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Method for efficiently producing rhizopus oryzae lipase by using aspergillus oryzae

A technology of Rhizomucor oryzae and Aspergillus oryzae, which is applied in the field of high-efficiency production of Rhizomucor oryzae lipase by Aspergillus oryzae, can solve the problems of agglomeration, reducing production efficiency, affecting mass transfer and mixing of fermented liquid, and achieving yield and Improvement of enzyme activity and effect of increasing effective volume

Pending Publication Date: 2022-01-14
EAST CHINA UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The production of RML by Aspergillus oryzae is a complex fermentation system, often due to excessive growth of mycelium, causing problems such as sticky walls of the fermentation tank and agglomeration above the fermentation liquid. On the one hand, an uneven fermentation environment is formed, which affects the mass transfer of the fermentation liquid. Mixing and other characteristics, and it is easy to produce liquid escape and other phenomena, causing the fermentation to end early; on the other hand, the effective volume of the fermentation broth is greatly reduced, thereby reducing production efficiency

Method used

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  • Method for efficiently producing rhizopus oryzae lipase by using aspergillus oryzae
  • Method for efficiently producing rhizopus oryzae lipase by using aspergillus oryzae
  • Method for efficiently producing rhizopus oryzae lipase by using aspergillus oryzae

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Embodiment 1

[0068] The present embodiment provides a method for efficient production of Rhizomucor miehei lipase by Aspergillus oryzae, comprising steps:

[0069] Seed culture: inoculate the spore suspension of Aspergillus oryzae in the seed medium according to the inoculation amount of 0.6%, and cultivate it for 28 hours at a temperature of 28°C and a rotation speed of 150rpm to obtain a seed liquid; wherein, the spore suspension The concentration of spores is 2×10 5 a / mL;

[0070]Fermentation culture: inoculate the seed liquid into the fermentation medium according to the inoculum amount of 10%, set the culture temperature to 28°C, the initial rotation speed to 300rpm, and the ventilation to 1vvm to maintain the pressure of the reaction vessel at 0.03-0.04MPa, and calibrate The DO electrode (dissolved oxygen electrode) is full, and fermented; after the fermentation, the fermentation broth is obtained, that is, the Rhizomucor miehei lipase is obtained.

[0071] In this embodiment, duri...

Embodiment 2

[0077] The present embodiment provides a method for efficient production of Rhizomucor miehei lipase by Aspergillus oryzae, comprising steps:

[0078] Seed culture: inoculate the spore suspension of Aspergillus oryzae in the seed medium according to the inoculation amount of 0.6%, and cultivate it for 28 hours at a temperature of 28°C and a rotation speed of 150rpm to obtain a seed liquid; wherein, the spore suspension The concentration of spores is 4×10 5 a / mL;

[0079] Fermentation culture: inoculate the seed liquid into the fermentation medium according to the inoculum amount of 10%, set the culture temperature to 28°C, the initial rotation speed to 300rpm, and the ventilation to 1vvm to maintain the pressure of the reaction vessel at 0.03-0.04MPa, and calibrate The DO electrode (dissolved oxygen electrode) is full, and fermented; after the fermentation, the fermentation broth is obtained, that is, the Rhizomucor miehei lipase is obtained.

[0080] In this embodiment, dur...

Embodiment 3

[0086] The present embodiment provides a method for efficient production of Rhizomucor miehei lipase by Aspergillus oryzae, comprising the steps of:

[0087] Spread the spores of Aspergillus oryzae on the potato dextrose agar (PDA) medium and culture them at 28°C for 2 days, pick a single colony with better growth and continue to culture it on a new PDA plate for 7 days, and some colonies with spores (green) Scrape the spores in a test tube filled with sterile water and glass beads to elute, filter through a sterile funnel, count the spores using a hemocytometer, and then dilute the obtained spore suspension with sterile water to the required spore concentration. Store in refrigerator at 4°C for later use.

[0088] Under sterile conditions, take 300 μL of the spore suspension and add it to the seed shake flask with the seed culture solution. The liquid volume of the shake flask is 20%, and cultivate it for 28 hours at a temperature of 28 ° C and a rotation speed of 150 rpm to ...

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Abstract

The invention discloses a method for efficiently producing rhizopus oryzae lipase by using aspergillus oryzae. The method comprises the following steps of: seed culture: inoculating a spore suspension of the aspergillus oryzae into a seed culture medium, and culturing under the conditions that the temperature is 26-30 DEG C and the rotating speed is 100-200rpm to obtain a seed solution; fermentation culture: inoculating the seed solution into a fermentation culture medium, fermenting under the conditions that the temperature is 26-30 DEG C and the initial rotating speed is 300rpm, and obtaining fermentation liquor after the fermentation is finished, namely obtaining the rhizopus oryzae lipase, wherein the initial nitrogen source amount in the fermentation culture medium is 10-15% of all nitrogen sources, and the nitrogen sources are supplemented in the subsequent fermentation process. By adopting the method disclosed by the invention, the bacterial form can be controlled, a stable fermentation system is obtained under the conditions of nitrogen source supplementation and rotating speed control, the effective volume of fermentation is obviously improved, and the RML yield and enzyme activity are improved.

Description

technical field [0001] The application relates to the technical field of microbial fermentation, in particular to a method for efficiently producing Rhizomucor miehei lipase by Aspergillus oryzae. Background technique [0002] Rhizomucor miehei lipase (RML) is a typical interfacial enzyme that can catalyze ester synthesis, transesterification, ester hydrolysis, alcoholysis and other reactions in a non-aqueous environment, and is widely used in food processing, medicine, etc. , Detergent and other industries. The wild-type Rhizomucor miehei strain has a low ability to produce Rhizomucor miehei lipase (RML), and the composition of the enzyme liquid is unstable, which is difficult to meet the industrial application of Rhizomucor miehei lipase (RML). Some researchers have used genetic engineering technology to construct heterologous expression systems for RML in other strains, mainly including Pichia pastoris and Aspergillus oryzae. Although there have been many reports on the...

Claims

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Application Information

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IPC IPC(8): C12N9/20C12N1/14C12R1/69
CPCC12N9/20C12Y301/01003C12N1/14
Inventor 储炬许逗田锡炜王永红杨倚铭田桂尾吴学智庄英萍
Owner EAST CHINA UNIV OF SCI & TECH
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