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Method for preparing human fibrinogen

A human fibrinogen and protein technology, which is applied in the field of human fibrinogen preparation, can solve the problems of plasma waste, backward product technology, low purity, etc., and achieves the effects of good removal of impurities, improved production efficiency, and uniform precipitation and crossover.

Active Publication Date: 2018-05-11
GUIZHOU TAIBANG BIOLOGICAL PROD
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] CN201510748334.1 discloses "A Method for Simultaneously Preparing High-Purity Human Coagulation Factor VIII and Human Fibrinogen". The process of the present invention simultaneously extracts FVIII and Fg contained in both raw materials, which greatly improves the yield of the two products. Among them, the human coagulation factor VIII can reach 200,000 IU / ton of plasma, and the human fibrinogen exceeds 2,000 vials / ton of plasma, both of which are much higher than the traditional process. The product obtained by it has poor purity, long reconstitution time, and poor yield , causing a waste of plasma
[0005] The preparation process of Fg has also achieved large-scale production in China, but compared with foreign products, the level of product technology in my country is backward
Low purity, low thermal stability, long reconstitution time and other quality levels are obviously behind

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  • Method for preparing human fibrinogen
  • Method for preparing human fibrinogen
  • Method for preparing human fibrinogen

Examples

Experimental program
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Effect test

Embodiment 1

[0032] A preparation method of human fibrinogen, comprising the following preparation steps:

[0033] 1) Plasma melting: the plasma is completely melted at 0°C;

[0034] 2) Ethanol precipitation: adjust the pH of the reacted plasma to 7.0 with acetic acid buffer, adjust the protein concentration of the reaction solution to 5.2%, cool down to -1°C, add 95% ethanol until the final ethanol concentration of the reaction solution is 8%, and adjust the temperature to -1°C, continue to stir for more than 30 minutes after adding ethanol, and collect F by centrifugation I precipitation;

[0035] 3) First extraction: F I Dissolve the precipitate with dissolving solution 1, filter through the filter element, and obtain a clarified filtrate;

[0036] 4) Chromatography: Adsorb the clarified filtrate obtained in step 3) with Toyopeal DEAE-650M gel, the sample loading flow rate is 1.3cm / min, pass through the ultraviolet detector, and after the protein peak is detected, take the flow-throu...

Embodiment 2

[0051] A preparation method of human fibrinogen, comprising the following preparation steps:

[0052] 1) Plasma melting: the plasma is completely melted at 3°C;

[0053] 2) Ethanol precipitation: adjust the pH of the reacted plasma to 7.3 with acetic acid buffer, adjust the protein concentration of the reaction solution to 5.3%, cool down to 0°C, add 95% ethanol until the final ethanol concentration of the reaction solution is 8%, and adjust the temperature to - 2°C, continue to stir for more than 30 minutes after adding ethanol, and collect F by centrifugation I precipitation;

[0054] 3) First extraction: F I Dissolve the precipitate with solution 1, filter through the filter element, and obtain a clarified filtrate;

[0055] 4) Chromatography: Adsorb the clarified filtrate obtained in step 3) with Toyopeal DEAE-650M gel, the sample loading flow rate is 1.4cm / min, pass through the ultraviolet detector, and after the protein peak is detected, take the flow-through protein ...

Embodiment 3

[0070] A preparation method of human fibrinogen, comprising the following preparation steps:

[0071] 1) Plasma melting: the plasma is completely melted at 0-6°C;

[0072] 2) Ethanol precipitation: adjust the pH of the reacted plasma to 7.5 with acetic acid buffer, adjust the protein concentration of the reaction solution to 5.5%, cool down to 1°C, add 95% ethanol until the final ethanol concentration of the reaction solution is 8%, and adjust the temperature to - 3°C, continue to stir for more than 30 minutes after adding ethanol, and collect F by centrifugation I precipitation;

[0073] 3) First extraction: F I Dissolve the precipitate with dissolving solution 1, and filter through the filter element to obtain a clarified filtrate;

[0074] 4) Chromatography: Adsorb the clarified filtrate obtained in step 3) with Toyopeal DEAE-650M gel, the sample loading flow rate is 1.5cm / min, pass through the ultraviolet detector, and after the protein peak is detected, take the flow-t...

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Abstract

The invention belongs to the technical field of biopharmaceuticals and blood products and particularly relates to a method for preparing human fibrinogen. According to the method, optimal conditions are obtained through optimizing ethanol precipitation and first-time extracting conditions, optimal dissolving conditions are obtained through screening a dissolving solution 1, thus, impurities can bebetter removed during subsequent adsorption, the Fg purity and yield of running-through liquid are relatively high and reach up to 96.3%. During glycine and sodium chloride precipitation, glycine andsodium chloride of different concentrations are adopted during twice precipitation, and dissolving is carried out at different temperatures, and thus, the yield of the obtained Fg is relatively high;further carrying out a freeze-drying protectant formula and a freeze-drying process, obtained freeze-dried products are uniform and loose in appearance and are free of macrocrystals, redissolution time of the products is 5 to 6 minutes, solutions are clarified and transparent and are free of visible foreign matters after redissolution, and the moisture content of the products is in line with regulations of pharmacopoeia.

Description

technical field [0001] The invention belongs to the technical field of biopharmaceuticals and blood products, and in particular relates to a preparation method of human fibrinogen. Background technique [0002] Human fibrinogen (Human Fibrinogen, Fg) (referred to as fibrin), also known as human coagulation factor Ⅰ (Human coagulation factor Ⅰ, FI), is mainly synthesized by liver parenchymal cells and is one of the main components of plasma proteins. Abundant, about 2 ~ 4g / L in normal human plasma, is one of the "central" proteins in the blood coagulation system, is the final substrate of the successive activation of coagulation factors in the blood coagulation process, has hemostatic function, except directly participates in the late stage of the blood coagulation process , also mediates platelet aggregation and affects blood viscosity. [0003] The methods for extracting and purifying fibrils include low-temperature ethanol precipitation and glycine precipitation. The two ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/75C07K1/36C07K1/34C07K1/30C07K1/16C07K1/14
CPCC07K14/75
Inventor 陈云华陈艾军向宏王勇符平松赵学梅刘欣晏杨刚
Owner GUIZHOU TAIBANG BIOLOGICAL PROD
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