CN109295152B - Method for catalyzing, esterifying and splitting 2-phenylpropionic acid enantiomer by Novozym435 lipase - Google Patents
Method for catalyzing, esterifying and splitting 2-phenylpropionic acid enantiomer by Novozym435 lipase Download PDFInfo
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- CN109295152B CN109295152B CN201811059512.XA CN201811059512A CN109295152B CN 109295152 B CN109295152 B CN 109295152B CN 201811059512 A CN201811059512 A CN 201811059512A CN 109295152 B CN109295152 B CN 109295152B
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- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P41/00—Processes using enzymes or microorganisms to separate optical isomers from a racemic mixture
- C12P41/003—Processes using enzymes or microorganisms to separate optical isomers from a racemic mixture by ester formation, lactone formation or the inverse reactions
- C12P41/005—Processes using enzymes or microorganisms to separate optical isomers from a racemic mixture by ester formation, lactone formation or the inverse reactions by esterification of carboxylic acid groups in the enantiomers or the inverse reaction
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- C12P7/40—Preparation of oxygen-containing organic compounds containing a carboxyl group including Peroxycarboxylic acids
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Abstract
The patent introduces a new chiral resolution method for obtaining (R) -2-phenylpropionic acid enantiomer, belonging to the technical field of chiral compound resolution by a biological method. The stereoselectivity of Novozym435 and the high catalytic efficiency of (S) -2-phenylpropionic acid are utilized to catalyze and esterify the 2-phenylpropionic acid enantiomer to synthesize the (S) -2-phenylpropionic acid n-hexyl ester, the residual unreacted substrate is high-optical purity (R) -2-phenylpropionic acid, the substrate conversion rate is 97.23%, the enantiomeric excess value is 89.20%, and the yield of the target enantiomer is 5.24%. Compared with other methods, the method has the advantages of simple reaction system, high reaction concentration, high reaction rate, high optical purity of the target product, environmental friendliness, simple post-treatment and the like.
Description
Technical Field
The invention belongs to a method for resolving chiral compounds by a biological method, and relates to a method for obtaining a 2-phenylpropionic acid enantiomer with a single configuration by stereoselectively catalyzing and esterifying a racemic 2-phenylpropionic acid enantiomer by using immobilized lipase Novozym 435. Novozym435 lipase is adopted to selectively catalyze racemic 2-phenyl propionic acid to synthesize (S) -2-phenyl propionic acid n-hexyl ester with single configuration, so as to obtain (R) -2-phenyl propionic acid with higher optical purity.
Background
2-Phenylpropionic acid (2-Phenylpropionic acid) is also called alpha-Phenylpropionic acid, is an aryl propionic acid substance with the simplest structure, is an intermediate for synthesizing aryl propionic acid anti-inflammatory analgesics and fine chemical products, and is widely applied to the synthesis of medicines, spices and pesticides. 2-phenylpropionic acid is an important drug synthesis intermediate, and can be used for synthesizing the calcium solenoofen.
At present, methods for splitting 2-phenylpropionic acid enantiomers mainly comprise a capillary electrophoresis resolution method, a chromatographic resolution method, a liquid-liquid extraction resolution method and a biological enzyme-catalyzed stereoselective hydrolysis of 2-phenylpropionic acid ester. The biological enzyme method selectively catalyzes one enantiomer to carry out hydrolysis or esterification reaction by utilizing the high stereospecificity of enzyme, and can achieve a more ideal resolution effect. Meanwhile, the synthesis efficiency of the biological enzyme method is high, the reaction condition is mild, the cost is low, the production energy consumption can be reduced, and a large amount of precious resources are saved. The biological enzyme method resolution in the organic solvent has the advantages of the aqueous solvent enzyme method resolution, and simultaneously has the advantages of high reaction concentration and easy enzyme recovery due to the good solubility of the organic solvent to reaction substrates and the characteristic of not dissolving the enzyme.
The invention discloses a method for stereoselectively catalyzing and esterifying an (S) configuration enantiomer in racemic 2-phenylpropionic acid by utilizing Novozym435 lipase in an organic solvent to obtain an unreacted (R) configuration enantiomer in a residual substrate. By the method disclosed by the invention, the (R) configuration enantiomer with high optical purity can be obtained. The invention utilizes the biological enzyme method to catalyze, esterify and split the racemic 2-phenylpropionic acid enantiomer in the organic solvent, solves the problems of low selectivity, low optical purity of the product, complex operation and equipment, long splitting time, environmental pollution and the like of the non-enzyme method, and also solves the problems of difficult improvement of reaction concentration, difficult enzyme recovery and the like of the aqueous solution enzyme method.
Disclosure of Invention
The invention aims to provide a method for separating 2-phenylpropionic acid enantiomer to obtain single configuration enantiomer aiming at the problems of low reaction concentration, slow reaction rate, low optical purity of the product and the like of other raceme resolution methods.
The technical scheme of the invention is as follows: the (S) -n-hexyl phenylpropionate is synthesized by esterification by utilizing the high selectivity and high catalytic efficiency of Novozym435 lipase on the (S) -2-phenylpropionate enantiomer, and the (R) -2-phenylpropionate with high optical purity is obtained. Adding a certain concentration of racemic 2-phenylpropionic acid enantiomer, a certain concentration of n-hexanol and a certain amount of immobilized Novozym435 lipase into n-hexane; the concentration of racemic 2-phenylpropionic acid is 10-150 mmol/L, the dosage of Novozym435 lipase is 10-150mg/mL, and the concentration of n-hexanol is 100-250 mmol/L; the reaction was heated in a 25mL reaction tube at a constant speed for a period of time and monitored. Qualitative and quantitative detection is carried out on the product through a high performance liquid chromatograph; the enantiomeric excess of the remaining 2-phenylpropionic acid was found to be 89.20%, the substrate conversion was 97.23%, and the yield of the target enantiomer was 5.24%.
Compared with the prior art, the invention has the following advantages:
the invention utilizes the high stereoselectivity of Novozym435 to the 2-phenylpropionic acid enantiomer and the high catalytic efficiency of the esterification of (S) -2-phenylpropionic acid and n-hexanol. The method has high reaction concentration, and the concentration of the used racemic 2-phenylpropionic acid reaches 100 mmol/L; the reaction condition is mild; the method is green and environment-friendly, a second organic solvent n-hexane is used, the reaction system is simple, and no toxic substance is used or generated in the production process; the immobilized biological enzyme is very convenient to recycle. The invention can obtain the (R) -phenylpropionic acid with high optical purity by controlling the conditions of reaction temperature, concentration of the 2-phenylpropionic acid, concentration of n-hexanol, enzyme dosage, reaction time and the like. The method has simple implementation process and simple and convenient operation and post-treatment process.
[ detailed description ] embodiments
The invention is further illustrated below with reference to examples of the invention:
first, testing and analyzing
The enantiomeric excess value and the substrate conversion rate of the separated product in the embodiment of the invention are calculated according to the detected concentration of each enantiomer of the 2-phenylpropionic acid. The concentration of each enantiomer was measured and analyzed by a Waters 1525 high performance liquid chromatograph, and the column was a Diamonsil C18 (250 mm. times.4.6 mm, 5 μm) column. The mobile phase composition was 80:20 (v/v) of a mixed solution of an aqueous phase (containing 25 mmol/L HP-beta-CD and 0.5% acetic acid (v/v)) and methanol, pH was adjusted to 4.0 with triethylamine, the flow rate was 0.8 mL/min, the injection volume was 10. mu.L, the detection wavelength was 225 nm, and the column temperature was 303K. (R) -2-phenylpropionic acid peaks before (S) -2-phenylpropionic acid.
Second, example
Example 1
100 mmol/L2-phenylpropionic acid and 140 mmol/L n-hexanol, 100 mg Novozym435 lipase were added to 2mL n-hexane; the reaction was heated at 400 rpm and 50 ℃ for 60 min in a 25mL reaction tube. At this time, the substrate conversion was 97.33%, the enantiomeric excess of the remaining 2-phenylpropionic acid enantiomer was 87.41%, and the yield of the target enantiomer was 5.01%.
Example 2
Adding 100 mmol/L2-phenylpropionic acid, 200 mmol/L n-hexanol and 300 mg Novozym435 lipase into 2mL n-hexane organic solvent; the reaction was heated at 400 rpm and 50 ℃ for 45 min in a 25mL reaction tube. At this time, the substrate conversion was 97.23%, the enantiomeric excess of the remaining 2-phenylpropionic acid enantiomer was 89.20%, and the yield of the target enantiomer was 5.24%.
Example 3
Adding 10 mmol/L2-phenylpropionic acid, 140 mmol/L n-hexanol and 60 mg Novozym435 lipase into 2mL n-hexane organic solvent; the reaction was heated at 400 rpm at 50 ℃ for 140 min in a 25mL reaction tube. At this time, the substrate conversion was 86.73%, the enantiomeric excess of the remaining 2-phenylpropionic acid enantiomer was 86.74%, and the yield of the target enantiomer was 24.77%.
Example 4
Adding 10 mmol/L2-phenylpropionic acid, 250mmol/L n-hexanol and 300 mg Novozym435 lipase into 2mL n-hexane organic solvent; the reaction was heated at 400 rpm and 50 ℃ for 90 min in a 25mL reaction tube. At this time, the substrate conversion was 97.22%, the enantiomeric excess of the remaining 2-phenylpropionic acid enantiomer was 92.93%, and the yield of the target enantiomer was 5.37%.
Example 5
Adding 100 mmol/L2-phenylpropionic acid, 140 mmol/L n-hexanol and 40mg Novozym435 lipase into 2mL n-hexane organic solvent; the reaction was heated at 400 rpm and 70 ℃ for 30 min in a 25mL reaction tube. At this time, the substrate conversion was 67.04%, the enantiomeric excess of the remaining 2-phenylpropionic acid enantiomer was 50.81%, and the yield of the target enantiomer was 49.71%.
The above examples merely represent some embodiments of the present invention, which are described in more detail and in more detail, but are not to be construed as limiting the scope of the invention. It should be noted that, for a person skilled in the art, several variations and modifications can be made without departing from the inventive concept, which falls within the scope of the present invention. Therefore, the protection scope of the present patent shall be subject to the appended claims.
Claims (1)
1. A method for catalyzing and esterifying 2-phenylpropionic acid enantiomer by applying Novozym435 stereoselectivity and preparing (R) -2-phenylpropionic acid with high optical purity,
comprises the following steps:
(1) selecting Novozym435 lipase as a biocatalyst;
(2) selecting normal hexane as an organic solvent;
(3) taking 2mL of n-hexane, weighing a certain amount of 2-phenylpropionic acid enantiomer, putting n-hexanol and Novozym435 lipase into a 25mL reaction tube, and heating and stirring at a certain temperature for reacting for a certain time;
the reaction temperature is 50-70 ℃; the dosage of the Novozym435 lipase is 10-150 mg/mL; the concentration of 2-phenylpropionic acid enantiomer is 10-100 mmol/L; the n-hexanol concentration was 140-250 mmol/L.
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| CN111041015B (en) * | 2019-12-31 | 2022-03-18 | 浙江工业大学 | Method for preparing (R) - (+) -N-acetyl-1-methyl-3-amphetamine at high temperature |
| CN113667704A (en) * | 2021-08-24 | 2021-11-19 | 浙江工业大学 | Method for preparing (R) -N- (2, 6-dimethylphenyl) methyl aminopropionate by two-step enzyme method |
Citations (4)
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| CN101892283A (en) * | 2010-06-03 | 2010-11-24 | 华东理工大学 | Method for catalytically preparing optically pure (S)-(+)-2-phenylpropionic acid with Rhodococcus ruber 4.1187 |
| CN104450852A (en) * | 2014-02-28 | 2015-03-25 | 宁夏大学 | Method for preparing homoallylic alcohol compound |
| CN107446960A (en) * | 2017-09-30 | 2017-12-08 | 湖南理工学院 | A kind of method that 2,3 diphenyl-propionic acid enantiomers are split using biological enzyme stereoselectivity |
| CN107988307A (en) * | 2017-12-27 | 2018-05-04 | 湖南理工学院 | The method that enzymatic stereoselectivity splits 2- (4- hydroxy phenyls) propionic acid enantiomer |
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Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101892283A (en) * | 2010-06-03 | 2010-11-24 | 华东理工大学 | Method for catalytically preparing optically pure (S)-(+)-2-phenylpropionic acid with Rhodococcus ruber 4.1187 |
| CN104450852A (en) * | 2014-02-28 | 2015-03-25 | 宁夏大学 | Method for preparing homoallylic alcohol compound |
| CN107446960A (en) * | 2017-09-30 | 2017-12-08 | 湖南理工学院 | A kind of method that 2,3 diphenyl-propionic acid enantiomers are split using biological enzyme stereoselectivity |
| CN107988307A (en) * | 2017-12-27 | 2018-05-04 | 湖南理工学院 | The method that enzymatic stereoselectivity splits 2- (4- hydroxy phenyls) propionic acid enantiomer |
Non-Patent Citations (3)
| Title |
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| 《Kinetic study on enantioselective resolution of (R,S)-2-phenylpropionic acid through Novozyme 435–catalyzed esterification》;Xin Yuan等;《Int J Chem Kinet》;20191231;第520-527页 * |
| 《微水相中 S-2-苯基丙酸的制备》;张波等;《过程工程学报》;20080830;第8卷(第4期);第799-801页 * |
| 《脂肪酶拆分手性药物布洛芬的工艺研究》;黄霜霜;《中国优秀硕士学位论文全文数据库工程科技Ⅰ》;20170615(第06期);第B016-244页 * |
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