Method for producing R-mandelic acid with biocatalysis and separating and coupling method

A biocatalysis, mandelic acid technology, applied in biochemical equipment and methods, microorganism-based methods, separation/purification of carboxylic acid compounds, etc., can solve the problems of batch decline, low cumulative product concentration, etc., and achieve catalytic yield The effect of improving, de-inhibiting, good prospects for industrial application

Active Publication Date: 2012-06-27
ZHEJIANG UNIV OF TECH
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  • Summary
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  • Description
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AI Technical Summary

Problems solved by technology

[0008] However, in the process of producing R-mandelic acid from the existing nitrilase biocatalyzed racemic mandelonitrile, although the reaction is mild and the reaction speed is fast, it is found through research that the product mandelic acid has a strong inhibition on nitrilase The effect is to reduce the batches used by the bacteria, and the cumulative concentration of the product is not high

Method used

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  • Method for producing R-mandelic acid with biocatalysis and separating and coupling method
  • Method for producing R-mandelic acid with biocatalysis and separating and coupling method
  • Method for producing R-mandelic acid with biocatalysis and separating and coupling method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1: Cultivation of Cells Containing Nitrilase

[0042] The composition of the final concentration of the seed medium: ammonium acetate 10g / L, yeast extract 6g / L, dipotassium hydrogen phosphate 5g / L, sodium chloride 1g / L, magnesium sulfate 0.2g / L, and the solvent is water. The CCTCC No: M 208168 strain was inoculated into the seed medium and cultured at 30°C for 28 hours to obtain the seed solution.

[0043] Final concentration composition of enzyme-producing fermentation medium: ammonium acetate 12.14g / L, yeast extract 7.79g / L, n-butyronitrile 3.29g / L, dipotassium hydrogen phosphate 5g / L, sodium chloride 1g / L, magnesium sulfate 0.2g / L, the solvent is water.

[0044] The seed solution was transferred to the above-mentioned fermentation medium with an inoculation amount of 6% (v / v), and the liquid content in a 15L fermenter was 8L. The culture temperature is 30° C., the rotation speed is 150 rpm, and the ventilation volume is 0.4 vvm. Cultivate for 20 hours and...

Embodiment 2

[0045] Example 2: Effect of different resin types on the catalytic yield of the catalyst.

[0046] Pretreatment of anionic resin: first wash with water to remove pigment and impurities, then soak with twice the volume of 1N HCl for 24 hours, wash with water until neutral, then soak with 1N NaOH for 24 hours, convert to OH type, wash with water until neutral, and use pumping The filter device is drained and set aside.

[0047] The concentration of wet bacteria added to the aqueous solution (adjusted with NaOH) of pH 8.0 was 5g / l, the substrate concentration was 20mM, and then 20g / L of different resins (purchased from Shanghai Huazhen Technology Co., Ltd.) , under the condition of 150rpm, the biocatalytic reaction was carried out for 1h. Measure the R-mandelic acid concentration and the ee value in the solution, and calculate the catalytic yield of the catalyst and the optical activity of its product, the results obtained are shown in the following table:

[0048] Res...

Embodiment 3

[0050] Embodiment 3: the influence of the catalytic yield of catalyst of different resin addition amount in conversion system

[0051] The concentration of wet bacteria added in the phosphate buffer solution of pH8.5 is 5g / l, and the substrate concentration is 20mM, then add 10, 20, 30, 40, 50g / L pretreated HZ202 wet resin respectively, in The reaction was carried out at 30° C. and 150 rpm, and samples were taken every 5 minutes to analyze the concentrations of products and substrates. Stop the reaction when the substrate is completely consumed and record the reaction time required for the conversion of the substrate. The results are shown in the following table:

[0052] Amount of added resin (g / L)

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Abstract

The invention provides a method for producing R-mandelic acid with biocatalysis and separating and coupling method. The method comprises: adding alkaliteit anion exchange resin of 10-50g / L into a reaction system with racemic mandelonitrile as substrate and nitrilase as catalyst after reaction begins 0-1 hour to perform hydrolysis reaction with pH being 8.0-8.5 and temperature being 20-60 DEG C; when the concentration of substrate racemic mandelonitrile in the conversion system is low, further adding racemic mandelonitrile of 10-100mmol / L and alkaliteit anion exchange resin of 10-50g / L into the reaction system to perform hydrolysis reaction of next batch with the same conditions. The invention provides a method for efficiently producing R-mandelic acid, R-mandelic acid is prepared by nitrilase biocatalysis and separating and coupling technology, thus effectively removing inhibitory action of the product; resin is used for absorbing R-mandelic acid to perform coupling conversion, thus the catalysis yield of the nitrilase biocatalyst is greatly enhanced and sequential separation is facilitated; therefore, the inventive technology has good industrial application prospect.

Description

(1) Technical field [0001] The invention relates to a method for producing R-mandelic acid by a coupling method of biocatalysis and separation. (2) Background technology [0002] R-mandelic acid is an important pharmaceutical intermediate, widely used in the synthesis of various drugs, such as cephalosporins, penicillins, anti-tumor agents, anti-obesity drugs, optically pure amino acids, angiotensin-converting enzyme inhibitors , Coenzyme A, etc., can also be used to make chiral solvents. Studies have shown that, compared with drugs synthesized by racemic mandelic acid or its derivatives, the drugs synthesized from single-configuration mandelic acid (or derivatives of mandelic acid) not only have higher efficacy, but more importantly, the side effects are reduced Therefore, it must be a single-type compound in many drug synthesis applications; R-mandelic acid is currently the most eye-catching acidic optical resolution agent due to its good biodegradability, which can make ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12P41/00C12P7/42C07C59/50C07C51/47B01D15/08C12R1/05
Inventor 郑裕国薛亚平柳志强沈寅初
Owner ZHEJIANG UNIV OF TECH
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