Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Method for preparing chiral 2-chloro-3,4-difluorophenethyl alcohol

A technology for difluorophenethyl alcohol and difluoroacetophenone is applied in the field of preparing chiral 2-chloro-3,4-difluorophenethyl alcohol, and achieves the advantages of avoiding harsh reaction conditions, high yield and good practical industrial application value. Effect

Active Publication Date: 2017-03-22
杭州酶易生物技术有限公司
View PDF4 Cites 9 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Through literature search, there are no examples of biological enzymes catalyzing this reduction reaction

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Method for preparing chiral 2-chloro-3,4-difluorophenethyl alcohol
  • Method for preparing chiral 2-chloro-3,4-difluorophenethyl alcohol
  • Method for preparing chiral 2-chloro-3,4-difluorophenethyl alcohol

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] The shake flask production technology of embodiment 1 ketoreductase (KRED) enzyme powder

[0027] The gene fragment (synthesized by Shanghai Jierui Bioengineering Co., Ltd.) containing the ketoreductase gene (SEQ ID NO.1 to 23) was ligated with the digested product of the pET28a plasmid, and transformed into competent E.coli BL21(DE3) The strains were screened to obtain positive clones, which were inoculated into 4 mL liquid LB medium containing ampicillin resistance and activated overnight (37° C., 200 rpm).

[0028] Take the overnight culture, transfer it to 100mL liquid LB medium containing ampicillin antibody at 1 / 100 inoculum size, culture at 37°C with shaking at 200rpm until OD 600 When the value reached 0.6, IPTG was added and the cultivation was continued overnight at 30°C. The cells were collected by centrifugation, and suspended in 10 mL of phosphate buffer (2 mM, pH 7.0). The cell suspension was ultrasonically disrupted in an ice bath for 10 minutes, centri...

Embodiment 2

[0029] Embodiment 2 Ketoreductase (SEQ ID NO.1~21) milligram level reaction

[0030] In a 5mL reaction flask, add 10mg of the substrate 2-chloro-3,4-difluoroacetophenone and 0.1mL of isopropanol, after the substrate is completely dissolved, add 1.2ml of TEA-HCl buffer solution (0.1M, pH7.0), 0.1mgNAD + , 0.1mgNADP + (dissolved in 0.1ml of buffer), then add 20mg of glucose and 2mg of glucose dehydrogenase, and finally, respectively add 2mg of the ketoreductase KRED enzyme powder obtained in Example 1 (0.5U / mg, dissolved in 0.1ml of buffer) , Shaking the reaction at 30°C for 20h.

[0031] Get the product after reaction and carry out HPLC analysis, the productive rate of detection product and product ee value, result is as shown in table 1:

[0032] Table 1 Yield and product ee value of different ketoreductase reactions

[0033]

[0034] Yield label description: + stands for 1%-20% yield, ++ stands for 20%-50% yield, +++ stands for 50%-80% yield, ++++ stands for 80%-95% yi...

Embodiment 3 100

[0038] Embodiment 3 100 milligram level preparation technology

[0039] In a 5ml reaction flask, add 150mg of the substrate 2-chloro-3,4-difluoroacetophenone and 0.3ml of isopropanol, after the substrate is completely dissolved, add 1.2ml of TEA-HCl buffer (0.1M, pH7.0), 7.5mg ketoreductase powder (SEQ ID NO.9) (dissolved in 0.1ml buffer), 0.75mgNAD + (dissolved in 0.1ml of buffer solution), at 30°C, a magnetic stirring reaction was performed, and two reaction times were set: 0h and 20h.

[0040] Sampling after reaction carries out HPLC analysis to product, and result is as follows: figure 1 and figure 2 shown;

[0041] figure 1For the analysis result of the product obtained after reacting for 0 hours, the substance at 7.67 minutes in the figure is the substrate;

[0042] figure 2 It is the analysis result of the product obtained after 20 hours of reaction. In the figure, the substance with a retention time of 6.47 minutes is the product, and the final conversion rate ...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses a method for preparing chiral 2-chloro-3,4-difluorophenethyl alcohol. According to the method, 2-chloro-3,4-difluoroacetophenone is used as a substrate, and ketoreductase is used for catalytic reduction of the substrate so as to generate the chiral 2-chloro-3,4-difluorophenethyl alcohol. Amino acid sequence of the ketoreductase is as shown in the SEQ ID NO.1-21. 2-chloro-3,4-difluoroacetophenone which is cheap and easily available is used as the substrate, and the biocatalyst ketoreductase is adopted so as to carry out an asymmetric reduction reaction to obtain 2-chloro-3,4-difluorophenethyl alcohol with high chiral purity. The method has advantages of high yield, mild reaction condition and simple operation. Problems of a chemical reduction method, such as strict reaction condition, complex preparation of a catalyst, high cost, inflammable property and low chiral purity of the product, are avoided. The method has good practical industrial application value.

Description

technical field [0001] The invention relates to the technical fields of biological pharmacy and biochemical industry, in particular to a method for preparing chiral 2-chloro-3,4-difluorophenethyl alcohol. Background technique [0002] Ticagrelor is a new small molecule anticoagulant drug developed by AstraZeneca. The drug has obvious inhibitory effect on platelet aggregation caused by ADP, and the effect is rapid after oral administration, so it can effectively improve the symptoms of patients with acute coronary heart disease. In multiple synthetic routes of ticagrelor (WO2008018822, WO2013150495), chiral 2-chloro-3,4-difluorophenethyl alcohol (3) is a key intermediate. For example, one of the synthetic routes is as follows (Chinese Journal of Pharmaceutical Industry, 2014, 45.4: 315-321): [0003] [0004] In the prior art, chiral 2-chloro-3,4-difluorophenethyl alcohol [(S)-3] is synthesized by chemical asymmetric reduction of 2-chloro-3,4-difluoroacetophenone, which ...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C12P7/22
CPCY02P20/584
Inventor 周硕赖敦岳王瑞玲张双玲陈振明
Owner 杭州酶易生物技术有限公司
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com