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Method for separating and purifying human serum albumin from Cohn component V supernatant

A technology for separation and purification of human serum albumin, applied in the preparation methods of serum albumin, albumin peptides, peptides, etc., can solve the problems of long operation time and large amount of buffer, and achieves low cost, reduced dosage, and improved The effect of plasma utilization

Pending Publication Date: 2019-04-16
INST OF PROCESS ENG CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, this method consumes a large amount of buffer and takes a long time to operate

Method used

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  • Method for separating and purifying human serum albumin from Cohn component V supernatant
  • Method for separating and purifying human serum albumin from Cohn component V supernatant
  • Method for separating and purifying human serum albumin from Cohn component V supernatant

Examples

Experimental program
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Effect test

Embodiment 1

[0049] This embodiment separates and purifies human serum albumin by the following method

[0050] Equilibrate the anion-exchange chromatography medium DEAE Sepharose Fast Flow with 20mM phosphate buffer with a pH value of 7.0, adjust the pH value of the Cohn component V supernatant to 7.0, and load it into the chromatography column. Rinse 3 CVs with phosphate buffered saline;

[0051] Use a pH value of 4.0 and a concentration of 20 mM citric acid-sodium citrate buffer to elute the chromatography column, collect the eluate, and concentrate the obtained elution peak by ultrafiltration with an ultrafiltration membrane with a molecular weight cut-off of 10 kDa to obtain human serum Albumin; the chromatography medium was finally washed with 20 mM citric acid-sodium citrate buffer, pH 4.0, containing 0.5 M NaCl.

[0052] The human serum albumin that embodiment 1 obtains is carried out 12%SDS-PAGE detects, and specific result is as follows figure 1 As shown, the result shows a sin...

Embodiment 2

[0055] This embodiment separates and purifies human serum albumin by the following method

[0056] Equilibrate the anion-exchange chromatography medium Q Sepharose Fast Flow with 20mM phosphate buffer with a pH value of 7.5, adjust the pH value of the Cohn component V supernatant to 7.5, and load it into the chromatography column. Rinse 5 CVs with phosphate buffered saline;

[0057] Use a pH value of 4.5 and a concentration of 50 mM citric acid-sodium citrate buffer to elute the chromatography column, collect the eluate, and concentrate the obtained elution peak by ultrafiltration with an ultrafiltration membrane with a molecular weight cut-off of 15 kDa to obtain human serum Albumin; the chromatography medium was finally washed with 50 mM citric acid-sodium citrate buffer, pH 4.5, containing 1 M sodium chloride.

Embodiment 3

[0059] This embodiment separates and purifies human serum albumin by the following method

[0060] Equilibrate the anion-exchange chromatography medium Capto DEAE with 20mM phosphate buffer with a pH value of 6.0, adjust the pH value of the Cohn component V supernatant to 6.0, and load it into the chromatography column. Wash with salt buffer for 3 CVs;

[0061] Use a pH value of 4.0 and a concentration of 20 mM citric acid-sodium citrate buffer to elute the chromatography column, collect the eluate, and concentrate the obtained elution peak by ultrafiltration with an ultrafiltration membrane with a molecular weight cut-off of 10 kDa to obtain human serum Albumin; the chromatography medium was finally washed with 30 mM citric acid-sodium citrate buffer, pH 4.0, containing 0.2 M NaCl.

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Abstract

The invention provides a method for separating and purifying human serum albumin from a Cohn component V supernatant, which comprises the following steps of: loading a supernatant of the Cohn component V into a chromatographic column containing an anion exchange chromatographic medium, eluting the chromatographic column with eluent, collecting the eluent to obtain human serum albumin; according tothe method, a novel idea of separating and purifying albumin by using the low-temperature ethanol precipitation component V supernatant as a raw material is put forward for the first time, ethanol isnot removed by adopting an ultrafiltration process, so that the using amount of a buffer solution is greatly reduced, the operation time is reduced, the method avoids the problem of denaturation andinactivation of the protein in a higher ethanol solution for a long time, and has the advantages of simple operation, low cost, short process time consumption, high protein activity yield and the like. The method can further recover the albumin in the supernatant of the component V, effectively improve the utilization rate of the plasma and reduce the cost, and has important value for practical application.

Description

technical field [0001] The invention belongs to the field of biotechnology and relates to a method for separating and purifying human serum albumin from Cohn component V supernatant. Background technique [0002] Human serum albumin (Human Serum Albumin, HSA) is the most abundant protein in plasma, accounting for about 60% of the total plasma protein content. In the human body, the main physiological function of serum albumin is to maintain blood osmotic pressure and carry a variety of ligands (including fatty acids, amino acids, steroids, metal ions and drugs) in the blood to exchange with tissues. It is clinically used for surgical blood transfusion and critically ill patients Rehydration, treatment of traumatic shock, fever, edema and hypoalbuminemia, etc., and can enhance the body's resistance, it is currently the most clinically used medicinal protein. [0003] At present, the preparation of human serum albumin mainly adopts the low-temperature ethanol precipitation pr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/765C07K1/18
CPCC07K14/765
Inventor 罗坚苏志国陈颖向杰
Owner INST OF PROCESS ENG CHINESE ACAD OF SCI
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