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A kind of method utilizing mixed bacteria fermentation to prepare dextran sucrase

A technology for fermentation of dextran and mixed bacteria, which is applied in the field of producing dextran sucrase, can solve the problems of high cost, many steps, and low enzyme activity, and achieve the effects of low cost, high enzyme activity, and high enzyme activity

Active Publication Date: 2018-05-01
SHANGHAI HUAMAO PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] The invention provides a method for preparing dextran sucrase by using mixed bacteria fermentation, aiming to overcome the shortcomings of traditional preparation of dextran sucrase, such as low enzymatic activity, many steps and high cost

Method used

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  • A kind of method utilizing mixed bacteria fermentation to prepare dextran sucrase
  • A kind of method utilizing mixed bacteria fermentation to prepare dextran sucrase
  • A kind of method utilizing mixed bacteria fermentation to prepare dextran sucrase

Examples

Experimental program
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Effect test

Embodiment 1

[0028] The strains used: Leuconostoc mesenteroides-0326, Penicillium aculeatum F1008.

[0029] 1. Preparation of Penicillium aculeatus seed solution

[0030]The mass percent content of each component in the seed medium of Penicillium aculeatus is: 1.5% dextran T70, 0.5% peptone, 0.05% MgSO4 7H2O, 0.05% KCl, 0.1% K 2 HPO 4 ·3H 2 O, 0.001% FeSO 4 ·7H 2 O, the balance is water; the seed culture medium of Penicillium aculeatus has a pH of 5.5 and is sterilized at 0.1 MPa for 20 minutes.

[0031] Preparation of the seed solution of Penicillium aculeatum: pick colonies from the culture dish where Penicillium aculeatum was preserved, put them in the culture medium, cultivate them at 28°C and 200r / min for 5 days, and the obtained fermentation liquid is Acanthosporium aculeatum The seed solution of Penicillium was stored at 4°C for later use.

[0032] 2. Preparation of the seed solution of Leuconostoc enterica

[0033] The mass percent content of each component in the seed cultu...

Embodiment 2

[0038] Example 2, Dextransucrase prepared by fermentation of Leuconostoc membranaceus

[0039] As a control group, the seed solution of Leuconostoc enterococcus was inserted into a 250mL Erlenmeyer flask containing 100mL of fermentation medium according to 3% of the medium volume, and fermented at 26°C and 120r / min for 48 hours. After centrifugation, The obtained supernatant is subjected to membrane fractionation filtration to obtain a crude enzyme liquid of dextran sucrase.

[0040] The enzymatic activity of the obtained dextran sucrase was detected by the DNS method. The specific steps were: mix 2 mL of crude enzyme solution and 2 mL of sucrose solution with a mass concentration of 10% to obtain a reaction solution, take out 500 μL of the reaction solution and add 375 μL of DNS reagent, and keep it in boiling water for 5 minutes After cooling to room temperature, add 5.375mL of distilled water and shake well, and measure the absorbance at a wavelength of 520nm. Taking the r...

Embodiment 3

[0043] The seed liquid of Leuconostoc enterococci, the seed liquid of Penicillium aculeatus and the fermentation medium used in this embodiment are the same as those in Example 1.

[0044] The present embodiment prepares dextran sucrase by mixed bacteria fermentation according to the following steps:

[0045] First, the seed liquid of Leuconostoc enterococcus is inserted into a 250mL Erlenmeyer flask containing 50mL of fermentation medium according to 3% of the fermentation medium volume, and simultaneously the seeds of Penicillium aculeatus accounting for 3% of the fermentation medium volume are 26 ° C, 120r / min mixed bacteria fermentation culture, 20mL of sucrose solution with a mass concentration of 15% was added at 5h, 10h and 20h of fermentation, and the fermentation was completed after 48 hours of mixed bacteria fermentation culture, and the above obtained after centrifugation The supernatant was filtered by membrane fractionation to obtain about 100 mL of crude enzyme s...

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Abstract

The invention discloses a method for preparing dextransucrase by using mixed fermentation. The method is characterized in that firstly seed solution of leuconostoc mesenteroides is inoculated into a fermentation medium containing saccharose with the mass concentration of 2 percent to 5 percent for fermentation, saccharose solution with the mass concentration of 15 percent is replenished in the fermentation process, seed solution of penicillium aculeatum is added within 0-36 hours of the fermentation of the leuconostoc mesenteroides for continuously performing mixed fermentation, and an obtained fermentation liquor is subjected to membrane classified filtering to obtain dextransucrase crude enzyme. The method disclosed by the invention has the advantages that relative to separately using the leuconostoc mesenteroides to ferment to prepare the dextransucrase, mixed fermentation can solve the problem of difficulty in separating the dextransucrase, and higher enzyme activity can be obtained with lower cost and less energy consumption; relative to using engineering bacteria to prepare the dextransucrase, mixed fermentation can achieve higher enzyme stability and simple and convenient operation, and the requirements on bacteria growth conditions and enzyme preparation facilities are low.

Description

technical field [0001] The invention belongs to the field of microbial fermentation, and in particular relates to a method for producing dextran sucrase by using mixed bacteria fermentation. Background technique [0002] Dextran sucrase, also known as Glucosyltransferase and Glucansucrases, can synthesize dextran of different structures using sucrose as a substrate. The enzyme is mostly produced by Leuconostoc mesenteroides, Streptococcus sp. and Lactobacillus sp. Dextran sucrase is currently mainly used to prepare dextran, which is widely used in pharmaceutical, food, chemical and other industries. In addition, immobilized dextran sucrase can be used to synthesize some functional oligosaccharides, which are often used as prebiotics in the food industry. Dextransucrase can also use different sugars as substrates to synthesize some products with special effects through reactions. [0003] The high-quality production of dextran and the generation of dextran with different m...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N9/10C12R1/01C12R1/80
CPCC12N9/1051C12Y204/01005
Inventor 张洪斌李若菡陈爽曹研研胡雪芹
Owner SHANGHAI HUAMAO PHARMA
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