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Preparation method of porcine small intestine antibacterial peptides PR39

A pig small intestine, antimicrobial peptide technology, applied in the field of biopharmaceuticals, can solve the problems of waste cost, complicated operation, low yield, etc., and achieve the effects of reducing pollution, high purity and simple method

Inactive Publication Date: 2012-12-12
ZHENGZHOU HOUYI PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Due to the complex structure of intestinal tissue and various types of miscellaneous proteins, the current extraction technology of antimicrobial peptides is complicated to operate, and often requires multiple separations and purifications to obtain purified antimicrobial peptides, which is a waste of cost and the yield is not high.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] The preparation method of the porcine small intestine antimicrobial peptide PR39 provided in this example, the specific steps are as follows:

[0021] 1) Take pig small intestine, remove fat and contents, weigh 20g of small intestinal mucosa, and rinse with sterilized physiological saline;

[0022] 2) According to the ratio of 20ml / g, add 5% acetic acid ice-water mixture by volume fraction, and make a homogenate through a tissue crushing machine;

[0023] 3) 100°C water bath for 10 minutes, then rapidly cooled to 0°C;

[0024] 4) Centrifuge at 8000rpm for 30min at 4°C, take the supernatant and put it on ice for 30s of ultrasonic cracking, stir and extract at 4°C, centrifuge at 8000rpm for 30min, take the supernatant;

[0025] 5) Mix the centrifuged precipitate with an equal volume of 5% acetic acid, stir and extract again, centrifuge at 8000rpm for 30min, and take the supernatant;

[0026] 6) Combine the supernatant obtained in step 4) and step 5), adjust the pH=6, ce...

Embodiment 2

[0030] The preparation method of the porcine small intestine antimicrobial peptide PR39 provided in this example, the specific steps are as follows:

[0031] 1) Take pig small intestine, remove fat and contents, weigh 20g of small intestinal mucosa, and rinse with sterilized physiological saline;

[0032] 2) According to the ratio of 20ml / g, add 4.5% volume fraction of acetic acid ice-water mixture, and make a homogenate through a tissue crushing machine;

[0033] 3) Water bath at 90°C for 10 minutes, then rapidly cool to 5°C;

[0034] 4) Centrifuge at 8000rpm for 30min at 4°C, take the supernatant and put it on ice for 30s of ultrasonic cracking, stir and extract at 4°C, centrifuge at 8000rpm for 30min, take the supernatant;

[0035] 5) Mix the centrifuged precipitate with an equal volume of acetic acid with a volume fraction of 4.5%, stir and extract again, centrifuge at 8000rpm for 30min, and take the supernatant;

[0036] 6) Combine the supernatant obtained in step 4) an...

Embodiment 3

[0040] The preparation method of the porcine small intestine antimicrobial peptide PR39 provided in this example, the specific steps are as follows:

[0041] 1) Take pig small intestine, remove fat and contents, weigh 20g of small intestinal mucosa, and rinse with sterilized physiological saline;

[0042] 2) According to the ratio of 20ml / g, add 4% acetic acid ice-water mixture by volume fraction, and make a homogenate through a tissue crushing machine;

[0043] 3) Water bath at 95°C for 10 minutes, then rapidly cool to 10°C;

[0044] 4) Centrifuge at 8000rpm for 30min at 4°C, take the supernatant and put it on ice for 30s of ultrasonic cracking, stir and extract at 4°C, centrifuge at 8000rpm for 30min, take the supernatant;

[0045] 5) Mix the centrifuged precipitate with an equal volume of 4% acetic acid, stir and extract again, centrifuge at 8000rpm for 30min, and take the supernatant;

[0046] 6) Combine the supernatant obtained in step 4) and step 5), adjust the pH=5, c...

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PUM

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Abstract

The invention discloses a preparation method of porcine small intestine antibacterial peptides PR39, which comprises the following specific steps of: 1, taking porcine small intestinal mucosa and flushing; 2, adding a mixture of acetic acid and ice water with the volume fraction of 4-5%, and smashing to be made into a homogenate; 3, carrying out a boiling water bath for 10min, and cooling to 0-10 DEG C; 4, centrifuging, cracking a supernatant by ultrasonic waves for 30s, stirring and extracting, centrifuging, and taking the supernatant; 5, mixing a precipitate with acetic acid with the volume fraction of 4-5%, stirring and extracting, centrifuging, and taking the supernatant; 6, combining the supernatants, centrifuging and taking the supernatant; 7, sampling the supernatant to a Sephadex G2100 gel column for chromatography, and collecting an eluate; and 8, detecting the antibacterial property of liquid in each collection tube by an agarose diffusion method, collecting the eluent with an antibacterial effect to obtain the porcine small intestine antibacterial peptides PR39, and storing by freezing and drying in a vacuum. The process is simple, the cost is low, most of impure proteins are removed, and the extraction rate is high.

Description

technical field [0001] The invention relates to a preparation method of porcine small intestine antimicrobial peptide PR39, which belongs to the technical field of biopharmaceuticals. Background technique [0002] In recent years, the number of pig diseases has been increasing, especially the abuse of drugs has made many bacteria resistant to drugs, and the emergence of some mutant strains has added difficulties to the prevention of complex pig diseases, and has greatly hindered the development of the pig industry. Piglet diarrhea and various viral diseases such as swine fever respiratory and reproductive syndrome have always been difficult diseases in the breeding industry. At present, the health care of sows and piglets is more and more recognized by people, but this is also a difficult problem in front of people. The application of antimicrobial peptides can solve this problem well. [0003] Antimicrobial peptides are a kind of small defensive peptides produced by organ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/47C07K1/16
Inventor 吴红云韩改会徐进
Owner ZHENGZHOU HOUYI PHARMA
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