Application of alpha-L-rhamnoside enzyme in directional synthesis of isoquercitrin by biological conversion of rutin

A technology of rhamnosidase and isoquercitrin, which is applied in directions such as being fixed on/in an organic carrier, fermentation, etc., can solve the problem of failing to achieve directional hydrolysis of rutin to prepare isoquercitrin, and not yet having high selectivity. Targeted preparation of isoquercitrin and other problems, to achieve good industrial application prospects, high catalytic efficiency and specificity, and high enzymatic conversion rate.

Active Publication Date: 2011-03-16
JIANGSU UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In addition, most of the reports on the preparation of isoquercitrin by using rhamnosidase to hydrolyze rutin focus on the enzymatic properties of the enzyme, gene recombination and other basic research (Archives of Biochemistry and Biophysics, 2003, 415: 235~244; Enzyme and Microbial Technology, 2007, 40: 1181~1187; Journal of Dalian Institute of Light Industry, 2007, 26 (2): 112~115; Process Biochemistry, 2010, 45: 1226~1235), there is no report on the highly selective and directional preparation of isoquercitrin by rhamnosidase biocatalyzed hydrolysis of rutin
In summary, none of the above hydrolysis methods can realize the directional hydrolysis of rutin to prepare isoquercitrin

Method used

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  • Application of alpha-L-rhamnoside enzyme in directional synthesis of isoquercitrin by biological conversion of rutin
  • Application of alpha-L-rhamnoside enzyme in directional synthesis of isoquercitrin by biological conversion of rutin
  • Application of alpha-L-rhamnoside enzyme in directional synthesis of isoquercitrin by biological conversion of rutin

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] (1) Selection of strains: Select Aspergillus niger CGMCC No.2588 ( Aspergillus niger LJ-1) as a strain producing α-L-rhamnosidase;

[0032] (2) Preparation of crude enzyme solution of α-L-rhamnosidase:

[0033] Enzyme production medium formula (weight ratio): 0.1% potassium hydrogen phosphate, 0.1% potassium dihydrogen phosphate, 0.1% ammonium sulfate, 0.01% sodium nitrate, 0.01% magnesium sulfate, 0.001% ferrous sulfate heptahydrate, zinc sulfate 0.001%, sucrose 1%, pH 5.0.

[0034] The strains were inoculated in the α-L-rhamnosidase enzyme production medium with a conventional amount, and fermented for 120 hours at 45°C with a shaker speed of 100 rpm, and then all the fermented products were mixed at a speed of 8000 rpm Centrifuge for 10 minutes, collect the supernatant to obtain the crude enzyme solution of α-L-rhamnosidase;

[0035] (3) α-L-rhamnosidase catalyzes the directional biosynthesis of isoquercitrin by hydrolyzing rutin:

[0036] A. The free enzyme of ...

Embodiment 2

[0043] (1) Selection of strains: Select Penicillium CGMCC No. 2590 ( Penicillium notatum LJ-2) as a strain producing α-L-rhamnosidase;

[0044] (2) Preparation of crude enzyme solution of α-L-rhamnosidase:

[0045] Enzyme production medium formula (weight ratio): 1.5% potassium dihydrogen phosphate, 1.5% potassium dihydrogen phosphate, 1.5% ammonium sulfate, 0.5% sodium nitrate, 0.5% magnesium sulfate, 0.5% ferrous sulfate heptahydrate, zinc sulfate 0.5%, sucrose 5%, pH 8.0.

[0046] The strains were inoculated in the α-L-rhamnosidase enzyme production medium in a conventional amount, and fermented for 12 hours at 25°C with a shaker rotation speed of 250 rpm, and then all the fermented products were mixed at a rotation speed of 4000 rpm Centrifuge for 30 minutes, collect the supernatant to obtain the crude enzyme solution of α-L-rhamnosidase;

[0047] (3) α-L-rhamnosidase catalyzes the directional biosynthesis of isoquercitrin by hydrolyzing rutin:

[0048] A. The free e...

Embodiment 3

[0055] (1) Selection of strains: Select Aspergillus niger CGMCC No.2588 ( Aspergillus niger LJ-1) as a strain producing α-L-rhamnosidase;

[0056] (2) Preparation of crude enzyme solution of α-L-rhamnosidase:

[0057] Enzyme production medium formula is (weight ratio): dipotassium hydrogen phosphate 1.0%, potassium dihydrogen phosphate 0.5%, ammonium sulfate 0.5%, sodium nitrate 0.05%, magnesium sulfate 0.05%, ferrous sulfate 0.01%, zinc sulfate 0.01% , pH 6.0.

[0058] The strains were inoculated in the α-L-rhamnosidase enzyme production medium in a regular amount, and fermented for 72 hours at 35°C with a shaker rotation speed of 160 rpm, and then all the fermented products were mixed at a rotation speed of 5000 rpm Centrifuge for 20 minutes, collect the supernatant to obtain the crude enzyme solution of α-L-rhamnosidase;

[0059] (3) α-L-rhamnosidase catalyzes the directional biosynthesis of isoquercitrin by hydrolyzing rutin:

[0060] A. The free enzyme of α-L-rhamnos...

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Abstract

The invention relates to application of an alpha-L-rhamnoside enzyme in the directional synthesis of isoquercitrin by the biological conversion of rutin. Because the crude enzyme preparation or the immobilized enzyme preparation of the alpha-L-rhamnoside enzyme is used for catalyzing and hydrolyzing the rutin and directionally and biologically synthesizing the isoquercitrin, the application has the advantages of wide source, easy preparation and low cost of catalysts, high stability, and high catalysis efficiency and specificity of the enzyme preparation, and is easy to store. Thus, the alpha-L-rhamnoside enzyme can greatly reduce the production cost of quercetin, the catalysis and conversion rate of the enzymes is high, and the products are single.

Description

[0001] 1. Technical field [0002] The invention belongs to the field of natural product biosynthesis, and in particular relates to a method for directional synthesis of isoquercitrin through enzymatic biotransformation of rutin. 2. Background technology [0003] Existing technology: Isoquercitrin (Quercetin-3-glucose) is a secondary glycoside produced by the hydrolysis of α-L-rhamnosyl in rutin molecule, which has anti-oxidation, blood pressure-lowering, anti-inflammatory and anti-cancer properties. It has similar or even higher bioactivity and bioavailability to rutin and quercetin in terms of inhibiting platelet aggregation and other aspects, and plays an important role in delaying human aging and preventing and curing diseases. The results of relevant pharmacological experiments show that isoquercitrin has the effects of promoting body fluid and quenching thirst, clearing heat and relieving heat, lowering blood pressure, strengthening the heart and prolonging life, and c...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P17/06C12N11/10
Inventor 王俊吴福安孙国霞马延龙安亚雄
Owner JIANGSU UNIV OF SCI & TECH
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