Method for purifying heparitin sulfate from heparin byproduct

A technology of heparan sulfate and heparin by-products, which is applied in the field of preparation of heparan sulfate, can solve the problems of high price, difficulty in obtaining heparan sulfate, difficulty in carrying out, etc., and achieves the effects of low cost, high product yield and simple process

Active Publication Date: 2010-11-17
SHENZHEN HEPALINK PHARMA GRP CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, it is extremely difficult to obtain heparan sulfate, and the price of HS from reagent companies is extremely expensive, which discourages many researchers.
Few researchers in China hav

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0015] Dissolve heparin by-products into a 2% solution, add potassium acetate to it to a concentration of 10%, stir to dissolve, adjust the pH to 5 with acetic acid, and precipitate at 2°C for 1 day. The supernatant was collected by centrifugation to obtain a sulfated polysaccharide solution after heparin was removed.

[0016] Add 0.5 times the volume of Banner's reagent and 0.05 times the volume of saturated sodium hydroxide to the aforementioned sulfated polysaccharide solution, stir and precipitate at room temperature for 10 minutes, and centrifuge to separate the supernatant and the precipitate. Add 0.5 times the volume of Banner's reagent to the precipitate, stir well to dissolve, then add 0.05 times the volume of saturated sodium hydroxide, stir and precipitate at room temperature for 10 minutes, and centrifuge to remove the precipitate.

[0017] Combine the supernatants obtained in the first two steps, dilute with 2000ml of purified water, put on a Q-SepharoseFF column,...

Embodiment 2

[0019] Dissolve heparin by-products into a 10% solution, add potassium acetate to it to a concentration of 80%, stir to dissolve, adjust the pH to 7 with acetic acid, and precipitate at 2°C for 7 days. The supernatant was collected by centrifugation to obtain a sulfated polysaccharide solution after heparin was removed.

[0020] Add 1.6 times the volume of Banner's reagent and 0.2 times the volume of saturated sodium hydroxide to the aforementioned sulfated polysaccharide solution, stir and precipitate at room temperature for 60 minutes, and centrifuge to separate the supernatant and the precipitate. Add 1.6 times the volume of Banner's reagent to the precipitate, stir well to dissolve, then add 0.2 times the volume of saturated sodium hydroxide, stir and precipitate at room temperature for 60 minutes, and centrifuge to remove the precipitate.

[0021] Combine the supernatants obtained in the first two steps, dilute with 2000ml purified water, put on a Q-Sepharose FF column, w...

Embodiment 3

[0023] Dissolve heparin by-products into an 8% solution, add potassium acetate to it to a concentration of 50%, stir to dissolve, adjust the pH to 6 with acetic acid, and precipitate at 5°C for 3 days. The supernatant was collected by centrifugation to obtain a sulfated polysaccharide solution after heparin was removed.

[0024] Add 1.2 times the volume of Benedict's reagent and 0.08 times the volume of saturated sodium hydroxide to the aforementioned sulfated polysaccharide solution, stir and precipitate at room temperature for 30 minutes, and centrifuge to separate the supernatant and the precipitate. Add 1.2 times the volume of Banner's reagent to the precipitate, stir well to dissolve, then add 0.08 times the volume of saturated sodium hydroxide, stir and precipitate at room temperature for 30 minutes, and centrifuge to remove the precipitate.

[0025] Combine the supernatants obtained in the first two steps, dilute with 2000ml purified water, put on a Q-SepharoseFF column...

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PUM

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Abstract

The invention discloses a method for purifying heparitin sulfate from a heparin byproduct. The method comprises the following steps of: using the heparin byproduct as a raw material; dissolving the heparin byproduct, and then adding potassium acetate into the dissolved heparin byproduct; adjusting the pH value of solution by using acetic acid; removing a deposit to obtain sulfate polysaccharide solution; adding a Ban's agent and saturated sodium hydroxide solution into the sulfate polysaccharide solution; centrifugally collecting clear solution; adding the Ban's agent and the saturated sodium hydroxide solution into the deposit again, and centrifugally collecting the clear solution; combining the clear solution; washing away copper ions by using a chromatography column with anion exchange properties and sodium chloride solution; performing linear gradient elution by using the sodium chloride solution; collecting eluant; and evaporating and concentrating the eluant, and depositing the eluant by using ethanol to obtain high-purity dermatan sulfate. The method has the advantages of simple purification process, high purification yield, low cost, easy amplification and suitability for industrial production.

Description

technical field [0001] The invention relates to a preparation method of heparan sulfate, in particular to a method for purifying heparan sulfate from heparin by-products. Background technique [0002] Heparan sulfate (HS for short) is a natural glycosaminoglycan composed of repeating disaccharide units, and the disaccharides are sulfated glucosamine and hexuronic acid. It is widely distributed in animal tissues and can be synthesized in almost all cells. Heparan sulfate differs from heparin in that it is less sulfated and has a smaller proportion of iduronic acid in the hexuronic acid. [0003] Because it has been reported that HS has anti-tumor function, the research of HS has attracted much attention in recent years. However, it is extremely difficult to obtain heparan sulfate, and the HS price of reagent companies is extremely expensive, which makes many researchers stop. Few researchers in China have conducted anti-tumor experiments on HS, and several research groups ...

Claims

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Application Information

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IPC IPC(8): C08B37/10
Inventor 李锂李坦
Owner SHENZHEN HEPALINK PHARMA GRP CO LTD
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