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Method for producing neutral phytase through immobilized bacillus fermentation

A technology of bacillus and phytase, applied in the direction of microorganism-based methods, immobilized on or in inorganic carriers, biochemical equipment and methods, etc., can solve the problems of low enzyme production efficiency, low equipment utilization, and production costs Advanced problems, to achieve the effect of improving fermentation efficiency, improving utilization rate, and increasing specific surface area

Pending Publication Date: 2020-04-24
陈东浩
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, the industrialized production of phytase mainly adopts the method of free cell batch fermentation or fed-batch fermentation, which has problems such as high production cost, high labor intensity, low equipment utilization rate, and low enzyme production efficiency. The research on fermentative production of phytase has not been reported at home and abroad

Method used

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  • Method for producing neutral phytase through immobilized bacillus fermentation
  • Method for producing neutral phytase through immobilized bacillus fermentation
  • Method for producing neutral phytase through immobilized bacillus fermentation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Embodiment 1 The selection of inorganic carrier

[0040] (1) Seed liquid preparation: Streak inoculation of the Bacillus megaterium strain on LB slant medium, culture at a constant temperature of 37°C for 36~48h, then inoculate in the seed medium, under the conditions of temperature 37°C and rotation speed 180r / min Under culture to the logarithmic growth phase; the composition of the seed medium is: 15g / L glucose, 15g / L peptone, 4g / L yeast extract, 5g / L sodium chloride, 2g / L calcium chloride, 0.4g / L Magnesium sulfate, 0.2g / L potassium dihydrogen phosphate, 0.03g / L ferrous sulfate, pH7.0.

[0041] (2) Pretreatment of the carrier: choose fly ash, activated carbon, zeolite and ceramsite with a diameter of 2~3mm as the carrier, wash with distilled water and dry at 105°C for 2 hours; put the ceramsite into 1.0mol / L sodium hydroxide solution In this method, the material-liquid ratio of ceramsite and sodium hydroxide solution is 1g:5mL, shake and adsorb at 150r / min for 30min,...

Embodiment 2

[0047] According to the method in Example 1, the alkali-modified ceramsite was selected as the carrier to prepare immobilized cells, and then the continuous fermentation culture was carried out according to the following steps:

[0048] (1) Pre-cultivation: Add the fermentation medium into the fixed-bed reactor, and carry out circulating flow culture at a flow rate of 1.5L / h. The culture time is 36h, the temperature is 37°C, and the ventilation rate is 1.8vvm, and then released into the reactor culture medium; the composition of the fermentation medium is: 35g / L bran, 5g / L glucose, 15g / L bean cake powder, 7g / L ammonium sulfate, 2.5g / L sodium phytate, 6g / L sodium chloride, 2g / L calcium chloride, 0.15g / L potassium dihydrogen phosphate, 0.3g / L magnesium sulfate, 0.03g / L ferrous sulfate, 50μmol / L H 2 o 2 , 7.5g / L n-dodecane, pH7.0.

[0049] (2) Continuous fermentation culture: add fresh fermentation medium to 0.06h -1 The dilution ratio flows into a fixed-bed reactor for conti...

Embodiment 3

[0063] A kind of method that immobilized bacillus fermentation produces neutral phytase comprises the steps:

[0064] (1) Seed liquid preparation: Streak inoculation of the Bacillus megaterium strain on LB slant medium, culture at a constant temperature of 37°C for 36~48h, then inoculate in the seed medium, under the conditions of temperature 37°C and rotation speed 180r / min Under culture to the logarithmic growth phase; the composition of the seed medium is: 15g / L glucose, 15g / L peptone, 4g / L yeast extract, 5g / L sodium chloride, 2g / L calcium chloride, 0.4g / L Magnesium sulfate, 0.2g / L potassium dihydrogen phosphate, 0.03g / L ferrous sulfate, pH7.0.

[0065] (2) Pretreatment of the carrier: put ceramsite with a diameter of 2~3mm into 1.0mol / L sodium hydroxide solution, the ratio of solid to liquid between ceramsite and sodium hydroxide solution is 1g:5mL, shake at 150r / min Adsorb for 30 minutes, then let it stand for 24 hours, rinse the carrier with distilled water until the pH...

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Abstract

The invention provides a method for producing neutral phytase through immobilized bacillus fermentation. The method comprises the following steps: (1) preparing a seed solution; (2) carrying out pretreatment of a carrier: modifying a ceramsite carrier with an alkali liquor; (3) preparing immobilized cells: filling the carrier into a fixed bed reactor, and immobilizing thallus cells on the carrierthrough an adsorption effect; (4) carrying out pre-culturing: culturing the immobilized cells for 30-40 hours by using a fermentation culture medium; and (5) carrying out continuous fermentation culture: enabling the fermentation culture medium to flow into a fixed bed reactor at a certain dilution rate for continuous fermentation. The bacillus is immobilized by adopting the alkali-modified carrier, and the method has the advantages of being low in cost, stable in property, non-toxic and good in adsorption effect. According to the method, the immobilized cell technology is combined with the continuous fermentation process, so that the purpose of continuously producing neutral phytase at high intensity is achieved, the fermentation efficiency and the phytase yield are greatly improved, andindustrial production is facilitated.

Description

technical field [0001] The invention relates to the fields of fermentation engineering and enzyme engineering, in particular to a method for producing neutral phytase by fermentation of immobilized bacillus. Background technique [0002] Phosphorus, as a mineral element abundant in animals, is closely related to life activities, mainly involved in the composition and growth of bones and muscles. The phosphorus in the animal body is mainly provided by the feed, and the phosphorus in the feed mainly exists in the form of phytate phosphorus or phytate. Phytic acid is an anti-nutritional factor, which has a strong chelating effect due to its special structure and physical and chemical properties. Phytic acid will form stable complexes with zinc, iron, copper, manganese, potassium, sodium, magnesium, calcium, etc., reducing the digestion and utilization of mineral elements in feed. Phytic acid will also form a phytate-protein binary complex with protein, which will precipitate ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N11/14C12N9/16C12R1/11
CPCC12N11/14C12N9/16C12Y301/03008C12Y301/03026
Inventor 陈东浩
Owner 陈东浩
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