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Fine heparin and method for preparing fine heparin

A crude heparin and heparin technology, applied in the field of high-quality heparin and the preparation of the high-quality heparin, can solve the problems of high economic cost and time cost, large amount of ethanol consumption, difficult separation, etc.

Active Publication Date: 2017-10-13
TSINGHUA UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The removal of galactosamine impurities is also one of the key issues in the purification of heparin. In the existing purification process, galactosamine impurities are often removed through multi-step fractional precipitation methods by adjusting different ethanol concentrations. Removal of polysaccharides, mainly in the form of dermatan sulfate, is particularly difficult
[0010] As mentioned above, the existing heparin purification method to remove galactosamine impurities (mainly chondroitin, CS-A, CS-C, DS) in heparin is mainly to carry out multi-stage precipitation with ethanol, because heparin and other galactosamine impurities The structure, molecular weight, and alcohol solubility of heparin are very close, so it is difficult to separate by ethanol precipitation alone, and multi-stage separation is required to achieve a relatively ideal separation effect, so the yield of heparin decreases, and the amount of ethanol is large, economic and time costs higher

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  • Fine heparin and method for preparing fine heparin
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  • Fine heparin and method for preparing fine heparin

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[0081] The mass yield obtained by the method for preparing refined heparin described above can be calculated by: (mass of refined heparin / mass of crude heparin)×100%.

[0082] The potency yield obtained by the method for preparing refined heparin described above can be calculated by: mass yield×(fine heparin aPTT / crude heparin aPTT)×100%.

[0083] The titer yield of the method for preparing high-quality heparin according to the present invention is usually above 70%.

[0084] The mass yield of the method for preparing high-quality heparin according to the present invention is usually above 35%.

[0085] The high-quality heparin herein can be obtained by the method described herein above, that is, the high-quality heparin with a sheep plasma titer of 150 IU / mg or more, preferably 160 IU / mg or more, and more preferably 170 IU / mg or more, and the high-quality heparin contained in the above-mentioned high-quality heparin The impurity of galactosamine is 5% by mass or less, prefer...

Embodiment 1

[0144] 40g of crude heparin sodium purchased from Sichuan Guangyuan Shenda Industrial Co., Ltd. was dissolved in 400ml of deionized water, and 0.3% (w / v) of sodium chloride and 0.22% (w / v) of calcium chloride were added to adjust the pH to 7. 5. Add 200 IU of chondroitinase B (MBP-ChonB) and react at 30°C for 0.5 hours, then add 200IU and react at 30°C for 1.5 hours. After the reaction, add 2.7% (W / V) NaCl, adjust the pH to 8.9, control the temperature at 48°C, add 0.6% (W / W) trypsin, adjust the pH to 8.9 every hour, and enzymolysis for 5.5 hours. After trypsinization, add 2.78% (W / V) CaCl 2 React at room temperature for 30 minutes, raise the temperature to 80-85°C, adjust the pH to 9.3 with 20% NaOH, and centrifuge at 10,000 g for 10 minutes. Cool the feed solution to below 40°C, adjust the pH to 10.3, and centrifuge at 10,000 g for 10 minutes. Add 3.0% (W / V) anhydrous sodium carbonate to the feed solution after centrifugation, react at 48°C for 30 minutes, and centrifuge a...

Embodiment 2

[0153] 40g of crude heparin sodium purchased from Sichuan Guangyuan Shenda Industrial Co., Ltd. was dissolved in 400ml of deionized water, and 0.3% (w / v) of sodium chloride and 0.22% (w / v) of calcium chloride were added to adjust the pH to 7. 5. Add 120 IU of chondroitinase AC (MBP-ChonAC) and react at 30°C for 0.5 hours, then add 120IU and react at 30°C for 1.5 hours. After the reaction, add 2.7% (W / V) NaCl, adjust the pH to 8.9, control the temperature at 48°C, add 0.6% (W / W) trypsin, adjust the pH to 8.9 every hour, and enzymolysis for 5.5 hours. After trypsinization, add 2.78% (W / V) CaCl 2 React at room temperature for 30 minutes, raise the temperature to 80-85°C, adjust the pH to 9.3 with 20% NaOH, and centrifuge at 10,000 g for 10 minutes. Cool the feed solution to below 40°C, adjust the pH to 10.3, and centrifuge at 10,000 g for 10 minutes. Add 3.0% (W / V) anhydrous sodium carbonate to the feed solution after centrifugation, react at 48°C for 30 minutes, and centrifuge...

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Abstract

This article relates to a high-quality heparin and a method for preparing the high-quality heparin. The method for preparing high-quality heparin includes: dissolving crude heparin in water to prepare a heparin solution, adding chondroitinase AC and / or chondroitinase B to the heparin solution, and performing an enzyme reaction with galactosamine impurities; removing the heparin solution Chondroitinase AC and / or Chondroitinase B in

Description

technical field [0001] This document relates to refined heparins and methods of preparing said refined heparins. Background technique [0002] Heparin is a mucopolysaccharide formed by hexuronic acid (L-iduronic acid, D-glucuronic acid) and D-glucosamine sulfate by alternating 1→4 glycosidic bonds, and has a linear structure of repeating units of six or eight sugars. Chain structure, its molecular weight is between 3000-37000Da, it is used as anticoagulant reagent and antithrombotic reagent in medicine. In addition, heparin also has various biological functions such as anti-inflammation, anti-allergy, anti-virus, anti-cancer, blood lipid regulation (Maurice Petitou, Benito Casu, UlfLindah.1976–1983, a critical period in the history of heparin: the discovery of the antithrombin binding site. Biochimie 85 (2003) 83–89). [0003] Crude heparin can be used for medicine after purification, which includes a series of general separation steps and steps to inactivate foreign subst...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C08B37/10
Inventor 邢新会吴敬君张翀
Owner TSINGHUA UNIV