Bacillus subtilis capable of producing chitinase and application thereof

A technology of Bacillus subtilis and bacterial preparations, applied in the field of chitinase-producing Bacillus subtilis, which can solve the problems of low biological activity, acid pollution, and difficult control

Inactive Publication Date: 2015-02-18
QINGDAO AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there are mainly two methods for the production of chitosan oligosaccharides: chemical degradation method and biological enzymatic method; however, the chemical degradation method is costly, difficult to control, and the product has a low degree of polymerization (low biological activity) and is mostly monosaccharides, which has little application value. High, and will bring serious acid pollution, its use is strictly restricted

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0013] Example 1: Breeding of Bacillus subtilis PLA119 and optimization of its enzyme production conditions

[0014] 1. The Bacillus subtilis of the starting strain was screened from the soil enriched with shrimp heads, and was identified and preserved by the Food Microbiology Laboratory of Qingdao Agricultural University. It has the ability to produce chitinase and protease.

[0015] 2. Medium

[0016] (1) Enrichment medium (g / L): beef extract 3, peptone 10, NaCl 5, pH 7.0.

[0017] (2) Isolation medium (g / L): colloidal chitin 2.5, MgSO 4 ·7H 2 O 0.5,K 2 HPO 4 , KH 2 PO 4 0.3, FeSO 4 ·7H 2 O 0.01, (NH 4 ) 2 SO 4 3, Agar 20. pH 7.0, 121°C, sterilize for 20min.

[0018] (3) Shake flask screening medium (enzyme production medium) (g / L): colloidal chitin 10, soybean meal 10, peptone 5, yeast extract 5, K 2 HPO 4 0.7, KH 2 PO 4 0.3, Glucose 1, MgSO 4 ·7H 2 O0.5, FeSO 4 ·7H 2 O 0.01, ZnSO 4 0.01. pH 7.0, 121°C, sterilize for 20min.

[0019] 3. Plasma mutagen...

Embodiment 2

[0026] Embodiment 2: Chitooligosaccharide preparation experiment of Bacillus subtilis PLA119

[0027] 1. Preparation of colloidal chitin

[0028] The fine powder chitin is prepared into a uniform solution with a concentration of 5% by using 1% acetic acid solution.

[0029] 2. Preparation of chitinase enzyme solution

[0030] The Bacillus subtilis PLA119 fermentation broth was centrifuged (5000r / min, 5min) to remove the bacteria, the supernatant was concentrated by low-temperature rotary evaporation, 70% ammonium sulfate was precipitated, Sephadex G25 was desalted, and the low-temperature rotary evaporation was concentrated to chitinase activity It is 10U / mL.

[0031] 3. Chitooligosaccharide preparation

[0032] Mix the chitinase solution with 5% colloidal chitin at a ratio of 1:10, heat and react for 10 hours at 37°C and pH 7.0, heat to inactivate the enzyme, centrifuge at 3000r / min for 10min, take the supernatant, Concentrate by rotary evaporation, and dry at 80°C to obt...

Embodiment 3

[0033] Embodiment 3: Bacillus subtilis PLA119 prepares experiment of shrimp head sauce without chemical preservatives

[0034] Take unpolluted shrimp heads, mix them with water at room temperature according to the weight ratio of 1:1.5, adjust the pH to 7.0, insert activated Bacillus subtilis PLA119, ferment and cultivate at 35°C for 48 hours, and obtain rich amino acids, short peptides and several Add 10% (v / v) rice wine, 16.8% sugar, 9.89% garlic, 1.5% ginger, 1.25% monosodium glutamate, and 15.98% salt to the shrimp head fermentation broth of butyl oligosaccharides. After 15 days of fermentation, the amino acid content is 0.067±0.03 g / g, the chitosan oligosaccharide content is 0.011±0.001g / g, and the shrimp head sauce without chemical preservatives is prepared,

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PUM

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Abstract

The invention aims to provide bacillus subtilis capable of producing chitinase and application thereof. The bacillus subtilis is obtained by plasma mutagenesis, has higher chitinase enzyme activity and protease enzyme activity, can be used as leavening agents, probiotics and preservatives, and can be widely applied to enzyme preparations and food industries. The preservation number of bacillus subtilis is CGMCC No.9166. The bacillus subtilis strain has relatively high chitinase enzyme activity and protease enzyme activity, is simple in nutrition requirement but has high fecundity, can be used as leavening agents, probiotics and preservatives, and has a wide application prospect.

Description

technical field [0001] The invention belongs to the technical field of microorganism application, and in particular relates to a strain of chitinase-producing bacillus subtilis and its application. Background technique [0002] Chitinase (chitinase) can catalyze the degradation of chitin (chitin) to generate chitooligosaccharides. Chitooligosaccharides are water-soluble and are upgraded products of chitin and chitosan products, and have a significant inhibitory effect on spoilage bacteria , is a natural food preservative with excellent performance. At the same time, chitosan oligosaccharides also have the ability to promote the growth of beneficial microorganisms (such as bifidobacteria, lactic acid bacteria), improve the intestinal microbial flora, improve the body's immunity, promote the formation of liver and spleen antibodies, promote the absorption of calcium and minerals, reduce Blood lipids, removing oxygen and anion free radicals, delaying aging, inhibiting tumor ce...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N9/42A23L1/33A23L1/24C12R1/125A23L17/40A23L27/60
CPCA23V2002/00A23L17/65A23L27/60A23L33/135C12N1/20C12N9/2442C12Y302/01014C12N1/205C12R2001/125A23V2200/10
Inventor 李静谭海刚王凤舞
Owner QINGDAO AGRI UNIV
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