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Bacteriocin generated by bacillus coagulans FM603 and preparing method thereof

A technology of Bacillus coagulans and bacteriocin, applied in the field of microorganisms and its cultivation and application, can solve the problem of lactic acid bacteria losing the protective function of mice, and achieve the effect of promoting animal production performance and improving breeding efficiency

Active Publication Date: 2015-12-23
沈阳丰美生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Bacteriocin is an important functional substance of probiotics. Researchers at University College Cork found that bacteriocin-producing lactic acid bacteria can effectively reduce the infection of Listeria monocytogenes in mice, but the bacteria obtained in Listeria monocytogenes Lactic acid bacteria lose their protective function in mice after immunization with bacteriocin

Method used

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  • Bacteriocin generated by bacillus coagulans FM603 and preparing method thereof
  • Bacteriocin generated by bacillus coagulans FM603 and preparing method thereof
  • Bacteriocin generated by bacillus coagulans FM603 and preparing method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Embodiment 1: Analysis of the antibacterial activity of Bacillus coagulans FM603 bacteriocin

[0031] Streak FM603 on LB agar plate, culture overnight at 30°C, pick a single colony and inoculate in 10mL LB medium, 150rpm, culture at 30°C for 24h, inoculate 0.2% inoculum in 1000mL modified tryptone medium, culture at 150rpm, 30°C After 24 hours, centrifuge at 10,000 rpm for 15 minutes to obtain the fermentation supernatant. Streak L.innocua, L.monocytogenes, S.aureus, B.cereus, and M.luteus on LB agar plates and culture overnight at 37°C, pick a single colony and inoculate it in LB medium, culture overnight at 37°C, and pipette 10ul The culture solution was inoculated on 10mL LB soft agar (LB medium containing 0.75% agar), spread evenly on the LB agar plate, placed in a sterilized Oxford cup after cooling, and added 100μl FM603 fermentation supernatant or two-fold gradient dilution The fermentation supernatant was cultured overnight at 30°C, the inhibition zone was obse...

Embodiment 2

[0034] Embodiment 2: the preparation of FM603 bacteriocin

[0035] 1. Fermentation culture

[0036] Streak culture of Bacillus coagulans FM603 on the LB plate, pick a single colony and inoculate it in 10mL liquid LB medium for overnight culture, inoculate 10mL overnight culture seed solution in 1000mL improved tryptone medium, its composition is: mass volume percentage, Tryptone 1.5%, glucose 1.5%, yeast extract powder 0.5%, corn steep liquor dry powder 0.2%, ammonium sulfate 0.05%, sodium chloride 0.1%, sodium dihydrogen phosphate 0.05%, distilled water 1000mL, pH7.0, culture condition is 30 Cultivate at 150 rpm for 20 hours.

[0037] 2. Preparation of Bacteriocin Crude Extract

[0038] The cultured FM603 fermentation broth was centrifuged at 10000×g for 15 min to take the supernatant for later use. Equilibrate the ion column with 50mM, pH 6.5 phosphate buffer, the fermentation supernatant passes through the ion column at a flow rate of 50mL / min, discard the supernatant th...

Embodiment 3

[0041] Embodiment 3: the biological characteristic of FM603 bacteriocin

[0042] 1) Tolerance to temperature: Treat the fermentation supernatant of Example 2 at 60, 70, 80, 90, and 100°C for 10 minutes respectively, take the sample without heat treatment as a control, and L.innocua as an indicator bacterium to detect heat treatment For the effect on bacteriocin activity, the antibacterial activity retention rate is represented by the ratio of the diameter of the inhibition zone of the treatment group to the diameter of the control group. The results are shown in Table 2. The bacteriocin activity is basically Unaffected, the activity remained above 90%.

[0043] 2) Sensitivity to enzymes: get the fermentation supernatant of Example 2 and add pepsin, trypsin, pronase, lipase, amylase respectively, the final concentration of enzyme is 1mg / mL, and the pepsin treatment pH is 2.5, The pH of other enzyme treatment was 6.5, and after 2 hours of treatment at 37°C, L.innocua was used a...

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Abstract

The invention provides bacteriocin and a preparing method thereof. The bacteriocin is generated by bacillus coagulans FM603 with the collection number of CGMCC NO.10221, and has antimicrobial activity on Gram-positive pathogenic bacteria such as Listeria monocytogenes, staphylococcus aureus, methicillin resistant staphylococcus aureus, clostridium perfringens and clostridium firmus. The molecular weight of the bacteriocin is 4276.45 Da, a part of the amino acid sequence is Ala-Gly-His-Dhb-Phe-Val-Dhb-Gly-Pro, and the bacteriocin is stable under treatment of heat, acid, pepsin or trypsin and can be easily degraded by pronase to be inactivated. The bacteriocin can be used as microorganism feed additives, the laying rate of laying hens can be increased, the feed-egg ratio can be reduced, and egg quality can be improved; the feed intake and the daily gain of piglets can be increased, and the feed-meat ratio can be reduced; the daily gain of broilers and the content of lysozyme can be increased, and the feed-meat ratio and the death rate can be reduced.

Description

technical field [0001] The invention relates to microorganisms and their cultivation and application technology, in particular to a bacteriocin produced by bacillus coagulans FM603 and a preparation method of the bacteriocin. Background technique [0002] Antibiotics are secondary metabolites produced by microorganisms that have antimicrobial activity against other microorganisms. In the 1940s, scientists discovered that feeding animals low doses of antibiotics could improve animal performance and increase economic efficiency. In the following decades, antibiotics were widely used in the animal husbandry industry, added to feed as a growth promoter, or used as a therapeutic agent in the breeding field. The use of antibiotics has significantly improved the growth efficiency and feed conversion rate of animals, which has played an important role in the development of the animal husbandry industry. But the use of antibiotics has also brought many problems, such as the spread ...

Claims

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Application Information

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IPC IPC(8): C07K14/32C12P21/00C07K1/18C07K1/20A23K1/17
CPCC07K14/32
Inventor 郭耀棋丁毅王丽娟曹岩峰唐日益
Owner 沈阳丰美生物技术有限公司
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