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Bacillus cereus and application thereof to preparation of nitrite reductase

A technology of bacillus cereus and nitrite, which is applied in the application field of preparing nitrite reductase, and achieves the effects of low fermentation production cost, good commercial prospects, and simple conditions

Active Publication Date: 2014-12-17
SOUTH CHINA UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In addition, when degrading nitrite in food and food sources, the first thing to consider is to ensure food safety, but the above-mentioned oxidation method must strengthen the oxidant, add iron filings in the reduction method, and Pseudomonas in microbial degradation must have certain toxicity. It will affect food safety, so it is imperative to seek a method that is safe to eat and has a good effect on degrading nitrite

Method used

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  • Bacillus cereus and application thereof to preparation of nitrite reductase
  • Bacillus cereus and application thereof to preparation of nitrite reductase
  • Bacillus cereus and application thereof to preparation of nitrite reductase

Examples

Experimental program
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Effect test

Embodiment 1

[0035] Pick a loop of bacteria from the slant of the LJ01 test tube into a 250mL Erlenmeyer flask containing 100mL of seed medium, and cultivate it on a constant temperature shaker at 30°C and 180r / min for 24h to obtain the LJ01 seed solution. The composition of the seed medium is: 10g of peptone, 3g of beef extract powder, 5g of NaCl, 1L of distilled water, and the final pH is 7.4±0.2.

[0036] Prepare a fermentation induction medium containing 50mg / mL ~ 100mg / mL sodium nitrite, adjust its pH to 7.4, sterilize and cool at 121°C, and insert the LJ01 seed solution according to the inoculation amount of 1% ~ 5% (volume percentage) After culturing at 30°C to 40°C and 150r / min to 250r / min for 18h to 30h, the fermented liquid obtained was centrifuged at 4°C and 9000r / min for 10min, the supernatant was removed, the sludge was washed with sterile water and placed in Centrifuge at 4°C and 9000r / min for 10min, repeat the washing and centrifuging operations twice, get bacteria and weigh...

Embodiment 2

[0043] The method for producing and preparing NiR by bacillus cereus

[0044] The fermentation induction medium (broth medium) containing 100mg / mL sodium nitrite and pH 7.4 was sterilized and cooled at 121°C, and then inserted into the LJ01 seed solution according to the inoculum size of 2% (volume percentage), at 30 After culturing for 24 hours at ℃ and 250r / min, the fermented liquid was centrifuged at 4℃ and 9000r / min for 10min, removed the supernatant, washed with sterile water, centrifuged at 4℃ and 9000r / min for 10min, washed and centrifuged The operation was repeated twice, the cells were obtained and weighed, and the enzyme extraction buffer (containing 3 μg / mL trypsin inhibitor and 0.5 μmol / mL DTT) was added to make the concentration of LJ01 cells 200 mg / mL, and lysozyme was added , so that the concentration of lysozyme was 20mg / mL, after mixing, the LJ01 cell wall was broken at 30°C and 180r / min for 3h, and then centrifuged at 8000r / min for 20min at 4°C, the supernata...

Embodiment 3

[0052]Prepare a fermentation induction medium containing 75 mg / mL sodium nitrite and adjust the pH to 7.4. After sterilizing and cooling at 121°C, insert the seed liquid at a 3% (volume percentage) inoculum size, and place it at 35°C and 180r / min. After culturing for 20 hours, the fermented liquid was centrifuged at 4°C and 9000r / min for 10min, the supernatant was removed, the bacteria sludge was washed with sterile water and then centrifuged at 4°C and 9000r / min for 10min, and the cleaning and centrifugation operations were repeated twice. The thalline was obtained and weighed, adding enzyme extracting buffer (containing 1 μg / mL trypsin inhibitor and 2 μmol / mL DTT) to make the concentration of LJ01 thallus 300 mg / mL, adding lysozyme to make the concentration of lysozyme 5 mg / mL, after mixing, break the LJ01 cell wall at 30°C and 180r / min for 4h, then centrifuge at 8000r / min for 20min at 4°C, the supernatant after centrifugation is NiR crude enzyme solution, and its NiR enzyme...

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Abstract

The invention discloses a Bacillus cereus and application thereof to preparation of nitrite reductase. The Bacillus is isolated from thick broad-bean sauce, and is identified as Bacillus cereus LJ01, which has been preserved in China General Microbiological Culture Collection Center in June 19, 2014 and has a preservation number CGMCC No.9360. The strain LJ01 is used for the preparation of nitrite reductase (NiR); NiR extracted from Bacillus cereus LJ01 has high activity; and conditions for NiR to degrade nitrite are simple. The conditions disclosed by the invention can reduce NiR enzyme activity loss to obtain high purity NiR; and NiR provides a novel method for solving the problem of nitrite in three fields of fermented vegetables, meat products and aquaculture.

Description

technical field [0001] The invention relates to the field of biological products, in particular to the use of bacillus cereus from traditional soybean paste and its application in the preparation of nitrite reductase. Background technique [0002] Nitrite is a potential carcinogen, which is easy to accumulate in the process of vegetable fermentation, bringing potential food safety problems to the product, and excessive intake of nitrite can induce methemoglobinemia, so strict control of nitrite in food The salt content is very important. Many studies have shown that nitrite is the precursor of nitrosamines, which are strong carcinogens that can induce various cancers in the digestive system, such as gastric cancer, intestinal cancer and liver cancer. In view of a series of problems such as the possibility of excessive nitrite pollution in food, the potential food safety problems of nitrite in meat products, and the potential food safety problems caused by excessive nitrite ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N9/06C12R1/085
Inventor 刘冬梅罗彤晖郭均吴晖李晓凤袁琨
Owner SOUTH CHINA UNIV OF TECH
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