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Bacillus licheniformis Bl22 for high-yield production of chitin deacetylase, and related products and application thereof

A technology of Bacillus licheniformis and deacetylase, applied in the field of microorganisms, can solve the problems of few sources of bacteria, low enzyme production activity, long fermentation time, etc., and achieve cost saving of energy and auxiliary materials, broad application prospects, and simple cultivation methods Effect

Active Publication Date: 2021-03-09
JILIN COFCO BIOCHEM +3
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the reported chitin deacetylases generally have problems such as long fermentation time, low enzyme activity, high catalytic temperature, and poor deacetylation effect, and most of them are enzymes from fungi, and few from bacteria.

Method used

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  • Bacillus licheniformis Bl22 for high-yield production of chitin deacetylase, and related products and application thereof
  • Bacillus licheniformis Bl22 for high-yield production of chitin deacetylase, and related products and application thereof
  • Bacillus licheniformis Bl22 for high-yield production of chitin deacetylase, and related products and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment approach

[0039] Wherein, the fermentation medium can be a common liquid LB medium, but according to a preferred embodiment of the present invention, the fermentation medium contains glucose, yeast powder, tryptone and K 2 HPO 4 , pH value 6.5-7.2; among them, glucose can be 35-45g / l, yeast powder can be 20-30g / l, tryptone can be 10-20g / l, K 2 HPO 4 It can be 2-4g / L, and the balance is water.

[0040]Wherein, the conditions of the fermentation culture may be as follows: the temperature is 30-37° C., and the time is 20-24 hours. It is further preferred to perform the cultivation under agitation (100-200 rpm). After being cultured, the content of chitin deacetylase in the fermentation liquid reaches 1800-2100U / ml through detection.

[0041] It can be seen that the Bacillus licheniformis Bl22 provided by the present invention can utilize common carbon sources (eg, glucose) and nitrogen sources (eg, yeast powder) for rapid culture and high-yield chitin deacetylase fermentation.

[0042...

Embodiment 1

[0078] This example is used to illustrate the method of using Bacillus licheniformis Bl22 to produce chitin deacetylase

[0079] (1) Activation of bacteria

[0080] Cultivation method: Take an appropriate amount of cryopreserved strains and streak them on the LB activation plate, and culture them in a constant temperature incubator at a temperature of 35±2°C for 20 hours.

[0081] Activation medium: yeast extract 5g / L, tryptone 10g / L, NaCl 10g / L, agar powder 20g / L, balance water, pH 6.5-7.2.

[0082] (2) Seed cultivation

[0083] Cultivation method: Pick a single colony from the activated plate and place it in the seed medium for constant temperature shaking culture, the stirring speed is 150rpm, the temperature is 35±2°C, and the time is 9h.

[0084] Seed medium: yeast extract 5g / L, tryptone 10g / L, NaCl 10g / L, the balance is water, pH 6.5-7.2.

[0085] (3) Fermentation culture

[0086] Cultivation method: Transfer the seed liquid to the fermentation medium with constant tem...

Embodiment 2

[0090] This example is used to illustrate the preparation of Bacillus licheniformis Bl22 chitin deacetylase crude enzyme solution

[0091] Collect 10ml of fermentation broth fermented for 23 hours, centrifuge at 12000rpm for 5min, discard the supernatant of the culture medium, take 10mL of ultrasonic disruption buffer (50mM Tris-cl, 20mM NaOH, pH 8.0) and pipette up and down to fully resuspend the bacteria; Divide the resuspended bacteria into 10 parts on average, 1ml each, transfer to 10 2mL centrifuge tubes, place each centrifuge tube in a fixed ice-water mixing bath, perform ultrasonic disruption separately, and set the amplitude bar of the ultrasonic breaker to 2 , the power is 20%, the ultrasonic frequency is 2s / 2s (that is, crushing for 2s, pausing for 2s), and the total crushing time is 5min; all crushed liquid is centrifuged at 12000rpm for 5min, and the supernatant is collected into a new 15ml centrifuge tube, which is lichen Bacillus deacetylase crude enzyme solution...

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Abstract

The invention relates to the field of microorganisms, and more specifically, relates to bacillus licheniformis Bl22 for high-yield production of chitin deacetylase, and related products and application thereof. The preservation number of the bacillus licheniformis is CGMCC No. 20490. The bacillus licheniformis Bl22 has the advantages that the provided bacillus licheniformis Bl22 culture method issimple, and a common carbon source and a nitrogen source can be used for rapid culture and fermentation to produce the chitin deacetylase with high yield. The optimum catalytic temperature of the produced chitin deacetylase is relatively low, is about 40 DEG C, and shows high catalytic activity in a wide pH range (pH value of 6-10); the cost of energy, auxiliary materials and the like can be savedin industrial application, and the application prospect is wide.

Description

technical field [0001] The present invention relates to the field of microorganisms. More specifically, the present invention relates to Bacillus licheniformis Bl22 with high chitin deacetylase production and related products and applications. Background technique [0002] Chitin, also known as chitin and chitin, is a polysaccharide composed of N-acetylamino-D-glucose monomers linked by β-1,4-glycosidic bonds. It is the second largest type of natural polymer organic compound after cellulose in nature, and widely exists in the exoskeleton of invertebrates (such as shrimp, crab and insects, etc.) and the cell walls of fungi and algae. Due to the poor solubility of chitin, it is insoluble in water, dilute acid, dilute alkali and organic solvents, etc., which greatly limits its utilization value. For example, deacetylation of chitin (deacetylation degree greater than 55%) can obtain chitosan (chitosan) with greatly improved solubility. Moreover, chitosan has various physiolog...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N9/80C12P19/26C12R1/10
CPCC12N1/20C12N9/80C12P19/26C12Y305/01041
Inventor 佟毅臧传刚张媛沈雪梅王小艳赵国淼王靖焦琳魏超王琪陈博李义周勇卢宗梅张钊商谈
Owner JILIN COFCO BIOCHEM
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