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Method for producing high-yield gamma-propalanine and application thereof

A high-yield technology of aminobutyric acid, which is applied in the fields of medical biology and food engineering, and can solve problems such as hidden safety and sanitation

Inactive Publication Date: 2011-09-07
TIANJIN UNIVERSITY OF SCIENCE AND TECHNOLOGY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Most of the strains disclosed in the prior art use Escherichia coli as the strain for fermenting GABA, but there are hidden dangers in safety and health

Method used

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  • Method for producing high-yield gamma-propalanine and application thereof
  • Method for producing high-yield gamma-propalanine and application thereof
  • Method for producing high-yield gamma-propalanine and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] (1) Screening of starting strains: From more than 40 samples of sauerkraut (soup) naturally pickled in northern my country, the double-layer plate method was used, that is, containing 5% calcium carbonate and GABA, cultured at 30°C for 48 hours, and selected. The colonies in the transparent circle were fermented and the GABA content was detected by high-performance liquid chromatography. Five strains of GABA-producing strains were obtained, and one strain of Lactobacillus with a yield of 1.4g / L was screened out, numbered S1, and transferred to an inclined plane and stored in a refrigerator at 4°C. .

[0060] (2) Ultraviolet-acridine orange mutagenesis: Take the well-grown Lactobacillus brevis S1 seed solution, irradiate with ultraviolet light for 90s, and then transfer 600 μL of the ultraviolet-mutated bacterial suspension into a test tube containing 5 mL of mutagenic medium Incubate in a dark water bath for 12 hours at 30°C with shaking. (The mutagenesis medium contains...

Embodiment 2

[0070] A kind of high-yielding Lactobacillus brevis Lactobacillus brevis The new process flow diagram of CGMCCNO.3414 production of γ-aminobutyric acid is shown in the attachment figure 1 .

[0071] will activate the Lactobacillus brevis CGM CCN 0 The .3414 strain was inoculated in the seed medium for cultivation, and was transferred to the fermentation medium when the bacterial mass reached the fermentation conditions.

[0072] Seed medium (g / L) and cultivation method: corn saccharification solution 20, peptone 10, yeast extract powder 5, ammonium citrate 2, magnesium sulfate 0.58, manganese sulfate 0.25, sodium acetate 2, potassium dihydrogen phosphate 2. Seed culture conditions: 30 ° C incubator, static culture for 12 hours. According to the inoculation amount of 10-15%, it is inserted into a 5L fermenter.

[0073] Fermentation medium (g / L) and culture method: corn saccharification liquid 20, yeast extract powder 20, corn steep liquor 10, sodium acetate 2, potassiu...

Embodiment 3

[0078] will activate Lactobacillus brevis CGM CCN 0 .3414 strain was inoculated in the seed medium, and cultured statically at 30°C for 10-12 hours.

[0079] Seed medium composition (g / L): peptone 10, yeast extract 10, glucose 20, sodium acetate 5, ammonium citrate 2, MnSO 4˙ h 2 O 0.05, MgSO 4˙ 7H 2 0 0.2, Tween 80 1mL, distilled water 1000 mL, pH control 5.0-6.0. Transfer to the fermentation medium containing 8% L-sodium glutamate at an inoculum size of 10-15%.

[0080] Fermentation medium (g / L): corn steep liquor 10, glucose 22, yeast powder 15, sodium acetate 2, magnesium sulfate 0.2, (NH 4 ) 2 SO 4 0.2, pH6.0. After 16 hours of cultivation, the growth of the bacteria entered the logarithmic phase, and the pH was adjusted to 4.6 and kept at 4.6. After 60 hours of cultivation, the GABA production reached a maximum of 76.52g / L. After culturing the cells for 60 h, the cells were collected, centrifuged for 10 min, 8000 rpm, 4 °C, washed twice with 0.2 mM sodium phos...

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Abstract

The invention relates to a method for producing high-yield gamma-propalanine and an application thereof, belonging to the technical field of biotechnology medicine and food engineering. The method comprises the following steps: carrying out strain separated screening, acridine orange-ultraviolet ray mutagenesis and N<+> injection mutagenesis to obtain a high-yield gamma-propalanine LactobacillusbrevisTCCC (CGMCC No.3414) strain; optimizing a fermentation medium and fermentation conditions; coupling fermentation of the gonotokont of the strain with resting cell biotransformation of the strain to produce the gamma-propalanine; employing the membrane filtration technology, the adsorbent resin decoloration technology, the strong acid resin cation exchange technology, the ethanol recrystallization technology, the preparative chromatography technology and the energy-saving and cost-reducing technology to separate the gamma-propalanine from the fermentation fluid and the biotransformation fluid; and then carrying out purification to obtain the crystallized gamma-propalanine with the purity of 99%. The method has the advantages of cheap raw materials, low energy consumption for production, low production cost, good product security and easy realization of industrialized production. Applied to glutamic acid fermentation waste liquor for producing the gamma-propalanine, the strain has good potential for generating social and economic benefits.

Description

technical field [0001] The invention belongs to the technical field of medical biology and food engineering, is an organic combination of modern biological and chemical engineering and traditional fermentation engineering technology, and is an improvement on the current production method of gamma-aminobutyric acid (GABA) by biotechnology. More specifically, Lactobacillus brevis isolated and bred from sauerkraut in northern my country Lactobacillus brevis CGMCCNO.3414, through the biotransformation of growing cells and resting cells, efficiently produces the internationally recognized safest API and health food - γ-aminobutyric acid. Background technique [0002] γ-Aminobutyric acid (GABA), also known as aminobutyric acid, is a natural amino acid that is not a protein component. It exists widely in nature and is an important inhibitory neurotransmitter that has been studied in depth. Metabolic activity has the effects of lowering blood pressure, regulating arrhythmia, impro...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12N13/00C12N15/01C12P13/00C12R1/24
CPCY02P20/10Y02P20/50
Inventor 高年发张颖高强宋磊冯宇张健王德培
Owner TIANJIN UNIVERSITY OF SCIENCE AND TECHNOLOGY
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