Method for preparing benzoyl formic acid and R-mandelic acid by coupling reaction of S- mandelic acid dehydrogenase and laccase

A mandelic acid dehydrogenase and coupling reaction technology, applied in the direction of fermentation, etc., can solve the problems of lack of ferricyanide experimental data, etc., and achieve the effects of convenient industrial production, low environmental pressure and high purity

Inactive Publication Date: 2013-05-08
NANJING UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Foreign Dietmar Haltrich et al. used cellobiose dehydrogenase and laccase to couple to produce lactobionic acid, but lacked experimental data on ferricyanide as a chemical medium

Method used

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  • Method for preparing benzoyl formic acid and R-mandelic acid by coupling reaction of S- mandelic acid dehydrogenase and laccase

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Reaction system: 50ml NaH with pH6.5 2 PO 4 - Citric acid (NaH 2 PO 4 Mixed with citric acid) buffer containing 10mmol / L racemic mandelic acid (commercially available), 0.3mmol / L sodium ferrocyanide, 50U S-mandelate dehydrogenase, 25U laccase. After shaking and reacting at 140rpm at 30°C for 10 hours, 49.9% of the racemic mandelic acid was consumed, and all the consumed S-mandelic acid was converted into acetophenone acid. The pH of the reaction solution was adjusted to below 2.0, and acetophenone acid and R- Mandelic acid, ether was distilled off, the mixture of acetophenone acid and R-mandelic acid was separated by vacuum distillation at 85°C and 13.33 pascals to obtain acetophenone acid and R-mandelic acid respectively.

Embodiment 2

[0021] Reaction system: 50ml NaH with pH6.5 2 PO 4 - Citric acid (NaH 2 PO 4 Mixed with citric acid) buffer containing 10mmol / L racemic mandelic acid (commercially available), 0.1mmol / L potassium ferrocyanide, 50U S-mandelate dehydrogenase, 25U laccase. After shaking and reacting at 140rpm at 30°C for 10 hours, 49.3% of the racemic mandelic acid was consumed, and all the consumed S-mandelic acid was converted into acetophenone acid. The pH of the reaction solution was adjusted to below 2.0, and acetophenone acid and R- Mandelic acid, ether was distilled off, the mixture of acetophenone acid and R-mandelic acid was separated by vacuum distillation at 85°C and 13.33 pascals to obtain acetophenone acid and R-mandelic acid respectively.

Embodiment 3

[0023] Reaction system: 50ml NaH with pH6.5 2 PO 4 - Citric acid (NaH 2 PO 4 Mixed with citric acid) buffer containing 5mmol / L racemic mandelic acid (commercially available), 1mmol / L sodium ferrocyanide, 50U S-mandelate dehydrogenase, 25U laccase. After reacting at 30°C for 12 hours, the consumption of racemic mandelic acid was 49.5%, and all the consumed S-mandelic acid was converted into acetophenone acid. Adjust the pH of the reaction solution to below 2.0, and diethyl ether extracted acetophenone acid and R-mandelic acid. , Distilled to remove ether, the mixture of acetophenone acid and R-mandelic acid was separated by vacuum distillation at 85°C 13.33 Pascals to obtain acetophenone acid and R-mandelic acid, adjust the pH of the reaction solution to below 2.0, and diethyl ether extracted phenethyl Keto acid and R-mandelic acid, ether was removed by distillation, the mixture of acetophenone acid and R-mandelic acid was separated by vacuum distillation at 85°C and 13.33 p...

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Abstract

The invention provides a method for catalyzing S-mandelic acid to be converted into benzoyl formic acid and R-mandelic acid by coupling reaction of S-mandelic acid dehydrogenase and laccase. The method is characterized by continuously regenerating reaction media by laccase to continuously regenerate coenzyme FMN (Flavin Mononucleotide) of S-mandelic acid dehydrogenase, so that the S-mandelic acid dehydrogenase and the laccase are coupled to carry out catalytic reaction. Therefore, the conversion efficiency of independently using the S-mandelic acid dehydrogenase is improved by sufficiently utilizing the high efficiency of enzyme catalysis and continuously catalyzing and oxidizing the S- mandelic acid through less reaction media; and the reaction cost is reduced. According to the method disclosed by the invention, the S-mandelic acid dehydrogenase and laccase are coupled to catalyze the S-mandelic acid to be converted into benzoyl formic acid; and the enantiomer excess of the product R-mandelic acid can reach 99.8%. Moreover, only a small quantity of ferrocyanide or ferricyanide is needed as the reaction medium; the process is simple; the cost is low; and the environment pressure is small; and besides, the method is convenient for industrial production and of a great application value to chiral resolution of racemic mandelic acid.

Description

technical field [0001] The invention relates to a method for coupling two enzymes to catalyze S-mandelic acid to generate acetophenone acid by using ferricyanide or ferrocyanide as a coupling agent, and at the same time realize the chiral resolution of racemic mandelic acid, specifically A coupling reaction involving S-mandelate dehydrogenase and laccase. Background technique [0002] S-mandelate dehydrogenase is a kind of oxidoreductase with flavin FMN as coenzyme, which can specifically catalyze the oxidation of S-mandelic acid to generate acetophenone acid, and its coenzyme FMN is reduced to FMNH 2 . Ferricyanide can oxidize FMNH 2 Generate FMN and ferrocyanide. Laccase is a kind of phenol oxidase containing copper ions, which uses oxygen to oxidize the substrate, and oxygen is reduced to water, and laccase has the characteristics of a wide range of substrate catalysis, and can catalyze the oxidation of ferrocyanide to ferricyanide compounds. Foreign Dietmar Haltrich...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P7/40C12P7/42
Inventor 李大力王鹏杨军方杨成丽
Owner NANJING UNIV OF SCI & TECH
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