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Method for producing psicose 3-epimerase through high-density fermentation

An epimerase, high-density fermentation technology, applied in the direction of racemase/epimerase, microbial-based methods, isomerase, etc., can solve the problem of psicose 3-epimerase Problems such as low expression, low purity, and high cost have achieved a wide range of industrial application prospects, separation and purification savings, and the effect of reducing burdens

Active Publication Date: 2022-05-13
HENAN ZHONGDA HENGYUAN BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] In order to overcome the deficiencies and shortcomings of the fermentation production of psicose 3-epimerase in the prior art, such as low expression, low purity, and high cost, the object of the present invention is to provide a high-density fermentation production of psicose 3- The method of epimerase, which greatly improves the fermentation density of bacteria, improves the expression and purity of psicose 3-epimerase, reduces production costs, and has important industrial application value

Method used

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  • Method for producing psicose 3-epimerase through high-density fermentation
  • Method for producing psicose 3-epimerase through high-density fermentation
  • Method for producing psicose 3-epimerase through high-density fermentation

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0057] Embodiment 1 High-density fermentation (using peptone and yeast powder as nitrogen source in fermentation medium and feeding medium)

[0058] (1) Culture medium preparation

[0059] A. Activation medium: The composition of the activation medium is peptone 10g / L, yeast powder 5g / L, sodium chloride 10g / L, after sterilizing at 121°C for 20min, add kanamycin to 50mg / L under sterile conditions.

[0060] B. Fermentation medium: The composition of the fermentation medium is peptone 10g / L, yeast powder 5g / L, potassium dihydrogen phosphate 2.5g / L, dipotassium hydrogen phosphate 15g / L, manganese chloride tetrahydrate 0.1g / L, glucose 6g / L, sterilized at 121°C for 20min.

[0061] C. Feed medium: the composition of the feed medium is 15% by mass of glucose and 20% by mass of yeast powder, and it is sterilized at 121° C. for 20 minutes.

[0062] (2) Activation of bacteria

[0063] Activation of primary strains: Take 100 μL of the preserved bacterial solution of recombinant Bacillu...

Embodiment 2

[0068] Embodiment 2 High-density fermentation (using peptone and yeast powder as nitrogen source in fermentation medium and feeding medium)

[0069] (1) Culture medium preparation

[0070] A. Activation medium: The composition of the activation medium is peptone 5g / L, yeast powder 1g / L, sodium chloride 8g / L, after sterilizing at 121°C for 20min, add kanamycin to 25mg / L under sterile conditions.

[0071] B. Fermentation medium: The composition of the fermentation medium is peptone 5g / L, yeast powder 1g / L, potassium dihydrogen phosphate 0.5g / L, dipotassium hydrogen phosphate 5g / L, manganese chloride tetrahydrate 0.02g / L, glucose 2g / L, sterilized at 121°C for 20min.

[0072] C. Feed medium: the composition of the feed medium is 10% by mass of glucose and 10% by mass of yeast powder, and it is sterilized at 121° C. for 20 minutes.

[0073] (2) Activation of bacteria

[0074] Activation of primary strains: Take 50 μL of the preserved bacterial liquid of recombinant Bacillus subt...

Embodiment 3

[0079] Embodiment 3 High-density fermentation (using peptone and yeast powder as nitrogen source in fermentation medium and feeding medium)

[0080] (1) Culture medium preparation

[0081]A. Activation medium: The composition of the activation medium is peptone 15g / L, yeast powder 5g / L, sodium chloride 10g / L, after sterilizing at 121°C for 20min, add kanamycin to 50mg / L under sterile conditions.

[0082] B. Fermentation medium: The composition of the fermentation medium is peptone 15g / L, yeast powder 5g / L, potassium dihydrogen phosphate 5g / L, dipotassium hydrogen phosphate 20g / L, manganese chloride tetrahydrate 0.2g / L, glucose 10g / L, sterilized at 121°C for 20min.

[0083] C. Feed medium: the composition of the feed medium is 30% by mass of glucose and 30% by mass of yeast powder, and it is sterilized at 121° C. for 20 minutes.

[0084] (2) Activation of bacteria

[0085] Activation of primary strains: Take 500 μL of the preserved bacterial solution of recombinant Bacillus...

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Abstract

The invention belongs to the technical field of microbial fermentation engineering, and particularly relates to a method for producing psicose 3-epimerase through high-density fermentation. Aiming at the problems of low expression quantity of psicose 3-epimerase in the current fermentation and the like, the fermentation OD value of the recombinant bacillus subtilis and the total enzyme activity of the psicose 3-epimerase in the fermentation liquor are remarkably improved by controlling the feeding speed in the fermentation process, increasing the culture temperature in the middle and later periods of the fermentation and the like; the expression quantity of the psicose 3-epimerase is remarkably increased, the production cost of psicose is reduced, and the psicose 3-epimerase has an extremely wide industrial application prospect.

Description

technical field [0001] The invention belongs to the technical field of microbial fermentation engineering, and in particular relates to a method for producing psicose 3-epimerase by high-density fermentation. Background technique [0002] D-psicose is a rare sugar with 70% of the sweetness of sucrose, but very low in calories and is a good substitute for sucrose. Moreover, the absorption rate of D-psicose is lower than that of other sweeteners, which can reduce the body's absorption of fructose and glucose, reduce the accumulation of fat, and thus reduce the risk of diseases such as type 2 diabetes and obesity. At present, studies have found that D-psicose also has the function of lowering blood lipids and blood sugar. The U.S. Food and Drug Administration (FDA) has certified D-psicose as GRAS (Generally recognized as safe), which can be used as a component of food additives. D-psicose has broad application prospects in food and health products. [0003] Allulose 3-epimer...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/90C12N1/21C12P19/24C12P19/02C12R1/125
CPCC12N9/90C12N1/20C12P19/24C12P19/02C12Y501/03C12N15/75C12N15/52C12R2001/125C12N1/205C12N2500/02C12N2500/34C12N2500/74C12N2523/00
Inventor 王三营李林正金子恒文雁君
Owner HENAN ZHONGDA HENGYUAN BIOTECH CO LTD
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