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Application of alkaline cyclodextrin glucosyltransferase in production of alpha-glucosylhesperidin

A technology based on glucosyl and cyclodextrin, which is applied in the direction of glycosyltransferase, transferase, and microorganism-based methods, can solve the problems of low substrate conversion rate and achieve high substrate conversion rate, high-efficiency preparation, and water-soluble Improved effect

Inactive Publication Date: 2020-04-10
ZHEJIANG UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The enzymatic method has strong specificity, mild conditions, and no "three wastes". Some cyclodextrin glucosyltransferases have been reported to modify hesperidin, but there is a disadvantage of low substrate conversion rate.

Method used

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  • Application of alkaline cyclodextrin glucosyltransferase in production of alpha-glucosylhesperidin
  • Application of alkaline cyclodextrin glucosyltransferase in production of alpha-glucosylhesperidin
  • Application of alkaline cyclodextrin glucosyltransferase in production of alpha-glucosylhesperidin

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Experimental program
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Embodiment 1

[0032] Embodiment 1, preparation produces the bacterial agent of α-glucosyl hesperidin

[0033] 1. Construction of Escherichia coli that highly expresses alkaline cyclodextrin glucosyltransferase

[0034]Referring to the amino acid sequence of cyclodextrin glucosyltransferase (NCBI Accession No. AAV38118) from Bacillus sp. reported in the NCBI database, the gene synthesis company (Hua Da Gene Qinglan Biotechnology Co., Ltd. ), through codon optimization, artificially synthesized cyclodextrin glucosyltransferase coding gene alkacgt (the nucleotide sequence is shown in SEQ ID NO.1, the amino acid sequence is shown in SEQ ID NO.2), and cloned into the expression vector Between NcoI / BamHI of pET22b, a recombinant plasmid pET22b-alkacgt was obtained.

[0035] Transform the recombinant plasmid pET22b-alkacgt constructed by the gene synthesis company into the expression host E.coli Rosetta TM (DE3), the specific operation is as follows: Take 2 μL of the recombinant plasmid pET22b-a...

Embodiment 2

[0049] Embodiment 2, the application of catalyst bacteria agent in the production of α-glucosyl hesperidin

[0050] 1. Activity detection of catalytic bacteria

[0051] Take 0.2g of hesperidin in a round bottom flask (with rotor), add 10ml of pre-configured 50mmol / L pH 10.0 glycine-sodium hydroxide buffer as the reaction medium, add 0.08g of sodium hydroxide solid, shake well , to completely dissolve the solid particles of hesperidin, and gradually add dropwise 11mol / L HCl aqueous solution to adjust the pH of the system to 10.0; Dilute with 50mmol / L glycine-sodium hydroxide buffer solution with a pH of 10.0 to obtain a bacterial suspension with a bacterial cell content of 10g / L, and obtain a catalytic bacterial agent after high-pressure homogenization and crushing; add 30mL of the catalytic bacterial agent to the above reaction system During the catalytic process, titrate the pH with 2mol / L NaOH aqueous solution to maintain it at 10.0, catalyze it in a magnetic stirrer in a c...

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Abstract

The invention discloses application of alkaline cyclodextrin glucosyltransferase in production of alpha-glucosylhesperidin. The amino acid sequence of the alkaline cyclodextrin glucosyltransferase isas shown in SEQ ID NO. 2. The alkaline cyclodextrin glucosyltransferase disclosed by the invention can realize high-level secretory expression in escherichia coli, so efficient preparation of the alkaline cyclodextrin glucosyltransferase is realized; the conversion rate of hesperidin with a concentration of 0.5-15% can reach 58-88% within 16 hours; and the alkaline cyclodextrin glucosyltransferasehas the characteristics of high substrate feeding amount, high catalytic rate and high substrate conversion rate and is suitable for industrial production of alpha-glucosylhesperidin.

Description

[0001] (1) Technical field [0002] The invention relates to the application of a cyclodextrin glucosyltransferase in the production of α-glucosyl hesperidin. [0003] (2) Background technology [0004] Hesperidin, also known as hesperidin and hesperidin, is a glycoside formed by hesperetin and rutinose, and is a dihydroflavone derivative. It is widely found in legumes, Labiatae, Disciformes, and Rutaceae citrus plants. It has the functions of maintaining normal osmotic pressure of blood vessels, reducing vascular fragility, enhancing capillary toughness, and shortening bleeding time. Hesperidin not only has the effect of lowering cholesterol in the human body, but also has the effect of anti-virus and improving immunity. [0005] Hesperidin has poor water solubility and a bitter taste, which greatly limits its application. Through glycosylation modification, its solubility in water can be increased by more than 10,000 times, greatly improving its solubility, and improving th...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P19/60C12P19/18C12N9/10C12N15/54C12N15/70C12N1/21C12R1/19
CPCC12N9/1074C12N15/70C12P19/18C12P19/60C12Y204/01019
Inventor 陈小龙江瑞妮朱林江许敏陆跃乐范永仙陈翰驰
Owner ZHEJIANG UNIV OF TECH
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