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A kind of extraction and purification method of capsicum antimicrobial peptide

A purification method and antibacterial peptide technology are applied in the field of extraction and purification of capsicum antibacterial peptides, and can solve the problems of unsatisfactory extraction methods and the like.

Active Publication Date: 2021-08-31
SOUTHWEST UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, current extraction methods are still not ideal

Method used

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  • A kind of extraction and purification method of capsicum antimicrobial peptide
  • A kind of extraction and purification method of capsicum antimicrobial peptide
  • A kind of extraction and purification method of capsicum antimicrobial peptide

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0135] 1. Test material

[0136] Pepper seeds: the varieties are Tianying Xiaojiao, Dianthus, Xinhua, Bullet, Indian Pepper and Guizhou Bell Pepper, purchased from Chongqing Murong Agricultural Products Sales Co., Ltd.

[0137] Tested strain: Aspergillus flavus isolated and screened from corn in the early stage, from the Grass Industry Teaching and Research Office of the School of Animal Science, Southwest University.

[0138] 2. Main reagents

[0139] Sample buffer: 10% SDS (W / V), 0.25mol / L Tris, 5% mercaptoethanol (V / V), 50% glycerol (W / V), 0.5% bromophenol blue (W / V), pH Adjusted to 6.8.

[0140] Tris-HCL buffer: 0.02mol / L Tris-HCL, pH adjusted to 7.6.

[0141] PBS buffer: 0.02mol / L, add distilled water to 1L, adjust pH to 7.6

[0142] EDTA buffer: 10mM Na 2 HPO 4 , 15mM NaH 2 PO 4 , 100mM KCl, 1.5% EDTA.

[0143] Acrylamide stock solution: Liquid A (49.5%T, 3%C): 48g of acrylamide, 1.5g of methylene bisacryloyl, distilled water to volume 100mL. Solution B (49.5%T...

Embodiment 2

[0149] Example 2 Antibacterial activity of different varieties of capsicum seed antimicrobial peptides to Aspergillus flavus

[0150] Pepper seeds: the varieties are Tianying Xiaojiao, Dianthus, Xinhua, Bullet, Indian Pepper and Guizhou Bell Pepper, purchased from Chongqing Murong Agricultural Products Sales Co., Ltd.

[0151] Tested strain: Aspergillus flavus isolated and screened from corn in the early stage, from the Grass Industry Teaching and Research Office of the School of Animal Science, Southwest University.

[0152] Pepper seeds were crushed and sieved, extracted with anhydrous ether overnight to remove crude fat. Add it to the EDTA buffer solution at a material-to-liquid ratio of 1:5, extract overnight at 4°C, filter the extract and centrifuge at 8,000 rpm for 15 minutes to take the supernatant, and measure the amount of solid ammonium sulfate required for 40% saturation. After grinding and pulverizing, add slowly, stir evenly to make it fully dissolved, stand stil...

Embodiment 3

[0157] Example 3 Optimization of Extraction Conditions for Capsicum Seed Antimicrobial Peptides——Selection of Buffer

[0158] Take 10 g of degreased chili seed powder, and use 50 mL of A. sterile water; B. PBS; C. EDTA; D. Tris-HCl; E. 75% ethanol. Soak overnight at 4°C, stir to homogenate, filter through 4 layers of gauze, and centrifuge at 12000 rpm for 20 min at 4°C. Ammonium sulfate was added to the supernatant to make the concentration reach 70%. After magnetic stirring for 30 minutes, it was allowed to stand at 4° C. for 4 hours, and then centrifuged at 4° C. at 12,000 rpm for 20 minutes. After the precipitate was redissolved with 10 mL of distilled water, 200 μL of the filtrate was taken to measure the zone of inhibition by the Oxford cup double-layer agar method with Aspergillus flavus as the indicator bacterium.

[0159] The effects of different buffer leaching on the extraction of antimicrobial peptides are as follows: figure 1 with figure 2 shown. Antimicrobial...

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Abstract

The invention discloses an optimized method for extracting and purifying capsicum antimicrobial peptides. Specifically, the inventors have obtained The optimal conditions for the extraction of capsicum antimicrobial peptides. Further, the components with good antibacterial effects were selected after ultrafiltration and crude separation, and the electrophoretic pure antibacterial peptide samples were obtained by Sephadex column chromatography. The extraction and purification method disclosed in the present invention is simple, and the bacteriostatic effect of the extracted antibacterial peptide is increased by 50.32% compared with that before optimization, and has a wider antibacterial spectrum, especially has a strong inhibitory effect on Aspergillus flavus, and can effectively inhibit Aspergillus flavus Spore germination and mycelium growth, and can inhibit the absorption and utilization of nutrients in the medium by Aspergillus flavus. The antimicrobial peptide extracted by the method disclosed in the invention is used as a corn antifungal agent, which can better maintain the quality of corn and prolong the service life, and the effect is better when used in combination with other antifungal agents.

Description

technical field [0001] The invention relates to the field of separation and purification of antimicrobial peptides, in particular to a method for extracting and purifying capsicum antimicrobial peptides. Background technique [0002] Antimicrobial peptides (AMPs), a naturally evolved innate immune barrier, are produced by all organisms. The first antimicrobial peptide discovered in the world was discovered by Swedish scientist G.Boman and others in the silkworm chrysalis of Xigubi, and then people successively discovered it from bacteria, fungi, amphibians, insects, higher plants, mammals and even humans. And isolate the polypeptide with antibacterial activity. As an important part of the innate immune system, antimicrobial peptides have natural antimicrobial effects and are not prone to drug resistance. Due to the protein characteristics, non-pollution, safety and antibacterial properties of antimicrobial peptides, it has a good advantage in research on the substitution o...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K1/36C07K1/34C07K1/30C07K1/16
CPCC07K1/16C07K1/303C07K1/34C07K1/36
Inventor 韩玉竹杨叶梅曾兵赵建军赵展展陈强廖欢科吴长凤
Owner SOUTHWEST UNIV
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