Method for purifying antibacterial protein in bacillus cereus fermentation broth

A technology of Bacillus cereus and fermentation liquid, which is applied in the field of bioengineering, can solve problems such as short shelf life, unstable amount of live bacteria, and short shelf life of products, and achieve the effects of convenient operation, low cost, and improved antibacterial ability

Active Publication Date: 2013-05-08
NANJING XINBAI PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this preparation, like other microecological preparations, has the following problems: factors such as temperature and tablet compression during the production process lead to a decrease in the number of viable bacteria during the validity period, and the number of viable bacteria is unstable during the validity period, and the product has a short shelf life and short shelf life.
However, Bacillus cereus preparations generally have problems such as short shelf life and short shelf life.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] (1) Extraction of antibacterial proteins: Bacillus cereusCGMCC4348, which was independently screened and identified, was used to culture a fermentation broth containing antibacterial proteins through microbial fermentation, and the fermentation broth was graded and precipitated with ammonium sulfate to extract antibacterial proteins Crude product, refer to the bacteriostatic protein component A extracted in Example 1 in the inventor's patent application (application number is 201210056365.7), its bacteriostatic rate to cotton Fusarium wilt pathogen is 62%;

[0023] (2) Column packing: Weigh Sephadex G-100 Sephadex G-100 with distilled water at a ratio of 16mL / g to swell for 3 hours before packing; put the treated Sephadex G-100 on the column by wet method; The diameter-to-height ratio is 1:15, and fully equilibrated with 0.02mol / L Tris-HCl buffer solution with a pH of 7.1 for 3 hours after being loaded on the column;

[0024] (3) Sample loading: Prepare the antibacteria...

Embodiment 2

[0028] (1) Extraction of antibacterial proteins: Bacillus cereusCGMCC4348, which was independently screened and identified, was used to culture a fermentation broth containing antibacterial proteins through microbial fermentation, and the fermentation broth was graded and precipitated with ammonium sulfate to extract antibacterial proteins Crude product; refer to the bacteriostatic protein component B extracted in Example 2 in the inventor's patent application (application number is 201210056365.7), and its bacteriostatic rate to cucumber wilt pathogen is 42%;

[0029] (2) Column packing: Weigh Sephadex G-150 Sephadex G-150 with distilled water at a ratio of 20mL / g to swell for 5 hours before packing; put the processed Sephadex G-150 on the column by wet method; The diameter-to-height ratio is 1:15, and fully equilibrated with 0.03mol / L Tris-HCl buffer solution with a pH of 7.5 for 4 hours after loading;

[0030] (3) Sample loading: Prepare the bacteriostatic protein obtained ...

Embodiment 3

[0034] (1) Extraction of antibacterial proteins: Bacillus cereusCGMCC4348, which was independently screened and identified, was used to culture a fermentation broth containing antibacterial proteins through microbial fermentation, and the fermentation broth was graded and precipitated with ammonium sulfate to extract antibacterial proteins Crude product; refer to the bacteriostatic protein component A extracted in Example 3 in the inventor's patent application (application number is 201210056365.7), its bacteriostatic rate to cotton Fusarium wilt pathogen is 59%;

[0035] (2) Column packing: Weigh Sephadex G-75 Sephadex G-75 and swell it with distilled water at a ratio of 18mL / g for 4 hours before packing; put the treated Sephadex G-75 on the column by wet method; The diameter-to-height ratio is 1:10, and fully equilibrated with 0.04mol / L Tris-HCl buffer solution with a pH of 8.5 for 2 hours after loading on the column;

[0036] (3) Sample loading: Prepare the bacteriostatic p...

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Abstract

The invention discloses a method for purifying antibacterial protein in bacillus cereus fermentation broth. The method comprises the following steps of: culturing the bacillus cereus fermentation broth through microbial fermentation by utilizing autonomously screened and identified Bacillus cereus (CGMCC (China General Microbiological Culture Collection Center) NO.4348), extracting an antibacterial protein crude product by utilizing a ammonium sulfate fractional precipitation method, further purifying the antibacterial protein crude product by utilizing a sephadex column and detecting the bacteriostatic activity on fungal diseases. The method disclosed by the invention has the advantages of convenience for operation, simple process, low cost and capability of remarkably increasing the antibacterial activity of the antibacterial protein in the fermentation broth.

Description

technical field [0001] The invention belongs to the field of bioengineering, and specifically relates to microbial fermentation, biopharmaceuticals, extraction, separation and purification processes of fermentation products, in particular to a method for purifying antibacterial proteins in Bacillus cereus fermentation liquid. Background technique [0002] Bacillus cereus (B.cereus for short) is widely distributed in nature, and Bacillus cereus can be isolated from air, dust, soil, water and plants. The live Bacillus cereus preparation is a microbial product officially approved by the Ministry of Agriculture of my country. It has the function of regulating the micro-ecological balance of the host and is widely used in agriculture, feed, medical care, food and other fields. However, this preparation, like other probiotics, has the following problems: the number of viable bacteria decreases due to factors such as temperature and tablet compression during the production process,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P21/00C07K1/36C07K1/30C07K1/16C12R1/085
Inventor 胡永红杨文革江心敏唐容容沈飞朱文俊冯旌
Owner NANJING XINBAI PHARMA
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