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Method for preparing alpha-ketobutyric acid

A technology of ketobutyric acid and sodium ketobutyrate, applied in the direction of microorganism-based methods, biochemical equipment and methods, separation/purification of carboxylic acid compounds, etc., to achieve high conversion rate, convenient operation, and simple separation and extraction Effect

Inactive Publication Date: 2010-12-29
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

There is no report on the method of converting α-hydroxybutyric acid into α-ketobutyric acid by using microbial cells containing α-hydroxyacid oxidase

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  • Method for preparing alpha-ketobutyric acid
  • Method for preparing alpha-ketobutyric acid

Examples

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Embodiment 1

[0039] (1) Preparation of complete cell suspension containing α-hydroxyacid oxidase: Pseudomonas putida KT2440 (ATCC 47054) was selected for routine culture, during which 2,4-dinitrophenylhydrazine (DNP) and α-ketone The method of generating brown-red 2,4-dinitrophenylhydrazone by the action of butyric acid was used to detect the activity of α-hydroxyacid oxidase in cells, and when the activity of α-hydroxyacid oxidase reached 150 units / liter, the fermentation culture was terminated; separated and collected The thalline was washed 2 times with a pH 7.2 potassium phosphate buffer solution, and the thalline was resuspended in deionized water, so that the thalline concentration reached 200 grams of wet cells / liter, and a complete cell suspension containing α-hydroxyacid oxidase was obtained. Store at 4°C for later use;

[0040] (2) Transformation: the complete cell suspension prepared in step (1) is mixed with an aqueous solution of sodium α-hydroxybutyrate, and the concentration...

Embodiment 2

[0048] (1) Preparation of complete cell suspension containing α-hydroxyacid oxidase: Pseudomonas aeruginosa (ATCC15442) was selected for routine culture, during which 2,4-dinitrophenylhydrazine (DNP) and α-ketobutyl The method of generating brown-red 2,4-dinitrophenylhydrazone by the action of acid detects the activity of α-hydroxyacid oxidase in cells, and when the activity of α-hydroxyacid oxidase reaches 160 units / liter, stop the fermentation culture; separate and collect the bacteria , wash the cells twice with pH 7.2 potassium phosphate buffer, resuspend the cells in deionized water, make the cell concentration reach 200 g wet cells / liter, and obtain a complete cell suspension containing α-hydroxyacid oxidase, at 4°C store for later use;

[0049] (2) Transformation: The complete cell suspension prepared in step (1) is mixed with an aqueous solution of sodium α-hydroxybutyrate, and the concentration of sodium α-hydroxybutyrate in the mixture is 200 mmol / L, the concentratio...

Embodiment 3

[0057] (1) Preparation of complete cell suspension containing α-hydroxyacid oxidase: select Pseudomonas stutzeri A1501 (CGMCC0351), and conduct routine culture, during which 2,4-dinitrophenylhydrazine (DNP) and α-ketone The method of generating brown-red 2,4-dinitrophenylhydrazone by the action of butyric acid was used to detect the activity of α-hydroxyacid oxidase in cells, and when the activity of α-hydroxyacid oxidase reached 160 units / liter, the fermentation culture was terminated; separated and collected The thalline was washed 2 times with a pH 7.2 potassium phosphate buffer solution, and the thalline was resuspended in deionized water, so that the thalline concentration reached 200 grams of wet cells / liter, and a complete cell suspension containing α-hydroxyacid oxidase was obtained. Store at 4°C for later use;

[0058] (2) Transformation: The complete cell suspension prepared in step (1) is mixed with an aqueous solution of sodium α-hydroxybutyrate, and the concentrat...

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Abstract

The invention discloses a method for preparing alpha-ketobutyric acid by converting alpha-hydroxybutyric acid by cells containing alpha-hydroxyacid oxidase, which comprises: (1) preparing suspension of complete cells containing alpha-hydroxyacid oxidase; allowing the complete cells to convert alpha-hydroxybutyric acid into alpha-ketobutyric acid; (3) pretreating liquid of the conversion; (4) decolorizing and concentrating the liquid of the conversion; (5) crystallizing the alpha-ketobutyric acid; and the like. The method has the characteristics that: the culture medium is simple; the growth period is short; the cost is low; the subsequent separation and extraction cost is low; the substrate concentration resistance is high; the purity of the alpha-ketobutyric acid product is high; and the like. The method lays a foundation for the efficiency production of alpha-ketobutyric acid.

Description

technical field [0001] The invention relates to a method for preparing α-ketobutyric acid, in particular to a method for preparing α-ketobutyric acid by converting α-hydroxybutyric acid with intact cells of microorganisms. Background technique [0002] α-ketobutyric acid (α-ketobutyric acid), also known as 2-oxobutyric acid (2-Oxobutyric acid), 2-oxobutanoic acid (2-Oxobutanoic acid), 2-ketobutyric acid (2-Ketobutyric acid). α-ketobutyric acid is the precursor of many useful compounds, and has a wide range of applications in the pharmaceutical and food synthesis industries, such as the synthesis of isoleucine, baolomycin and furanone, etc. "Production of 2-ketobutyric acid from 1 , 2-butanediol by resting cells of Rhodococcusequi IFO 3730. Biotechnol. Lett. 1994, 16: 263-268. ". [0003] The production methods of α-ketobutyric acid include chemical synthesis and biotechnology. The chemical synthesis of α-ketobutyric acid can be carried out by the mixed hydrolysis of dieth...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P7/40C07C59/185C07C51/42C12R1/01C12R1/38C12R1/385C12R1/40
Inventor 马翠卿高超张文许平
Owner SHANDONG UNIV
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