A method for enzymatically synthesizing Danshensu

Active Publication Date: 2022-06-21
SHANDONG YANGCHENG BIOLOGY TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The purpose of the present invention is to provide a new method of enzymatically synthesizing Danshensu to solve the problem of shortage of raw materials in the prior art. The method has high efficiency, low pollution and low cost

Method used

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  • A method for enzymatically synthesizing Danshensu
  • A method for enzymatically synthesizing Danshensu
  • A method for enzymatically synthesizing Danshensu

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Embodiment 1 A kind of method for enzymatic synthesis of Danshensu, concrete steps are as follows:

[0049] 1. Heterologous expression of tyrosine ammonia lyase TAL-made, phenylpyruvate reductase PPR-made and glucose dehydrogenase GDH-made:

[0050] 1. Heterologous expression and enzyme activity assay of tyrosine ammonia lyase TAL-made:

[0051] (1) Obtaining TAL-made gene fragments

[0052] According to the tyrosine ammonia lyase (TAL) gene sequence (GenBank accession number: CP015288, SEQ ID NO.1), the TAL-made gene (SEQ ID NO.1) was synthesized by the company after codon optimization suitable for expression in E. coli. 2), the amino acid sequence of the gene is exactly the same as that of the TAL gene, and restriction sites of BamHI and XhoI are introduced upstream and downstream of the gene sequence to facilitate subsequent vector construction.

[0053] (2) Construction of pET28a-TAL-made expression vector

[0054] The synthesized TAL-made gene and pET28a vector ...

Embodiment 2

[0074] Except that the following steps are different, the remaining steps are the same as those in Example 1.

[0075] According to the method in step 1, the expression cells of TAL, PPR and GDH were obtained, and the following drugs were added to the 250ml conical flask to prepare the catalytic system: 1.0 mol of levodopa, 20 g / L of amino acid deaminase wet cells, phenylpyruvate Reductase wet cell 20g / L, glucose dehydrogenase wet cell 20g / L, adjusting the pH of the reaction system to 8.0, and transforming at 30°C for 16h, the obtained Danshensu content was 58.0g / L, and the molar conversion rate was greater than 58.1 %.

Embodiment 3

[0077] Except that the following steps are different, the remaining steps are the same as those in Example 1.

[0078] According to the method in step 1, the expression cells of TAL, PPR and GDH were obtained, and the following drugs were added to the 250ml conical flask to prepare the catalytic system: 1.5mol of levodopa, 30g / L of amino acid deaminase wet cells, phenylpyruvate Reductase wet cell 30g / L, glucose dehydrogenase wet cell 30g / L, adjusting the pH of the reaction system to 8.0, and transforming at 30°C for 16h, the obtained Danshensu content was 60.7g / L, and the molar conversion rate was greater than 60.8 %.

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Abstract

The invention discloses a novel method for synthesizing danshensu. The method uses tyrosine ammonia lyase, phenylpyruvate reductase and glucose dehydrogenase to catalyze levodopa to produce danshensu. Specifically, 0.5-1.5mol of levodopa (3,4-dihydroxyphenylalanine), 10-30g / L of wet bacterium of tyrosine ammonia lyase, and 10-30g of wet bacterium of phenylpyruvate reductase / L, 10-30g / L of wet cells of glucose dehydrogenase were placed in a reactor and mixed evenly, the pH value was adjusted to 7.0, the temperature was 30°C, and the catalytic reaction was carried out for 16-20h to obtain Danshensu. For the first time, the method uses tyrosine ammonia lyase, phenylpyruvate reductase and glucose dehydrogenase to produce danshensu enzymatically with levodopa as a substrate, saving raw materials and avoiding the pollution of chemical synthesis.

Description

technical field [0001] The invention belongs to the technical field of biosynthesis, in particular to a method for synthesizing Danshensu by an enzymatic method. Background technique [0002] Danshensu, the chemical name is β-(3,4-dihydroxyphenyl)lactic acid, is named Danshensu because it is the active ingredient of Chinese herbal medicine Salvia. Pharmacological studies have found that Danshensu can reduce the scope of myocardial infarction and relieve At the same time, it has a protective effect on myocardial ischemia and reperfusion injury, and can remove exogenous oxygen free radicals and protect the function of mitochondria; and Danshensu can significantly inhibit the aggregation of platelets, which is effective for coronary heart disease and helps the body. Rehabilitation of tissues and correction of serious complications such as adult respiratory distress syndrome; Danshensu also has certain effects in antibacterial, anti-inflammatory and enhancing immune function. Cl...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/70C12N1/21C12R1/19
CPCC12N9/0006C12N9/88C12N15/70C12P7/42C12Y101/01047C12Y403/01025
Inventor 张娟冯志彬段春利
Owner SHANDONG YANGCHENG BIOLOGY TECH CO LTD
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