Shiitake mushroom mycelium-based chicken feed and its preparation method

Abstract

This invention discloses a shiitake mushroom spawn-based chicken feed and its preparation method based on a compound microbial agent. The invention provides a compound microbial agent for preparing chicken feed, composed of *Rhizopus oryzae*, *Lactobacillus acidophilus*, and *Candida tropicalis*. The invention involves pretreating shiitake mushroom spawn through physical and chemical steps, then mixing it with wheat bran and soybean meal to obtain fermentation raw materials. After inoculation with the compound microbial agent, gradient fermentation is performed to obtain the shiitake mushroom spawn-based chicken feed. The method utilizes the ligninase secreted by *Rhizopus oryzae* to degrade lignin, the lactic acid produced by *Lactobacillus acidophilus* to inhibit other bacteria, and the ability of *Candida tropicalis* to synthesize protein from the degradation products. These components exert their respective effects at different stages of gradient fermentation, degrading the lignin and cellulose in the shiitake mushroom spawn raw materials into absorbable nutrients or using the degradation products to synthesize protein, effectively increasing the protein content of the product and achieving high-efficiency conversion and high added-value utilization of the shiitake mushroom spawn, meeting the nutritional needs of chickens at different growth stages.

Description

Technical Field

[0001] This invention relates to a chicken feed, and more particularly to a chicken feed based on a compound microbial agent made from shiitake mushroom spawn and its preparation method, belonging to the field of chicken feed preparation. Background Technology

[0002] As the main waste product after shiitake mushroom cultivation, the annual discharge of shiitake mushroom substrate exceeds 10 million tons. It contains cellulose (18%-22%), hemicellulose (12%-15%), lignin (6%-9%) and residual mycelial protein (7%-11%). However, due to its dense fiber structure, high content of anti-nutritional factors (phytic acid 1.3%-1.9%, tannin 0.4%-0.6%), and a certain amount of lignin-protein complex, conventional processing technology is difficult to efficiently convert it.

[0003] While existing processing technologies can degrade some of the fiber in shiitake mushroom substrate, the lignin degradation rate is less than 20%, and more than 3% glucose needs to be added as an exogenous carbon source, resulting in high costs. In addition, although the blasting method can destroy the fiber structure, the energy consumption is 60%-70% higher than that of room temperature treatment and the loss rate of amino acids in the substrate exceeds 15%. Furthermore, most processes have a fermentation cycle of 6-8 days and do not adjust the feed formula according to the nutritional needs of chickens at different growth stages, resulting in limited product applicability. Summary of the Invention

[0004] One objective of this invention is to provide a compound microbial inoculant for preparing shiitake mushroom bran chicken feed; The second objective of this invention is to provide a method for preparing a compound microbial agent; The third objective of this invention is to provide a method for preparing shiitake mushroom bran chicken feed based on compound microbial agents.

[0005] The above-mentioned objective of this invention is achieved through the following technical solution: To address the problems of low lignin degradation rate, high dependence on exogenous carbon sources, high energy consumption, and limited product applicability in existing mushroom-fed chicken feed, this invention first provides a compound microbial agent for preparing mushroom-fed chicken feed, composed of Rhizopus oryzae agent, Lactobacillus acidophilus agent, and Candida tropicalis agent.

[0006] In a preferred embodiment of the present invention, the compound microbial agent, calculated by the number of live bacteria, has a ratio of Rhizopus oryzae agent, Lactobacillus acidophilus agent, and Candida tropicalis agent of (2-5):(1-3):(1-3); preferably, the ratio of Rhizopus oryzae agent, Lactobacillus acidophilus agent, and Candida tropicalis agent is 3:2:2; Among them, the aforementioned Rhizopus oryzae ( 米根霉The preferred strain was strain CGMCC No. 20247 (CN 112011467A), purchased from the China General Microbiological Culture Collection Center. It exhibits ligninase activity (laccase activity ≥8 U / g, manganese peroxidase activity ≥12 U / g, GB / T30738-2014), with an optimal growth temperature of 26-30℃ and pH of 4.5-5.5. The aforementioned Lactobacillus acidophilus ( 嗜酸乳杆菌 The preferred strain is the strain with accession number CGMCC1.1854, purchased from the China General Microbiological Culture Collection Center. It produces ≥18 g / L of lactic acid in 24 hours, has an inhibition zone diameter of ≥14 mm against Salmonella, and has an optimal growth temperature of 36-39℃ and a pH of 3.8-5.0. The aforementioned Candida tropicalis ( 热带假丝酵母 The preferred strain is strain CGMCC No. 21583 (CN 113088459A), purchased from the China General Microbiological Culture Collection Center. It can utilize lignin degradation products as a carbon source, with an optimal growth temperature of 28-32℃ and pH of 4.2-6.0.

[0007] This invention combines three strains—Rhizopus oryzae producing ligninase to degrade lignin, Lactobacillus acidophilus producing lactic acid to inhibit other bacteria, and Candida tropicalis using the degradation products to synthesize proteins—to achieve synergistic effects. These strains play a key role in different stages of the gradient fermentation of chicken feed prepared from shiitake mushroom substrate, fully degrading the lignin and cellulose in the shiitake mushroom substrate into components that chickens can absorb and utilize, and using the degradation products to synthesize proteins, effectively increasing the protein content of chicken feed products.

[0008] Another aspect of the present invention provides a method for preparing a compound microbial agent, comprising: preparing Rhizopus oryzae seed liquid, Lactobacillus acidophilus seed liquid and Candida tropicalis seed liquid; mixing the various seed liquids to obtain a mixture, and adding a protectant to the mixture; preferably, the protectant is maltodextrin; the added protectant accounts for 1.5% of the total mass of the mixture.

[0009] In a preferred embodiment of the present invention, *Rhizopus oryzae* spores, *Lactobacillus acidophilus* colonies, and *Candida tropicalis* colonies obtained by slant agar activation are mixed in a viable cell ratio of 3:2:2 (*Rhizopus oryzae*: *Lactobacillus acidophilus*: *Candida tropicalis*) to prepare a seed liquid for a compound microbial agent. 1.0-2.0% (mass fraction) of maltodextrin is added as a protectant, and the mixture is activated at 24°C for 1.5 hours to produce the compound agent, with a total viable cell count ≥1.2 × 10⁻⁶. 10 CFU / g, shelf life ≥45 days when refrigerated at 2-8℃, live bacteria count decrease rate ≤8%.

[0010] Another aspect of the present invention provides a method for preparing chicken feed, comprising: inoculating a mixture of fermentation raw materials for chicken feed with the aforementioned compound microbial agent for fermentation, thereby obtaining the feed; wherein the mixture of fermentation raw materials for chicken feed mainly includes shiitake mushroom substrate, wheat bran, and soybean meal; the basic ratio by mass is shiitake mushroom substrate: wheat bran: soybean meal = (6-7):(1.5-2.5):(1-2); preferably, the ratio of shiitake mushroom substrate: wheat bran: soybean meal = 6.5:2:1.5; for laying hens, the proportion of soybean meal can be reduced to 1%, and 0.5% limestone powder (calcium content ≥38%) can be added; for broiler fattening, the proportion of wheat bran can be increased to 2.5%, and the proportion of soybean meal can be reduced to 1%.

[0011] In a preferred embodiment of the present invention, the shiitake mushroom substrate in the fermentation raw material mixture undergoes segmented pretreatment, including physical pretreatment and chemical pretreatment; wherein, the physical pretreatment method includes: crushing the shiitake mushroom substrate to a particle size of 3-5 mm using a pulverizer, and then ultra-fine pulverizing it to a particle size of 0.3-0.5 mm using an air jet mill (particle size qualification rate ≥96%), thereby destroying the lignin-cellulose structure through ultra-fine pulverization and improving the efficiency of subsequent enzymatic hydrolysis; The chemical pretreatment method includes: crushing shiitake mushroom spawn to a particle size of 0.3-0.5 mm by mass fraction, adding 0.2% oxalic acid solution to adjust the pH to 4.8-5.2, soaking at room temperature (22-26℃) for 20 min, and using ultrasonic-assisted treatment during soaking; preferably, the parameters of ultrasonic-assisted treatment are: power 300W, frequency 28kHz, treatment once every 5 min, each treatment lasting 30 s, to promote the combination of oxalic acid and lignin and destroy the lignin structure.

[0012] In a preferred embodiment of the present invention, the preparation method of the fermentation raw material mixture is as follows: pretreated shiitake mushroom spawn is mixed with wheat bran and soybean meal, and deionized water is added to adjust the moisture content to 48%-52% to obtain a mixture; potassium dihydrogen phosphate is added to the mixture, stirred evenly, and allowed to stand for 25 minutes, stirring once every 10 minutes during the standing period to ensure the uniformity of the material; preferably, the added potassium dihydrogen phosphate accounts for 0.1% of the mass percentage of the mixture.

[0013] In a preferred embodiment of the present invention, a pretreated material is obtained by inoculating a mixture of fermentation raw materials with a compound microbial agent and stirring it evenly. The compound microbial agent is then added to the pretreated material at an inoculation rate of 1.5-2.0% (mass fraction) and stirred evenly before fermentation using a gradient fermentation method. The specific parameters of the gradient fermentation are as follows: The first stage (0-16h, lignin degradation period): temperature 28-30℃, ventilation rate 0.4 L / (kg·h), dissolved oxygen level controlled at 8%-12%, to promote ligninase production by Rhizopus oryzae and degrade lignin; The second stage (16-32h, fiber degradation and acid production period): temperature 34-36℃, ventilation 0.6 L / (kg·h), dissolved oxygen controlled at 15%-18%, Rhizopus oryzae produces cellulase (filter paper enzyme activity ≥16 U / g) to degrade the fiber, and Lactobacillus acidophilus produces lactic acid to inhibit other bacteria. The third stage (32-48h, protein synthesis period): temperature 30-32℃, ventilation 0.7 L / (kg·h), dissolved oxygen controlled at 12%-15%, tropical Candida uses degradation products to synthesize proteins. Throughout the fermentation process, the material status was checked every 8 hours. When the moisture content was below 46%, deionized water was added, with the amount added calculated as "0.8% water for every 1% decrease in moisture content". When the pH was below 4.6, 0.08% sodium carbonate solution was added to adjust the pH to ensure it was maintained between 4.8 and 5.2.

[0014] The fermentation endpoint is determined as follows: fermentation ends when the following conditions are met: ① Lignin degradation rate ≥35%; ② Crude protein content ≥19% (laying hen during egg production) or ≥21% (broiler during fattening); ③ Phytic acid content ≤0.25%; ④ Fermentation time 40-50 hours.

[0015] In a preferred embodiment of the present invention, the material status is checked every 8 hours during the fermentation process. When the moisture content is lower than 46%, 0.8% water is added for every 1% decrease in moisture content; and the pH is maintained at 4.8-5.2.

[0016] In a preferred embodiment of the present invention, after fermentation, the fermentation product is processed by vacuum freeze-drying or low-temperature hot air drying. Preferably, the parameters for vacuum freeze-drying are: temperature -40℃ to -35℃, vacuum degree ≤10Pa, drying time 8h, moisture content ≤11% after drying, and heat-sensitive nutrient retention rate: vitamin E ≥90%, B vitamins ≥85%. The parameters for low-temperature hot air drying are: temperature 55-58℃, air velocity 1.2m / s, drying time 2.5h, and moisture content ≤11.5% after drying. After the above drying treatment, the fermentation product is pulverized using a grading pulverizer. For laying hens, the feed is pulverized to a particle size of 1.2-1.5mm; for broiler fattening feed, the feed is pulverized to a particle size of 0.6-0.8mm, with a particle size qualification rate ≥97%.

[0017] In a preferred embodiment of the present invention, the fermentation product is mixed with different additives to prepare laying hen feed or broiler feed respectively; wherein, the preparation method of the laying hen feed is as follows: the fermentation product is dried and pulverized, and then 0.25% vitamin premix (vitamin A 8000 IU / kg, vitamin D3 2000 IU / kg, vitamin E 30 mg / kg, vitamin K3 3 mg / kg), 0.4% mineral premix (calcium 12 g / kg, phosphorus 4 g / kg, zinc 90 mg / kg, selenium 0.4 mg / kg) and 0.1% methionine are added to obtain the laying hen feed.

[0018] The preparation method of the broiler fattening feed is as follows: the fermentation product is dried and pulverized, and then added to 0.2% vitamin premix (vitamin A 6000 IU / kg, vitamin D3 1500 IU / kg, vitamin E 25 mg / kg), 0.35% mineral premix (calcium 8 g / kg, phosphorus 3.5 g / kg, iron 70 mg / kg, manganese 60 mg / kg) and 0.15% lysine to obtain the broiler fattening feed.

[0019] The mixing process preferably employs a three-dimensional mixer, with a mixing time of 20 minutes and a mixing uniformity variation coefficient ≤ 4%.

[0020] This invention first screens strains with lignin degradation capabilities to obtain a compound microbial agent. After physical and chemical pretreatment, shiitake mushroom substrate is mixed with wheat bran and soybean meal to obtain pretreated fermentation raw materials. The compound microbial agent is then inoculated into the pretreated fermentation raw materials for gradient fermentation to obtain shiitake mushroom substrate chicken feed. The compound microbial agent of this invention utilizes the lignin-degrading enzymes of Rhizopus oryzae, the lactic acid-producing ability of Lactobacillus acidophilus to inhibit other bacteria, and the ability of Candida tropicalis to synthesize proteins using the degradation products. These components play a crucial role in different stages of the gradient fermentation of the shiitake mushroom substrate chicken feed, fully degrading the lignin and cellulose in the shiitake mushroom substrate into components that chickens can absorb and utilize, and using the degradation products to synthesize proteins, effectively increasing the protein content of the chicken feed product. By utilizing the ligninase activity of Rhizopus oryzae and ultrasound-assisted oxalic acid pretreatment, the lignin degradation rate is effectively improved, eliminating the need for exogenous carbon sources and reducing costs by 15%-20%. Segmented pretreatment of fermentation raw materials eliminates the need for high temperatures, resulting in an amino acid loss rate of ≤5%. Gradient temperature-controlled fermentation keeps the fermentation cycle within 40-50 hours, shortening it by 40%-50% compared to existing processes. The resulting fermentation products can be formulated for both laying hens and broilers, increasing egg production by 8%-10% and broiler weight gain by 12%-15%. The product exhibits high safety: heavy metals (lead ≤0.08mg / kg, cadmium ≤0.05mg / kg) and mycotoxins (aflatoxin B1 ≤5μg / kg) are all below the GB13078-2017 standard, and there are no antibiotic residues. Attached Figure Description

[0021] Figure 1 This is a flowchart of the preparation method of shiitake mushroom bran chicken feed based on compound microbial inoculants according to the present invention. Detailed Implementation

[0022] The present invention will be further described below with reference to specific embodiments, and the advantages and features of the present invention will become clearer as a result. However, these embodiments are merely exemplary and do not constitute any limitation on the scope of the present invention. Those skilled in the art should understand that modifications or substitutions to the details and form of the present invention can be made without departing from the spirit and scope of the invention, but all such modifications and substitutions fall within the protection scope of the present invention.

[0023] Example 1: Preparation of Compound Microbial Agent Rhizopus oryzae ( Rhizopus oryzae Purchased from China General Microbiological Culture Collection Center, it has ligninase activity (laccase activity ≥8 U / g, manganese peroxidase activity ≥12 U / g, GB / T 30738-2014), with an optimal growth temperature of 26-30℃ and pH of 4.5-5.5; Lactobacillus acidophilus ( Lactobacillus acidophilus(Purchased from China General Microbiological Culture Collection Center, producing ≥18 g / L of lactic acid in 24 hours, with an inhibition zone diameter of ≥14 mm against Salmonella, and an optimal growth temperature of 36-39℃ and pH of 3.8-5.0.) Tropical Candida ( Candida tropicalis (Purchased from China General Microbiological Culture Collection Center, with cell protein content ≥52%, can utilize lignin degradation products as a carbon source, optimal growth temperature 28-32℃, pH 4.2-6.0.)

[0024] 1. Inclined plane activation: Rhizopus oryzae: Inoculate onto potato-dextrose-agar medium (potato 200g / L, glucose 20g / L, agar 18g / L, KH2PO4 3g / L, pH 5.0, sterilize at 121℃ for 20min), and incubate at 28℃ for 40h until the slant is covered with gray spores; Lactobacillus acidophilus: Inoculate into MRS medium (10 g / L peptone, 8 g / L beef extract, 4 g / L yeast extract, 20 g / L glucose, 5 g / L sodium acetate, 2 g / L diammonium citrate, pH 6.0, sterilize at 121℃ for 20 min), and anaerobic culture at 37℃ (anaerobic gas N2:CO2=90:10) for 22 h, until the slant is covered with milky white colonies; Tropical Candida: Inoculate onto malt extract-yeast extract medium (malt extract 10°P, yeast extract 3g / L, agar 18g / L, pH 5.0, sterilized at 121℃ for 20min), and incubate at 30℃ for 32h until the slant is covered with milky white colonies.

[0025] 2. Seed culture medium: Rhizopus oryzae: Spores were scraped and inoculated into liquid culture medium (same as slant, without agar), and cultured at 28℃ and 160 rpm for 32 h. The viable count was ≥6 × 10⁻⁶. 8 CFU / mL; Lactobacillus acidophilus: Scrape colonies and inoculate into liquid MRS medium, incubate at 37℃ and 140 rpm in an anaerobic shaker for 16 h, viable count ≥ 1.2 × 10⁻⁶ 9 CFU / mL; Candida tropicalis: Scrape colonies and inoculate into liquid malt extract-yeast extract medium, incubate at 30℃ and 180 rpm for 20 h, viable count ≥ 9 × 10⁻⁶ 8 CFU / mL.

[0026] 3. Preparation of compound microbial inoculant: Seed culture was prepared by mixing *Rhizopus oryzae*, *Lactobacillus acidophilus*, and *Candida tropicalis* in a ratio of 3:2:2 (viable cell count). 1.5% (mass fraction) of maltodextrin (GB / T 29877-2013) was added as a protectant. The mixture was then activated at 24℃ for 1.5 hours to produce a compound inoculant with a total viable cell count ≥1.2 × 10⁻⁶. 10 CFU / g, shelf life ≥45 days when refrigerated at 2-8℃, live bacteria count decrease rate ≤8%.

[0027] Example 2 Preparation of compound microbial inoculant In the preparation of the compound microbial agent, the seed liquid was mixed according to the ratio of Rhizopus oryzae: Lactobacillus acidophilus: Candida tropicalis = 2:1:1 (viable cell ratio), and all other steps were exactly the same as in Example 1, to prepare the compound agent with a total viable cell count ≥ 1.2 × 10⁻⁶. 10 CFU / g, shelf life ≥45 days when refrigerated at 2-8℃, live bacteria count decrease rate ≤8%.

[0028] Example 3 Preparation of compound microbial inoculant In preparing the compound microbial agent, the seed culture was mixed according to the ratio of Rhizopus oryzae: Lactobacillus acidophilus: Candida tropicalis = 5:3:3 (viable cell ratio), and all other steps were exactly the same as in Example 1, to prepare the compound agent with a total viable cell count ≥ 1.2 × 10⁻⁶. 10 CFU / g, shelf life ≥45 days when refrigerated at 2-8℃, live bacteria count decrease rate ≤8%.

[0029] Example 4 Preparation of compound microbial inoculant In preparing the compound microbial agent, the seed culture was mixed according to the ratio of Rhizopus oryzae: Lactobacillus acidophilus: Candida tropicalis = 5:1:1 (viable cell ratio), and all other steps were exactly the same as in Example 1, to prepare the compound agent with a total viable cell count ≥ 1.2 × 10⁻⁶. 10 CFU / g, shelf life ≥45 days when refrigerated at 2-8℃, live bacteria count decrease rate ≤8%.

[0030] Example 5 Preparation of compound microbial inoculant In the preparation of the compound microbial agent, the seed liquid was mixed according to the ratio of Rhizopus oryzae: Lactobacillus acidophilus: Candida tropicalis = 2:1:1 (viable cell ratio), and all other steps were exactly the same as in Example 1, to prepare the compound agent with a total viable cell count ≥ 1.2 × 10⁻⁶. 10 CFU / g, shelf life ≥45 days when refrigerated at 2-8℃, live bacteria count decrease rate ≤8%.

[0031] Example 6: Preparation of Shiitake Mushroom Bran Chicken Feed Pretreatment of raw materials: Take 65 kg of shiitake mushroom spawn (60% moisture content), ultrafine grind it to 0.4 mm, add 1.3 kg of 0.2% oxalic acid solution to adjust the pH to 4.8-5.2, and soak it with ultrasonic assistance for 20 min (the parameters of ultrasonic assistance are: power 300W, frequency 28kHz, treatment once every 5 min, each treatment for 30 s). Then add 20 kg of wheat bran, 10 kg of soybean meal and 5 kg of stone powder, adjust the moisture content to 50%, add 0.1 kg of potassium dihydrogen phosphate, and let it stand for 25 min to obtain the pretreated raw materials. Fermentation: The pretreated raw materials were inoculated with the compound microbial agent prepared in Example 1 at an inoculation rate of 1.8% (mass fraction), and then fermented. (Fermentation can be carried out using dynamic fermentation equipment, such as a 100L volume, 304 stainless steel structure, equipped with a gradient temperature control system and a ventilation system (0.1-1.5 L / (kg·h)). The gradient temperature control fermentation parameters are as follows: 0-16h, fermentation temperature 29℃, ventilation 0.4 L / (kg·h), dissolved oxygen controlled at 8%; 16-32h, fermentation temperature 35℃, ventilation 0.6 L / (kg·h), dissolved oxygen controlled at 15%; 32-46h, 31℃, ventilation 0.7 L / (kg·h), dissolved oxygen controlled at 12%. During fermentation, the material status was checked every 8 hours. When the moisture content was below 46%, deionized water was added, calculated at a rate of 0.8% for every 1% decrease in moisture content. The pH was maintained below 4.6. When adding sodium carbonate solution (GB 1886.1-2015), adjust the pH to ensure it is maintained between 4.8 and 5.2.

[0032] After fermentation, the fermentation products were tested, and the lignin degradation rate reached 39%, and the crude protein content was 24.5%.

[0033] Example 7 Preparation of Shiitake Mushroom Bran Chicken Feed Pretreatment of raw materials: Take 65 kg of shiitake mushroom spawn (moisture content 62%), ultrafine grind to 0.5 mm, add 1.2 kg of 0.2% oxalic acid solution to adjust pH to 4.8-5.2, and ultrasonically assisted soak for 25 min (the parameters of ultrasonic assisted treatment are: power 300W, frequency 28kHz, treatment once every 5 min, each time for 30 s). Then add 25 kg of wheat bran and 10 kg of soybean meal, adjust the moisture content to 51%, add 0.1 kg of potassium dihydrogen phosphate, and let stand for 25 min to obtain the pretreated raw materials. Fermentation: After inoculating the pretreated raw materials with the compound microbial agent prepared in Example 2 at an inoculation rate of 1.6% (mass fraction), fermentation is carried out (a dynamic fermentation device can be used, such as a 100L volume, 304 stainless steel material, equipped with a gradient temperature control system and a ventilation system (0.1-1.5 L / (kg·h))). The gradient temperature control fermentation parameters are as follows: 0-16h 30℃, 0.4L / (kg·h), dissolved oxygen controlled at 12%; 16-32h 36℃, 0.6 L / (kg·h), dissolved oxygen controlled at 18%; 32-42h 32℃, 0.7 L / (kg·h), dissolved oxygen controlled at 15%. During fermentation, the material status is checked every 8 hours. When the moisture content is below 46%, deionized water is added, with the amount added calculated as "0.8% water for every 1% decrease in moisture content". When the pH is below 4.6, 0.08% sodium carbonate solution is added to adjust the pH to ensure that it is maintained between 4.8 and 5.2.

[0034] After fermentation, the fermentation products were tested, and the lignin degradation rate reached 37%, while the crude protein content was 21.2%. Post-processing: Dry with low-temperature hot air to a moisture content of 11%, pulverize to 0.5 mm, add 0.2 kg of vitamin premix (vitamin A 6000 IU / kg, vitamin D3 1500 IU / kg, vitamin E 25 mg / kg), 0.35 kg of mineral premix (calcium 8 g / kg, phosphorus 3.5 g / kg, iron 70 mg / kg, manganese 60 mg / kg), and 0.15 kg of lysine. Mix in a three-dimensional mixer for 20 min (mixing uniformity coefficient of variation 3.2%). Vacuum package (vacuum degree -0.098 MPa), 20 kg / bag, with an actual weight deviation of ±0.12 kg per bag. Store in a warehouse at 20℃ and 55% relative humidity. After 6 months, the viable bacterial count is 9.5 × 10⁻⁶. 7 CFU / g, moisture content 11.8%, no mold growth.

[0035] Example 8 Preparation of Shiitake Mushroom Bran Chicken Feed Pretreatment of raw materials: Take 70 kg of shiitake mushroom spawn (moisture content 60%), ultrafine grind to 0.3 mm, add 1.4 kg of 0.2% oxalic acid solution to adjust pH to 4.8-5.2, and ultrasonically assisted soak for 26 min (the parameters of ultrasonic assisted treatment are: power 300W, frequency 28kHz, treatment once every 5 min, each treatment for 30 s). Then add 15 kg of wheat bran and 10 kg of soybean meal, adjust the moisture content to 52%, add 0.16 kg of potassium dihydrogen phosphate, and let stand for 30 min to obtain the pretreated raw materials. Fermentation: The pretreated raw materials were inoculated with the compound microbial agent prepared in Example 3 at 2.0% (mass fraction) and then fermented (a dynamic fermentation device, such as a 100L volume, 304 stainless steel device equipped with a gradient temperature control system and a ventilation system (0.1-1.5 L / (kg·h)), was used). The gradient temperature control fermentation parameters were as follows: 0-16h 28℃, 0.4L / (kg·h), dissolved oxygen controlled at 10%; 16-32h 34℃, 0.6 L / (kg·h), dissolved oxygen controlled at 16%; 32-42h 30℃, 0.7 L / (kg·h), dissolved oxygen controlled at 14%. After fermentation, the lignin degradation rate was 34%, and the crude protein content was 20.8%. During fermentation, the material status is checked every 8 hours. When the moisture content is below 46%, deionized water is added, with the amount added calculated as "0.8% water for every 1% decrease in moisture content". When the pH is below 4.6, 0.08% sodium carbonate solution is added to adjust the pH to ensure that it is maintained between 4.8 and 5.2.

[0036] After fermentation, the fermentation products were tested, and the lignin degradation rate reached 38%, while the crude protein content was 22.6%. Post-processing: Dry with low-temperature hot air to a moisture content of 12%, pulverize to 0.4 mm, add 0.25 kg of vitamin premix (vitamin A 8000 IU / kg, vitamin D3 2000 IU / kg, vitamin E 30 mg / kg, vitamin K3 3 mg / kg), 0.40 kg of mineral premix (calcium 12 g / kg, phosphorus 4 g / kg, zinc 90 mg / kg, selenium 0.4 mg / kg), and 0.1 kg of methionine. Mix in a three-dimensional mixer for 20 min (mixing uniformity coefficient of variation 3.2%). Vacuum package (vacuum degree -0.098 MPa), 20 kg / bag, with an actual weight deviation of ±0.12 kg per bag. Store in a warehouse at 20℃ and 55% relative humidity. After 6 months, the viable bacterial count is 9.48 × 10⁻⁶. 7 CFU / g, moisture content 11.2%, no mold growth.

[0037] Example 9 Preparation of Shiitake Mushroom Bran Chicken Feed Pretreatment of raw materials: Take 62 kg of shiitake mushroom spawn (60% moisture content), ultrafine grind to 0.5 mm, add 1.3 kg of 0.2% oxalic acid solution, adjust the pH to 4.8-5.2, and ultrasonically assisted soak for 22 min (the parameters for ultrasonic assisted treatment are: power 300W, frequency 28kHz, treatment once every 5 min, each treatment lasts 30 s). Then add 18 kg of wheat bran and 12 kg of soybean meal, adjust the moisture content to 48%, add 0.12 kg of potassium dihydrogen phosphate, and let stand for 28 min to obtain the pretreated raw materials. Fermentation: After inoculating the pretreated raw materials with the compound microbial agent prepared in Example 4 at 1.8% (mass fraction), fermentation is carried out (a dynamic fermentation device can be used, such as a 100L volume, 304 stainless steel material, equipped with a gradient temperature control system and a ventilation system (0.1-1.5 L / (kg·h))). The gradient temperature control fermentation parameters are as follows: 0-16h, 29℃, 0.4L / (kg·h), dissolved oxygen controlled at 12%; 16-32h, 35℃, 0.6 L / (kg·h), dissolved oxygen controlled at 18%; 32-42h, 31℃, 0.7 L / (kg·h), dissolved oxygen controlled at 15%. During fermentation, the material status is checked every 8 hours. When the moisture content is below 46%, deionized water is added, with the amount added calculated as "0.8% water for every 1% decrease in moisture content". When the pH is below 4.6, 0.08% sodium carbonate solution is added to adjust the pH to ensure that it is maintained between 4.8 and 5.2.

[0038] After fermentation, the fermentation products were tested, and the lignin degradation rate reached 37%, while the crude protein content was 23.2%. Post-processing: Dry with low-temperature hot air to a moisture content of 11%, pulverize to 0.6 mm, add 0.2 kg of vitamin premix (vitamin A 6000 IU / kg, vitamin D3 1500 IU / kg, vitamin E 25 mg / kg), 0.35 kg of mineral premix (calcium 8 g / kg, phosphorus 3.5 g / kg, iron 70 mg / kg, manganese 60 mg / kg), and 0.15 kg of lysine. Mix in a three-dimensional mixer for 20 min (mixing uniformity coefficient of variation 3.2%). Vacuum package (vacuum degree -0.098 MPa), 20 kg / bag, with an actual weight deviation of ±0.12 kg per bag. Store in a warehouse at 20℃ and 55% relative humidity. After 6 months, the viable bacterial count is 9.42 × 10⁻⁶. 7 CFU / g, moisture content 10.7%, no mold growth.

Claims

1. A compound microbial inoculant for preparing chicken feed, characterized in that, It consists of Rhizopus oryzae inoculum, Lactobacillus acidophilus inoculum, and Candida tropicalalis inoculum.

2. The compound microbial agent according to claim 1, characterized in that, Based on the number of live bacteria, the ratio of Rhizopus oryzae agent, Lactobacillus acidophilus agent and Candida tropicalis agent is (2-5):(1-3):(1-3); preferably, the ratio of Rhizopus oryzae agent, Lactobacillus acidophilus agent and Candida tropicalis agent is 3:2:

2.

3. The method for preparing the composite microbial agent according to claim 1 or 2, characterized in that, include: Prepare Rhizopus oryzae seed culture, Lactobacillus acidophilus seed culture and Candida tropicalis seed culture; The mixture is obtained by mixing the various sub-liquids, and a protective agent is added to the mixture to obtain the final product; preferably, the protective agent is maltodextrin; the added protective agent accounts for 1.5% of the total mass of the mixture.

4. A method for preparing chicken feed, characterized in that, include: The chicken feed is fermented by inoculating a mixture of fermentation raw materials with a compound microbial agent; wherein the mixture of fermentation raw materials mainly includes shiitake mushroom spawn, wheat bran and soybean meal; and the compound microbial agent is the compound microbial agent as described in claim 1 or 2.

5. The preparation method according to claim 4, characterized in that, According to the mass ratio, in the fermentation raw material mixture, the ratio of shiitake mushroom substrate: wheat bran: soybean meal is (6-7):(1.5-2.5):(1-2); preferably, the ratio of shiitake mushroom substrate: wheat bran: soybean meal is 6.5:2:1.

5. The shiitake mushroom substrate is pretreated shiitake mushroom substrate. The pretreatment method includes: crushing the shiitake mushroom substrate to a particle size of 0.3-0.5 mm, adding oxalic acid solution to adjust the pH to 4.8-5.2, and then soaking it. During the soaking period, ultrasonic-assisted treatment is used. Preferably, the parameters of the ultrasonic-assisted treatment are: power 300W, frequency 28kHz, treatment once every 5 minutes, and each treatment lasts for 30 seconds.

6. The preparation method according to claim 5, characterized in that, The preparation method of the fermentation raw material mixture is as follows: pretreated shiitake mushroom spawn is mixed with wheat bran and soybean meal, and the moisture content is adjusted to 48%-52% to obtain a mixture; potassium dihydrogen phosphate is added to the mixture, stirred evenly, and allowed to stand to obtain the mixture; preferably, the added potassium dihydrogen phosphate accounts for 0.1% of the mass percentage of the mixture.

7. The preparation method according to claim 4, characterized in that, After inoculating the fermentation raw material mixture with the compound microbial agent and stirring evenly, gradient fermentation is carried out; the gradient fermentation parameters are as follows: Fermentation 0-16h: temperature 28-30℃, ventilation rate 0.4 L / (kg·h), dissolved oxygen level controlled at 8%-12%; Fermentation period 16-32 hours: temperature 34-36℃, ventilation rate 0.6 L / (kg·h), dissolved oxygen level controlled at 15%-18%; Fermentation period 32-48 hours: temperature 30-32℃, ventilation rate 0.7 L / (kg·h), dissolved oxygen level controlled at 12%-15%.

8. The preparation method according to claim 7, characterized in that, During fermentation, the material condition was checked every 8 hours. When the moisture content was below 46%, 0.8% water was added for every 1% decrease in moisture content. The pH was maintained between 4.8 and 5.

2.

9. The preparation method according to claim 7, characterized in that, After fermentation, the fermentation product is processed by vacuum freeze-drying or low-temperature hot air drying. Preferably, the parameters for vacuum freeze-drying are: temperature -40℃ to -35℃, vacuum degree ≤10Pa, and drying time 8h; the parameters for low-temperature hot air drying are: temperature 55-58℃, air velocity 1.2m / s, and drying time 2.5h.

10. Chicken feed prepared by any one of claims 5-9.

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