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Bacillus subtilis for degrading aflatoxin and application thereof

A technology of Bacillus subtilis and aflatoxin, applied in the direction of bacteria, microorganism-based methods, microorganisms, etc., can solve the problems of long degradation time and incomplete degradation degradation effect, so as to improve economic benefits, solve pollution problems, and strains safe effect

Active Publication Date: 2020-03-13
SHANXI UNIV +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In order to overcome the deficiencies in the prior art, solve the AF degradation time is long, can not completely degrade AF before the food is discharged from the intestinal tract and the existing strains are sensitive to high concentration AFB1 The technical problem of poor degradation effect, the present invention provides a kind of Bacillus subtilis that degrades aflatoxin with fast degradation rate and high degradation rate and does not destroy the nutritional components in the feed and its application

Method used

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  • Bacillus subtilis for degrading aflatoxin and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1 Isolation and identification of Bacillus subtilis ASAG212 that degrades AF

[0033] 1. Isolation of bacteria

[0034] (1) Multiple soil samples were randomly collected from grain planting areas in Xinzhou, Shanxi Province, and placed in ziplock bags, and the collection name, location, time and other information were indicated. Dilute 1 g of soil sample in 10 mL of sterile distilled water to prepare soil suspension, and use the concentration gradient method to gradually dilute 10, 100, 1000 and 1000 times.

[0035] (2) Spread soil sample suspensions diluted in different concentrations on LB solid plates and incubate at 37°C for 36 hours. , pick strains with different morphological features, colors, and sizes on the plate for plate streak purification, then perform AF degradation test on the purified strains, and analyze the strain with the highest degradation efficiency, the strain number is ASAG212.

[0036] 2. Bacteria identification

[0037] (1) Strain ...

Embodiment 2

[0065] Example 2 Degradation of AF by Bacillus subtilis ASAG212

[0066] 1. AF determination method

[0067] AF was detected by high performance liquid chromatography under the following conditions, Agilent C 18 Column (4.6 mm×150 mm×5 μm); mobile phase: methanol:water (50:50); flow rate: 1.0 mL / min; column temperature: 35˚C; detection wavelength: λex=360 nm, λem=440 nm ;Injection volume: 20 μL. The AF content was obtained by comparing the peak area with the standard curve.

[0068] 2. Degradation of AF by Bacillus subtilis ASAG212

[0069] (1) Inoculate 1 mL of preserved Bacillus subtilis ASAG212 into 50 mL of sterilized LB medium (10 g of tryptone, 5 g of yeast extract powder, 10 g of sodium chloride, 1000 mL of distilled water, pH 7.2-7.6) at 37 After culturing at 160 rpm for 18 h (OD value 0.8), take 900 µL of fermentation broth (concentration of 10 9 CFU / mL) with 100 µL AFB 1 standard mix to make the AFB 1 The final concentration of the standard is 10 μg / mL; at t...

Embodiment 3

[0076] Example 3 Degradation of AFB by Bacillus subtilis ASAG212 1 Product Safety Test

[0077] The TA98 standard strain (Moltox Company, USA) was activated, and TA98 was inoculated into 5 mL of sterilized broth medium, and cultured at 37°C, 150 r / min for 10 h for later use (cultivated in the dark). TA98 has undergone strain identification. These include: identification of histidine deficiency, identification of lipopolysaccharide barrier deficiency, identification of ampicillin resistance, identification of tetracycline resistance, UV-sensitivity properties and spontaneous back-change assay.

[0078] Divide 2.0 mL of the top layer agar medium containing 0.5 mmol / L histidine-0.5 mmol / L biotin solution into test tubes, incubate in a 45˚C water bath, and then add 0.1 mL of 0 h control group (900 µL PBS+100 µL 10 µg / mL AFB 1 reaction 0 h), 4 h control group (900µL PBS+100µL 10 μg / mL AFB 1 reaction for 4 h), 0 h degradation solution (900 µL Bacillus subtilis ASAG212 fermentati...

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Abstract

The invention discloses Bacillus subtilis for efficiently degrading aflatoxin and application thereof, and belongs to the field of biotechnology. The Bacillus subtilis is named ASAG212, and was preserved in the China General Microbiological Culture Collection Center with the preservation number of CGMCC No. 18367. The optimum degradation conditions are as follows: the initial concentration of Bacillus subtilis is 109 CFU / mL, the pH value is 7.2-7.6, the culture temperature is 35-40 DEG C, and the rotational speed is 120 rpm-180 rpm. 10 ug / mL of aflatoxin B1 is degrade into a non-toxic productwithin 4 hours by the strain of the invention, and the degradation rate reaches more than 90%. The invention also provides a bacterial preparation containing the strain and for degrading aflatoxin andan application of the bacterial preparation. The preparation method of the bacterial preparation used by the invention is simple, and the bacterial preparation does not contain antibiotic components,has no toxic or side effects, has no drug resistance, has no pollution, and has remarkable economic benefits and practical application value.

Description

technical field [0001] The invention belongs to the technical field of degradation of beneficial microorganisms, and specifically relates to a strain of bacillus subtilis capable of efficiently degrading aflatoxin and an application thereof. Background technique [0002] Aflatoxins (AF) are a group of fungi mainly composed of Aspergillus, such as Aspergillus flavus ( Aspergillus flavus ), Aspergillus parasitica ( A. parasiticus ) produced toxic secondary metabolites, which are highly toxic, carcinogenic and mutagenic, and seriously threaten animal production and human health. At present, more than 20 forms of AF have been discovered, and AF in agricultural products is mainly represented by B 1 , B 2 , G 1 and G 2 Wait. Among them, aflatoxin B 1 (AFB 1 ) is classified as a Group I carcinogen by the International Agency for Research on Cancer (IARC). According to the estimates of the Food and Agriculture Organization of the United Nations, 25% of the world's crops ar...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20A23L5/20C12R1/125
CPCC12N1/20A23L5/28C12R2001/125C12N1/205Y02P60/87
Inventor 贾如刘文彬范婵娟
Owner SHANXI UNIV
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