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Method for producing high-temperature-resistant chymosin by fermenting bacillus methanolicus in distiller's yeast

A technology of bacillus and rennet, applied in the biological field, can solve the problems of short fermentation period, easy spread of mold and low production cost, etc.

Active Publication Date: 2017-11-24
BEIJING TECHNOLOGY AND BUSINESS UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the characteristics of easy to spread, easy to pollute, and difficult to control growth conditions, the development of mold is limited.
In contrast, the liquid state fermentation of bacteria to produce chymosin has a short fermentation cycle, easy control of growth and enzyme production process, and the produced chymosin is an extracellular enzyme that is easy to extract and separate, and the production cost is low, so it has good research and development. Prospect, and there is no research report on the production of chymosin by fermentation of Bacillus methanolica in the literature

Method used

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  • Method for producing high-temperature-resistant chymosin by fermenting bacillus methanolicus in distiller's yeast
  • Method for producing high-temperature-resistant chymosin by fermenting bacillus methanolicus in distiller's yeast
  • Method for producing high-temperature-resistant chymosin by fermenting bacillus methanolicus in distiller's yeast

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] Embodiment 1, the acquisition and identification of Bacillus methanolica LB-1

[0054] 1. Isolation and purification of Bacillus methanolica LB-1

[0055] Weigh 0.5 g of pulverized wheat koji (rice koji for rice wine) and add it into a test tube containing 4.5 mL of sterile saline, mix well in a shaker, so that the wheat koji is evenly dispersed in sterile saline, and the prepared sample Dilution of 10 -1 . The samples were then serially diluted 10-fold until the dilution was 10 -7. Take 100μL each 10 -3 to 10 -7 Smear the diluted solution of the yeast extract on LB (yeast extract powder 5g, tryptone 10g, sodium chloride 10g, agar powder 15g, distilled water 1L), YEPD (yeast powder 10g, peptone 20g, glucose 20g, agar powder 15g, distilled water 1L) and PDA (potato powder 15g, glucose 3g, agar powder 15g, distilled water 1L) three solid medium, the experiment was repeated three times. Then put the culture medium upside down in a constant temperature incubator at 3...

Embodiment 2

[0072] Example 2, Determination of curdling activity and proteolytic activity of LB-1 bacterial strain

[0073] 1. Determination of milk curd activity of LB-1 strain

[0074] 1. Preparation of LB-1 strain fermentation broth

[0075] Get a small amount of LB-1 bacterial strain and inoculate in LB liquid medium, at 30 ℃, shake 12h under the condition of 120r / min, as activated seed liquid, then inoculate in LB liquid medium with the inoculation amount of 3% (volume fraction), The liquid filling volume is 40% (v / v), and the culture is shaken at 30° C. and 120 r / min to obtain LB-1 fermentation broth. When culturing 0h, 12h, 24h, 36h, 48h, 60h and 72h respectively, the LB-1 fermentation liquid was taken as the bacterial liquid to be tested. Experiments were repeated three times.

[0076] 2. Determination of curdling activity of LB-1 fermented broth

[0077] Dissolve skim milk (purchased from Fonterra Trading Shanghai Co., Ltd.) in CaCl with a concentration of 0.01mol / L 2 Prepar...

Embodiment 3

[0102] The optimization of embodiment 3, LB-1 strain culture condition

[0103] 1. Optimization of medium components

[0104] 1. Effect of different carbon sources on curdling activity

[0105] (1) Preparation of different carbon source medium

[0106] The following different types of carbon sources were selected respectively: fructose (Beijing Pinellia Science and Technology Development Co., Ltd., product number a3125), soluble starch (Sinopharm Chemical Reagent Co., Ltd., product number 20120405), glucose (Beijing Pinellia Science and Technology Development Co., Ltd., product number B2657), α-lactose (Sinopharm Chemical Reagent Co., Ltd., product number is 20111025), sucrose (Beijing Banxia Science and Technology Development Co., Ltd., product number is b3075), galactose (Beijing Pinellia Science and Technology Development Co., Ltd., product number is b2650) Add it to the original culture medium (yeast extract powder 5g, tryptone 10g, sodium chloride 10g, distilled water 1...

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Abstract

The invention discloses a method for producing high-temperature-resistant chymosin by fermenting bacillus methanolicus in distiller's yeast. The bacillus methanolicus is separated and screened out from yellow rice wine wheat starter and has the preservation number of CGMCC (China General Microbiological Culture Collection Center) No. 14424. The invention also provides the method for preparing high-activity chymosin by carrying out liquid state fermentation on the bacillus methanolicus. The method comprises the steps of activating a strain, optimizing the culture conditions of the strain by using a response surface method, extracting the chymosin under the best enzyme production condition, and researching the enzymological properties of the chymosin. The test results prove that after the culture conditions are optimized, the novel high temperature tolerance type chymosin with high chymosin activity can be prepared, and the chymosin activity reaches up to 3.2*10<6>+ / -0.35SU / mg. The production method of the chymosin is simple and easy to implement and low in cost, thus being suitable for industrial production.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for producing high-temperature-resistant rennet by fermentation of Bacillus methanolica in distiller's yeast. Background technique [0002] The use of koji in the brewing process is a feature of Chinese wine, and it is also one of the great inventions of China. The essence of traditional koji is a crude enzyme preparation, and the main function is the microorganisms in the koji. They can not only give the wine a unique aroma but also Various enzymes are produced, including amylases and proteases. The rennet in rice wine koji is the metabolite of microorganisms in koji, which belongs to microbial rennet. Using the rennet produced by microbial fermentation in rice wine koji to make cheese can ensure the food safety of the product. [0003] Coagulation of milk is a fundamental step in cheese making and plays a huge role in the maturation of cheese. The earliest re...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/54A23C19/032C12R1/07
CPCA23C19/032C12N9/54C12Y304/21039
Inventor 杨贞耐郑喆李柳赵笑赵娟郝晓娜
Owner BEIJING TECHNOLOGY AND BUSINESS UNIVERSITY