitation; 6), precipitations are collected and dried; the method belongs to the field of mucopolysaccharide pharmaceuticals.

Description

technical field [0001] The invention discloses a method for extracting high-purity heparan sulfate, in particular to a method for extracting high-purity heparan sulfate from heparin production waste, belonging to the field of mucopolysaccharide biopharmaceuticals. Background technique [0002] Heparin sodium is a commonly used mucopolysaccharide. In addition to the target product heparin sodium, other by-products of mucopolysaccharides are usually produced during the production process, such as heparan sulfate, dermatan sulfate, and chondroitin sulfate. The chondroitin sulfate in the by-product is easy to remove , while dermatan sulfate and heparan sulfate have similar properties, it is difficult to separate and purify the two by ion exchange chromatography and organic solvent fractional precipitation, and generally exist in the form of an unseparated mixture of the two, or stored in each production unit In the warehouse, or directly discarded, this leads to a serious waste ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C08B37/10
CPCC08B37/0075
Inventor 黄洪陈海苑
Owner 深圳市格利科生物科技有限公司
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