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Method for extracting and purifying polyglutamic acid

A technology of polyglutamic acid and polyglutamic acid serum, applied in the biological field, can solve the problems of low product purity, high cost, heavy metal pollution, etc., and achieve the effect of improving product quality

Inactive Publication Date: 2019-10-25
SYNCOZYMES SHANGHAI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In order to overcome the problems of large amount of organic solvent, high cost, heavy metal pollution and low product purity in the existing production technology, the present invention provides a new method for extracting and purifying polyglutamic acid without alcohol, low cost and high product purity. The method is characterized in that in the presence of ammonium sulfate, the extraction and purification of polyglutamic acid are realized by adjusting the pH value for fractional precipitation

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Embodiment 1 The preparation of the Bacillus subtilis fermented liquid containing polyglutamic acid

[0020] Strain selection: Bacillus subtilis, isolated from natto food.

[0021] Primary seed culture: Inoculate 50 μl of strains stored in glycerol tubes into 50 ml of seed medium, shake and cultivate in a constant temperature shaker at 37°C with a rotation speed of 160 rpm, and cultivate for 15 hours as the primary seed solution.

[0022] Secondary seed culture: Inoculate the primary seed liquid into 400ml of seed culture medium at 5% inoculum amount, shake and cultivate in a constant temperature shaker at 37°C with a rotation speed of 160rpm, and cultivate for 15h as the secondary seed liquid.

[0023] Fermentation culture: inoculate the secondary seed liquid into the fermentation medium in the fermenter according to the inoculum amount of 5%, and stir and cultivate. The fermentation temperature was 37°C, the initial fermentation pH was 7.5, the ventilation rate was 1...

Embodiment 2

[0027] Extraction and purification of embodiment 2 polyglutamic acid

[0028] Get 5L of the fermented liquid fermented by Bacillus subtilis, add 2.5kg of ammonium sulfate while stirring, and adjust the pH of the fermented liquid to be 6.0. After the ammonium sulfate is completely dissolved, let it stand at room temperature for 30 minutes. The resulting mixture was centrifuged at 4000 g for 5 min with a desktop centrifuge, the supernatant was taken, and the precipitate was discarded. While stirring the supernatant, adjust the pH to 2.5 with 6M sulfuric acid solution, let it stand for 30 minutes, pour off the supernatant, and collect the precipitate. Add 5 L of pure water to the precipitate, stir to redissolve, adjust the pH to 7.2 with 1M NaOH solution, and stir for 1 h. The resulting polyglutamic acid complex solution was ultrafiltered with a 6KDa polyethersulfone roll-type ultrafiltration membrane, and when the remaining 2.5L of the retentate was added, 2.5L of pure water w...

Embodiment 3

[0029] Extraction and purification of embodiment 3 polyglutamic acid

[0030] Get 100L of the fermented liquid fermented by Bacillus subtilis, add 30kg of ammonium sulfate while stirring, and adjust the pH of the fermented liquid to be 8.0. After the ammonium sulfate is completely dissolved, let it stand at room temperature for 30 minutes. The resulting mixed solution was filtered with a plate and frame filter press. The plate and frame filter press is first pre-coated with 500g of 400 mesh diatomite, and the filter cake is formed. When the pressure is 0.02MPa, the above mixture is subjected to plate and frame filter press, and the pressure is controlled at 0.05-0.2MPa. Finally, 10L of 30% The ammonium sulfate solution was used to press out the residual polyglutamic acid, and the filtrate was collected. While stirring, adjust the pH to 3.5 with 6M sulfuric acid solution, let it stand for 30 minutes, pour off the supernatant, and collect the precipitate. Add 100L of pure wat...

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Abstract

The invention discloses a method for extracting and purifying polyglutamic acid by adjusting the pH value to carry out fractional precipitation in the presence of ammonium sulfate. The method comprises the following steps: adding 30-60% (W / V) of ammonium sulfate into a fermentation liquid containing polyglutamic acid, adjusting the pH value to 6.0-8.0, removing the precipitate after stirring, andcollecting the glutamic acid clear solution; adjusting the pH value of the polyglutamic acid clear solution to be acidic, and collecting the glutamic acid precipitate; then re-dissolving the polyglutamic acid precipitate with water, adjusting the pH value to be alkalescent by using an alkali, carrying out ultrafiltration until conductivity of the permeate liquid is smaller than 5 ms*cm, and collecting the retained liquid; and adding activated carbon into the retained liquid for decolorization to obtain a colorless clear liquid, and carrying out drying to obtain a white polyglutamic acid solid.According to the method, polyglutamic acid can be effectively separated with other impurities, and use of organic solvents such as ethanol and the like is avoided. The purity of the obtained polyglutamic acid is high, and production cost is effectively reduced.

Description

Technical field: [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for extracting and purifying polyglutamic acid. Background technique: [0002] γ-polyglutamic acid is a water-soluble extracellular glutamic acid polymer produced by microbial fermentation [γ-polyglutamic acid, referred to as γ-PGA], generally referred to as polyglutamic acid in Chinese, and its relative molecular weight is generally between 100-2000KDa . Depending on the molecular weight, its application fields are also different, such as in soil modification, water flocculation, cosmetics, medicine, etc. Generally, the larger the molecular weight, the higher the viscosity. Due to the high viscosity of polyglutamic acid, separation and purification are extremely difficult. The purification methods reported so far mainly include organic solvent precipitation method, heavy metal precipitation method, alcohol-free purification method, etc. The organic solvent p...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C08G69/10C08G69/46
CPCC08G69/10C08G69/46
Inventor 张文明竺伟
Owner SYNCOZYMES SHANGHAI
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