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Method for preparing (S)-(4-chlorphenyl)-(pyridine-2-yl)-methanol by utilizing microbial catalysis

A technology of microorganisms and chlorophenyl, applied in the field of biocatalysis, can solve the problems of unfavorable application in the pharmaceutical industry, complicated operation process, and unsatisfactory yield, and achieve product safety, high enantioselectivity, and environmental friendliness

Active Publication Date: 2013-11-20
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] From the above data, it can be seen that the chemical reduction preparation ( S )-(4-chlorophenyl)-(pyridin-2-yl)-methanol was not effective, yield and e.e. The value is low, and the operation process is complicated. Some chiral catalysts will introduce heavy metal elements in the reaction process, which is not conducive to the application in the pharmaceutical industry.
The only report of a biological method, using ketoreductase (commercial enzyme) reduction preparation ( S )-(4-chlorophenyl)-(pyridin-2-yl)-methanol reaction yield and e.e. not ideal

Method used

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  • Method for preparing (S)-(4-chlorphenyl)-(pyridine-2-yl)-methanol by utilizing microbial catalysis
  • Method for preparing (S)-(4-chlorphenyl)-(pyridine-2-yl)-methanol by utilizing microbial catalysis
  • Method for preparing (S)-(4-chlorphenyl)-(pyridine-2-yl)-methanol by utilizing microbial catalysis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026]Example 1 Screening of microbial strains with asymmetric reduction of 4-chlorophenyl-(pyridin-2-yl)-methanone

[0027] The experimental strains come from 4 soil samples of Changguangxi Wetland Park in Wuxi, Jiangsu Province. By using enrichment medium and screening plate medium, several microorganisms with certain tolerance to the substrate can be isolated. The substrate (4-chlorophenyl)-(pyridin-2-yl)-methanone and other reagents used in the experiment are commercially available analytically pure, chemically pure or biochemical reagents.

[0028] The main media used in the experiment include:

[0029] Enrichment medium (I): Glucose 50, yeast extract 20, potassium dihydrogen phosphate 4, magnesium sulfate heptahydrate 1.5, adjust pH to 6.0; make substrate into 500 g / L tetrahydrofuran solution, add 1 mL of substrate solution.

[0030] Screening plate medium (MSM) (Ⅱ): 0.2 ammonium sulfate, 0.2 potassium dihydrogen phosphate, 0.1 sodium chloride, 0.02 magnesium sulfate h...

Embodiment 2

[0036] Influence of pH on CPMK asymmetric reduction reaction in the aqueous phase reaction system of embodiment 2

[0037] Suspend 1 g of wet cells in different buffer solutions containing 5% (W / V) glucose and 2 g / L CPMK (including: 0.2 M sodium acetate buffer (pH 4.5-5), potassium phosphate buffer (pH 6.0-7.5), Tris-HCl buffer (pH 8.0-8.5)), and set the volume to 10 mL, react at 30°C for 36 h, perform HPLC detection and calculate the reaction e.e. The values ​​and yields are shown in Table 2.

[0038] pH Yield (%) e.e. (%) 4.5 12.5 78.9 5.0 12.5 72.6 5.5 17.0 71.1 6.0 68.1 70.9 6.5 76.9 70.9 7.0 81.3 74.6 7.5 79.7 71.4 8.0 76.7 66.1 8.5 47.2 66.4

[0039] It can be seen from Table 2 that the pH value of the reaction system has a significant impact on the activity of carbonyl reductase in the cells, and the cells maintain a high activity in the pH range of 6.0–8.0. The highest yield and e.e. values, ...

Embodiment 3

[0040] Embodiment 3 The influence of conversion reaction temperature on CPMK asymmetric reduction reaction

[0041] Suspend 1 g of wet cells in 0.2 M PBS (pH 7.0) containing 5% (W / V) glucose, 2 g / L CPMK and dilute to 10 mL. , 40 ℃, 45 ℃ for 36 h after the HPLC detection and calculation of the reaction e.e. The values ​​and yields are shown in Table 3.

[0042] temperature(℃) Yield (%) e.e. (%) 25 75.4 78.2 30 79.2 78.2 35 3.79 -13.2 40 2.34 -22.4 45 0.923 -22.1

[0043] It can be seen from Table 3 that in the temperature range of 25 °C to 30 °C, e.e. Values ​​(78.2%-78.3%) and yields (75.4%-79.2%) are relatively stable, and when the temperature rises to 35 ℃, e.e. value . and the yield dropped sharply to -13.2% and 3.79%, respectively, which indicated that the reaction temperature should be around 30 ℃.

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Abstract

The invention relates to a method for preparing (S)-(4-chlorphenyl)-(pyridine-2-yl)-methanol by utilizing microbial catalysis, and belongs to the technical field of biological catalysis. According to the method, 4-chlorphenyl-(pyridine-2-yl)-ketone is subjected to asymmetrical reduction by utilizing kluyveromycessp (CCTCCM 2011385) whole cells to synthesize the (S)-(4-chlorphenyl)-(pyridine-2-yl)-methanol. The method comprises the following steps of: sieving a microbe which has high-stereoselectivity carbonyl reductase activity on a prochiral ketone substrate, determining a series of conditions of asymmetrical reduction reaction, such as reaction temperature, pH, the concentration of cells, the concentration of the substrate and reaction time and additives (including various secondary solvents, polyethyleneglycol (PEG), phosphates and the like), wherein the enantiomer excess value and yield of the (S)-(4-chlorphenyl)-(pyridine-2-yl)-methanol serving as a product can reach 86.7 percent e.e and 92.1. The product is separated and extracted initially by silicagel column chromatography, so that the purity of the separated product is 99.2 percent, and the extraction yield is 56.7 percent.

Description

technical field [0001] A method for preparing (S)-(4-chlorophenyl)-(pyridin-2-yl)-methanol by catalytic asymmetric reduction of whole cells of microorganisms, that is, using whole cells of microorganisms to asymmetrically reduce 4-chlorophenyl-( Pyridin-2-yl)-methanone is an important chiral intermediate for the synthesis of betahistines ( S )-(4-chlorophenyl)-(pyridin-2-yl)-methanol belongs to the technical field of biocatalysis. Background technique [0002] Betahistine drugs are a class of second-generation antihistamine drugs with high efficiency and low toxicity and side effects. They can be used to treat allergic rhinitis and other allergic diseases. They have the advantages of rapid action, high selectivity, low side effects, and significant curative effect. . Due to the rapid increase in the incidence of allergic diseases in recent decades, the demand for betahistines, which play an important role in the treatment of allergic diseases, is increasing. The same seco...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/16C12R1/645
Inventor 倪晔周婕妤孙志浩
Owner JIANGNAN UNIV
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