Production method of feed additive rich in heat-resistant protease and Cordycepin by means of solid state fermentation with Cordyceps militaris

Abstract

The invention relates to a preparation method of a Cordyceps feed additive. According to the method, Cordyceps militaris which produces heat-resistant protease and Cordycepin in high yield is used as a solid state fermentation production strain. Through the process steps of preparation of a fermentation strain, solid state fermentation with conical flasks, solid state fermentation with koji trays, proteolysis after fermentation and the like, the Cordyceps feed additive is prepared. The protein enzyme activity in dry culture obtained by the solid state fermentation with the koji trays reaches 12941 U / g, and the Cordycepin content is 780 mg / 100 g. According to the feed additive prepared from fermentation products, the active functional components of Cordyceps are maintained to the utmost extent, besides, the feed additive also contains highly active heat-resistant protease, functional active polypeptides and amino acids, wherein the functional active polypeptides and the amino acids are obtained by hydrolysis of the protease. By means of the additive, the growth of animals can be better promoted, and the conversion rate of the feed is improved, so that the cost is reduced; moreover, the immunity and disease resistance of the animals can be improved, and the survival rate is improved, so that the cost is reduced.

Description

technical field

[0001] The invention belongs to the technical field of processing Cordyceps militaris, in particular to a feed additive method for producing heat-resistant protease and cordycepin by solid-state fermentation of Cordyceps militaris. Background technique

[0002] In recent years, with the continuous improvement of people's living standards, the demand for livestock products is also increasing. However, due to the keen pursuit of the output of animal products, the quality and safety of animal products are often overlooked, especially the problem that the residues of veterinary drugs and drug additives in animal products are harmful to human health. Therefore, green and natural feed additives, such as natural non-toxic effects, no drug residues, no drug resistance, can promote animal growth, can effectively and economically improve animal production performance, improve feed utilization and animal product quality, and have high breeding benefits. This type of fe...

Examples

Embodiment 1

[0030] Embodiment 1, the screening of Cordyceps militaris bacterial strain

[0031] Isolation and screening of the above-mentioned Cordyceps militaris (Cordyceps militaris) bacterial classification:

[0032] 1. Isolation and screening of bacterial strains

[0033] In August 2015, wild Cordyceps militaris was collected in Yimeng Mountain, Yinan County, Shandong Province. The collection site was in a forest with loose soil and rich humus. The ambient temperature was 20-28°C and the air humidity was 70-80%. A total of 9 wild Cordyceps strains were collected, and the Cordyceps was slowly peeled off from the layer, and the surface of the Cordyceps was rinsed repeatedly with clean water until it was cleaned. Blot the surface moisture of Cordyceps dry with sterile filter paper. Then disinfect the surface with 75% ethanol or 0.1 L mercury, and rinse with sterile water again. Then, carefully cut out a small piece from the Cordyceps militaris subunit, put it on the activated solid me...

Embodiment 2

[0057] Embodiment 2: compound mutagenesis and breeding method of Cordyceps militaris bacterial strain

[0058] (1) Activation culture of strains The wild strain XZCM6 isolated in Example 1 was transferred to PDA solid slant medium for activation culture, and cultured at a constant temperature of 25-28°C for 4-5 days. The strains were activated twice as above.

[0059] (2) Preparation of strain mycelium Take 1 cm from the activated mycelium solid plate medium 2 The mycelium block was inoculated into a 250mL Erlenmeyer flask filled with 40mL PDA mycelium medium, and cultured with constant temperature shaking at 24-28°C and 160-200 r / min for 3-7d. Mycelia liquid medium formula: glucose 20-30g, peptone 5-8g, 20% potato extract 200ml, (NH 4 ) 2 S0 4 3~5g, KH 2 P0 4 1~2g, MgS0 4 ·7H 2 0 1~2g, yeast powder 4~6g, add water to make up to 1L, pH 6.5~7.0, sterilize at 121℃ for 20min.

[0060] (4) Protoplast preparation MgS0 4 ·7H 20. Mannitol was dissolved in double-distille...

Embodiment 3

[0064] Embodiment 3 Preparation of solid-state fermentation strain

[0065] This embodiment 3 illustrates the method for preparing a heat-resistant protease-rich and cordycepin feed additive by using Cordyceps militaris HYCM12 as a strain solid-state fermentation method, including the following steps:

[0066] Step 1, strain activation: first, inoculate the original seed of the preserved Cordyceps militaris HYCM12 test tube strain on the activation medium of a petri dish, and incubate in an incubator at 20-26° C. in the dark for 5-7 days to obtain first-grade seeds.

[0067] Activated slant medium: 200 g of potatoes, washed and chopped, add 1000 mL of water and boil until boiling for 20 min, filter with gauze, add 20 g of glucose to the filtrate, 1 mL of inorganic salt solution, 15 g of agar and continue to boil, fully dissolve and then pack into test tubes , each test tube is about 4 mL, and the culture medium is sterilized at 121°C for 20 min. After the sterilization is comp...

technical field

[0001] The invention belongs to the technical field of processing Cordyceps militaris, in particular to a feed additive method for producing heat-resistant protease and cordycepin by solid-state fermentation of Cordyceps militaris. Background technique

[0002] In recent years, with the continuous improvement of people's living standards, the demand for livestock products is also increasing. However, due to the keen pursuit of the output of animal products, the quality and safety of animal products are often overlooked, especially the problem that the residues of veterinary drugs and drug additives in animal products are harmful to human health. Therefore, green and natural feed additives, such as natural non-toxic effects, no drug residues, no drug resistance, can promote animal growth, can effectively and economically improve animal production performance, improve feed utilization and animal product quality, and have high breeding benefits. This type of fe...

Examples

Embodiment 1

[0030] Embodiment 1, the screening of Cordyceps militaris bacterial strain

[0031] Isolation and screening of the above-mentioned Cordyceps militaris (Cordyceps militaris) bacterial classification:

[0032] 1. Isolation and screening of bacterial strains

[0033] In August 2015, wild Cordyceps militaris was collected in Yimeng Mountain, Yinan County, Shandong Province. The collection site was in a forest with loose soil and rich humus. The ambient temperature was 20-28°C and the air humidity was 70-80%. A total of 9 wild Cordyceps strains were collected, and the Cordyceps was slowly peeled off from the layer, and the surface of the Cordyceps was rinsed repeatedly with clean water until it was cleaned. Blot the surface moisture of Cordyceps dry with sterile filter paper. Then disinfect the surface with 75% ethanol or 0.1 L mercury, and rinse with sterile water again. Then, carefully cut out a small piece from the Cordyceps militaris subunit, put it on the activated solid me...

Embodiment 2

[0057] Embodiment 2: compound mutagenesis and breeding method of Cordyceps militaris bacterial strain

[0058] (1) Activation culture of strains The wild strain XZCM6 isolated in Example 1 was transferred to PDA solid slant medium for activation culture, and cultured at a constant temperature of 25-28°C for 4-5 days. The strains were activated twice as above.

[0059] (2) Preparation of strain mycelium Take 1 cm from the activated mycelium solid plate medium 2 The mycelium block was inoculated into a 250mL Erlenmeyer flask filled with 40mL PDA mycelium medium, and cultured with constant temperature shaking at 24-28°C and 160-200 r / min for 3-7d. Mycelia liquid medium formula: glucose 20-30g, peptone 5-8g, 20% potato extract 200ml, (NH 4 ) 2 S0 4 3~5g, KH 2 P0 4 1~2g, MgS0 4 ·7H 2 0 1~2g, yeast powder 4~6g, add water to make up to 1L, pH 6.5~7.0, sterilize at 121℃ for 20min.

[0060] (4) Protoplast preparation MgS0 4 ·7H 20. Mannitol was dissolved in double-distille...

Embodiment 3

[0064] Embodiment 3 Preparation of solid-state fermentation strain

[0065] This embodiment 3 illustrates the method for preparing a heat-resistant protease-rich and cordycepin feed additive by using Cordyceps militaris HYCM12 as a strain solid-state fermentation method, including the following steps:

[0066] Step 1, strain activation: first, inoculate the original seed of the preserved Cordyceps militaris HYCM12 test tube strain on the activation medium of a petri dish, and incubate in an incubator at 20-26° C. in the dark for 5-7 days to obtain first-grade seeds.

[0067] Activated slant medium: 200 g of potatoes, washed and chopped, add 1000 mL of water and boil until boiling for 20 min, filter with gauze, add 20 g of glucose to the filtrate, 1 mL of inorganic salt solution, 15 g of agar and continue to boil, fully dissolve and then pack into test tubes , each test tube is about 4 mL, and the culture medium is sterilized at 121°C for 20 min. After the sterilization is comp...

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