Glucosyltransferase and application thereof in producing vanillin-alpha-D-glucoside

A technology of glucosyl and glucoside, applied in the field of glucosyl transferase and its production of vanillin-α-D-glucoside, can solve problems such as premature release of flavor, and achieve the effects of high conversion rate and high product concentration

Active Publication Date: 2019-04-16
杭州佳嘉乐生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

And its melting point is only about 80°C, and it will decompose in advance under the high t...

Method used

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  • Glucosyltransferase and application thereof in producing vanillin-alpha-D-glucoside
  • Glucosyltransferase and application thereof in producing vanillin-alpha-D-glucoside

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

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

[0023] 1. Extraction of Xanthomonas campestris genomic DNA

[0024] Xanthomonas campestrispv. campestris (Xanthomonas campestrispv. campestris) CGMCC No.13990 (disclosed in patent application CN107446844A) was activated by streaking on NYGA medium, cultured at 28°C for 36h, and a single colony was picked on NYGB medium in 30°C and 180rpm for 24 hours; use a 2mL centrifuge tube to take 1.5-3mL of bacterial liquid, centrifuge at 10000rpm for 2min, discard the supernatant, and wash the bacterial cells with deionized water for 2-3 times; 600 μL of cell lysate (40mM Tris-acetic acid buffer pH 7.8, 20mM sodium acetate, 1mM EDTA, 1% SDS) was pipetted sufficiently, and placed in ice water for 5min; then added 200μL of 5mol Invert / L NaCl back and forth 8-10 times, a white precipitate forms; to remove impurities and cell debris, centrifuge at 12,000 rpm at 4°C for 10 min; after centrifugation...

Embodiment 2

[0037] The application of embodiment 2 bacterial agents in the production of vanillin-α-D-glucoside

[0038] 1. Activity detection of bacterial agents

[0039] Get 10 mL of the bacterial agent cultured in Example 1, collect the cells by centrifugation at 5000 × g, resuspend 0.3 g of the cells in 10 mL of 50 mM phosphate buffer at pH 7.0; add vanillin and final concentration of 7.6 g / L Maltose with a concentration of 400g / L was catalyzed on a water bath shaker at 30°C and 150rpm for 30min. The reaction solution was used for HPLC analysis.

[0040] 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=35:65 (volume ratio); flow rate: 1.0mL min -1 ; Det...

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Abstract

The invention discloses glucosyltransferase and application thereof in producing vanillin-alpha-D-glucoside. The amino acid sequence of glucosyltransferase is shown in SEQ NO.2. According to the application, fermentation liquid which is obtained by conducting fermentation cultivation on engineering bacteria containing glucosyltransferase coding genes serves as a bactericide, maltose and vanillin serve as a substrate, a conversion reaction is carried out with the pH value of 6.0-8.5 and at the temperature of 20-40 DEG C, and after the reaction is carried out completely, reaction liquid is separated and purified to obtain vanillin-alpha-D-glucoside. According to glucosyltransferase and the application thereof, vanillin-alpha-D-glucoside is produced through biological catalysis of the bactericide, after the reaction is carried out for 12 h, a vanillin-alpha-D-glucoside solution with the concentration lager than 10% can be obtained, the substrate conversion rate is larger than 60%, the yield concentration of vanillin-alpha-D-glucoside is high, and the conversion rate is high.

Description

(1) Technical field [0001] The invention relates to a glucosyltransferase and its application in producing vanillin-α-D-glucoside. (2) Background technology [0002] Vanilla orchid is one of the most important food spices, widely used in high-end food, beverage and cosmetic industries, and its main flavor component is vanillin. Has a strong and distinctive vanilla bean aroma, naturally occurring in vanilla pods, as well as clove oil, oakmoss oil, Peru balsam, tolu balsam and benzoin. Vanillin is generally converted from vanillin glucoside in mature pods through a long fermentation process. Vanillin is gradually oxidized in the air, decomposes when exposed to light, and has unstable physical and chemical properties. And its melting point is only about 80°C, and it will decompose in advance under the high temperature during the tobacco combustion process, causing the fragrance to be released prematurely. As a flavor precursor, its glycoside has a relatively large molecular ...

Claims

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Application Information

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IPC IPC(8): C12N9/10C12N15/54C12N15/70C12N1/21C12P19/44C12R1/19
CPCC12N1/20C12N9/1051C12N15/70C12P19/44
Inventor 朱林江徐安杰陈翰驰陈小龙陆跃乐范永仙
Owner 杭州佳嘉乐生物技术有限公司
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