Method for extracting l-4-hydroxyisoleucine from fermentation catalytic liquid

Abstract

The invention relates to the technical field of separation and purification, in particular to a method for extracting L-4-hydroxyisoleucine from fermentation catalyst liquid. The steps of micro-filtration membrane filtering, super-filtration membrane filtering, ion exchange and the like are used for purifying and separating the L-4-hydroxyisoleucine in the L-4-hydroxyisoleucine fermentation catalyst liquid; the extraction system is simple and efficient; the flow process is simple; the implementation is easy; 732 cation resin adsorption is performed on filtration membrane clear liquid; ammoniawater with the ammonia content being 1.2 to 1.5 weight percent is used for elution; the ammonia salt content in the finished product can be reduced; the purity is improved; the alcohol sedimentation is used in primary crystallization; the yield of the primary coarse product can be improved by 50 percent or higher; in secondary crystallization, the alcohol sedimentation is used, the yield of a finished product can be improved by 50 percent or higher; in addition, the product liquid phase purity can be obviously improved; the final product total yield can reach 50 percent; the product purity isnot less than 95 percent. The method has the advantages that the efficiency is high; the cost is low; the large-scale industrial production is easy.

Description

Technical field

[0001] The present invention relates to the field of separation and purification, and more particularly to a method of extracting L-4-hydroxyobine extracting a fermentation catalyst. Background technique

[0002] L-4-hydroxyisoleucine (4-hydroxyisyisoleucine) is a non-protein amino acid present in nature. The study found that L-4-hydroxybiobiline has many aspects of promoting insulin secretion, improving insulin tolerance and regulation of blood lipids. The L-4-hydroxyisineluminine can not only promote the intake of glucose, but also reduce liver glucose output to improve insulin tolerance of the outer peripheral tissue. In contrast, insulin synthesis of L-4-hydroxybiobiline regulatory insulin-controlled insulin synthesis is strictly dependent on glucose concentration compared to current drugs that have been used to treat T2DM. Therefore, L-4-hydroxybiobine does not cause side effects such as hypoglycemia. At the same time, the L-4-hydroxybiobine can also effectiv...

Examples

Embodiment 1

[0057] Method for extracting L-4-hydroxybiobine in fermentation catalysts comprising the steps of:

[0058] (1) L-4-hydroxyisineluminine fermentation catalytic solution was prepared by the following method:

[0059] 1 After the concentration of Escherichia coli resection of L-4-hydroxyisoleine, it is used in seed tank medium containing final concentration of 0.1 g / L amp, at 37 ° C, pH 7.0, and solubol control. Cultured in more than 25% of the OD of the bacteria 600 The value is 15 to obtain a seed liquid;

[0060] 2 The seed fluid obtained by step (1) is 10% in the fermentation tank medium according to the volume percentage of 15%, and the liquid is deeply ventilated under 36 ° C and then cooling to 32 ° C and the final concentration is 0.1 mmol / L. IPTG continues to culture until fermentation is completed, wherein the fermentation process residue is 0.05% by weight, and the oxygen is controlled after 20 ~ 35%, pH 7.0, and the amount of glucose is carbon. source;

[0061] 3 fer...

Embodiment 2

[0068] Method for extracting L-4-hydroxybiobine in fermentation catalysts comprising the steps of:

[0069] (1) L-4-hydroxyisineluminine fermentation catalytic solution was prepared by the following method:

[0070] 1 After the concentration of Escherichia coli resection of L-4-hydroxyisoleine, it is used in seed tank medium containing final concentration of 0.1 g / L amp, at 37 ° C, pH 7.0, and solubol control. Cultured in more than 25% of the OD of the bacteria 600 The value is 14 to obtain a seed liquid;

[0071] 2 The seed fluid obtained by step (1) is incorporated into the fermentor medium according to the volume percentage of 15%, and the liquid deep ventilated fermentation was cultured under 35 ° C and then cooled to 30 ° C and the final concentration was 0.1 mmol / L. IPTG continues to culture until fermentation is completed, wherein the fermentation process residue is 0.05% by weight or less, and the oxygen is controlled after 20 ~ 35%, pH 6.5, and the base glycoose is ap...

Embodiment 3

[0079] Method for extracting L-4-hydroxybiobine in fermentation catalysts comprising the steps of:

[0080] (1) L-4-hydroxyisineluminine fermentation catalytic solution was prepared by the following method:

[0081]1 After the concentration of Escherichia coli resection of L-4-hydroxyisoleine, it is used in seed tank medium containing final concentration of 0.1 g / L amp, at 37 ° C, pH 7.0, and solubol control. Cultured in more than 25% of the OD of the bacteria 600 The value is 16, resulting in a seed liquid;

[0082] 2 The seed liquid obtained by step (1) is 10% in the fermenter medium according to the volume percentage of 15%, and the liquid is deeply ventilated under 40 ° C, and then cool down to 32 ° C and the final concentration is 0.1 mmol / L. IPTG continues to culture until fermentation is completed, wherein the fermentation process residue is 0.05% or less, and the oxygen is controlled after 20 ~ 35%, pH 7.5, and the base glycoose is used as a carbon source. ;

[0083] 3...

Technical field

[0001] The present invention relates to the field of separation and purification, and more particularly to a method of extracting L-4-hydroxyobine extracting a fermentation catalyst. Background technique

[0002] L-4-hydroxyisoleucine (4-hydroxyisyisoleucine) is a non-protein amino acid present in nature. The study found that L-4-hydroxybiobiline has many aspects of promoting insulin secretion, improving insulin tolerance and regulation of blood lipids. The L-4-hydroxyisineluminine can not only promote the intake of glucose, but also reduce liver glucose output to improve insulin tolerance of the outer peripheral tissue. In contrast, insulin synthesis of L-4-hydroxybiobiline regulatory insulin-controlled insulin synthesis is strictly dependent on glucose concentration compared to current drugs that have been used to treat T2DM. Therefore, L-4-hydroxybiobine does not cause side effects such as hypoglycemia. At the same time, the L-4-hydroxybiobine can also effectiv...

Examples

Embodiment 1

[0057] Method for extracting L-4-hydroxybiobine in fermentation catalysts comprising the steps of:

[0058] (1) L-4-hydroxyisineluminine fermentation catalytic solution was prepared by the following method:

[0059] 1 After the concentration of Escherichia coli resection of L-4-hydroxyisoleine, it is used in seed tank medium containing final concentration of 0.1 g / L amp, at 37 ° C, pH 7.0, and solubol control. Cultured in more than 25% of the OD of the bacteria 600 The value is 15 to obtain a seed liquid;

[0060] 2 The seed fluid obtained by step (1) is 10% in the fermentation tank medium according to the volume percentage of 15%, and the liquid is deeply ventilated under 36 ° C and then cooling to 32 ° C and the final concentration is 0.1 mmol / L. IPTG continues to culture until fermentation is completed, wherein the fermentation process residue is 0.05% by weight, and the oxygen is controlled after 20 ~ 35%, pH 7.0, and the amount of glucose is carbon. source;

[0061] 3 fer...

Embodiment 2

[0068] Method for extracting L-4-hydroxybiobine in fermentation catalysts comprising the steps of:

[0069] (1) L-4-hydroxyisineluminine fermentation catalytic solution was prepared by the following method:

[0070] 1 After the concentration of Escherichia coli resection of L-4-hydroxyisoleine, it is used in seed tank medium containing final concentration of 0.1 g / L amp, at 37 ° C, pH 7.0, and solubol control. Cultured in more than 25% of the OD of the bacteria 600 The value is 14 to obtain a seed liquid;

[0071] 2 The seed fluid obtained by step (1) is incorporated into the fermentor medium according to the volume percentage of 15%, and the liquid deep ventilated fermentation was cultured under 35 ° C and then cooled to 30 ° C and the final concentration was 0.1 mmol / L. IPTG continues to culture until fermentation is completed, wherein the fermentation process residue is 0.05% by weight or less, and the oxygen is controlled after 20 ~ 35%, pH 6.5, and the base glycoose is ap...

Embodiment 3

[0079] Method for extracting L-4-hydroxybiobine in fermentation catalysts comprising the steps of:

[0080] (1) L-4-hydroxyisineluminine fermentation catalytic solution was prepared by the following method:

[0081]1 After the concentration of Escherichia coli resection of L-4-hydroxyisoleine, it is used in seed tank medium containing final concentration of 0.1 g / L amp, at 37 ° C, pH 7.0, and solubol control. Cultured in more than 25% of the OD of the bacteria 600 The value is 16, resulting in a seed liquid;

[0082] 2 The seed liquid obtained by step (1) is 10% in the fermenter medium according to the volume percentage of 15%, and the liquid is deeply ventilated under 40 ° C, and then cool down to 32 ° C and the final concentration is 0.1 mmol / L. IPTG continues to culture until fermentation is completed, wherein the fermentation process residue is 0.05% or less, and the oxygen is controlled after 20 ~ 35%, pH 7.5, and the base glycoose is used as a carbon source. ;

[0083] 3...

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