Escherichia coli immobilizing method and method for producing L-lysine through fed-batch fermentation of immobilized escherichia coli

A technology of Escherichia coli, fed-feed fermentation, applied in microorganism-based methods, biochemical equipment and methods, immobilized on/in organic carriers, etc., can solve the problems of small force and easy shedding, Escherichia coli restriction, etc., Achieve high production efficiency, stable fermentation, and reduce production costs.

Active Publication Date: 2017-12-08
NANJING TECH UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This method is simple to operate and low in cost, but the number of Escherichia coli immobilized is limited by the type of carrier used and its surface area
Because the weak force between Escherichia coli and the carrier material is easy to fall off, few people use the adsorption method as a fixation method for Escherichia coli

Method used

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  • Escherichia coli immobilizing method and method for producing L-lysine through fed-batch fermentation of immobilized escherichia coli
  • Escherichia coli immobilizing method and method for producing L-lysine through fed-batch fermentation of immobilized escherichia coli
  • Escherichia coli immobilizing method and method for producing L-lysine through fed-batch fermentation of immobilized escherichia coli

Examples

Experimental program
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Effect test

Embodiment 1

[0039] (1) Cut the synthetic sponge into 1*1*1cm 3 The small cubes were soaked in 75% ethanol aqueous solution for 1 to 3 hours, washed in deionized water for 3 to 5 times, and then added to a 500ml Erlenmeyer flask according to the carrier addition amount of 10g / L. The Erlenmeyer flask was equipped with pre- Fermentation medium 50ml, then add MOPS of 6 different concentrations of 0, 50, 75, 100, 125, 150mM respectively, at least 3 parallel samples for each concentration, sterilize by high temperature and high pressure steam at 115°C for 15min;

[0040] Pre-fermentation medium formula: glucose 20g / L, peptone 1.5g / L, ammonium sulfate 0.5g / L, dipotassium hydrogen phosphate 0.5g / L, yeast extract 1g / L, manganese sulfate 0.2g / L, ferrous sulfate 0.4g / L, magnesium sulfate 0.1g / L, solvent is water, pH6.8~7.2.

[0041] (2) Wash the L-lysine producing strain Escherichia coli preserved on the slant with sterile physiological saline, and insert the bacterium suspension into the sterilize...

Embodiment 2

[0048] (1) Cut the synthetic sponge into 1*1*1cm 3 Small cubes were soaked in 75% ethanol aqueous solution for 1 to 3 hours, washed in deionized water for 3 to 5 times, and then added to the In the shake flask pre-fermentation medium (50ml seed medium / 500ml Erlenmeyer flask), at least 3 parallel samples are added for each carrier, sterilized by high temperature and high pressure steam at 115°C for 15min; the formula of the shake flask pre-fermentation medium is the same as in the example 1.

[0049] (2) The activation culture process of Escherichia coli is the same as step (2) in Example 1.

[0050] (3) Insert the inoculum amount of activated Escherichia coli into the shake flask pre-fermentation medium of step (1) according to the volume ratio of 10%, the culture temperature is 37°C, the shaking table speed is 200r / min, and the culture time is 20 ~24h. At the end of the cultivation, use the SBA-40E biosensor to detect the amount of L-lysine. For the test results, see fig...

Embodiment 3

[0052] (1) Shake flask seed culture: wash the L-lysine producing strain Escherichia coli preserved on the slant with sterile physiological saline, and insert the bacterial suspension into a 500ml triangle containing 50ml seed medium according to the inoculum size of 5% by volume Cultivate in the bottle, the culture temperature is 35°C, the shaker speed is 190r / min, the culture time is 20-24h, the formula of the shake flask seed medium is the same as

[0053] Example 1.

[0054] (2) Pre-fermentation culture: when the OD600 of the culture solution in the shake flask reaches 5, the inoculation amount of 5% by volume is inserted into a pre-fermentation tank that has sterilized bacteria in advance and is equipped with a pre-fermentation medium, and the ventilation rate is controlled during the pre-fermentation process 0.03~0.08L / (min L), stirring speed 100~1000rpm, adjust ventilation volume and stirring speed to control dissolved oxygen DO≥25%, temperature 35℃, ammonia water contro...

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Abstract

The invention discloses an escherichia coli immobilizing method which comprises the following steps: feeding the activated escherichia coli into a container with a culture medium, a pretreated porous mesh material and MOPS, and performing immobilizing culture so as to adsorb the escherichia coli on the porous mesh material in the immobilizing culture process. The invention also discloses a method for producing L-lysine through fed-batch fermentation of immobilized escherichia coli, which comprises a fermentation process and a fed-batch process, wherein the nutritional ingredients adopted in the fed-batch process are glucose and ammonium sulfate. The MOPS can enhance the adhesion effect of escherichia coli on the surface of the immobilizing material; the L-lysine is fermented by use of the immobilized escherichia coli; the L-lysine concentration is increased to 136g / L from 90g / L of free cell fermentation; by repeating the fermentation by 8 batches, higher production efficiency of L-lysine still can be maintained, and the final L-lysine yield of each batch is 150g / L on average.

Description

technical field [0001] The invention relates to fermentative production of L-lysine by using immobilized bacteria, in particular to an immobilization method of Escherichia coli and a method for producing L-lysine by fed-batch fermentation of immobilized Escherichia coli. Background technique [0002] L-lysine (Lysine) is an aliphatic basic amino acid, chemical name is 2,6-diaminocaproic acid, there are L-type (left-handed), D-type (dextral) and DL-type (racemic ) Three spin isomers, only the L-form can be absorbed and utilized by humans and animals. L-lysine is one of the eight essential amino acids required for the growth and development of humans and animals. It is important for regulating the metabolic balance in the body, improving the absorption of cereal protein in the body, improving human dietary nutrition and animal nutrition, and promoting growth and development. effect. [0003] L-Lysine was initially isolated from protein hydrolyzate. The protein hydrolysis met...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N11/02C12N11/08C12N11/12C12P13/08C12N1/20C12R1/19
CPCC12N1/20C12N11/02C12N11/08C12N11/12C12P13/08
Inventor 应汉杰张欣陈勇余斌赵南
Owner NANJING TECH UNIV
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