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Fermentation preparation method, and separation and purification method of heat-resistant and acid-resistant glucose oxidase

A glucose oxidase and heat-resistant technology, applied in the field of microorganisms, can solve the problems of difficult promotion and use of glucose oxidase, limitation of the scope of use of glucose oxidase, and relatively little research on glucose oxidase, so as to save manpower and financial resources and effectively Facilitate transportation, easy extraction and separation

Active Publication Date: 2017-07-07
CHONGQING FULING ZIZHU FOOD CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] A patent application number 201410228211.0 published by the State Intellectual Property Office on August 6, 2014, titled: Preparation method of glucose oxidase preparation and its utilization, this patent proposes the following technical scheme: inoculate Penicillium inoculated into , nitrogen source, inorganic salt solution and trace elements of the fermentation medium in the fermenter are aerated and fermented to make a fermented liquid, and the fermented liquid is separated into solid and liquid to make a glucose oxidase preparation. The glucose oxidase prepared by this strain has the following technology Problem: First, the glucose oxidase produced by the fermentation of this strain has poor thermal stability, and is easily inactivated in an acidic environment, and does not have acid resistance; second, the enzyme of glucose oxidase produced by the fermentation of this strain Low activity and low production efficiency; these two problems greatly limit the scope of use of the glucose oxidase prepared by this strain, therefore, the glucose oxidase prepared by this preparation method is difficult to promote and use in the market
[0004] At present, there have been many research reports on glucose oxidase at home and abroad, most of which are derived from microorganisms. In microorganisms, there are generally more studies on the enzyme production of mesophilic bacteria, but relatively few studies on the production of thermostable glucose oxidase by thermophilic fungi.

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  • Fermentation preparation method, and separation and purification method of heat-resistant and acid-resistant glucose oxidase
  • Fermentation preparation method, and separation and purification method of heat-resistant and acid-resistant glucose oxidase
  • Fermentation preparation method, and separation and purification method of heat-resistant and acid-resistant glucose oxidase

Examples

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Embodiment 1

[0047] A fermentation preparation method for heat-resistant and acid-resistant glucose oxidase produced by Paecilomyces thermophila, the specific steps are as follows:

[0048] (1) Streak and activate Paecilomyces thermophila species on the PDA activation medium plate, and place the PDA activation medium plate in a constant temperature incubator at 35° C. for 50 h to obtain Paecilomyces thermophila filaments;

[0049] (2) Scrape the Paecilomyces thermophila filaments prepared in step (1) with an inoculation loop and inoculate them on the Paecilomyces thermophila spore medium, and place the Paecilomyces thermophila spore medium at 35°C Cultivate for 3 days to obtain spores;

[0050] (3) The prepared spores in the step (2) are cleaned with 0.75% sterile saline containing 0.01% (v / v) Tween 80, and the cleaned spores are put into 50mL sterile saline and Place in a conical flask with sterile glass beads and fully shake for 40 minutes. After the spores are fully dispersed, prepare ...

Embodiment 2

[0063] A fermentation preparation method for heat-resistant and acid-resistant glucose oxidase produced by Paecilomyces thermophila, the specific steps are as follows:

[0064] (1) Streak and activate Paecilomyces thermophila species on the PDA activation medium plate, and place the PDA activation medium plate in a constant temperature incubator at 30°C for 48 hours of activation culture to obtain Paecilomyces thermophila filaments;

[0065] (2) Use the inoculation loop to scrape the prepared Paecilomyces thermophila filaments in step (1) and inoculate them on the Paecilomyces thermophila spore medium, and place the Paecilomyces thermophila spore medium at 30°C Under culture for 2 days, the spores are obtained;

[0066](3) The prepared spores in the step (2) are cleaned with 0.75% sterile saline containing 0.01% (v / v) Tween 80, and the cleaned spores are put into 50mL sterile saline and Place in a conical flask with sterile glass beads and fully shake for 30 minutes. After th...

Embodiment 3

[0079] A fermentation preparation method for heat-resistant and acid-resistant glucose oxidase produced by Paecilomyces thermophila, the specific steps are as follows:

[0080] (1) Streak and activate Paecilomyces thermophila species on a PDA activation medium plate, and place the PDA activation medium plate in a constant temperature incubator at 40°C for 72 hours of activation culture to obtain Paecilomyces thermophila filaments;

[0081] (2) Use the inoculation loop to scrape the prepared Paecilomyces thermophila filaments in step (1) and inoculate them on the Paecilomyces thermophila spore medium, and place the Paecilomyces thermophila spore medium at 40°C Under culture for 4 days, the spores are obtained;

[0082] (3) The prepared spores in the step (2) are cleaned with 0.75% sterile saline containing 0.01% (v / v) Tween 80, and the cleaned spores are put into 55mL sterile saline and Place in a conical flask with sterile glass beads and fully shake for 45 minutes. After the...

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Abstract

The invention discloses a fermentation preparation method, and a separation and purification method of heat-resistant and acid-resistant glucose oxidase, and belongs to the technical field of microorganisms. The glucose oxidase prepared by fermentation has high heat resistance and acid resistance, and the defect of low enzyme activity caused by use in food and feed in a high temperature and acidic environment is overcome. The fermentation preparation method comprises the following specific steps: culturing a bacterial strain into a spore, cleaning the spore and then preparing a mono-spore suspension; introducing the mono-spore suspension into a seed culture medium for fermentation culture; introducing seed culture fluid into a shaking flask fermentation medium for fermentation culture, adding a fermentation supplementary nutrient solution into fermentation liquid; adding dextrin, soluble starch and potassium sorbate into the fermentation liquid, and uniformly stirring; freezing, drying and crushing the fermentation liquid to obtain the glucose oxidase. After crude enzyme fluid of the glucose oxidase obtained by the fermentation is subjected to super-heating treatment, ammonium sulfate-ethanol-compound precipitation, ion exchange chromatography and molecular sieve gel filtration chromatography, the glucose oxidase with high recovery rate and high purity is obtained.

Description

technical field [0001] The invention relates to the technical field of microorganisms, in particular to a fermentation preparation and separation and purification method of heat-resistant and acid-resistant glucose oxidase. Background technique [0002] Glucose oxidase is a very important enzyme in the field of enzyme technology. It can use molecular oxygen as an electron acceptor to specifically catalyze the oxidation of β-D-glucose into gluconic acid and hydrogen peroxide. Because of its close relationship with the vital substances glucose and oxygen, it has been widely used in scientific research, medicine and food industry production. In the food industry, glucose oxidase can be used to remove glucose in food or medicine, thereby preventing browning of food or obtaining oligosaccharides with higher purity; at the same time, glucose oxidase can be used as a relatively An ideal potassium bromate substitute, used as a safer flour improver; in the fermentation industry, the...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/04C12R1/79
CPCC12N9/0006C12Y101/03004
Inventor 高兆建张铁柱孙会刚李同祥刘恩岐曹建冬顾强沈彬彬
Owner CHONGQING FULING ZIZHU FOOD CO LTD
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