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Application of glucosyltransferase in production of ethyl vanillin-alpha-D-glucoside

A technology based on ethyl vanillin and glucosyl, which is applied in the application field of glucosyltransferase in the production of ethyl vanillin-α-D-glucoside, which can solve the problems of disgust, poor stability, retention, etc. , to achieve the effect of high product concentration, high catalytic conversion efficiency and high conversion rate

Active Publication Date: 2019-05-17
SHENZHEN SHINESKY BIOLOGICAL TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, vanillin and ethyl vanillin, no matter they are synthetic or natural, have their own shortcomings that are difficult to overcome, that is, poor stability, and if they are used improperly, they will lose their ability to send out fragrance, which will cause the perfuming substances to produce harmful substances. disgusting smell
When used in cigarettes, the aroma is often pressed, causing retention

Method used

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  • Application of glucosyltransferase in production of ethyl vanillin-alpha-D-glucoside
  • Application of glucosyltransferase in production of ethyl vanillin-alpha-D-glucoside

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] Embodiment 1, preparation produces the bacterial agent of ethyl vanillin-α-D-glucoside

[0023] 1. Construction of Escherichia coli that highly expresses the glucosyltransferase of Xanthomonas tree fern

[0024] Referring to the amino acid sequence of the glucosyltransferase of Xanthomonas arborii reported in the NCBI database (NCBINO.AKU48857), send the sequence to the gene synthesis company (Wuxi Qinglan Biotechnology Co., Ltd.), requesting the codon-optimized gene sequence xar-agl (the nucleotide sequence is shown in SEQ ID NO.2, the amino acid sequence of the encoded protein (ie, glucosyltransferase) is shown in SEQ ID NO.1) is artificially synthesized, and cloned into the EcoRI / Between HindIII, the recombinant plasmid pET28a-xar-agl was obtained.

[0025] Transformation of E.coli Rosetta with Recombinant Plasmid pET28a-xar-agl TM (DE3) The cell method is as follows: take 5 μL of the recombinant plasmid pET28a-xar-agl and add it to 100 μL of E.coli Rosetta TM (D...

Embodiment 2

[0031] The application of embodiment 2 bacterial agents in the production of ethyl vanillin-α-D-glucoside

[0032] 1. Activity detection of bacterial agents

[0033] Take 10 mL of the bacteria agent cultured in Example 1, collect the cells by centrifugation at 5000 × g, resuspend 0.1 g of the cells in 10 mL of 50 mM phosphate buffer at pH 7.0; add ethyl vanillin at a final concentration of 8.3 g / L and maltose with a final concentration of 400g / L, catalyzed on a water bath shaker at 30°C and 150rpm for 30min. The reaction solution was used for HPLC analysis.

[0034] Liquid chromatography detection conditions. Sample pretreatment: Add 100 μL of reaction solution to 900 μL of 0.01mol / L dilute hydrochloric acid; centrifuge at 12000×g for 5 minutes, filter with a 0.22 μm filter membrane, and add the filtrate to a liquid phase vial; chromatographic column: C l8 Column, 250×4.6mm; column temperature: 25°C; mobile phase: CH 3 OH (methanol): H 2 O=45:55 (volume ratio); flow rate:...

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Abstract

The invention discloses application of glucosyltransferase in production of ethyl vanillin-alpha-D-glucoside. The amino acid sequence of the glucosyltransferase is shown as SEQ ID NO.2. According to the application, the fermentation liquor obtained by fermentation culture of engineering bacteria containing a glucosyltransferase coding gene is taken as a microbial agent, maltose and ethyl vanillinare taken as substrates, and the pH value is 6.0-8.5, conversion reaction is executed under the condition of 20-40 DEG C, and after the reaction is completed, the reaction liquid is separated and purified to obtain the ethyl vanillin-alpha-D-glucoside. The microbial agent disclosed by the invention is used for biocatalytic production of the ethyl vanillin-alpha-D-glucoside, the substrate conversion rate is 80% or above, and the product concentration of the ethyl vanillin-alpha-D-glucoside is high, the conversion rate is high, and recovery and purification the ethyl vanillin-alpha-D-glucoside prepared by a biological method are facilitated.

Description

(1) Technical field [0001] The invention relates to the application of a glucosyltransferase in the production of ethyl vanillin-α-D-glucoside. (2) Background technology [0002] The chemical structural formula of ethyl vanillin-α-D-glucoside is as follows figure 1 shown. Ethyl vanillin is made from o-ethoxyphenol or safrole oil. It is a broad-spectrum spice and one of the most important synthetic spices in the world today. It is an indispensable and important raw material in the food additive industry. Its aroma is 3-4 times that of vanillin, it has a strong vanilla bean aroma, and the fragrance lasts for a long time. Widely used in food, chocolate, ice cream, beverages and daily cosmetics to enhance and fix fragrance. In addition, ethyl vanillin can also be used as a feed additive, a brightener in the electroplating industry, and an intermediate in the pharmaceutical industry. However, vanillin and ethyl vanillin, no matter they are synthetic or natural, have their own...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/10C12P19/44C12P19/18
Inventor 陆跃乐徐安杰陈小龙朱林江陈翰驰范永仙
Owner SHENZHEN SHINESKY BIOLOGICAL TECH CO LTD
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