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Method for producing polysialic acid by using fermentation process, and extraction and refining method for polysialic acid

A technology of polysialic acid and fermentation method, which is applied in the field of bioengineering to achieve the effects of saving costs, reducing energy consumption, and reducing production costs

Pending Publication Date: 2020-10-02
HIERAND BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

But using this method, the highest yield of poly-N-acetylneuraminic acid can only reach 6g / L

Method used

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  • Method for producing polysialic acid by using fermentation process, and extraction and refining method for polysialic acid
  • Method for producing polysialic acid by using fermentation process, and extraction and refining method for polysialic acid
  • Method for producing polysialic acid by using fermentation process, and extraction and refining method for polysialic acid

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Embodiment 1

[0034] The fermentation strain is Escherichia coli SA-8 screened by mutagenesis in the Industrial Microbiology Laboratory of the Institute of Microbiology, Chinese Academy of Sciences, and its patent preservation number is CGMCC NO.5585.

[0035] A method for producing polysialic acid by fermentation, comprising the following steps:

[0036](1) Escherichia coli SA-8 with a volume percentage of 2% was inoculated in the basal medium of the seed tank, and the seed culture liquid was obtained by fermentation culture, and the seed tank was fermented in 1000 liters. The basal medium used is glucose 25g / L, ammonium sulfate 5g / L, casein peptone 15g / L, dipotassium hydrogen phosphate 20g / L, magnesium sulfate 0.4g / L, after mixing evenly, steam sterilization. After reaching 121°C, maintain the tank pressure at 0.09MPa and sterilize for 30 minutes. The sterilization process requires continuous stirring at 200rpm. After cooling, inoculate 9 liters of bacterial cell culture solution and in...

Embodiment 2

[0041] The fermentation broth obtained in Example 1 was used. Carry out the extraction and refinement of polysialic acid according to the following steps:

[0042] (1) Separation of bacterial liquid: 200nm ceramic membrane removes bacterial cells, and the clear liquid first passes through a weakly basic anion exchange resin to change ammonium polysialate into sodium polysialate;

[0043] (2) Removal of small molecular impurities: the ion-exchanged sodium polysialate solution is ultrafiltered with a 70,000 Dalton membrane system to concentrate macromolecules; and the concentrated sodium polysialate solution is washed with pure water to a pH below 7.2, and the filtered Discard the waste liquid and keep the concentrate.

[0044] (3) Ethanol precipitation of PSA: Add three times the volume of absolute ethanol to the concentrated and washed polysialic acid sodium solution, let it stand and cool down to below 8°C, and after maintaining it for 1 hour, collect the polysialic acid pre...

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Abstract

The invention discloses a method for producing polysialic acid by using a fermentation process, and an extraction and refining method for polysialic acid. The method comprises the following steps: inoculating an escherichia coli strain SA-8 (CGMCC No. 5585) into a basic culture medium; transferring seeds into final fermentation; after entering a logarithmic phase, introducing a mixture of pure oxygen and air for ventilation; and after fermentation for 10 hours, gradually adjusting and lowering a pH value and a temperature, adjusting the pH value to 8.0 or above with Ca(OH)2 in the later periodof fermentation, stopping stirring and cooling within 18-19 hours, only continuously introducing common air for 1.8-2.2 hours, and finally conducting fermenting to obtain the polysialic acid which can be purified with anion exchange resin. The method is simple in process, saved in cost and reduced in energy consumption; the obtained polysialic acid is high in purity, does not contain endotoxin, can meet the production requirement that the polysialic acid is further hydrolyzed into monomer sialic acid (N-acetylneuraminic acid), and is applicable as a raw material for producing food, cosmeticsand medicines.

Description

technical field [0001] The invention relates to the technical field of bioengineering, in particular to a fermentation method for producing polysialic acid and its extraction and refining method. Background technique [0002] There have been many studies and attempts on the production of polysialic acid (PSA) by microbial fermentation. It can be done by using the natural strain of Escherichia coli. Polysialic acid itself has various uses such as drug sustained-release agent, immune adjuvant, and drug precursor. Moreover, further hydrolysis can obtain monomer N-acetylneuraminic acid (Neu5Ac), which is an important food and pharmaceutical raw material. [0003] The production of polysialic acid by fermentation has been reported in the literature. Patent CN 103361283 discloses a method for producing poly-N-acetylneuraminic acid by fermentation of glucose using strain CGMCC No.5585. But using this method, the highest yield of poly-N-acetylneuraminic acid can only reach 6g / L. ...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12P19/26C08B37/00C12R1/19
CPCC12N1/20C12P19/26C08B37/0003
Inventor 刘晓鹏董黎爱李海涛
Owner HIERAND BIOTECH CO LTD
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