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Method for analyzing small molecule metabolite change in VC (Vitamin C)-producing strain passage process

A technology for small molecule metabolites and strain production, which is applied in the analysis of materials, material analysis by electromagnetic means, measurement devices, etc., and can solve problems such as unclear mechanism of action

Active Publication Date: 2014-04-16
TIANJIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Through subculture of mixed bacteria, the ability of mixed bacteria to ferment and produce 2-keto-L-gulonic acid is improved, but the mechanism of action is still unclear

Method used

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  • Method for analyzing small molecule metabolite change in VC (Vitamin C)-producing strain passage process
  • Method for analyzing small molecule metabolite change in VC (Vitamin C)-producing strain passage process
  • Method for analyzing small molecule metabolite change in VC (Vitamin C)-producing strain passage process

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Embodiment 1

[0039] A method for analyzing changes in small molecule metabolites in the process of subculture of vitamin C production strains, characterized in that it includes the following steps:

[0040] (1) Subculture of mixed bacteria:

[0041] ① Solid culture:

[0042] Take 100 μL of liquid nitrogen to inoculate Gluconobacter oxydans (Gluconobacter oxydans) preserved in 20% glycerol aqueous solution and 100 μL of Bacillus megaterium preserved in 20% glycerin aqueous solution respectively on solid On the culture medium, culture at 30°C for 36 hours;

[0043] (2) Seed cultivation:

[0044] Transfer the Bacillus megaterium and Gluconobacter oxidans cultured in step (1) into the seed culture medium respectively, shake and cultivate them at 30°C and 240 rpm for 36 hours, and obtain the Bacillus megaterium seed liquid and Gluconobacter oxidans seed liquid respectively;

[0045] Inoculate a new seed medium with Bacillus megaterium and Gluconobacter oxydans so that the density of Bacillus...

Embodiment 2

[0066] 1. A method for analyzing changes in small molecule metabolites in the process of subculture of vitamin C production strains, comprising the steps of:

[0067] (1) Subculture of mixed bacteria:

[0068] ① Solid culture:

[0069] Inoculate 10 μL of Gluconobacter oxydans stored in 15% aqueous glycerol and 10 μL of Bacillus megaterium stored in 15% aqueous glycerol stored in liquid nitrogen Incubate on solid medium at 28°C for 48 hours;

[0070] ②Seed cultivation:

[0071] Transfer the Bacillus megaterium and Gluconobacter oxidans cultivated in step (1) into the seed culture medium respectively, and shake and cultivate them at 28°C and 200 rpm for 48 hours to obtain the Bacillus megaterium seed liquid and Gluconobacter oxidans seed liquid respectively;

[0072] Inoculate a new seed medium with Bacillus megaterium and Gluconobacter oxydans so that the density of Bacillus megaterium is 2 x 10 7 CFU / mL, so that the density of Gluconobacter oxidans is 2×10 8 CFU / mL, at 2...

Embodiment 3

[0090] A method for analyzing changes in small molecule metabolites in the process of subculture of vitamin C production strains, comprising the steps of:

[0091] (1) Subculture of mixed bacteria:

[0092] ① Solid culture:

[0093] Take 500 μL of Gluconobacter oxydans stored in 30% aqueous glycerol solution and 500 μL of Bacillus megaterium stored in 30% aqueous glycerol solution and inoculate them respectively On solid medium, culture at 35°C for 24 hours;

[0094] ②Seed cultivation:

[0095] Transfer the Bacillus megaterium and Gluconobacter oxidans cultured in step (1) into the seed culture medium respectively, and culture them on a shaker at 35°C and 280 rpm for 24 hours to obtain the Bacillus megaterium seed liquid and Gluconobacter oxidans seed liquid respectively;

[0096] Inoculate a new seed medium with Bacillus megaterium and Gluconobacter oxydans so that the density of Bacillus megaterium is 2 x 10 10 CFU / mL, so that the density of Gluconobacter oxidans is 2×10...

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Abstract

The invention discloses a method for analyzing small molecule metabolite change in a VC (Vitamin C)-producing strain passage process, which comprises the steps of: 1, subculturing mixed strains; 2, preparing and measuring an intracellular small molecule metabolite sample; 3, analyzing main components; and 4, analyzing a process, to be specific, making the content of a small molecule metabolite marker into a chart according to different passage times, observing and analyzing the rule of the change of the small molecule metabolite marker, detecting the change of the intracellular small molecule metabolite in the VC-producing strain passage process. According to the method provided by the invention, through respective quantitative and qualititative analysis of bacillus megatherium, gluconobacter oxydans and the small molecule metabolite of the mixed strains, with different passage times, metabolite molecules and metabolic pathways, relevant to interaction of two reinforced VC-producing strains, are found, the direction is provided for improving the strain reconstruction and the culture condition optimization by using 2-keto-L-gulonic acid hydrate as a target, and the foundations of increasing VC yield and optimizing the fermenting process are laid.

Description

technical field [0001] The invention belongs to the field of industrial microbes, and relates to a method for analyzing the change of small molecule metabolites in the process of subculture of vitamin C production bacterial strain mixed bacteria. Background technique [0002] With the development of economy and the improvement of people's living standards, the demand for vitamin C (VC) in food and medicine is increasing year by year. At present, most of our country produces vitamin C by "two-step fermentation method". The first step of fermentation uses Acetobacter black to convert sorbitol into L-sorbose, and the second step of fermentation is a mixed fermentation of Bacillus megaterium and Gluconobacter oxydans to convert sorbose into 2-keto-L, a precursor of vitamin C. - Gulonic Acid. Among them, in the second step of fermentation, Bacillus megaterium is an associated bacterium, and Gluconobacter oxydans is an acid-producing bacterium. In the process of mixed fermentat...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N27/62
Inventor 元英进李霞邹旸胡梦龙
Owner TIANJIN UNIV