Method for biological catalysis preparation of (R)-2-hydroxyl-4-phenyl ethyl butyrate

A technology of ethyl phenylbutyrate and biocatalysis, applied in biochemical equipment and methods, methods based on microorganisms, microorganisms, etc., can solve the problem of lack of high enantioselectivity and high yield microbial enzyme sources, low conversion rate, etc. problem, to achieve high enantiomeric value, high optical purity, high reaction conversion effect

Inactive Publication Date: 2008-12-03
重庆博腾精细化工有限公司
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  • Abstract
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  • Application Information

AI Technical Summary

Problems solved by technology

[0010] Use whole cells of microorganisms to catalyze reactions such as using Rhodotorula minuta IFO 0920 and Candida holmii KPY 124020 to convert ethyl 2-carbonyl-4-phenylbutyrate in water / organic phase, ee value Reach 95% and 94%, shortcoming is that transformation rate is not high, the highest 58% (Shinobu Oda etc., Biosci.Biochem.62:1762-1767,1998); Symmetric catalytic conversion of 2-carbonyl-4-phenylbutyric acid ethyl ester to obtain product enantiomeric value of 82.25%, yield rate of 75.82% (Chemical Research and Application, 2006, 18 (4): 426-430; Food and Biology Journal of Technology, 2006, 25(2): 66-69)
[0011] Judging from the reports and studies on the biocatalytic synthesis of ethyl (R)-2-hydroxy-4-phenylbutyrate at present, there is a lack of microbial enzyme sources with high enantioselectivity and high yield. Therefore, microorganisms need to be screened , find a stable, more economical strain with higher catalytic efficiency, and select appropriate reaction conditions to maximize enantioselectivity and specific yield

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Candida boidinii AS 2.2159 was selected as the catalytic strain

[0042] Slope culture: the medium is 100mL wort juice (containing 10g of malt powder), 2g of agar, pH 6.0, sterilized at 121°C for 15 minutes, cooled and inoculated after sterilization, and cultured at 28°C for 2 days as the activated seeds on the slope.

[0043] Seed cultivation and fermentation: Yeast extract 10g / L, peptone 20g / L, glucose 20g / L, pH6.5, liquid volume is 250mL triangular bottle 50mL, sterilized at 120°C for 20 minutes, cooled and inoculated with slant seeds after sterilization, 180rpm shaker, cultured at 28°C for 48 hours, used as seeds or fermented enzyme liquid.

[0044] The amount of wet bacteria in each 50mL fermented enzyme liquid was 1.5g, and the bacteria were collected by centrifugation for 10 minutes (8,000 rpm), washed twice with potassium phosphate buffer (0.1M, pH6.5), and 1.5g of the above The bacterium was transferred to 5 mL of the same potassium phosphate buffer containing...

Embodiment 2

[0046] Candida boidinii AS 2.2160 was selected as the catalytic strain

[0047] Slope culture: the medium is 100mL wort juice (containing 10g of malt powder), 2g of agar, pH 6.0, sterilized at 121°C for 15 minutes, cooled and inoculated after sterilization, and cultured at 28°C for 2 days as the activated seeds on the slope.

[0048] Seed cultivation and fermentation: Yeast extract 10g / L, peptone 20g / L, glucose 20g / L, pH 6.0, liquid volume is 250mL triangular bottle 50mL, sterilized at 120°C for 20 minutes, cooled to inoculate slant seeds after sterilization, 180rpm shaker, cultured at 28°C for 48 hours, used as seeds or fermented enzyme liquid.

[0049] The amount of wet bacteria in every 50mL fermented enzyme liquid was 1.5g, and the bacteria were collected by centrifugation for 10 minutes (8,000 rpm), washed twice with potassium phosphate buffer (0.1M, pH6.7), and 1.2 grams of the above The cells were transferred to 5 mL of the same potassium phosphate buffer solution cont...

Embodiment 3

[0051] Sporobolomyces salmonicolor AS 2.2121 was selected as the catalytic strain

[0052] Slope culture: the medium is 100mL wort juice (containing 10g of malt powder), 2g of agar, pH 6.0, sterilized at 121°C for 15 minutes, cooled and inoculated after sterilization, and cultured at 28°C for 2 days as the activated seeds on the slope.

[0053] Seed cultivation and fermentation: Yeast extract 10g / L, peptone 20g / L, glucose 20g / L, pH6.5, liquid volume is 250mL triangular bottle 50mL, sterilized at 120°C for 20 minutes, cooled and inoculated with slant seeds after sterilization, 180rpm shaker, cultured at 28°C for 48 hours, used as seeds or fermented enzyme liquid.

[0054] The amount of wet bacteria in each 50mL fermented enzyme liquid was 1.5g, and the bacteria were collected by centrifugation for 10 minutes (8,000 rpm), washed twice with potassium phosphate buffer (0.1M, pH6.0), and 1.5g of the above The cells were transferred to 5 mL of the same potassium phosphate buffer co...

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PUM

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Abstract

The invention relates to a method for preparing (R)-2-hydroxyl-4- phenylbutyric ethyl butyrate by adopting biological catalysis, which belongs to the technology field of the biocatalytic anisomerous reducing preparation medical chiral intermediate. The method comprises the following steps: utilizing produced bacteria Candida boidinii of high antimer selective carbonyl reductase screened etc.; fermenting and producing enzyme under the optimizing condition; taking 2-hydroxyl-4-phenylbutyric ethyl butyrate as a substrate in a single water phase system, a water / organic two-phase system or a water / resin two-phase system; adding grape sugar to wet thalli; and preparing (R)-2- hydroxyl-4- phenylbutyric ethyl butyrate. When the concentration of the substrate of 2-hydroxyl-4-phenylbutyric ethyl butyrate is 1 to 50g / L, (R)-2- hydroxyl-4- phenylbutyric ethyl butyrate is subjected to transformation for 1h to 48h, so as to ensure that the enantiomeric excess of the (R)-2- hydroxyl-4- phenylbutyric ethyl butyrate reaches 84.9 to 98.88 percent, the transformation rate of Moore reaches 72.0 to 84.6 percent, and coenzyme is not needed during the whole reaction process.

Description

technical field [0001] The invention relates to a biocatalytic method for preparing ethyl (R)-2-hydroxy-4-phenylbutyrate, and belongs to the technical field of biocatalytic asymmetric reduction preparation of medical chiral intermediates. Background technique [0002] Chiral (R)-2-hydroxy-4-phenylbutanoic acid ethyl ester (Benzenebutanoic acid, a-hydroxy-, ethylester, (aR)-), the molecular formula is C 12 h 16 o 3 , molecular weight 208.25, CAS number: 90315-82-5. (R)-2-Hydroxy-4-phenylbutyric acid ethyl ester is a key chiral intermediate in the synthesis of many angiotensin-converting enzyme inhibitors (ACEI) pril drugs. Due to the importance of ethyl (R)-2-hydroxy-4-phenylbutyrate in the synthesis of this class of drugs, many people have been attracted to conduct extensive research on its preparation, and a large number of preparation methods have been reported. [0003] At present, the production of ethyl (R)-2-hydroxy-4-phenylbutyrate can be carried out by chemical o...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P7/62C12R1/01C12R1/645C12R1/72
Inventor 林文清孙志浩胡勋刘昊谭小兵
Owner 重庆博腾精细化工有限公司
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