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Method for separating and purifying serum albumin by using liquid-solid extraction system

A serum albumin, separation and purification technology, which is applied in the field of liquid-solid extraction system to purify serum albumin from human plasma or bovine plasma, can solve the problems of low product purity, difficult automation, and great harm to staff. To achieve the effect of clear phase system, easy to scale up and low cost

Inactive Publication Date: 2012-06-13
SOUTH CENTRAL UNIVERSITY FOR NATIONALITIES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although widely used in industrial production, the ethanol precipitation method also has some shortcomings: ethanol is a non-specific precipitant, although it has a significant effect on the production of human serum albumin HSA, the product purity is not very high; ethanol as an organic solvent may also make Aggregation and denaturation of proteins or changes in secondary conformation; production is not easy to automate, and the production cycle is long; organic solvents are also more harmful to workers in a non-closed environment, and may also cause product contamination due to unclean environment
In recent years, many laboratories and companies have successively tried to obtain HSA through genetic engineering methods, but they also encountered the same separation and purification problems

Method used

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  • Method for separating and purifying serum albumin by using liquid-solid extraction system
  • Method for separating and purifying serum albumin by using liquid-solid extraction system
  • Method for separating and purifying serum albumin by using liquid-solid extraction system

Examples

Experimental program
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Embodiment 1

[0058] Embodiment one: carbon sixteen long-chain dibasic fatty acid polyethylene glycol (C 16 Separation and Purification of HSA by PEG Modified Mixed Tween 80-Potassium Phosphate-Water-Liquid-Solid Extraction System

[0059] 1. Extraction method

[0060] Take 2.0 ml HSA solution, 0.4 g C 16 PEG modification, 0.8 ml of Tween 80 stock solution, 6.3 g of potassium phosphate solution in a colorimetric tube, pH in the range of 6.5-9.5, add water to make up to 10.0 mL, oscillate in an electric shaker for 5 minutes, mix well, After standing still, the system is divided into a polymer solid phase and a brine phase, pour the brine phase completely, and use distilled water to fix the volume of the polymer solid phase to 10.0mL, so that the acidity of the system is in the range of pH 3-6, and add 6.0 grams of phosphoric acid Potassium, placed on an electric shaker and vibrated for 5 minutes, removed and inverted for 20 minutes, the system was re-divided into a polymer solid phase and ...

Embodiment 2 16

[0070] Example 2C 16 Separation and Purification of HSA by PEG Modified Mixed Tween 80-Sodium Citrate-Water-Liquid-Solid Extraction System

[0071] 1. Extraction method

[0072] Take 2.0 ml HSA solution, 0.6 g C 16PEG modification, 0.75 ml of Tween 80 stock solution, 2.8 g of sodium citrate solution in a colorimetric tube, pH in the range of 4.1-6.2, add water to make up to 10.0 mL, shake in an electric shaker for 10 minutes, mix well , after standing for 10 minutes, the system was divided into polymer solid phase and brine phase, and the brine phase was completely poured out, and the polymer solid phase was fixed to 10.0 mL with distilled water, so that the acidity of the system was in the range of pH 3.0-5.2, and 2.4 gram of sodium citrate, placed on an electric shaker and vibrated for 10 minutes, removed and inverted for 10 minutes, the system was re-divided into a polymer solid phase and a saline liquid phase, and serum albumin was back-extracted; then the serum extracte...

Embodiment 3 12

[0075] Example 3C 12 Separation and Purification of HSA by PEG Modified Mixture 80-Potassium Carbonate Salt-Water-Liquid-Solid Extraction System

[0076] Take 2.0 ml HSA solution, 0.55 g C 12 PEG modification, 0.9 ml of Tween 80 stock solution, 1.7 g of potassium carbonate salt solution in a colorimetric tube, pH 7.0-8.5, add water to make up to 10.0 mL, shake in an electric shaker for 2 minutes, mix evenly, and After standing for 15 minutes, the system is divided into a polymer solid phase and a brine phase, and the brine phase is completely poured out, and the polymer solid phase is fixed to a volume of 10.0 mL with distilled water, so that the acidity of the system is within the pH range of 3-6, and 1.7 g of carbonic acid is added Potassium salt, placed on an electric shaker and vibrated for 2 minutes, removed and inverted for 15 minutes, the system was re-divided into a polymer solid phase and a saline liquid phase, and serum albumin was back-extracted into the saline pha...

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Abstract

The invention relates to a method for separating and purifying serum albumin by using a liquid-solid extraction system. The method comprises the following steps of: mixing a long-chain aliphatic dibasic acid polyethylene glycol modifier and Tween-80 polymer and inorganic salt to form a liquid-system extraction system and regulating a pH value of the system; adding a blood sample into the liquid-solid extraction system, horizontally placing the liquid-solid extraction system on an electric shaking machine for shaking, taking off the liquid-solid extraction system and placing the liquid-solid extraction system upside down, wherein the system is divided into a polymer solid phase and a salt solution liquid phase; and the serum albumin is extracted into the solid phase; after the solid phase is dissolved in distilled water, regulating the pH value and adding the inorganic salt into the system, horizontally placing the system on the electric shaking machine for shaking, and taking off the system and placing the system upside down, wherein the system is re-divided into the polymer solid phase and the salt solution liquid phase; and the serum albumin is back-extracted into the liquid phase; performing dialysis on the serum albumin which is extracted into the liquid phase in a dialysis bag; and freeze-drying the serum albumin which remains in the dialysis bag to obtain serum albumin dry powder. Due to the adoption of the method, the unpolymerized serum albumin which has high purity and unchanged structure is obtained by purifying, and steps such as pretreatment, crude separation and fine separation which are always performed on plasma are finished at one time.

Description

technical field [0001] The invention relates to a method for separating and purifying serum albumin from blood by using a liquid-solid extraction system, more particularly to a method for purifying serum albumin from human plasma or bovine plasma by using a liquid-solid extraction system. More specifically, serum albumin is extracted from blood by molecular recognition mechanism, using polyethylene glycol (PEG) dibasic fatty acid modified mixed polymer-salt-water-liquid-solid affinity extraction method. Background technique [0002] Serum albumin is a highly water-soluble protein with a stable structure and can bind a variety of small molecules. It has a wide range of applications in clinical medicine and biology. The separation and purification principle of protein is mainly based on the difference in solubility, charge difference, molecular size, hydrophobic interaction, and biological affinity difference to achieve separation. As a biological molecule, the separation and...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/765C07K1/14
Inventor 沈静茹雷灼霖刘波
Owner SOUTH CENTRAL UNIVERSITY FOR NATIONALITIES