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Pseudomonad esterase and application in preparing optical pure mandel and derivative thereof

A technology for pseudomonas and derivatives, which is applied in the field of Pseudomonas esterase and its use for preparing optically pure mandelic acid and derivatives thereof, achieving high optical purity, good stability and wide industrial application prospects Effect

Inactive Publication Date: 2009-09-23
EAST CHINA UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there has been no report on the preparation of optically pure mandelic acid by using ester hydrolase to catalyze the hydrolysis of the 2-hydroxyacetylated product of mandelic acid

Method used

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  • Pseudomonad esterase and application in preparing optical pure mandel and derivative thereof
  • Pseudomonad esterase and application in preparing optical pure mandel and derivative thereof
  • Pseudomonad esterase and application in preparing optical pure mandel and derivative thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] The cultivation of embodiment 1 bacterial strain CGMCC 2872

[0024] The strain CGMCC 2872 was inoculated into the seed liquid containing rich medium, and cultured at 30°C and 180rpm for 12 hours; the rich medium consisted of 15g of glycerol, 5g of peptone, 5g of yeast extract, 1g of NaCl, MgSO 4 0.2g, KH 2 PO 4 0.5g, K 2 HPO 4 0.5g, tap water 1000ml, adjust the pH value to 7.0. The seed solution was inoculated into the rich medium with an inoculation amount of 5% v / v, and cultured at 30° C. and 180 rpm for 18 hours. After the cultivation, centrifuge at 10,000rpm to obtain enzyme-containing bacteria.

Embodiment 2

[0025] Example 2 Different candidate strains for the hydrolysis of acetoxyphenylacetic acid

[0026] According to the method described in Example 1, the wet cells of different candidate strains were washed 3 times with physiological saline with a concentration of 0.85% w / v, and added to the substrate 2-acetoxy In a phosphate buffer solution with a pH of 7.0 and a phenylacetic acid concentration of 20 mmol / L, react at 30° C. and 180 rpm for 12 hours, and then add 0.1% v / v concentrated sulfuric acid to terminate the reaction. Add sodium chloride and ethyl acetate, shake and extract for 1 minute, centrifuge to take the supernatant, add anhydrous sodium sulfate and let stand overnight, and use liquid chromatography to detect the conversion rate of the reaction and the optical purity of the product.

[0027] Table 1 Catalytic performance comparison of candidate strains

[0028]

[0029] The results in Table 1 show that the 8 strains screened from the soil all have high enantios...

Embodiment 3

[0030] The influence of embodiment 3 different substrate concentration on CGMCC 2872 reaction characteristics

[0031] According to the method described in Example 1, the wet cells of CGMCC 2872 were cultivated, washed 3 times with physiological saline with a concentration of 0.85% w / v, and added to the substrate 2-acetoxy Phenylacetic acid concentrations were 5, 10, 20, 40, 60 and 100mmol / L pH7.0 phosphate buffer solution, reacted for a certain period of time at 30°C and 180rpm, after confirming that the reaction was complete, add 0.1% v / v of concentrated sulfuric acid to terminate the reaction. Add sodium chloride and ethyl acetate, shake for 1 minute, centrifuge to take the supernatant, add anhydrous sodium sulfate and let it stand overnight, and use liquid chromatography to detect the conversion rate of the reaction and the optical purity of the product.

[0032] Table 2 Effects of different substrate concentrations on the reaction characteristics of CGMCC 2872

[0033]...

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Abstract

The invention discloses a pseudomonad which produces esterase and an application in preparing (S)-mandel, (R)-mandel or derivative thereof by using the enantiomorphous selective hydrolization of esterase catalysis 2-acetoxyl group-phenylacetic acid or the derivative thereof produced by the pseudomonad. The esterase is the bacteria body cultured by using soil isolated bacteria-pseudomonas sp.ECU1011 (preserving number is CGMCC No.2872). The technique of producing (S)-mandel and (R)-mandel by adopting the disclosed pseudomonad esterase to catalyzing enantiomorphous selective hydrolization of 2-acetoxyl group-phenylacetic acid is new, the reaction condition is mild, the enantiomorph of the product and the substrate have high purity, the enantiomeric excess values (ee) are 98.1% and more than 99% respectively. The biological catalyst has extensive substrate spectrum, can be used for chiral separation of various mandel derivative, and has excellent industrial application prospect.

Description

technical field [0001] The invention relates to a strain of pseudomonas and the use of the esterase produced by the strain to catalyze the enantioselective hydrolysis (deacetylation) of 2-acetoxyphenylacetic acid to produce optical mandelic acid or its derivatives. technical background [0002] Mandelic acid (α-hydroxyphenylacetic acid) is an important chemical and pharmaceutical intermediate, among which optically pure mandelic acid has unique physiological functions and is mainly used as a key chiral building block in the pharmaceutical industry, (S) -Mandelic acid can be used to synthesize drugs for treating urinary incontinence, and (R)-mandelic acid can be used to synthesize β-lactam blockers. Optically pure mandelic acid is also a commonly used chiral compound resolution reagent, which has very important commercial value and broad application prospects. Therefore, how to obtain the pure single enantiomer of mandelic acid has become an important subject of people's att...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12P7/42C12R1/38
Inventor 许建和鞠鑫潘江
Owner EAST CHINA UNIV OF SCI & TECH
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