Biocatalysis method for preparing Alpha ketoglutarate from L-soda glutamate

A technology of ketoglutaric acid and sodium glutamate, applied in the biological field, can solve problems such as difficult expansion of industrial production

Active Publication Date: 2013-10-16
JIANGNAN UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In recent years, more and more people have turned their attention to microorganisms. Although a lot of work and efforts have been made, most of the research results are limited to laboratory research, and it is difficult to expand to industrial production.

Method used

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  • Biocatalysis method for preparing Alpha ketoglutarate from L-soda glutamate
  • Biocatalysis method for preparing Alpha ketoglutarate from L-soda glutamate
  • Biocatalysis method for preparing Alpha ketoglutarate from L-soda glutamate

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Embodiment 1, utilize single factor experiment to determine the optimal concentration of maltose.

[0019] After determining the optimum carbon source as maltose, the concentration gradient of 5g / L, 10g / L, 15g / L, 20g / L, 25g / L, 30g / L, 35g / L was selected, and the effect of maltose concentration on bacteria was studied. Effects on body growth and transformation ability. After 48 hours of culture in shake flasks, take 30ml of fermentation broth in a 50ml centrifuge tube and centrifuge at 8000rpm for 5min, discard the supernatant, resuspend the bacteria with 30ml of transformation solution and pour them into shake flasks. After 24 hours of down-conversion, samples were taken every 6 hours to determine the production of α-KG by HPLC.

Embodiment 2

[0020] Embodiment 2, under optimal culture medium and culture condition, measure the growth of bacterium and the situation of enzyme production.

[0021] In order to further improve the transformation ability of the bacteria, the growth characteristics and enzyme production characteristics of the bacteria under the optimal nutritional conditions and culture conditions were studied. Selected 9 shake flasks (500L in size) and divided them into three groups. The volume of each shake flask was 50ml, and 5ml of seed liquid was inserted into them. They were cultivated at 200r / m and 30°C for 48h, and samples were taken every few hours and collected. Measurement data.

Embodiment 3

[0022] Embodiment 3, the impact of different bacterial cell concentrations on transformation

[0023] Due to the relatively stable transformation performance of the bacteria, the bacteria concentration (dry weight) of 2g / L, 4g / L, 6g / L, 8g / L, and 12g / L was set to study the effect of the bacteria concentration on the conversion rate and α-KG output Impact. Measure 15ml, 30ml, 45ml, 60ml, 90ml of fermentation broth after 48 hours of fermentation (cell concentration is 4g / L), centrifuge at 8000rpm for 5min, resuspend cells with 30ml of transformation solution and pour into shake flask at 30℃, 200rpm The transformation was carried out for 6 hours under the conditions, and after 6 hours, a sample was taken to measure the yield of α-KG by HPLC.

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Abstract

The invention expounds a biocatalysis method for preparing Alpha ketoglutarate from L-soda glutamate, and belongs to the biotechnological field. The method comprises the following steps: utilizing strains (Rhodococcus opacus) producing L-amino acid oxidase, performing fermentation for 48 hours, collecting thallus, weighing and pouring 12 g / L (calculating by dry weight) thallus into a 10 g / L L-soda glutamate contained conversion solution, performing conversion at the temperature of 35 DEG C and the pH of 8.4 for 10 hours, adding 4.3 g / L L-soda glutamate into a reaction system every 6 hours, and performing continuous conversion for 24 hours, where the output of Alpha-ketoglutarate reaches 16.8 g / L, and the conversion rate reaches more than 90 percent.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to a novel process for producing α-ketoglutaric acid by microorganism catalysis. Specifically, the present invention relates to a new method for catalyzing L-sodium glutamate into α-ketoglutarate by a strain producing L-amino acid oxidase (Rhodococcus opacus), which is characterized by high efficiency, economy, environmental protection and low energy consumption. Background technique [0002] α-ketoglutarate (α-KG), also known as α-glutaric acid, 2-oxoglutarate or α-oxoglutarate, the molecular formula is C 5 h 6 o 5 , the relative molecular weight is 146.1, α-ketoglutaric acid is white or off-white crystal or crystalline powder, colorless, easily soluble in water, plays an important role in the metabolism of microbial cells, is the tricarboxylic acid cycle (TCA ) is one of the important intermediate products, and participates in the synthesis of amino acids, proteins, vitamins and ener...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P7/50C12R1/01
Inventor 刘立明徐继嗣董晓翔牛盼清
Owner JIANGNAN UNIV
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