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Fermentation process of Clostridium thermocellum with high cellulase production

A cellulase and fermentation process technology, applied in the directions of enzymes, bacteria, enzymes, etc., can solve the problems of high cost, low fermentation enzyme activity, and limited wide application, achieve optimization of fermentation culture conditions, improve cellulase activity, fermentation The effect of process optimization

Pending Publication Date: 2020-06-26
浙江科峰生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The second point is that due to factors such as its characteristics and culture conditions, the currently used bacterial strains make the cellulase fermentation time very long and the efficiency is low, resulting in high cost. Currently, the production of cellulase by liquid fermentation is suitable for large-scale production. But at present fermentation enzyme activity is low, causes the cellulase product price of liquid method production to be on the high side, has limited its wide application in industry, so the present invention has chosen high cellulase activity bacterium --- fusothermal bacillus, and to Its fermentation process is optimized to achieve cost control

Method used

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  • Fermentation process of Clostridium thermocellum with high cellulase production
  • Fermentation process of Clostridium thermocellum with high cellulase production
  • Fermentation process of Clostridium thermocellum with high cellulase production

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Experimental program
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Effect test

Embodiment 1

[0019] 1. Seed medium: cellobiose 10g / L, corn steep liquor 12g / L, tripotassium citrate 2g / L, citric acid monohydrate 1.25g / L, sodium sulfate 1g / L, potassium dihydrogen phosphate 1g / L , sodium bicarbonate 2.5g / L, ammonium chloride 1.5g / L, urea 5g / L, yeast extract 1g / L, magnesium chloride hexahydrate 50g / L, ferrous chloride tetrahydrate 0.1g / L, dihydrate Calcium chloride 0.2g / L, cysteine ​​hydrochloride monohydrate lg / L, pyridoxamine dihydrochloride 0.02g / L, p-aminobenzoic acid 0.004g / L, D-biotin 0.002g / L, vitamin B 12 0.02g / L, vitamin B l 0.002g / L, the pH value is natural.

[0020] 2. Strain activation: Inoculate the laboratory-preserved strains into the seed medium at an inoculation amount of 1%, activate and cultivate at 60°C, activate 2-3 generations in the same way, and continue to cultivate in the seed medium to obtain thalassemia bacillus bacteria.

[0021] 3. Fermentation medium: under the basic conditions of the seed medium, add 10g / L microcrystalline cellulose.

...

Embodiment 2

[0024] Embodiment 2: optimization of fermentation culture conditions

[0025] Keeping other conditions unchanged, different fermentation temperatures were set, respectively 30, 40, 50, 55, and 60°C, and the cellulase activity was measured after the fermentation was completed. The results are shown in Table 1. It was found that the enzyme activity of the bacteria was the highest at 55°C, so 55°C was selected as its fermentation temperature. Control other conditions unchanged, set different culture time, respectively 24h, 48h, 60h, 72h, 84h, 96h, measure the cellulase activity after the end of fermentation. The results are shown in Table 2. With the increase of fermentation time, the enzyme activity showed an upward trend, reached the peak at 60 hours of fermentation, and began to decline at 96 hours. Therefore, the fermentation is stopped after 60 hours, which can greatly save the fermentation cost.

[0026] Control other conditions unchanged, set different initial pH of medi...

Embodiment 3

[0034] Embodiment 3: process enlargement

[0035] Pick up 200 μL of the glycerol tube to store the strain, transfer it to 25 mL of seed medium, culture it at a constant temperature of 55°C, and activate the strain, transfer the activated strain to a 50mL fermentation medium according to the inoculation amount of 2%, and cultivate it at a constant temperature of 55°C for 24 hours to obtain First-class seed liquid.

[0036] Seed medium: cellobiose 10g / L, corn steep liquor 12g / L, tripotassium citrate 2g / L, citric acid monohydrate 1.25g / L, sodium sulfate 1g / L, potassium dihydrogen phosphate 1g / L, carbonic acid Sodium Hydrogen 2.5g / L, Ammonium Chloride 1.5g / L, Urea 5g / L, Yeast Extract 1g / L, Magnesium Chloride Hexahydrate 50g / L, Ferrous Chloride Tetrahydrate 0.1g / L, Chloride Dihydrate Calcium 0.2g / L, cysteine ​​hydrochloride monohydrate lg / L, pyridoxamine dihydrochloride 0.02g / L, p-aminobenzoic acid 0.004g / L, D-biotin 0.002g / L, vitamin B 12 0.02g / L, vitamin B l 0.002g / L, the pH ...

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Abstract

The invention discloses a fermentation process of Clostridium thermocellum with high cellulase production, and belongs to the technical field of fermentation engineering. The method comprises the following steps: inoculating Clostridium thermocellum into a seed culture medium according to an inoculation amount of 1%, activating and culturing at 60 DEG C, activating for 2-3 generations in the samemanner, and continuing culturing in the seed culture medium to obtain a seed liquid of Clostridium thermocellum; and inoculating the seed liquid into a fermentation medium according to an inoculationamount of 2%, and fermenting and culturing for 48-96 hours at 40-60 DEG C to obtain a fermentation product. The method optimizes fermentation culture conditions of Clostridium thermocellum, and feedsthe cells in time according to growth conditions of bacterial cells, can greatly improve cellulase activity of Clostridium thermocellum and save costs, and is suitable for industrial production.

Description

technical field [0001] The invention belongs to the technical field of fermentation engineering, and in particular relates to a fermentation process of fusothermal bacteria with high cellulase production. Background technique [0002] Lignocellulosic raw materials come from a wide range of sources and are renewable resources with abundant reserves. Cellulase is a general term for a group of enzymes that can degrade cellulose into glucose. Lignocellulose can be hydrolyzed by cellulase to produce sugar, which can be directly used in microbial fermentation and can convert cellulose into valuable primary metabolism. The product, cellulase, is often used in food and feed industry, paper industry, medicine, bioenergy and other industries, and has a good application prospect. [0003] Microorganisms that produce cellulase include bacteria, fungi, actinomycetes, etc. Among these microorganisms is Fusobacterium (a type of bacteria), known for its high cellulolytic activity, and the ...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12N9/42C12R1/01
CPCC12N1/20C12N9/2437
Inventor 章亭洲郎晓红
Owner 浙江科峰生物技术有限公司
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