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Biosynthesis method of ethyl vanillin

A technology for ethyl vanillin, biosynthesis, applied in microorganism-based methods, biochemical equipment and methods, microorganisms, etc., to achieve mild reaction conditions, easy operation, and little environmental pollution.

Inactive Publication Date: 2012-08-15
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

No natural sources of ethyl vanillin have been reported

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Dehydrogenase catalytic process:

[0025] (1) Seed culture: E. coli BL21 (DE3) containing pET30a-mdlB was inoculated on agar slant medium, cultured at 37°C for 24h to activate, and the activated strain was transferred to LB seed culture containing kanamycin Base, after 12h of culture;

[0026] (2) Fermentation culture: Inoculate the seed culture overnight in a fermentation medium containing kanamycin LB with an inoculum amount of 5%, ferment for 2h at 37℃, add IPTG to make the final concentration 0.2mM, induce at 30℃ 6h;

[0027] (3) Fermentation reaction: collect the bacteria in the fermentation broth by centrifugation at 5000 rpm for 25 min, remove the supernatant, wash twice with sterile water, and suspend in 0.5% 3-ethoxy-4-hydroxy phenylglycolic acid Phosphate buffer (Na 2 HPO 4 1.12%, NaH 2 PO 4 2.14%, MgSO 4 0.01%, pH=7), 30° C., 200 rpm, and fermentation for 40 hours to obtain a solution containing 3-ethoxy-4-hydroxyacetophenone acid. Centrifuge the fermentation ...

Embodiment 2

[0032] This example is different from Example 1 only in that the strain containing acetophenone acid decarboxylase used in the decarboxylase catalytic process is Pseudomonas aeruginosa. Specific steps are as follows:

[0033] Dehydrogenase catalytic process:

[0034] (1) Seed culture: E. coli BL21 (DE3) containing pET30a-mdlB was inoculated on agar slant medium, cultured at 37°C for 24h to activate, and the activated strain was transferred to LB seed culture containing kanamycin Base, after 12h of culture;

[0035] (2) Fermentation culture: Inoculate the seed culture overnight in a fermentation medium containing kanamycin LB with an inoculum amount of 5%, ferment for 2h at 37℃, add IPTG to make the final concentration 0.2mM, induce at 30℃ 6h;

[0036] (3) Fermentation reaction: collect the bacteria in the fermentation broth by centrifugation at 5000 rpm for 25 min, remove the supernatant, wash twice with sterile water, and suspend in 0.5% 3-ethoxy-4-hydroxy phenylglycolic acid Phos...

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PUM

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Abstract

The invention relates to a biosynthesis method of ethyl vanillin. The biosynthesis method comprises the steps of inoculating strains containing mandelic acid dehydrogenase genes and escherichia coli BL21(DE3) with plasmids pET30a-md1B into a fermentation medium, adding an inducer and then 3-ethoxy-4-hydroxy mandelic acid into the fermentation medium, and fermenting to obtain fermentation liquor of 3-ethoxy-4-hydroxy benzoylformic acid; and inoculating strains containing benzoylformic acid decarboxylase genes into fermentation supernate, and fermenting to obtain fermentation liquor containing the ethyl vanillin, wherein the strains are pseudomonas putida or pseudomonas aeruginosa with the preservation number being ATCC (American Type Culture Collection) 12633. The biosynthesis method provides a path for synthesizing the ethyl vanillin through a microbial fermentation method for the first time, has the advantages of being mild in reaction condition, low in environment pollution, simple and convenient to operate and the like, and is a novel ethyl vanillin synthesis method.

Description

Technical field [0001] The invention relates to a method for synthesizing ethyl vanillin, especially a method for biosynthesis of ethyl vanillin. Background technique [0002] The chemical name of ethyl vanillin is 3-ethoxy-4-hydroxybenzaldehyde. It has a sweet chocolate aroma and a strong aroma unique to vanillin. The aroma is 3 to 4 times stronger than vanillin, and the aroma is It is more refreshing and elegant. It has the advantages of less dosage, sufficient aroma, and no color change in flavoring. It is an important spectral high-end flavor, widely used in food, tobacco, and daily necessities, and is also an important pharmaceutical intermediate. [0003] At present, ethyl vanillin is chemically synthesized, mainly using glyoxylic acid and o-ethoxy phenol as starting materials, through condensation, oxidation and decarboxylation reactions, but there are problems such as difficulty in catalyst separation at the end of the oxidation reaction. . As consumers become more vigila...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P7/22A23L1/23C12R1/19C12R1/40A23L27/24
Inventor 冯骉潘晓霞李大力张晓鸣贾承胜何文森李静静
Owner JIANGNAN UNIV
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