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Preparation method and application of bacillus antibacterial peptide

A technology of bacillus and antibacterial peptides, applied in the field of feed microbial additives, can solve the problem of less antibacterial peptides, and achieve the effects of reducing fecal odor, promoting the use of growth-promoting drugs, and reducing drug use

Inactive Publication Date: 2017-01-04
BEIJING DAWEIJIA BIOTECH SHARE CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, antimicrobial peptides are rarely used as feed additives. This study is an antimicrobial peptide developed for feed additives.

Method used

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  • Preparation method and application of bacillus antibacterial peptide
  • Preparation method and application of bacillus antibacterial peptide
  • Preparation method and application of bacillus antibacterial peptide

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Embodiment one: bacillus starch liquefaction ( B. amylolique ) Determination of antibacterial activity of ES-2-4 metabolite antimicrobial peptide WJ-1

[0019] 1. Preparation of Culture Supernatant

[0020] from Bacillus amyloliquefaciens ( B. amylolique ) ES-2-4 preserved slant picked a ring of bacterial lawn and inoculated it in the broth medium, 37 ° C, 180 rpm shaking culture for 22 h, took 1 mL of bacterial liquid centrifuged at 12000 rpm for 1 min, and took the supernatant.

[0021] 2. Preparation of Antibacterial Activity Assay Plate

[0022] Pour 15 mL of completely melted LB agar medium into a petri dish, and place it vertically in an Oxford cup after it solidifies; take another completely melted LB agar medium, add indicator bacteria and mix it evenly when it is cooled to 55°C, and take Pour 10 mL of medium mixed with indicator bacteria into a petri dish with an Oxford cup, and pull out the Oxford cup after it is completely solidified to make a test plate...

Embodiment 2

[0025] Embodiment two: bacillus starch liquefaction ( B. amylolique ) Quantitative determination of ES-2-4 metabolite antimicrobial peptide WJ-1

[0026] from Bacillus amyloliquefaciens ( B. amylolique ) ES-2-4 preserved slant picked a ring of bacterial lawn and inoculated it in the broth medium, 37 ° C, 180 rpm shaking culture for 22 h, took 1 mL of bacterial liquid centrifuged at 12000 rpm for 1 min, and took the supernatant. Antimicrobial peptide powder was dissolved in deionized aqueous solution according to 5%. The content of subtilisin antimicrobial peptide WJ-1 was determined by high performance liquid chromatography. Chromatographic column: Poroshell 120 EC-C8 2.7μm 3.0*50mm; chromatography method: flow rate 0.8mL / min, column pressure 200bar; mobile phase A: water + 0.1% TFA, mobile phase B: 80% acetonitrile + 0.085% TFA; column Temperature: 30°C; injection volume is 50μl, each sample is tested 3 times, and the internal control product is tested again after the sa...

Embodiment 3

[0029] Embodiment three: bacillus starch liquefaction ( B. amylolique ) ES-2-4 metabolite antimicrobial peptide WJ-1 is resistant to artificial gastrointestinal digestion.

[0030] from Bacillus amyloliquefaciens ( B. amylolique ) After the ES-2-4 fermentation broth was centrifuged, the fermentation metabolite antimicrobial peptide WJ-1 was obtained. Take 10 mL of fermentation metabolite antimicrobial peptide WJ-1 and connect them to 90 mL of artificial gastric juice with pH 3=3.5 and artificial pancreatic juice with pH=6.5 respectively. Then cultured at 37°C for 2 hours, and measured the antibacterial activity of the culture at 0, 0.5, 1, 1.5, and 2 h by the inhibition zone method, that is, the diameter of the inhibition zone. Antibacterial activity of antimicrobial peptides.

[0031] The results showed that Bacillus amyloliquefaciens ( B. amylolique) ES-2-4 metabolite antimicrobial peptide WJ-1 still has antibacterial activity after artificial gastric juice and pancre...

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Abstract

The invention relates to a preparation method and application of bacillus antibacterial peptide. A Bacillus amylolique strain ES-2-4 used in the invention can produce antibacterial peptide WJ-1 which exerts antibacterial effect on Escherichia coli, Salmonella, Micrococcus luteus and Staphylococcus aureus and has a bacterium inhibition zone diameter of 20 to 30 mm. The Bacillus amylolique strain ES-2-4 undergoes inoculation, preparation of shake-flask seeds, production of primary seed liquid, secondary fermentation and solid-liquid separation; then obtained supernatant is mixed with a carrier and spray drying is carried out on the obtained mixture so as to obtain dried active bacterium powder; and carrier dilution is carried out so as to obtain a feed additive product. The additive is applicable to livestock and poultry in the whole course of culture, and can improve the productivity of livestock and poultry, promote growth of livestock and poultry, enhance immunity of livestock and poultry, reduce the mortality of young animals, decrease the excretion of manure, reduce the odor of manure and improve a culturing environment.

Description

technical field [0001] The invention relates to a preparation method of a bacillus antibacterial peptide and its application in livestock and poultry breeding, especially suitable for livestock and poultry feed additives, and belongs to the technical field of feed microbial additives. Background technique [0002] The food safety of livestock and poultry products brought about by the long-term use of antibiotics in livestock and poultry feed has attracted more and more attention, because the abuse of antibiotics leads to the destruction of the normal flora balance in the animal intestines, and at the same time, the abuse of antibiotics leads to resistance. The emergence of drug properties and drug residues have brought great threats and harm to the health of animals and humans. Therefore, seeking effective, safe, green and natural alternatives to antibiotics in feed has become a hot issue in the livestock and poultry industry. . [0003] Antimicrobial peptides are natural a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P21/02A23K1/16A23K1/18C12R1/07
Inventor 乔琳崔栩金忠辉张晓华王兰孙菡聪
Owner BEIJING DAWEIJIA BIOTECH SHARE CO LTD
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