Method for preparing feed by fermenting wheat bran with edible fungus mycelia

Abstract

The invention relates to a method for preparing feed by fermenting wheat bran with edible fungus mycelia, which comprises the following steps: (1) inoculating edible fungus mycelia into a plane culture medium to prepare an activated strain plate; (2) selecting a strain block from the activated strain flat plate, and culturing in a liquid culture medium to prepare a liquid seed solution; (3) inoculating the liquid seed solution into a solid wheat bran culture medium, fermenting in a dark place, and turning over during fermentation to prepare a wheat bran fermentation base material; and (4) drying the wheat bran fermentation base material to obtain the feed product. The preparation process is simple and low in cost, and the obtained fermented wheat bran feed is high in crude protein and amino acid content and high in digestibility.

Description

Technical Field

[0001] The invention relates to the technical field of microbial fermentation, and in particular to a method for preparing feed by fermenting wheat bran with edible fungus mycelium and a feed product prepared therefrom. Background Art

[0002] Wheat bran is a byproduct of wheat flour production and is an economical feed raw material. However, the crude protein content of wheat bran is 11.77%~17.02%, which is of poor quality and has a low essential amino acid content, which cannot fully meet the nutritional needs of livestock and poultry. Wheat bran contains a variety of anti-nutritional factors that will intercept nutrients and interfere with digestion and absorption. At the same time, wheat bran has strong water absorption and is prone to caking and mildew. These problems limit the scale of wheat bran use in feed.

[0003] Solid-state fermentation is fermentation without (or almost without) free water. Solid-state fermentation has a low moisture content, generally does not require wastewater treatment, and has less environmental pollution. Solid-state fermentation does not require a sterile environment. Low moisture means that fermentation can only be carried out by a limited number of microorganisms, mainly yeast and fungi. Therefore, most of the fermentation strains used in the study of solid-state fermentation of wheat bran are fungi. Different strains can promote different effects of solid-state fermentation of wheat bran. For example, solid-state fermentation of wheat bran with Mucor, Trichoderma, and Bacillus can produce some enzymes we need and increase the yield of enzymes. Solid-state fermentation of wheat bran with Bacillus can produce xylanase, solid-state fermentation of Aspergillus niger can release ferulic acid with antioxidant and anti-inflammatory capabilities from wheat bran, and solid-state fermentation of wheat bran with Mucor can increase the content of phytase.

[0004] In recent years, the application of edible fungus mycelium fermentation has received widespread attention. Compared with traditional microorganisms, edible fungus mycelium has a stronger ability to decompose and utilize hemicellulose, cellulose and lignin. It can produce a variety of enzymes such as cellulase, hemicellulase, esterase, laccase and amylase during the fermentation process, which has a great influence on the structure, biological activity and bioavailability of wheat bran. During the fermentation process, edible fungus mycelium can convert soluble sugars and other nutrients into other nutrients such as organic acids, which brings a unique flavor while increasing its nutritional quality. It can also produce a large amount of metabolites, which is beneficial to the intestinal health of farmed animals. Summary of the invention

[0005] The invention aims to provide a fermented wheat bran feed with simple preparation process, low cost, high crude protein and amino acid content and high digestibility and a preparation method thereof.

[0006] The present invention adopts the following technical solution: A method for preparing feed by fermenting wheat bran using edible fungus mycelium comprises the following steps: (1) Inoculating edible fungus mycelium into a flat culture medium to prepare an activated fungus plate; (2) Pick up bacterial blocks from the activated bacterial plate and culture them in liquid culture medium to prepare liquid seed solution; (3) inoculating the liquid seed liquid into a solid wheat bran culture medium for fermentation in the dark, turning the medium during the fermentation period, and preparing a wheat bran fermentation base material; (4) Drying the wheat bran fermentation base to obtain feed products.

[0007] Furthermore, the edible fungi include at least one of Pleurotus geesteranus, Pleurotus ostreatus, Pleurotus eryngii, Oyster mushroom, Pleurotus citrinum, Pleurotus citrinum or Enoki mushroom.

[0008] Furthermore, the edible fungus mycelium is preserved in a slant PDA culture medium, which includes: 5g potato powder, 20g glucose, 20g agar, 1.5g magnesium sulfate, 1.5g potassium dihydrogen phosphate, 30mg vitamin B1, 0.1g chloramphenicol, 1000mL deionized water, and natural pH.

[0009] Furthermore, the planar culture medium in step (1) comprises: 5 g potato powder, 20 g glucose, 20 g agar, 0.1 g chloramphenicol, 1000 mL deionized water, natural pH, and high temperature sterilization at 115-125°C for 20-30 min. In a sterile environment, pour it into a culture dish to prepare a planar culture medium.

[0010] Furthermore, in step (1), a bacterial block is picked up with an inoculation spatula and inoculated in the middle of a flat culture medium, and cultured upside down at 27° C. in a dark environment until the mycelium just covers the surface of the culture medium.

[0011] Furthermore, the liquid culture medium in step (2) comprises: 30 g malt extract, 3 g soy peptone, 1000 mL deionized water, natural pH, and high temperature sterilization at 115-125°C for 20-30 min.

[0012] Furthermore, in step (2), a 0.3*0.3 cm 2 The bacterial block was inoculated into a shake flask containing 100 mL of liquid culture medium and cultured in the dark at 100 rpm and 27°C for 7 days.

[0013] Furthermore, the solid wheat bran culture medium in step (3) is prepared by the following method: the wheat bran and water are mixed in a mass volume ratio of 1:1-2 and stirred evenly, and sterilized at 0.2 MPa, 115-125°C and high temperature and high pressure for 20-30 min, and cooled.

[0014] Preferably, the solid wheat bran culture medium in step (3) is prepared by the following method: the wheat bran and water are mixed in a mass volume ratio of 1:1.2 and stirred evenly, and sterilized at 0.2 MPa, 115-125°C and high temperature and high pressure for 20-30 min, and cooled.

[0015] Furthermore, in step (3), the liquid seed solution is inoculated into the solid wheat bran culture medium at a rate of 10% v / w, stirred evenly, the temperature is controlled at 25-30° C., and the fermentation time is 30-42 days; during the fermentation period, the material is turned over every 10 days.

[0016] Furthermore, in step (4), the fermented wheat bran base is dried to a moisture content of 8-12% and then crushed.

[0017] A feed product prepared by the method.

[0018] The beneficial effects of the present invention are: The present invention selects wheat bran, a by-product in wheat processing production, as a raw material, which is low in price and has unique advantages when used in feed.

[0019] The present invention adopts solid-state fermentation, which generally does not require wastewater treatment and has less environmental pollution. Solid-state fermentation does not require a sterile environment.

[0020] The present invention uses edible fungus mycelium with high nutritional value, which is rich in protein, fat, carbohydrate and various physiologically active substances, such as polyphenols, polysaccharides and the like. During the fermentation process, the edible fungus mycelium can produce various enzyme systems, such as cellulase, hemicellulase, esterase, laccase and amylase, etc., which greatly improve the structure, biological activity and bioavailability of wheat bran. It can also convert soluble sugar and other nutrients into other nutrients such as organic acid, which brings unique flavor and increases its nutritional quality, and can also produce a large amount of metabolites, which is beneficial to the intestinal health of raised animals.

[0021] The present invention proposes a method for preparing high-protein fermented wheat bran feed using edible fungus mycelium. The method has the characteristics of simple, low-cost raw materials and high yield, simple preparation process, high crude protein and essential amino acid content, etc. The crude protein content is increased from 16.17% of wheat bran to 31.16%, the essential amino acid content is increased from 2.003% to 5.862%, and the rumen degradation rate of dry matter is increased from 75.93% to 91.22%. The product quality is excellent, and it is applicable to multiple edible fungus mycelium strains, has no harsh requirements on the environment, and can be promoted and used on a large scale. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 The growth of mycelium of 15 edible fungi in solid wheat bran medium.

[0023] Among them, the first row shows the growth of 15 kinds of edible fungi mycelium fermentation for 7 days, from left to right: shiitake mushroom, black fungus, platinum special 6, white jade mushroom, seafood mushroom, shiitake mushroom, morel 26, king oyster mushroom, black skin chicken mushroom, black boletus, golden needle mushroom, cordyceps, elm yellow mushroom, oyster mushroom, pleurotus geesteranus; the second row shows the growth of 14 kinds of edible fungi mycelium fermentation for 14 days, from left to right: shiitake mushroom, black fungus, platinum special 6, white jade mushroom , seafood mushroom, oyster mushroom, morel 26, king oyster mushroom, black-skinned chicken mushroom, black boletus, golden needle mushroom, cordyceps, umbel mushroom, oyster mushroom, and Pleurotus geesteranus; the third row shows the growth of 14 edible fungi mycelium fermentation for 28 days, from left to right are shiitake mushroom, black fungus, platinum special no. 6, white jade mushroom, seafood mushroom, oyster mushroom, morel 26, king oyster mushroom, black-skinned chicken mushroom, black boletus, golden needle mushroom, cordyceps, umbel mushroom, oyster mushroom, and Pleurotus geesteranus.

[0024] Figure 2 The growth of different bacterial species in different liquid culture media.

[0025] Figure 3 The present invention is a schematic diagram of the process of preparing wheat bran feed fermented by edible fungus mycelium. DETAILED DESCRIPTION

[0026] The experimental methods described in the following examples are conventional methods unless otherwise specified; the reagents and materials described are commercially available unless otherwise specified.

[0027] The 15 edible fungi mycelium strains used in the following examples were donated by Beijing Academy of Agriculture and Forestry Sciences.

[0028] In the following embodiments, the crude protein content was determined by the national standard method GB / T6432-2018, the crude fat content was determined by the national standard method GB / T6433-2006, the neutral detergent fiber content was determined by the national standard method GB / T 20806-2006, the hemicellulose content was determined by the industry standard method NY / T 3494-2019, the ash content was determined by the ignition method, and the amino acid content was determined by referring to GB / T18246-2019.

[0029] The present invention is further described in detail below in conjunction with specific embodiments. The given examples are only for illustrating the present invention, but not for limiting the scope of the present invention.

[0030] Example 1 Growth of different strains on solid wheat bran medium Take 15 kinds of edible fungi mycelium strains under the same storage conditions and inoculate them into a flat culture medium (5g potato extract powder, 20g glucose, 20g agar, 0.1g chloramphenicol, 1000mL deionized water, natural pH), in a dark environment at 27℃, invert and culture until the mycelium just covers the surface of the culture medium to make an activated culture plate. On the activated culture plate, pick up a 0.3*0.3cm 2 The fungus blocks were inoculated into a test tube containing solid wheat bran medium (wheat bran and water were mixed in a mass volume ratio of 1:1.5 and stirred evenly, sterilized at 0.2 MPa, 115-125℃ high temperature and high pressure for 20-30min, and cooled) at 27℃ in a dark environment to verify the growth activity of 15 edible fungi mycelium in solid wheat bran medium ( Figure 1 ).

[0031] By measuring the growth depth of mycelium in the fermentation tube and measuring the growth rate of mycelium, it was found that the mycelium of Pleurotus ostreatus, Pleurotus citrinopileatus, Pleurotus geesteranus, Pleurotus eryngii, Pleurotus velutipes, and Pleurotus eryngii grew faster and denser, as shown in Table 1. This shows that solid wheat bran medium is suitable for the growth of some strains.

[0032] The 28-day fermentation products of the six strains with better growth were dried at 65℃, and the crude protein content of different fermentation products was determined. The results are shown in Table 2. It was found that the crude protein content of the fermentation product of Pleurotus ostreatus was the highest, followed by the fermentation products of Pleurotus citrinopileatus, Pleurotus geesteranus and Pleurotus velutipes. This shows that solid wheat bran culture medium can effectively promote the secretion and production of mycelium-related protease system of Pleurotus ostreatus, Pleurotus citrinopileatus, Pleurotus geesteranus and Pleurotus velutipes.

[0033] Table 1 Statistics of growth rates of different strains in solid wheat bran medium .

[0034] Table 2 Determination of crude protein content of fermentation products of different strains .

[0035] Example 2 Effect of liquid culture medium on liquid seed solution concentration According to the method of Example 1, the spawn of Pleurotus ostreatus, Pleurotus citrinopileatus, Pleurotus geesteranus and Pleurotus eryngii were prepared into activated spawn plates, and 0.3*0.3 cm 2 The bacterial blocks were inoculated into shake flasks containing 100 mL of liquid medium 1, liquid medium 2, liquid medium 3, and liquid medium 4, respectively, and cultured at 27°C, 100 rpm, in the dark for 7 days (see Figure 2). The mycelium in the liquid seed solution was filtered with filter paper, dried at 65°C, weighed, and the concentration of the liquid seed solution was determined, calculated based on the dry weight of the mycelium. The results are shown in Table 3. It was found that the seed solution concentrations of the four strains in liquid culture medium 3 were significantly higher than those in other liquid culture media, indicating that the four strains had higher activity in liquid seed solution 3.

[0036] Liquid culture medium 1: 2 g soluble starch, 2 g maltose, 0.5 g magnesium sulfate, 0.25 g potassium dihydrogen phosphate, 1 g sodium nitrate, 0.5 g ammonium sulfate, 0.5 g calcium chloride, 1000 mL deionized water, natural pH.

[0037] Liquid medium 2: 20 g glucose, 5 g yeast extract, 1 g potassium dihydrogen phosphate, 0.5 g magnesium sulfate, 1000 mL deionized water, natural pH.

[0038] Liquid culture medium 3: 30 g malt extract, 3 g soy peptone, 1000 mL deionized water, natural pH.

[0039] Liquid culture medium 4: 80 g corn syrup powder, 0.8 g yeast extract, 0.08 g soy peptone, 0.8 g malt extract, 1000 mL deionized water, natural pH.

[0040] Table 3 Effect of different liquid culture media on liquid seed solution concentration .

[0041] Example 3 Effect of the material-water ratio of solid wheat bran culture medium on the crude protein content of fermentation products On the basis of the preferred liquid culture medium in Example 2, liquid seed solutions of Pleurotus ostreatus, Pleurotus citrinopileatus, Pleurotus geesteranus and Pleurotus eryngii were prepared respectively, and the liquid seed solutions were inoculated at a 10% v / w inoculation rate and inoculated into solid wheat bran culture media with different material-water ratios (wheat bran and water at a mass volume ratio of 1:0.7-2.2), and cultured at 27°C in a dark environment for 35 days. The crude protein content of the fermentation products was detected. The results are shown in Table 4.

[0042] Table 4 Effect of different feed-water ratios on crude protein content of fermentation products .

[0043] It can be seen from Table 4 that when other fermentation conditions are the same, the crude protein content of the fermentation products of solid wheat bran culture medium with different feed-water ratios is significantly different. When the feed-water ratio of solid wheat bran culture medium is 1:1.2, the crude protein content of the fermentation products of Pleurotus ostreatus, Pleurotus geesteranus, Enoki mushroom and Pleurotus citrinopileatus is the highest, that is, 1:1.2 is the optimal feed-water ratio of solid wheat bran culture medium.

[0044] Example 4 Preparation method of fermented wheat bran feed Combined with the experimental results of Examples 1 to 3, a method for preparing feed by fermenting wheat bran with edible fungus mycelium is obtained. The method flow is as follows: Figure 3 shown.

[0045] (1) Inoculate the mycelium of edible fungi into a flat culture medium to prepare an activated culture plate.

[0046] Use an inoculation spatula to pick up 0.5*0.5cm of the PDA culture medium on the slant 2 The oyster mushroom block is inoculated in the middle of the flat culture medium and cultured upside down at 27℃ in a dark environment until the mycelium just covers the surface of the culture medium. This is the culture activation plate.

[0047] The slant PDA culture medium includes: 5 g potato powder, 20 g glucose, 20 g agar, 1.5 g magnesium sulfate, 1.5 g potassium dihydrogen phosphate, 30 mg vitamin B1, 0.1 g chloramphenicol, 1000 mL deionized water, and natural pH.

[0048] The planar culture medium includes: 5 g potato extract powder, 20 g glucose, 20 g agar, 0.1 g chloramphenicol, 1000 mL deionized water, and natural pH.

[0049] Among them, the mycelium of edible mushrooms includes oyster mushroom, citronella mushroom, enoki mushroom and golden needle mushroom.

[0050] (2) Pick up bacterial blocks from the activated bacterial plate and culture them in liquid culture medium to make liquid seed solution.

[0051] Use an inoculation shovel to pick up a 0.3*0.3cm 2 The bacterial block was inoculated into a shaking flask containing 100 mL of liquid culture medium, and cultured in the dark at 100 rpm and 27°C for 7 days to obtain liquid seed solution.

[0052] The liquid culture medium includes: 30 g malt extract, 3 g soy peptone, 1000 mL deionized water, and natural pH.

[0053] (3) The liquid seed solution is inoculated into a solid wheat bran culture medium for fermentation in the dark, and the medium is turned over during the fermentation period to prepare a wheat bran fermentation base material.

[0054] Inoculate the liquid seed solution into the solid wheat bran culture medium at a 10% (v / w) inoculation rate, stir evenly, control the temperature at 27°C, and ferment for 35 days; turn the material over every 10 days during the fermentation period.

[0055] The solid wheat bran culture medium is prepared by the following method: the wheat bran and water are mixed in a mass volume ratio of 1:1.2 and stirred evenly, sterilized at 0.2 MPa, 115-125°C and high temperature and high pressure for 20-30 minutes, and cooled.

[0056] (4) Dry the fermented wheat bran base material to a moisture content of 8-12%, and then grind it to obtain a feed product.

[0057] Effect Example 1 Comparison of nutritional indicators between fermented wheat bran feed and wheat bran In order to verify that the fermented wheat bran feed prepared by the method provided by the present invention has obvious effects, the crude protein, crude fat, neutral detergent fiber, hemicellulose and crude ash of the prepared fermented wheat bran feed were measured according to the method of Example 4. The results are shown in Table 5.

[0058] Table 5 Statistical table of nutritional components of fermentation products of different strains .

[0059] The results show that the crude protein and crude fat contents of the wheat bran feed fermented with edible fungi mycelium prepared by the method of the present invention are significantly increased compared with wheat bran, and the contents of neutral detergent fiber and hemicellulose are significantly reduced. The crude protein content is increased from 16.17% to 26.46%~31.16%, that is, the crude protein content is increased by 1.64~1.93 times compared with that before fermentation. The crude protein content of the wheat bran feed fermented with edible fungi mycelium is greater than 25%, the crude fiber content is less than 12%, and the crude ash content is less than 10%, that is, the nutritional components can meet the nutritional requirements of the concentrate supplements for calves, growing cattle, beef cattle, and dairy cows at various stages listed in the standard Q / SGS03-2018 "Concentrate Supplements for Ruminants".

[0060] Effect Example 2 Comparison of amino acid content between fermented wheat bran feed and wheat bran In order to verify that the edible fungus mycelium fermented wheat bran feed prepared by the method provided by the present invention is a high-quality protein feed, the contents of 17 kinds of amino acids in the prepared fermented wheat bran feed were determined according to the method of Example 4. The results are shown in Table 6.

[0061] Table 6 Statistics of amino acid mass fractions in different experimental groups .

[0062] The results show that the total amino acid content and essential amino acid content of the wheat bran feed fermented with edible fungi mycelium are significantly improved compared with wheat bran. The total amino acid content is increased from 5.312% to 11.733~17.704%, and the essential amino acid content is increased from 2.003% to 4.380~5.862%. Analysis shows that the reason is that the present invention adopts high-protein edible fungi mycelium strain fermentation to convert the protein in wheat bran into high-quality edible fungi mycelium microbial protein.

[0063] Effect Example 3 Comparison of rumen fluid degradation rate of fermented wheat bran feed and wheat bran In order to further verify that the edible fungus mycelium fermented wheat bran feed prepared by the method provided by the present invention has good practical effect, according to the method of Example 4, an in vitro degradation experiment of the fermented wheat bran feed in cattle rumen fluid was carried out.

[0064] The specific experimental method is as follows: 3 healthy Holstein cows with similar lactation age (137±37) days, body weight (550±50) kg, milk production (25.3±2.8) kg and permanent fistula were selected. Rumen fluid was collected through the rumen fistula 1 hour before morning feeding on the day of the experiment and placed in a preheated (39℃) and CO2-filled thermos bottle and quickly brought back to the laboratory. After filtering through 4 layers of medical gauze (while continuously passing CO2), it was immediately used in the experiment.

[0065] The results of the nutrient degradation rate of the fermented wheat bran feed before and after degradation in vitro in the rumen fluid of cattle are shown in Table 7.

[0066] Table 7 Statistical table of the degradation rate of various nutrients in fermented wheat bran feed .

[0067] The results show that the dry matter and crude protein rumen degradation rate of the wheat bran feed fermented by edible fungi mycelium prepared by the present invention are significantly higher than those of wheat bran, which proves that the crude protein content of the wheat bran feed fermented by edible fungi mycelium prepared by the present invention is increased by 1.64 to 1.93 times compared with wheat bran, and it is easy to digest and can be used as a high-quality concentrate supplement for ruminants.

Claims

1. A method for preparing feed by fermenting wheat bran using edible fungus mycelium, characterized in that: It includes the following steps: (1) Inoculating edible fungus mycelium into a flat culture medium to prepare an activated fungus plate; (2) Pick up bacterial blocks from the activated bacterial plate and culture them in liquid culture medium to prepare liquid seed solution; (3) inoculating the liquid seed liquid into a solid wheat bran culture medium at a ratio of 10% (v / w) for light-protected fermentation, turning the material during the fermentation period, and preparing a wheat bran fermentation base material; the solid wheat bran culture medium is obtained by mixing wheat bran and water in a mass volume ratio of 1:1-2, sterilizing the mixture at high temperature and high pressure, and cooling the mixture; the liquid seed culture medium contains malt extract and soy peptone in a mass ratio of 10:1; (4) Drying the wheat bran fermentation base to obtain feed products.

2. The method according to claim 1, characterized in that The edible fungi include at least one of Pleurotus geesteranus, Pleurotus ostreatus, Pleurotus eryngii, Shiitake mushroom, Pleurotus citrinum, Pleurotus eryngii, Lentinula edodes, Black fungus, Platinum edodes, White jade mushroom, Seafood mushroom, Morel, Black-skinned Chicken mushroom, Black Boletus, Cordyceps sinensis, and Pleurotus geesteranus.

3. The method according to claim 1, characterized in that The planar culture medium in step (1) comprises: 5 g potato extract powder, 20 g glucose, 20 g agar, 0.1 g chloramphenicol, 1000 mL deionized water, natural pH, 0.2 MPa, and after high temperature and high pressure sterilization at 115-125°C, pour it into a culture dish in a sterile environment to prepare a planar culture medium.

4. The method according to claim 1, characterized in that: In step (1), a bacterial block is picked up with an inoculation spatula and inoculated in the middle of a flat culture medium, and cultured upside down at 27°C in a dark environment until the mycelium just covers the surface of the culture medium.

5. The method according to claim 1, characterized in that: The liquid culture medium in step (2) includes: 30 g malt extract, 3 g soy peptone, 1000 mL deionized water, natural pH, 0.2 MPa, and high temperature and high pressure sterilization at 115-125°C.

6. The method according to claim 1, characterized in that In step (2), use an inoculation shovel to pick up a 0.3*0.3 cm 2 The bacterial block was inoculated into a shake flask containing 100 mL of liquid culture medium and cultured in the dark at 100 rpm and 27°C for 7 days.

7. The method according to claim 1, characterized in that The solid wheat bran culture medium in step (3) is prepared by the following method: the wheat bran and water are mixed in proportion and stirred evenly, sterilized at 0.2 MPa, 115-125°C for 20-30 min, and cooled; the wheat bran and water are mixed in a mass volume ratio of 1:0.7, 1:1, 1:1.2, 1:1.5, 1:1.7, 1:2, 1:2.

2.

8. The method according to claim 1, characterized in that: In step (3), the liquid seed solution is inoculated into the solid wheat bran culture medium at a rate of 10% v / w, stirred evenly, the temperature is controlled at 25-30° C., and the fermentation time is 30-42 days; during the fermentation period, the material is turned over every 10 days.

9. The method according to claim 1, characterized in that: In step (4), the fermented wheat bran base is dried to a moisture content of 8-12% and then crushed.

10. The feed product obtained by the method for preparing feed by fermenting wheat bran with edible fungus mycelium according to claims 1-9.

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