Bacillus pumilus for degrading aflatoxin M1 and active proteins secreted by Bacillus pumilus

A technology of Bacillus pumilus and aflatoxin, which is applied in the direction of bacteria, microorganism-based methods, biochemical equipment and methods, etc., can solve the problems of no clear degradation active substances and long time, and achieve significant degradation effect and short reaction time Short, mild effect

Inactive Publication Date: 2016-02-24
HEBEI AGRICULTURAL UNIV.
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Application number 201410292766.1, publication number CN104087528A, "A Bacillus pumilus and its application in degrading aflatoxin", although its 1 There is a certain degradation effect, but the Bacillus pumilus applied for is screened from the plant material lily, and the main products targeted are grain, soy sauce and feed, and the optimal degradation effect can only be achieved after contacting for 96 hours, which takes a long time. And there is no clear main active substance for its degradation

Method used

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  • Bacillus pumilus for degrading aflatoxin M1 and active proteins secreted by Bacillus pumilus
  • Bacillus pumilus for degrading aflatoxin M1 and active proteins secreted by Bacillus pumilus
  • Bacillus pumilus for degrading aflatoxin M1 and active proteins secreted by Bacillus pumilus

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Experimental program
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Effect test

Embodiment 1

[0034] Embodiment 1 The cultivation method of bacillus pumilus of the present invention

[0035]Take Bacillus pumilus, the preservation number is CGMCCNO.10228, 0.3ml (the concentration of viable bacteria is 109CFU / ml), inoculated in 100ml medium for shake flask fermentation culture, the fermentation temperature is 37°C, the fermentation time is 14h, and the pH value is 7.2 , speed 130r / min.

[0036] Wherein, the shake flask fermentation medium is composed of the following components: 6 g of maltose, 8 g of yeast powder, 10 g of sodium chloride, 1000 mL of distilled water, and the pH value is 7.2.

Embodiment 2

[0037] Embodiment 2 active protein preparation method of the present invention

[0038] Take the bacterium liquid after the fermentation of Example 1, centrifuge to remove the bacterium, extract the active protein in the supernatant with 75% ammonium sulfate, precipitate, centrifuge at 5000r / min for 20min, dissolve the obtained precipitate in PBS buffer, freeze-dry and concentrate That is the active protein.

Embodiment 3

[0039] Example 3 bacterial liquid to aflatoxin M 1 Degradation

[0040] Take 900 μL of the bacterial solution prepared in Example 1 and add it to a sterilized centrifuge tube, add 100 μL of AFM with a concentration of 400 ng / mL 1 , mixed evenly and placed in a constant temperature incubator with a temperature of 37°C, sampling for 12 hours, and using high performance liquid chromatography to determine aflatoxin M 1 At the same time, the sterile fermentation medium was used as a blank control, and the detection conditions: the chromatographic column was C18 (250mm×4.6mm, 10 μm); the mobile phase was acetonitrile:water (25:75), and the flow rate was 1ml / min. Excitation wavelength is 360nm, emission wavelength is 410nm, injection volume is 10μL, AFM at 12h 1 The degradation rate is 92.5%.

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Abstract

The invention relates to a Bacillus pumilus for degrading aflatoxin M1, collected under CGMCC NO. 10228 and active proteins secreted by the Bacillus pumilus. The Bacillus pumilus is obtained by separating and screening from excrement of herbivores, and the active proteins are extracellular active proteins. The invention also provides a bacterial culture solution of the Bacillus pumilus or a method of using the active proteins secreted by the Bacillus pumilus to degrade aflatoxin M1, and application of the method. By using the method, the bacterial culture solution of the Bacillus pumilus has a degrading rate for aflatoxin M1 not less than 90% in 12 h at 37 DEG C, and the active proteins have a degrading rate for aflatoxin M1 not less than 90% under the same conditions. The Bacillus pumilus and the active proteins secreted thereby have high detoxification activity, high specificity, high acting speed and mild conditions and are applicable to degrading and eliminating aflatoxin M1 in animal-origin foods, and the Bacillus pumilus has the advantages of high bioactivity and probiotic performance.

Description

technical field [0001] The invention relates to Bacillus pumilus, in particular to a method for degrading aflatoxin M 1 Bacillus pumilus and its secreted active protein. Background technique [0002] Aflatoxin (AF) is a kind of highly toxic secondary metabolites with similar structure and physicochemical properties produced by Aspergillus flavus and Aspergillus parasiticus in the genus Aspergillus. Aflatoxin B has been found 1 , B 2 , G 1 , G 2 , M 1 , M 2 There are more than 20 kinds. AFB 1 and AFM 1 In 1993 and 2002, it was identified as a class I carcinogen by the International Agency for Research on Cancer of the World Health Organization. AFM 1 Mainly consumed by animals containing AFB 1 It is metabolized and transformed in the body after the feed, so it mainly appears in animal-derived foods, such as milk and milk products. AFM was detected in a batch of dairy products produced by a dairy industry in 2011 1 The content exceeds the standard, and its harm i...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12P21/02A23L5/20C12R1/07
Inventor 桑亚新崔玉琦武瑞霞桑雨芃
Owner HEBEI AGRICULTURAL UNIV.
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