A fermentation inoculant for preparing functional feed by fermenting traditional Chinese medicine residues and a preparation method of the functional feed

Abstract

The present invention belongs to the field of feeds, and particularly relates to a fermentation bacterium agent for fermenting Chinese medicine residues to prepare functional feeds and a preparation method of the functional feeds. The fermentation bacterium agent contains a composite bacterium, and the composite bacterium contains Kluyveromyces marxianus and Bacillus coagulans. Among them, the Kluyveromyces marxianus is isolated from Bupleurum chinense DC. residues, and the preservation number is CGMCC No. 24455. The Bacillus coagulans is isolated from the intestinal contents of healthy piglets, and the preservation number is CGMCC No. 22951. The present invention constructs a fermentation bacterium agent with a Kluyveromyces marxianus isolated from Bupleurum chinense DC. residues and excellent strains previously discovered by the applicant, improves the adaptability to the fermentation of Chinese medicine residues, reduces the crude fiber content, and significantly increases the contents of polysaccharides, crude proteins and organic acids, so that it is more suitable for feeding livestock and poultry.

Description

Technical Field

[0001] The present invention belongs to the field of feeds, and particularly relates to a fermentation bacterium agent for fermenting Chinese medicine residues to prepare functional feeds and a preparation method of the functional feeds. Background Art

[0002] The abuse of antibiotics in livestock production has caused serious harms: it has accelerated the generation of drug-resistant strains, led to the decline of the animal body's immunity, and induced the occurrence and prevalence of various diseases; the antibiotics remaining in livestock and poultry products enter the human body through the food chain, seriously endangering human health. Especially the antibiotics used for preventing diseases and promoting animal growth, with large usage doses and long usage times, are the main reasons for the antibiotic residues in animals and the generation of drug resistance in bacteria.

[0003] Currently, emerging antibiotic substitutes represented by Chinese herbal medicines and probiotics have become research hotspots. These "antibiotic substitution products" have the functions of promoting feed digestion and absorption, improving animal production performance, regulating the balance of animal intestinal flora, enhancing animal immunity, and inhibiting viruses. At the same time, they have the advantages of no toxic side effects, no drug residues, no induction of drug resistance in strains, high efficiency, safety, and practicability.

[0004] In recent years, with the continuous increase in the production of Chinese herbal medicines, the output of Chinese medicine residues has also been increasing day by day. Chinese medicine residues are rich in nutritional components, containing a large number of nutrients such as crude protein, crude fiber, crude fat, starch, polysaccharides, amino acids, alkaloids, vitamins, trace elements, and various pharmacodynamic active components, and have certain medicinal and feeding values.

[0005] The Chinese invention patent application with the publication number of CN107897523A discloses a fermented feed for livestock and poultry using Chinese medicine residues, which ferments Chinese medicine residues such as cassia seed residues, scutellaria baicalensis residues, and coptis chinensis residues by using a conventional bacterium agent to prepare the fermented feed. Through research, it is found that the adaptability of the conventional bacterium agent in fermenting Chinese medicine residues is limited, and the fermentation effect on Chinese medicine residues also needs to be improved. Summary of the Invention

[0006] The purpose of the present invention is to provide a fermentation bacterium agent for fermenting Chinese medicine residues to prepare functional feeds, thereby improving the fermentation effect on Chinese medicine residues.

[0007] The second purpose of the present invention is to provide a preparation method of functional feeds based on the above fermentation bacterium agent.

[0008] In order to achieve the above purposes, the technical solutions adopted by the present invention are as follows:

[0009] A fermentation inoculant for preparing functional feed by fermenting traditional Chinese medicine residues. The fermentation inoculant contains a composite bacterium, and the composite bacterium contains Kluyveromyces marxianus and Bacillus coagulans. Among them, the Kluyveromyces marxianus is isolated from Bupleurum chinense DC. residues, and the preservation number is CGMCC No. 24455. The Bacillus coagulans is isolated from the intestinal contents of healthy piglets, and the preservation number is CGMCC No. 22951.

[0010] In the present invention, a Kluyveromyces marxianus isolated from Bupleurum chinense DC. residues is combined with excellent strains previously discovered by the applicant to construct a fermentation inoculant, which improves the adaptability to the fermentation of traditional Chinese medicine residues, reduces the crude fiber content, and significantly increases the contents of polysaccharides, crude proteins and organic acids, so that it is more suitable for feeding livestock and poultry.

[0011] To further obtain a functional feed with a sweet taste, preferably, the fermentation inoculant is a composite bacterial liquid, and the composite bacterial liquid contains the Kluyveromyces marxianus bacterial liquid and the Bacillus coagulans bacterial liquid. The viable count of the Kluyveromyces marxianus bacterial liquid is 3 - 4×10 8 cfu / mL, and the viable count of the Bacillus coagulans bacterial liquid is 3 - 4×10 9 cfu / mL; the inoculation amount of the Kluyveromyces marxianus bacterial liquid is 0.5 - 6%, and the inoculation amount of the Bacillus coagulans bacterial liquid is 0.5 - 6%.

[0012] Preferably, the composite bacterium further contains a Bacillus subtilis bacterial liquid. The Bacillus subtilis is isolated from the aerobic pond sewage of a pharmaceutical factory, and the preservation number is CGMCC No. 15560.

[0013] More preferably, the fermentation inoculant is a composite bacterial liquid. The fermentation inoculant is a composite bacterial liquid, and the composite bacterial liquid contains the Kluyveromyces marxianus bacterial liquid, the Bacillus coagulans bacterial liquid and the Bacillus subtilis bacterial liquid. The viable count of the Kluyveromyces marxianus bacterial liquid is 3 - 4×10 8 cfu / mL, and the viable count of the Bacillus coagulans bacterial liquid is 3 - 4×10 9 cfu / mL, and the viable count of the Bacillus subtilis bacterial liquid is 6 - 7×10 8 cfu / mL; the inoculation amount of the Kluyveromyces marxianus bacterial liquid is 0.5 - 6%, the inoculation amount of the Bacillus coagulans bacterial liquid is 0.5 - 6%, and the inoculation amount of the Bacillus subtilis bacterial liquid is 2 - 5%.

[0014] A preparation method of a functional feed, comprising the following steps: sterilize the traditional Chinese medicine residues, add a nitrogen source, a phosphorus source and magnesium sulfate to make a solid fermentation medium, mix the above fermentation inoculant and the solid fermentation medium evenly, and culture at a temperature of 25 - 37°C for 3 - 15 days.

[0015] The preparation method of the functional feed of the present invention uses Chinese medicine residues as the main raw material to ferment and prepare the functional feed. The obtained functional feed has a low crude fiber content, and the contents of polysaccharides, crude protein and organic acids are significantly increased. Feeding animals with this functional feed can effectively increase the average feed intake and average daily weight gain of animals, reduce the feed-to-meat ratio, and significantly reduce the odor of animal feces.

[0016] Preferably, the uniformly mixed material is filled into a fermentation bag with a one-way air release valve for the cultivation.

[0017] Preferably, the Chinese medicine residues are Scutellaria baicalensis residues, Qianbai Biyanpian residues or Xiao'er Kechuanling residues. Xiao'er Zhike Ling, Qianbai Biyanpian and Scutellaria baicalensis are all commonly used Chinese medicines, and a large amount of residues are generated during their production. Flos Lonicerae, Bupleuri Radix, Isatidis Radix, Scutellaria baicalensis, Ephedrae Herba and Senecio scandens contained in these Chinese medicines or Chinese medicine prescriptions have the effects of clearing heat and drying dampness, purging fire and detoxifying, etc., and have good effects on treating upper respiratory tract infections, pulmonary heat cough and other symptoms, and are also very beneficial to the health of livestock and poultry. Subsequent experimental results show that the present invention has achieved good fermentation effects on the above-mentioned residues.

[0018] Preferably, the nitrogen source is urea or ammonium sulfate, and the addition amount of the nitrogen source relative to the Chinese medicine residues is 0.9 - 1.3 g / Kg; the phosphorus source is potassium dihydrogen phosphate or dipotassium hydrogen phosphate, and the addition amount of the phosphorus source relative to the Chinese medicine residues is 0.9 - 1.3 g / Kg, and the addition amount of magnesium sulfate relative to the Chinese medicine residues is 1.3 - 1.9 g / Kg.

[0019] Preferably, the fermentation inoculant is a composite bacterial liquid composed of Kluyveromyces marxianus bacterial liquid and Bacillus coagulans bacterial liquid, or a composite bacterial liquid composed of Kluyveromyces marxianus bacterial liquid, Bacillus coagulans bacterial liquid and Bacillus subtilis bacterial liquid; wherein, the viable count of Kluyveromyces marxianus bacterial liquid is 3 - 4×10 8 cfu / mL, the viable count of Bacillus coagulans bacterial liquid is 3 - 4×10 9 cfu / mL, and the viable count of Bacillus subtilis bacterial liquid is 6 - 7×10 8 cfu / mL;

[0020] The Chinese medicine residue is Scutellaria baicalensis Georgi residue, and the compound bacterial liquid is composed of Bacillus subtilis bacterial liquid, Bacillus coagulans bacterial liquid and Kluyveromyces marxianus bacterial liquid, and the inoculation amounts are 2-3%, 4-5%, 1-2% respectively; or the Chinese medicine residue is Qianbai Biyanpian residue, and the compound bacterial liquid is composed of Bacillus coagulans bacterial liquid and Kluyveromyces marxianus bacterial liquid, and the inoculation amounts are 1-2%, 4-5% respectively; or the Chinese medicine residue is Xiao'er Kechuanling residue, and the compound bacterial liquid is composed of Bacillus subtilis bacterial liquid, Bacillus coagulans bacterial liquid and Kluyveromyces marxianus bacterial liquid, and the inoculation amounts are 2-3%, 1-2%, 1-2% respectively. The Qianbai Biyanpian residue is mainly composed of the residues of Senecio scandens Buch.-Ham., Selaginella tamariscina (Beauv.) Spring, Cassia obtusifolia L. and Ephedra sinica Stapf. The Xiao'er Kechuanling residue is mainly composed of the residues of Ephedra sinica Stapf., Lonicera japonica Thunb., Armeniaca vulgaris Lam. var. ansu (Maxim.) Yü et Lu, Isatis indigotica Fort., Glycyrrhiza uralensis Fisch. and Trichosanthes kirilowii Maxim.

[0021] Preferably, the functional feed is admixed into the common feed for use, and the admixing amount is 20-30%. Detailed implementation mode

[0022] The present invention uses a Kluyveromyces marxianus strain isolated from Bupleurum chinense DC. residue and excellent strains previously discovered by the applicant to form a fermentation inoculant for fermenting Chinese medicine residue to produce functional feed. Using the above fermentation inoculant for fermentation production can effectively improve the palatability of the fermented feed of Chinese medicine residue. The crude protein, polysaccharide and organic acid indexes of the fermented functional feed are excellent, and it is suitable for mixing with common feed to feed animals, so as to give full play to the dual effects of Chinese medicine components and probiotics.

[0023] In addition to crude protein, amino acids, cellulose, lignin, hemicellulose, crude fat, minerals, oils, vitamins and various trace elements such as nitrogen, phosphorus and potassium, the Chinese medicine residue also contains bioactive substances such as polysaccharides, terpenoids, alkaloids, volatile oils, quinones, flavonoids, steroids and their glycosides, tannins, and some unknown growth-promoting substances. These nutritional components and bioactive substances have the functions of balancing animal nutrition, accelerating animal growth and development, regulating physiological functions, and improving the yield and quality of livestock and poultry products. Xiao'er Zhike, Qianbai Biyanpian and Scutellaria baicalensis Georgi are all commonly used Chinese medicines, and a large amount of medicine residues are generated during the production process. Among them, Lonicera japonica Thunb., Bupleurum chinense DC., Isatis indigotica Fort., Scutellaria baicalensis Georgi, Ephedra sinica Stapf. and Senecio scandens Buch.-Ham. all have the effect of clearing heat and detoxifying, which is of great benefit to the health of livestock and poultry.

[0024] Probiotics such as yeasts, bacilli and lactic acid bacteria inherently have the health care functions of promoting the growth and development of the body and improving immunity. During the fermentation process, probiotics produce a large amount of cellulase, amylase and lipase, which can promote the absorption and utilization of functional components in the Chinese medicine residue and enhance the efficacy performance of the Chinese medicine residue. The Chinese medicine residue is beneficial to the growth and reproduction of microorganisms, and the two complement each other.

[0025] The situations of three typical strains will be described below.

[0026] 1. Screening and Identification of Kluyveromyces marxianus

[0027] Strain screening: The Bupleurum residue was diluted with sterile water at different multiples, and the diluted solutions were respectively spread on the broth agar medium plates and cultured overnight at 37°C. The white round colonies grown on the broth agar medium plates were picked and inoculated onto fresh broth PDA agar medium plates by the method of streaking and cultured for 36 h. The separation and purification were carried out 3 times according to the above method to obtain pure strains. The composition of the broth agar medium (g / 100 mL): peptone 1, beef extract 0.3, sodium chloride 0.5, agar 1.5, the solvent was deionized water, and the pH value was 7.4.

[0028] Strain identification: The colonies of the strain were round, milky white, convex, transparent, with a smooth surface and neat edges. The cells were oval, about 4 - 8 mm in size, reproduced by budding, and formed strings singly, in pairs or in multiples.

[0029] The purified strain was inoculated into a common broth medium (the other components were the same as the broth agar medium except without agar), cultured at 28°C for 36 - 48 h, the bacterial solution was centrifuged at 5000 r / min for 20 min, and the obtained precipitate (mycelium) was stored frozen at -20°C for 24 h.

[0030] The frozen mycelium was thawed and shaken well, ultrasonically vibrated for 10 min, and then centrifuged at 5000 r / min for 10 min. After adding benzyl chloride extraction buffer (pH = 9.0) to the precipitate, it was shaken well, water-bathed at 50°C for 1 - 2 h, shaken once every 10 min. After the mycelium was completely dissolved, 0.1 M sodium acetate solution (pH = 5.2) was added, and then ice-bathed for 15 min. After ice-bathing, it was centrifuged at 5000 r / min for 15 min. Isopropanol was added to the supernatant and mixed well, and then precipitated at room temperature for 20 min, and then centrifuged at 5000 r / min for 10 min. The precipitate was washed twice with 80% ethanol and dissolved with 1×TE to obtain a crude DNA product. Equal volume of phenol / chloroform / isopentyl alcohol was added to the crude DNA product, gently inverted up and down 30 times, centrifuged at 18°C and 12000 r / min for 5 min, and the supernatant was pure DNA.

[0031] Using the bacterial DNA as a template, the 18S rRNA gene fragment of the strain was amplified by PCR with the 18S rRNA universal primers NS1 (5’-GTAGTCATATGCTTGTCTC-3’) and NS2 (5’-GGCTGCTGGCACCAGACTTGC-3’). The 20 μL PCR amplification system contained 0.5 μL of each upstream and downstream primer, 0.5 μL of the extracted DNA template (10 - 100 ng / μL), 8.5 μL of sterile ultrapure water, and 10 μL of 2×Ex Tap enzyme. The PCR amplification program was: (94°C, 5 min) + {(94°C, 30 s) + (55°C, 30 s) + (72°C, 1 min)} × 30 + (72°C, 7 min). After verifying the purity of the PCR product by electrophoresis on a 1% agarose gel, it was sent to Beijing Tsingke Biotechnology Co., Ltd. for sequencing.

[0032] The sequence peak map of the sequencing result was viewed through BioEdit 7.2.6. The base sequence with clear and continuous peaks and valleys was selected. Through the BLAST system in the NCBI database, the target sequence was homologously aligned with the sequences in the nucleic acid sequence library GenBank, and it was determined that the strain was Kluyveromyces marxianus. It was deposited in the China General Microbiological Culture Collection Center, with the deposit number: CGMCC No. 24455, the strain name: JBH-BJM4, the deposit date: March 1, 2022, and the deposit address: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing.

[0033] 2. Bacillus coagulans and Bacillus subtilis

[0034] Bacillus coagulans is a strain preserved in this laboratory, with the deposit number CGMCC No. 22951, and it is described in detail in the Chinese patent application with the publication number CN113652373A.

[0035] Bacillus subtilis is a strain preserved in this laboratory, with the deposit number CGMCC No. 15560, and it is described in detail in the Chinese patent application with the publication number CN109022318A.

[0036] The following specifically describes the specific implementation process of the present invention in combination with specific embodiments.

[0037] Example 1 Fermentation Bacterial Agent

[0038] The fermentation inoculant for preparing functional feed by fermenting traditional Chinese medicine residues in this embodiment is a compound bacterial liquid, and the compound bacteria therein are composed of Kluyveromyces marxianus, Bacillus subtilis, and Bacillus coagulans. The preservation number of Kluyveromyces marxianus is: CGMCC No. 24455, the preservation number of Bacillus subtilis is: CGMCC No. 15560, and the preservation number of Bacillus coagulans is: CGMCC No. 22951.

[0039] The preparation process of the fermentation inoculant is as follows:

[0040] (1) Culture medium of strains

[0041] For the activation and enlarged cultivation of Kluyveromyces marxianus and Bacillus subtilis, broth culture medium is used, and its composition is: 2 g of tryptone, 2 g of NaCl, 1 g of yeast extract, 100 mL of purified water, and pH 6.8.

[0042] The activation culture medium (YPD medium) of Bacillus coagulans has the following composition: 2 g of glucose, 2 g of peptone, 1 g of yeast extract, 100 mL of purified water, and pH 6.8. The enlarged culture medium uses MRS medium, and its composition is: 1 g of peptone, 1 g of beef extract, 0.5 g of yeast powder, 0.2 g of K2HPO4, 0.2 g of diammonium citrate, 0.5 g of sodium acetate, 2 g of glucose, 0.1 mL of Tween 80, 0.058 g of MgSO4·7H2O, 0.025 g of MnSO4·4H2O, 100 mL of distilled water, and pH 6.3.

[0043] (2) Cultivation method

[0044] Cultivation method of yeast: Inoculate the Kluyveromyces marxianus slant strain into 100 mL of activation culture medium, and cultivate at 30 °C and 150 r / min for 48 h; then add 8 mL of seed liquid into 200 mL of enlarged culture medium, and cultivate at 30 °C and 150 r / min for 48 h to obtain yeast fermentation broth, and the viable count of the yeast fermentation broth is 3.5×10 8 cfu / mL.

[0045] Cultivation method of bacillus: Inoculate the Bacillus subtilis slant strain into 100 mL of activation culture medium, and cultivate at 37 °C and 180 r / min for 24 h; then add 8 mL of seed liquid into 200 mL of enlarged culture medium, and cultivate in a shaker for 24 h, and the cultivation conditions are 37 °C and 180 r / min to obtain bacillus fermentation broth, and the viable count of the bacillus fermentation broth is 6.9×10 8 cfu / mL.

[0046] Method for activating lactic acid bacteria: Inoculate the Bacillus coagulans slant strain into 100 mL of activation medium, and culture it at 37 °C and 180 r / min for 24 h; then add 8 mL of seed liquid to 200 mL of enlarged medium, and statically culture it at 37 °C for 24 h to obtain the lactic acid bacteria fermentation broth. The viable count of the lactic acid bacteria fermentation broth is 3.6×10 9 cfu / mL.

[0047] Example 2 Preparation of functional feed using Scutellaria baicalensis residue

[0048] The preparation method of the functional feed in this example is specifically described as follows:

[0049] Add 1.1 g of urea and 1.1 g of dipotassium hydrogen phosphate to 1.2 kg of pulverized and sterilized Scutellaria baicalensis residue (water content 30 - 40%) to prepare a solid fermentation medium. Pack the solid fermentation medium into fermentation bags with a one-way air release valve, 200 g per bag, add the bacterial liquid into the bags according to different ratios, seal the bags after stirring evenly, and culture them in an incubator at 30 °C for 7 days. During the culture period, turn the materials once a day to make the materials mix evenly.

[0050] Table 1 shows the addition ratios of different bacterial liquids. The total amount of bacterial liquid in each group is 15 mL, and the water content of the residue after adding the bacterial liquid is 46.25%. The fermentation results are shown in Table 1.

[0051] Table 1 Composition of fermentation inoculant strains and fermentation results

[0052]

[0053] (Note: Bacillus, lactic acid bacteria, and yeast are Bacillus subtilis, Bacillus coagulans, and Kluyveromyces marxianus involved in Example 1.)

[0054] In Table 1, after fermentation with the 3rd (group) formula, the odor of the product is better and has a sweet taste. For the products fermented with other formulas, the odor is average, without a sweet taste, and there is no mildew phenomenon after being placed for 30 days. Select the 3rd formula for in-depth research, and conduct batch culture and feeding tests.

[0055] To verify the effect of the strains used in the present invention, a strain substitution test was carried out according to the 3rd formula. The fermentation agent in Test Group 1 used the 3rd formula in Table 1. In Test Group 2, commercially available feed yeast was used to replace Kluyveromyces marxianus. In Test Group 3, Bacillus subtilis produced by Shandong Sukahan Bioengineering Co., Ltd. was used to replace the Bacillus subtilis used in the present invention. In Test Group 4, Bacillus coagulans produced by Shandong Zhongke Jiayi Biotechnology Co., Ltd. was used to replace the Bacillus coagulans used in the present invention. All other conditions were the same. It was found that the odor and final pH of the products obtained in several test groups were not obvious.

[0056] The components of the materials before and after fermentation were analyzed. The polysaccharide content was determined by the 3,5-dinitrosalicylic acid method, the crude fiber by the acid-base washing method, the crude protein by the Kjeldahl method, and the organic acid content by the acid-base titration method. Each sample was analyzed 3 times and the average value was taken. The results are shown in Table 2. Statistical analysis and variance analysis were performed using SAS (Statistical Analysis) version 7.2, and the results were considered significant when P<0.05. The significant differences were represented by the letter marking method: First, all the average values were arranged in descending order, and then the letter a was marked on the largest average value; and this average value was compared with the following average values. Those with non-significant differences were all marked with the letter a until a significantly different average value was encountered, and the letter b was marked; then, taking the average value marked with b as the standard, it was compared with the larger average values above it. Those with non-significant differences were also all marked with the letter b; then, taking the largest average value marked with b as the standard, it was compared with the following unmarked average values. Those with non-significant differences were continuously marked with the letter b until a significantly different average value was encountered and marked with c.

[0057] Table 2 Changes in the main components of the materials before and after fermentation

[0058]

[0059] As can be seen from the results in Table 2, after fermentation with the fermentation inoculant of the example, in the test group 2 using commercially available feed yeast, it was significantly inferior to the example in terms of crude protein, crude fiber, and organic acid indicators; in the test group 3 using commercially available Bacillus subtilis, it was significantly inferior to the example in terms of polysaccharide, crude fiber, and organic acid indicators; in the test group 4 using commercially available Bacillus coagulans, it was significantly inferior to the example in terms of crude fiber indicator.

[0060] Example 3 Preparation of functional feed using the residues of Qianbai Biyan tablets

[0061] The preparation method of the functional feed in this example includes the following steps:

[0062] 1.1 g of urea and 1.1 g of dipotassium hydrogen phosphate were added to 1.2 kg of crushed and sterilized residues of Qianbai Biyan tablets (with a water content of 30-40%) to prepare a solid fermentation medium. The solid fermentation medium was filled into a fermentation bag with a one-way air release valve, the bacterial liquid was added to the bag in different proportions, stirred evenly and sealed, and cultured at 30°C for 7 days. Other conditions were the same as in Example 2.

[0063] Table 3 shows the addition ratios of different bacterial liquids. The total amount of bacterial liquid in each group was 10 mL, and the water content of the residues after adding the bacterial liquid was 44.25%. The fermentation results are shown in Table 3.

[0064] Table 3 Composition of the fermentation inoculant strains and fermentation results

[0065]

[0066] (Note: Bacillus, Lactobacillus, and Saccharomyces are Bacillus subtilis, Bacillus coagulans, and Saccharomyces kluyveri involved in Example 1.)

[0067] After fermentation with the 4th (group) formula, the odor is good, with a sweet taste, and there is no mildew after 30 days of storage. For the products fermented with other formulas, the odor is average, without a sweet taste, and there is slight mildew after 30 days of storage. The self-grown fungi may be harmful bacteria. Therefore, the 4th formula is selected for in-depth research, and batch culture and feeding tests are carried out.

[0068] Design a strain substitution test according to Example 2, and conduct component analysis of the materials before and after fermentation. For the fermentation agent in Test Group 1, the 4th group formula in Table 3 is used. In Test Group 2, commercially available feed yeast is used to replace Saccharomyces kluyveri. In Test Group 3, Bacillus coagulans from Shandong Zhongke Jiayi Biotechnology Co., Ltd. is used to replace the Bacillus coagulans of the present invention. All other conditions are the same. It is found that the odor and final pH of the products obtained in several test groups are not obvious. The results of the nutritional components are shown in Table 4.

[0069] Table 4 Changes in the main components of the materials before and after fermentation

[0070]

[0071] From the results in Table 4, it can be seen that after fermentation with the fermentation agent of the example, in Test Group 2 where commercially available feed yeast is used, it is significantly inferior to the example in terms of the crude protein index. In Test Group 3 where commercially available Bacillus coagulans is used, it is significantly inferior to the example in terms of the crude fiber and organic acid indexes.

[0072] Example 4 Preparation of functional feed using the residue of Xiao'er Kechuanling

[0073] The preparation method of the functional feed in this example includes the following steps:

[0074] Take 1.2 kg of the residue after extraction of Xiao'er Kechuanling (with a water content of 30 - 40%) after pulverization, add 1.1 g of urea and 1.1 g of dipotassium hydrogen phosphate to prepare a solid fermentation medium. Fill the solid fermentation medium into a fermentation bag with a one-way air release valve, add the bacterial liquid into the bag in different proportions, stir evenly and seal, and culture at 30°C for 7 days.

[0075] Table 5 shows the addition ratios of different bacterial liquids. The total amount of bacterial liquid in each group is 10 mL, and the water content of the residue after adding the bacterial liquid is 44.25%. The fermentation results are shown in Table 5.

[0076] Table 5 Composition of the fermentation agent strains and fermentation results

[0077]

[0078]

[0079] (Note: Bacillus, Lactobacillus, and Saccharomyces are Bacillus subtilis, Bacillus coagulans, and Saccharomyces kluyveri involved in Example 1.)

[0080] From the results in Table 5, it can be seen that after fermentation with the second formula, the smell is good, with a sweet taste, and there is no mildew phenomenon after 30 days of storage. After fermentation with other formulas, the products have no sweet taste, a strong sour taste, and a general smell. After 30 days of storage, there is a slight mildew phenomenon. Therefore, the second formula was selected for in-depth research, and batch culture and feeding tests were carried out.

[0081] Design a strain substitution test according to Example 2, and conduct component analysis of the materials before and after fermentation. In the first test group, the fermentation agent uses the second formula in Table 5. In the second test group, the commercially available feed yeast is used to replace Saccharomyces kluyveri. In the third test group, the Bacillus subtilis produced by Shandong Sukhan Biotechnology Co., Ltd. is used to replace the Bacillus subtilis used in the present invention. In the fourth test group, the Bacillus coagulans of Shandong Zhongke Jiayi Biotechnology Co., Ltd. is used to replace the Bacillus coagulans of the present invention. All other conditions are the same. It is found that the smell and final pH of the products obtained in several test groups are not obvious. The results of the nutrient components are shown in Table 6.

[0082] Table 6 Changes in the main components of the materials before and after fermentation

[0083]

[0084] From the results in Table 6, it can be seen that after fermentation with the fermentation agent of the example, in the second test group, the indexes of polysaccharides, crude fiber, and organic acids are significantly inferior to those of the example. In the third test group, the indexes of crude fiber and organic acids are significantly inferior to those of the example. In the fourth test group, the indexes of polysaccharides, crude fiber, and organic acids are significantly inferior to those of the example.

[0085] Evaluation of the functional feed prepared from Scutellaria baicalensis residue in Experimental Example 5

[0086] Conduct animal feeding tests on each test group involved in Example 2.

[0087] The feeding method is as follows: Select 500 piglets with similar body weights (17.5 ± 1.0 kg), adopt a single-factor completely randomized experimental design, divide them into five treatment groups, with 10 replicates in each group and 10 piglets in each replicate. The test period is 30 days. The test groups are fed with the fermented feed of Example 2 containing 20 wt% and 80 wt% of the commercially available feed, and the control group is fed with 100% commercially available feed. All other feeding conditions are the same.

[0088] Before the start of the experiment and at the end of the experiment, the average feed intake (ADFI), average daily gain (ADG), and feed-to-gain ratio (F / G) of each group of pigs were measured respectively. To investigate the effect of the fermented feed of the present invention on the growth performance of fattening pigs, and statistical analysis was carried out according to the method described in Example 2. At the same time, the diarrhea rate and the odor of feces were investigated, and the experimental results are shown in Table 7:

[0089] Table 7 Effect of Fermented Feed on the Growth Performance of Fattening Pigs

[0090]

[0091] As can be seen from the results in Table 7, after adding the feed prepared with fermented medicinal residues, the average feed intake and average daily gain (ADG) of fattening pigs increased, the feed-to-gain ratio decreased, and the diarrhea rate of the example group was reduced most effectively and the fecal odor was significantly reduced.

[0092] Experimental Example 6 Evaluation of Functional Feed Prepared with Qianbai Biyanpian Medicinal Residues

[0093] Animal feeding experiments were carried out on each test group involved in Example 3.

[0094] The feeding method is as follows: 160 Holstein beef cattle were selected and randomly divided into 4 groups according to the principle of the same age, weight, health status, and gender ratio, with two replicates in each group and 20 heads in each group. Test group 1 was fed with 20 wt% of the fermented feed of Example 3 and 80 wt% of commercially available fattening feed, test group 2 was fed with 20 wt% of the fermented feed of test group 2 and 80 wt% of commercially available fattening feed, test group 3 was fed with 20 wt% of the fermented feed of test group 3 and 80 wt% of commercially available fattening feed, and the control group was fed with 100% commercially available fattening feed, and the remaining feeding conditions were the same.

[0095] Before the start of the experiment and at the end of the experiment, the average feed intake (ADFI), average daily gain (ADG), and feed-to-gain ratio (F / G) of each group of cattle were measured respectively. To investigate the effect of the fermented feed of the present invention on the growth performance of beef cattle. At the same time, the odor of feces was investigated, and statistical analysis was carried out according to the method described in Example 2. The experimental results are shown in Table 8:

[0096] Table 8 Effect of Fermented Feed on the Growth Performance of Beef Cattle

[0097]

[0098] As can be seen from the results in Table 8, after adding the feed prepared with fermented medicinal residues, the average feed intake and average daily gain (ADG) of beef cattle increased. Among them, the average feed intake and average daily gain (ADG) of beef cattle in test group 1 increased significantly, and the feed-to-gain ratio also decreased significantly. And the diarrhea rate of the example group was reduced most effectively and the fecal odor was significantly reduced.

[0099] Evaluation of Functional Feed Prepared from Pediatric Antitussive Medicinal Residue in Example 7

[0100] Animal feeding tests were conducted on each test group involved in Example 4.

[0101] The fattening pigs were fed with the fermented feed of Example 4, and the method was the same as that of Example 5. The results are shown in Table 9.

[0102] Table 9 Effects of Fermented Feed on Growth Performance of Fattening Pigs

[0103]

[0104] It can be seen from the results in Table 9 that after adding the feed prepared from fermented medicinal residues, the average feed intake and average daily gain (ADG) of fattening pigs increased, the feed-to-meat ratio decreased, and the diarrhea rate reduction effect of the example group was the best. In addition, the odor of the feces in the test group was significantly reduced.

Claims

1. A preparation method of a functional feed, characterized in that, Using a fermentation inoculant to ferment traditional Chinese medicine residues to prepare a functional feed, the fermentation inoculant is a composite bacterial liquid composed of Kluyveromyces marxianus bacterial liquid and Bacillus coagulans bacterial liquid, or a composite bacterial liquid composed of Kluyveromyces marxianus bacterial liquid, Bacillus coagulans bacterial liquid and Bacillus subtilis bacterial liquid; wherein, the viable count of Kluyveromyces marxianus bacterial liquid is 3-4×10 8 cfu / mL, the viable count of Bacillus coagulans bacterial liquid is 3-4×10 9 cfu / mL, and the viable count of Bacillus subtilis bacterial liquid is 6-7×10 8 cfu / mL; The Chinese medicine residue is Scutellaria baicalensis Georgi residue, and the composite bacterial liquid is composed of Bacillus subtilis bacterial liquid, Bacillus coagulans bacterial liquid and Kluyveromyces marxianus bacterial liquid, and the inoculation amounts are 2-3%, 4-5%, and 1-2% respectively; or the Chinese medicine residue is Qianbai Biyanpian residue, and the composite bacterial liquid is composed of Bacillus coagulans bacterial liquid and Kluyveromyces marxianus bacterial liquid, and the inoculation amounts are 1-2% and 4-5% respectively; or the Chinese medicine residue is Xiao'er Kechuanling residue, and the composite bacterial liquid is composed of Bacillus subtilis bacterial liquid, Bacillus coagulans bacterial liquid and Kluyveromyces marxianus bacterial liquid, and the inoculation amounts are 2-3%, 1-2%, and 1-2% respectively. Among them, the Kluyveromyces marxianus is isolated from Bupleurum chinense DC. residue, and the preservation number is CGMCC No. 24455; the Bacillus coagulans is isolated from the intestinal contents of healthy piglets, and the preservation number is CGMCC No. 22951; the Bacillus subtilis is isolated from the aerobic pond sewage of a pharmaceutical factory, and the preservation number is CGMCC No. 15560.

2. A preparation method of a functional feed, characterized in that, It includes the following steps: After sterilizing the Chinese medicine residue, add nitrogen source and phosphorus source to make a solid fermentation medium, mix the fermentation inoculant described in Claim 1 and the solid fermentation medium evenly, and culture at a temperature of 25-37 °C for 3-15 days.

3. The preparation method of the functional feed according to claim 2, characterized in that, Load the evenly mixed material into a fermentation bag with a one-way air release valve for the above-mentioned culture.

4. The preparation method of the functional feed according to claim 2, characterized in that, The nitrogen source is urea or ammonium sulfate, and the addition amount of the nitrogen source relative to the Chinese medicine residue is 0.9-1.3 g / Kg; the phosphorus source is potassium dihydrogen phosphate or dipotassium hydrogen phosphate, and the addition amount of the phosphorus source relative to the Chinese medicine residue is 0.9-1.3 g / Kg.

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