Method for producing chymosin from paenibacillus damxungensissp.nov. in bran culture medium

Abstract

The invention discloses a method for producing chymosin by fermenting paenibacillus damxungensissp.nov. CGMCC (China General Microbiological Culture Collection Center) No.8333 in a bran culture medium. The method comprises the following steps: inoculating paenibacillus damxungensissp.nov. seed liquid to the bran culture medium, and fermenting at 28-32 DEG C for 16-20 hours to obtain fermenting liquid with chymosin activity, wherein the standard liquid loading volume of a conical flask of 250mL is 20mL to 40mL. Through measurement, highest chymosin activity is 6733.81SU / mL. A paenibacillus damxungensissp.nov. fermenting liquid extract with relatively high chymosin activity can be obtained by virtue of optimized culture conditions, and has wide application prospects in cheese production and processing industry.

Description

technical field

[0001] The invention belongs to the field of biotechnology, and in particular relates to a method for producing chymosin by fermenting Paenibacillus damxungensis sp.nov. CGMCC No.8333 in a bran medium. Background technique

[0002] In recent years, the cheese industry has developed rapidly. Curdling is a key step in the production of original cheese. Rennet plays a vital role in this process and is directly related to the quality and yield of cheese. Therefore, as the world's cheese production continues to grow, so does the demand for rennet. The annual demand for curd agent in the world is 25×10 6 L, the output accounts for about 20% of the total enzyme preparations in the world, becoming the largest enzyme preparation. The main function of chymosin is to specifically hydrolyze the κ-casein polypeptide chain in milk to form paracasein and hydrophilic glycomacropeptide. When the casein is hydrolyzed to a certain extent, the calcium ions in the system pass t...

Examples

Embodiment 1

[0025] The preparation of embodiment 1 seed liquid

[0026] Take 30mL TYC liquid culture medium and place them in 100mL Erlenmeyer flasks, seal with multi-layer (8-12 layers) gauze, seal with kraft paper, and sterilize with high-temperature steam at 121°C for 15 minutes;

[0027] Pick the fresh culture of Paenibacillus damxungensis sp.nov. CGMCC No.8333 cultured on TYC solid medium (agar) for 24 hours, inoculate it into TYC liquid medium, and culture it on a shaker at 30°C for 20 hours to obtain seeds liquid.

Embodiment 2

[0029] The seed liquid that embodiment 1 prepares is inoculated by liquid medium 5% (percentage by volume), in 20mL bran culture medium (the bran content of described bran culture medium is 3%, all the other are deionized water) For fermentation, set the temperature of the shaker at 30° C. and the rotation speed of 300 r / min, and shake the fermentation for 20 hours.

[0030] After culturing for 20 hours, take 5 mL of fermentation broth, centrifuge at 4°C and 9000 r / min, and the supernatant obtained is the crude enzyme solution. The obtained crude enzyme solution was diluted 5 times, and 0.5 mL of the diluted solution was taken to determine the MCA value.

[0031] The experimental results showed that the chymosin activity of 20mL bran cultured crude enzyme liquid of Paenibacillus damxungensis sp.nov. CGMCC No.8333 was shaken and fermented at 30°C for 20 hours, and the activity of rennet was 4760.71SU / mL.

Embodiment 3

[0033] The seed liquid that embodiment 1 prepares is inoculated by liquid medium 5% (percentage by volume), in 40mL bran culture medium (the bran content of described bran culture medium is 3%, all the other are deionized water) For fermentation, set the temperature of the shaker at 30° C. and the rotation speed of 300 r / min, and shake the fermentation for 20 hours.

[0034]After culturing for 20 hours, take 5 mL of fermentation broth, centrifuge at 4°C and 9000 r / min, and the supernatant obtained is the crude enzyme solution. The obtained crude enzyme solution was diluted 5 times, and 0.5 mL of the diluted solution was taken to determine the MCA value.

[0035] The experimental results showed that the rennet activity of 40mL bran cultured crude enzyme solution of Paenibacillus damxungensis sp.nov. CGMCC No.8333 was shaken and fermented at 30°C for 20 hours, and the rennet activity was 4578.64SU / mL.

technical field

[0001] The invention belongs to the field of biotechnology, and in particular relates to a method for producing chymosin by fermenting Paenibacillus damxungensis sp.nov. CGMCC No.8333 in a bran medium. Background technique

[0002] In recent years, the cheese industry has developed rapidly. Curdling is a key step in the production of original cheese. Rennet plays a vital role in this process and is directly related to the quality and yield of cheese. Therefore, as the world's cheese production continues to grow, so does the demand for rennet. The annual demand for curd agent in the world is 25×10 6 L, the output accounts for about 20% of the total enzyme preparations in the world, becoming the largest enzyme preparation. The main function of chymosin is to specifically hydrolyze the κ-casein polypeptide chain in milk to form paracasein and hydrophilic glycomacropeptide. When the casein is hydrolyzed to a certain extent, the calcium ions in the system pass t...

Examples

Embodiment 1

[0025] The preparation of embodiment 1 seed liquid

[0026] Take 30mL TYC liquid culture medium and place them in 100mL Erlenmeyer flasks, seal with multi-layer (8-12 layers) gauze, seal with kraft paper, and sterilize with high-temperature steam at 121°C for 15 minutes;

[0027] Pick the fresh culture of Paenibacillus damxungensis sp.nov. CGMCC No.8333 cultured on TYC solid medium (agar) for 24 hours, inoculate it into TYC liquid medium, and culture it on a shaker at 30°C for 20 hours to obtain seeds liquid.

Embodiment 2

[0029] The seed liquid that embodiment 1 prepares is inoculated by liquid medium 5% (percentage by volume), in 20mL bran culture medium (the bran content of described bran culture medium is 3%, all the other are deionized water) For fermentation, set the temperature of the shaker at 30° C. and the rotation speed of 300 r / min, and shake the fermentation for 20 hours.

[0030] After culturing for 20 hours, take 5 mL of fermentation broth, centrifuge at 4°C and 9000 r / min, and the supernatant obtained is the crude enzyme solution. The obtained crude enzyme solution was diluted 5 times, and 0.5 mL of the diluted solution was taken to determine the MCA value.

[0031] The experimental results showed that the chymosin activity of 20mL bran cultured crude enzyme liquid of Paenibacillus damxungensis sp.nov. CGMCC No.8333 was shaken and fermented at 30°C for 20 hours, and the activity of rennet was 4760.71SU / mL.

Embodiment 3

[0033] The seed liquid that embodiment 1 prepares is inoculated by liquid medium 5% (percentage by volume), in 40mL bran culture medium (the bran content of described bran culture medium is 3%, all the other are deionized water) For fermentation, set the temperature of the shaker at 30° C. and the rotation speed of 300 r / min, and shake the fermentation for 20 hours.

[0034]After culturing for 20 hours, take 5 mL of fermentation broth, centrifuge at 4°C and 9000 r / min, and the supernatant obtained is the crude enzyme solution. The obtained crude enzyme solution was diluted 5 times, and 0.5 mL of the diluted solution was taken to determine the MCA value.

[0035] The experimental results showed that the rennet activity of 40mL bran cultured crude enzyme solution of Paenibacillus damxungensis sp.nov. CGMCC No.8333 was shaken and fermented at 30°C for 20 hours, and the rennet activity was 4578.64SU / mL.

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