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Method for producing 2-keto-l-gulonic acid

A production method and technology of gulonic acid, applied in the production field of 2-keto-L-gulonic acid, can solve problems such as low acid production, insufficient nutrients, and hindered microbial growth

Active Publication Date: 2012-07-04
ANHUI BBCA FERMENTATION TECH ENG RES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This method makes the initial liquid volume lower, and in the later stage of fermentation, the volume of the fermented liquid increases with the addition of sugar, the concentration of bacteria decreases, and the nutrients (mainly nitrogen source) in the fermented liquid are insufficient, and the growth of microorganisms is hindered, resulting in middle and late production. The acid is low, and the high acid production capacity of the two-step fermentation cannot be fully utilized, so the concentration of acid production is relatively low

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Bacteria: Gluconobacter oxidans (small bacteria) and Bacillus megaterium (big bacteria).

[0018] Eggplant bottle slant medium: yeast extract 0.1%, peptone 0.2%, corn steep liquor 1.5%, calcium carbonate 0.15%, magnesium sulfate 0.1%, urea 0.1%, glucose 0.1%, sorbose 2.0%, agar 2.0%.

[0019] The volume of the eggplant bottle slope is 80ml. Use an inoculation loop to take 3 rings of small bacteria and half rings of large bacteria and evenly spread them on the slope of the eggplant bottle, and incubate at 29°C for 80 hours.

[0020] Shake flask seed medium: yeast extract 0.1%, corn steep liquor 1.5%, calcium carbonate 0.15%, magnesium sulfate 0.1%, urea 0.1%, glucose 0.1%, sorbose 2.0%.

[0021] The specification is 1000ml seed shaker bottle, the capacity is 100ml, eggplant bottle slant seeds are inoculated, and each eggplant bottle slant seeds are inoculated with 8-10 bottles. 200r / min, shake culture at 30°C for 20-22 hours.

[0022] The initial liquid volume in the 3...

Embodiment 2

[0029] 140m 3 The initial liquid volume in the fermenter is 30m 3 , the initial sorbose concentration was 25 mg / ml, and the inoculum size was 30% of the initial volume.

[0030] The content of each component in the fermentation medium is: 1.5% of corn steep liquor, 0.2% of potassium dihydrogen phosphate, 0.1% of magnesium sulfate, 0.5% of urea and 2.5% of sorbose.

[0031] When the residual sorbose sugar drops to 20mg / ml, start feeding the sorbose sugar solution with a concentration of 220mg / L, and control the sugar content to 19±1mg / ml during the feeding process; start feeding the nutrient solution after 20 hours, and the ingredients in the nutrient solution The content is: corn steep liquor 110g / L, potassium dihydrogen phosphate 6g / L, magnesium sulfate 1.5g / L, urea 15g / L, stop feeding after 40 hours, the total feeding amount is 2.5m 3 , Dissolved oxygen is controlled at 40±5% during feeding.

[0032] The initial pH was 6.5±0.1, and it was controlled at 6.8±0.1 during the ...

Embodiment 3

[0035] 140m 3 The initial liquid volume in the fermenter is 35m 3 , the initial sorbose concentration was 20 mg / ml, and the inoculum size was 30% of the initial volume.

[0036] The content of each component in the fermentation medium is as follows: 2% of corn steep liquor, 0.2% of potassium dihydrogen phosphate, 0.2% of magnesium sulfate, 1% of urea and 2% of sorbose.

[0037] When the residual sorbose sugar drops to 15mg / ml, start to feed the sorbose sugar solution with a concentration of 240mg / L, and control the sorbose content to 16±1mg / ml during the fed-batch process; start to feed the nutrient solution within 24 hours. The content of ingredients is: corn steep liquor 100g / L, potassium dihydrogen phosphate 5g / L, magnesium sulfate 2g / L, urea 10g / L, stop feeding after 45 hours, the total feeding amount is 2m 3 , Dissolved oxygen is controlled to be 35±5% during feeding.

[0038] The initial pH was 6.6±0.1, and it was controlled at 6.8±0.1 during the fermentation process....

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Abstract

The invention provides a method for producing 2-keto-l-gulonic acid, including the following steps of: feeding nitrogen source nutrient solution from the 20th to the 26th hour in the fermentation process to compensate nutritional demands of microorganism growth in the fermentation process, finishing feeding solution in the 40th to the 45th hour, wherein the dissolved oxygen is controlled to be 30-50% in the nutrient solution feeding process. The invention utilizes a method of feeding nutrient solution at middle and later stages of fermentation, overcomes disadvantage that biomass grows and generates acid slowly, combines control of factors of temperature, pH, dissolved oxygen, low initial glucose, fed-batch glucose compensation and the like, and optimizes the growth relations of biomasseswith different sizes in microbial mixed fermentation. The gulonic acid content of the invention is 105-115mg / ml, and the glucose acid convert ratio is 92-95%.

Description

technical field [0001] The invention relates to a production method of 2-keto-L-gulonic acid, which belongs to the field of microorganism production. Background technique [0002] At present, domestic production of gulonic acid mostly adopts a two-step fermentation method. The two-step fermentation method uses mixed bacteria fermentation. Among them, the small bacteria are acid-producing bacteria, and the intracellular secretion of L-sorbosone oxidase can oxidize sorbose into 2-keto-L-gulonic acid, but the growth is slow when cultured alone, and the acid-producing ability is low; the large bacteria cannot produce acid, but It can be used as the companion bacteria of small bacteria, which can promote the growth and acid production of small bacteria. [0003] It has been confirmed by research that both the somatic fluid and extracellular fluid of large bacteria can promote the growth of small bacteria and shorten the lag period of small bacteria growth. In the mixed culture ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P39/00C12P7/60C12R1/11C12R1/01
Inventor 李荣杰薛培俭尚海涛穆晓玲符秀迪王芳
Owner ANHUI BBCA FERMENTATION TECH ENG RES
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