Preparation method and application of fermented liver-protecting tablet dreg feed additive

Abstract

The invention discloses a preparation method and application of a fermented liver-protecting tablet dreg feed additive, and belongs to the technical field of feed additive preparation. The preparation method of the fermented liver-protecting tablet dreg feed additive comprises the following steps: mixing and inoculating liver-protecting tablet dregs, bacillus subtilis liquid and saccharomycetes liquid, and culturing to obtain the fermented liver-protecting tablet dreg feed additive. The liver protection tablet residues are developed into the feed additive according to the principle of reduction, harmlessness and recycling, the traditional Chinese medicine residues can be turned into wealth while the environment is protected, and the feed additive is green, free of toxic and side effects and has certain technical advantages, a new idea is provided for recycling of the traditional Chinese medicine residues, and the feed additive is worthy of popularization and application. And a theoretical basis and a practical basis are provided for application of the traditional Chinese medicine residues in livestock and poultry feed additives.

Description

Technical Field

[0001] The invention belongs to the technical field of feed additive preparation, and more specifically relates to a preparation method and application of a fermented liver protection tablet residue feed additive. Background Art

[0002] The rapid development of the traditional Chinese medicine industry has led to an increasing output of Chinese medicine residues. Due to the lack of financial, material and human resources, many Chinese medicine companies have not reprocessed the Chinese medicine residues, resulting in a large amount of Chinese medicine residues being piled up, landfilled or incinerated as waste, which not only wastes resources but also causes serious harm to the environment.

[0003] At present, researchers have applied Chinese herbal medicine residues to many fields, including edible fungus cultivation, organic fertilizer, papermaking, pyrolysis and gasification. Chinese herbal medicine residues are rich in bioactive substances, crude fiber, crude protein, amino acids, minerals, vitamins and trace elements. Studies have shown that whether Chinese herbal medicine residues are fed directly or after fermentation, they can have certain benefits for animals. However, direct feeding of Chinese herbal medicine residues has problems such as poor palatability and low digestibility of animals.

[0004] Therefore, the use of Chinese medicine residue in animal production will also play an important role in the development of green animal husbandry in my country. It is of great significance to develop a feed additive prepared using Chinese medicine residue. Summary of the invention

[0005] The purpose of the present invention is to provide a preparation method and application of a fermented liver protection tablet residue feed additive to solve the problems existing in the above-mentioned prior art, improve the nutritional value of Chinese medicine residue, reduce the content of anti-nutritional factors, enhance the balance of microflora and the generation of metabolites, and realize the efficient utilization of Chinese medicine residue.

[0006] To achieve the above object, the present invention provides the following solutions:

[0007] One of the technical solutions of the present invention is to provide a method for preparing a fermented liver-protecting tablet residue feed additive, comprising the following steps:

[0008] The liver protection tablet residues, Bacillus subtilis liquid and yeast liquid are mixed, inoculated and then cultured to obtain the fermented liver protection tablet residue feed additive.

[0009] The present invention utilizes probiotics (Bacillus subtilis and yeast) to ferment the residues of liver protection tablets, which can further promote the increase of bioactive ingredients such as polyphenols, polysaccharides, and flavonoids. The residues of liver protection tablets are fermented as feed additives. The prepared feed additives can not only improve animal growth performance to a certain extent, enhance animal immunity, improve anti-stress and antibacterial effects, and reduce the use of antibiotics, but also help pharmaceutical companies solve the problem that the treatment of traditional Chinese medicine residues is difficult and expensive. In addition, this recycling method has great application potential in reducing environmental pollution, reducing the cost of livestock and poultry breeding enterprises, and improving food safety and economic benefits.

[0010] In view of the anti-nutritional factors in the liver protection tablets residue, the present invention utilizes the enzyme system of yeast and Bacillus subtilis to degrade and transform these anti-nutritional factors, further reducing their impact on the digestion and absorption of poultry animals. After detection and analysis, no drug residues were detected in the fermented liver protection tablets residue feed additive prepared by the present invention, and the heavy metal content was also within the qualified range, meeting the requirements of the feed hygiene standard GB 13078-2017.

[0011] In the above technical scheme, there is mutual symbiosis and synergy between the yeast and Bacillus subtilis. Yeast decomposes Chinese medicine residues to produce soluble oligosaccharides and other metabolites, providing energy and substrates for Bacillus subtilis, providing suitable environmental conditions and metabolites, and promoting the growth and cellulose degradation of Bacillus subtilis. Bacillus subtilis decomposes complex polysaccharides and cellulose, produces organic acids and enzyme substances, and organic acids reduce the pH value to provide suitable acidity and alkalinity conditions for yeast, while Bacillus subtilis regulates the oxygen concentration by consuming oxygen, provides a lower oxygen environment for yeast, and promotes the generation of products such as acid production and alcohol production. Yeast and Bacillus subtilis provide nutrients, enzymes and other auxiliary factors to each other, jointly promote growth and metabolic activity, and enhance the overall fermentation effect.

[0012] The Bacillus subtilis of the present invention can produce a variety of hydrolases, can effectively degrade macromolecular substances such as crude fiber and protein in traditional Chinese medicine residues, and also has antibacterial effect, can effectively inhibit diarrhea, infection and other problems caused by pathogenic bacteria such as Escherichia coli and Salmonella in the intestines of animals; after the Bacillus subtilis and yeast are compounded and acted on the residues of liver protection tablets, they can be degraded into small molecular active substances, enrich the dietary structure of poultry animals, improve palatability, and at the same time enable microorganisms to colonize in the intestines, can inhibit harmful bacteria, promote beneficial bacteria, regulate the intestinal flora structure, improve intestinal health, stabilize the intestinal environment, and thus promote the body's digestion and absorption of nutrients.

[0013] Preferably, the liver protection tablet residues are dried and sieved before inoculation; the mesh size of the liver protection tablet residues is ≥ 60 meshes;

[0014] Furthermore, the moisture content of the dried liver protection tablet residue is preferably 50-60wt%, and more preferably 54wt%.

[0015] Preferably, the liver protection tablet residues are the residues remaining after the effective ingredients are extracted from the Chinese medicinal materials during the production process of the liver protection tablets.

[0016] Preferably, the dosage ratio of the liver protection tablet residue, Bacillus subtilis solution and yeast solution is 20g:3-15mL:3-15mL, the mass ratio of Bacillus subtilis in the Bacillus subtilis solution and the yeast in the yeast solution is 1-5:1-5; and / or the live bacteria concentration of the Bacillus subtilis solution and the yeast solution is independently 1.0×10 8 ~9.0×10 9 CFU / mL.

[0017] Preferably, the liver protection tablet residues, Bacillus subtilis liquid and yeast liquid are mixed and inoculated into a fermentation bag.

[0018] Preferably, the culture conditions are set as follows: anaerobic conditions, a moisture content of 50-60 wt%, a temperature of 25-40° C., a C / N ratio of 25, and a time of 3-6 d.

[0019] Furthermore, the culture temperature is more preferably 25-35°C, more preferably 30°C.

[0020] In order to enhance the function of the fermentation process of Chinese medicine residue, the present invention selects probiotic strains (Bacillus subtilis and yeast) with good fermentation performance and metabolic activity. The application of probiotics not only improves the fermentation efficiency, but also increases the generation of beneficial metabolites, which has a positive impact on the intestinal health of poultry animals. The inventor systematically studies and optimizes multiple fermentation parameters such as temperature, inoculation amount, water content and fermentation time, successfully improves the growth and metabolic activity of strains, and realizes a more efficient and stable fermentation process; when the water content is too high, it will lead to an anaerobic environment, inhibit the growth of beneficial bacteria, and when it is too low, it will affect the activity of strains and reduce fermentation efficiency; when the temperature is too high, the strain will die or lose activity, and when it is too low, the fermentation rate will be slowed down, affecting the generation of metabolites; when the C / N ratio is too high, it will lead to insufficient nitrogen source, inhibit the growth of strains, and when it is too low, it will cause excess nitrogen, leading to the generation of undesirable metabolites; when the time is too long, it will lead to the decomposition of fermented products and the loss of nutrients, and when it is too short, the fermentation will be incomplete, affecting the quality and function of the final product.

[0021] Preferably, the C / N value is adjusted by glucose.

[0022] The second technical solution of the present invention is to provide a fermented liver protection tablet residue feed additive prepared by the above preparation method.

[0023] The third technical solution of the present invention is to provide the use of the above-mentioned fermented liver protection tablet residue feed additive in the preparation of poultry feed.

[0024] Preferably, the poultry animals include chickens and / or ducks; and the feed includes brooding feed or growing feed.

[0025] Preferably, the amount of the fermented liver protection tablet residue feed additive added to the feed is 1-5wt%.

[0026] The present invention discloses the following technical effects:

[0027] 1. The present invention develops the residues of liver protection tablets into feed additives in accordance with the principle of "reduction, harmlessness, and resource utilization". While protecting the environment, it can also turn the residues of traditional Chinese medicine into treasure. It is green and has no toxic side effects. It has certain technical advantages, provides new ideas for the reuse of traditional Chinese medicine residues, and provides a theoretical basis and practical basis for the application of traditional Chinese medicine residues in livestock and poultry feed additives.

[0028] 2. The feed additive prepared by the present invention makes full use of active ingredients such as polysaccharides, flavonoids, polyphenols and saponins in traditional Chinese medicine residues, reduces harmful bacteria such as Escherichia coli and Salmonella, and increases the content of beneficial bacteria such as Bacillus subtilis and yeast, thereby improving the growth performance of poultry animals to a certain extent, enhancing animal immunity, and improving anti-stress and antibacterial effects.

[0029] 3. The Bacillus subtilis of the present invention can produce a variety of hydrolases, which can effectively degrade macromolecular substances such as crude fiber and protein in traditional Chinese medicine residues. It also has antibacterial effects and can effectively inhibit diarrhea, infection and other problems caused by pathogenic bacteria such as Escherichia coli and Salmonella in the intestines of animals; after the compounding of Bacillus subtilis and yeast, they can be degraded into small molecular active substances after acting on the residues of liver-protecting tablets, enriching the dietary structure of poultry animals and improving palatability. At the same time, it allows microorganisms to colonize in the intestines, inhibit harmful bacteria, promote beneficial bacteria, regulate the structure of intestinal flora, improve intestinal health, stabilize the intestinal environment, and then promote the body's digestion and absorption of nutrients. After adding 3wt% of the fermented liver-protecting tablet residue feed additive obtained by the present invention to broiler feed and feeding for one week, the diarrhea symptoms of chicks improved and turned into normal-shaped feces.

[0030] 4. The present invention significantly improves the utilization rate of feed. Under the same feeding conditions, adding 1 to 5 wt% of the fermented liver protection tablet residue feed additive to broiler feed can reduce the amount of feed used during the feeding period and reduce the cost of poultry breeding. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 The contents of the main bioactive substances in the fermented liver protection tablets residue feed additives obtained in Example 3 and Comparative Examples 1 to 7, wherein a: polysaccharide content, b: polyphenol content, c: flavonoid content, d: saponin content;

[0032] Figure 2 This is a graph showing the effect of the fermented liver protection tablet residue feed additive obtained in Example 2 on the liver tissue structure of broiler chickens;

[0033] Figure 3 This is a graph showing the effect of the fermented liver protection tablet residue feed additive obtained in Example 2 on the kidney tissue structure of broiler chickens;

[0034] Figure 4 The gas chromatography analysis chart of the organochlorine pesticide content in the fermented liver protection tablets residue feed additive obtained in Example 2;

[0035] Figure 5 The gas chromatography analysis chart of the organophosphorus pesticide content in the fermented liver protection tablets residue feed additive obtained in Example 2;

[0036] Figure 6 This is an analytical graph of the pyrethroid insecticide content in the fermented liver protection tablet residue feed additive obtained in Example 2 detected by gas chromatography. DETAILED DESCRIPTION

[0037] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as limiting the present invention, but should be understood as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0038] It should be understood that the terms described in the present invention are only for describing a particular embodiment and are not intended to limit the present invention. In addition, for the numerical range in the present invention, it should be understood that each intermediate value between the upper and lower limits of the scope is also specifically disclosed. The intermediate value in any stated value or stated range, and each smaller range between any other stated value or intermediate value in the described range is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the scope.

[0039] Unless otherwise indicated, all technical and scientific terms used herein have the same meanings as those generally understood by those skilled in the art. Although the present invention describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may also be used in the implementation or testing of the present invention. All documents mentioned in this specification are incorporated by reference to disclose and describe the methods and / or materials associated with the documents. In the event of a conflict with any incorporated document, the content of this specification shall prevail.

[0040] It will be apparent to those skilled in the art that various modifications and variations may be made to the specific embodiments of the present invention description without departing from the scope or spirit of the present invention. Other embodiments derived from the present invention description will be apparent to those skilled in the art. The present invention description and examples are exemplary only.

[0041] The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

[0042] The liver-protecting tablets dregs used in the examples and comparative examples of the present invention are provided by Wuchang Sunflower Pharmaceutical Group Co., Ltd. in Heilongjiang Province, and include Artemisia capillaris, Bupleurum Root and Radix Isatidis, with a C / N of 19 and an initial moisture content of 80 wt %. The liver-protecting tablets dregs obtained from other sources do not affect the implementation of the technical solution of the present invention.

[0043] The live bacteria concentration of the Bacillus subtilis solution and the yeast solution used in the present invention is 1×10 8 CFU / mL.

[0044] The configuration of the Bacillus subtilis solution used in the examples and comparative examples of the present invention is as follows:

[0045] A single colony of Bacillus subtilis BU412 was inoculated into a 3 mL LB liquid culture medium shaking tube, cultured at 30°C, 170 r / min overnight, and inoculated into 300 mL LB liquid culture medium again at a volume ratio of 1:100, and cultured at 30°C, 170 r / min for 24 h to obtain a Bacillus subtilis liquid;

[0046] The Bacillus subtilis BU412 was provided by Heilongjiang Bayi Agricultural University, has been disclosed in the documents "CN108929866A" and "CN109096379A", and was deposited in China Center for Type Culture Collection on March 23, 2016, with the deposit number of CCTCC NO: M2016142;

[0047] The LB liquid culture medium was purchased from Qingdao Haibo Biotechnology Co., Ltd.

[0048] The configuration of the plant lactobacillus liquid used in the comparative example of the present invention is as follows:

[0049] A single colony of plant lactobacillus was inoculated into a conical flask containing 10 mL of MRS liquid culture medium and cultured in a constant temperature incubator at 37°C for 4 h. It was then inoculated again into 300 mL of MRS liquid culture medium at a volume ratio of 1:100 and cultured in a constant temperature incubator at 37°C for 48 h.

[0050] The plant lactobacillus was stored and provided by the Microbiology Laboratory of the School of Food Science and Engineering of Heilongjiang Bayi Agricultural University.

[0051] The configuration of the yeast solution used in the examples and comparative examples of the present invention is as follows:

[0052] Commercially available Angel Yeast, peptone and glucose were added into sterile water at concentrations of 7 g / 100 mL, 4 g / 100 mL and 4 g / 100 mL, respectively. After mixing well, the mixture was placed in a 37°C constant temperature incubator for fermentation for 72 h to obtain yeast solution.

[0053] The commercially available Angel Yeast was purchased from Angel Yeast Co., Ltd.

[0054] The fermentation bags used in the examples and comparative examples of the present invention were purchased from Pingyang County Hongxu Plastic Packaging Bag Business Department.

[0055] Unless otherwise specified, other raw materials used in the examples and comparative examples of the present invention are commercially available products.

[0056] The basic feed used for performance verification of the present invention is purchased from Harbin Baishicheng Animal Husbandry Co., Ltd., and the property indicators are crude protein content ≥21wt%, crude fiber content ≤6wt%, crude ash content ≤8wt%, calcium content 0.5-1.5wt%, total phosphorus content ≥0.6wt%, salt content 0.2-1wt%, methionine 0.45-0.9wt%, and lysine content 1.10wt%.

[0057] The Staphylococcus aureus ATCC29213, Escherichia coli ATCC25922, Salmonella xy202310, and Pseudomonas aeruginosa ATCC27853 used in the performance verification of the present invention are stored and provided by the Molecular Pharmacology Laboratory of the College of Life Science and Technology of Heilongjiang Bayi Agricultural University.

[0058] Example 1

[0059] The liver protection tablet residues are dried and sieved through a 60-mesh sieve, and the sieve residues are taken to obtain the liver protection tablet residues, which are placed in a fermentation bag, and then Bacillus subtilis liquid and yeast liquid are inoculated into the liver protection tablet residues to ensure that the dosage ratio of the liver protection tablet residues, Bacillus subtilis liquid and yeast liquid is 20g:9mL:15mL, and the mass ratio of Bacillus subtilis to yeast is 3:5. Fermentation is carried out for 144h (6d) under the conditions of anaerobic, temperature 25°C, water content 50wt%, and C / N of 25 to obtain a fermented liver protection tablet residue feed additive.

[0060] Example 2

[0061] The liver protection tablet residues are dried and sieved through a 60-mesh sieve, and the sieve residues are taken to obtain the liver protection tablet residues, which are placed in a fermentation bag, and then Bacillus subtilis liquid and yeast liquid are inoculated into the liver protection tablet residues to ensure that the dosage ratio of the liver protection tablet residues, the Bacillus subtilis liquid and the yeast liquid is 20g:9mL:15mL, and the mass ratio of Bacillus subtilis to the yeast is 3:5. The mixture is fermented for 127h under the conditions of anaerobic, temperature of 30°C, water content of 54wt%, and C / N of 25 to obtain a fermented liver protection tablet residue feed additive.

[0062] Example 3

[0063] The liver protection tablet residues are dried and sieved through a 60-mesh sieve, and the sieve residues are taken to obtain the liver protection tablet residues, which are placed in a fermentation bag, and then Bacillus subtilis liquid and yeast liquid are inoculated into the liver protection tablet residues to ensure that the dosage ratio of the liver protection tablet residues, Bacillus subtilis liquid and yeast liquid is 20g:9mL:15mL, and the mass ratio of Bacillus subtilis to yeast is 3:5. The liver protection tablet residues are fermented for 72h (3d) under the conditions of anaerobic, temperature of 35°C, water content of 60wt%, and C / N of 25 to obtain a fermented liver protection tablet residue feed additive.

[0064] Comparative Example 1

[0065] The difference from Example 3 is that no bacteria are added. Specifically, the moisture content of the liver protection tablet residue is adjusted to 60wt%, placed in a fermentation bag, and fermented for 72h (3d) under anaerobic conditions at a temperature of 35°C and a C / N of 25 to obtain a fermented liver protection tablet residue feed additive.

[0066] Comparative Example 2

[0067] The difference from Example 3 is that the Bacillus subtilis solution and the yeast solution are replaced with equal amounts of live bacteria with a concentration of 1×10 8 CFU / mL of plant acid lactobacillus liquid, and the rest is the same as in Example 3.

[0068] Comparative Example 3

[0069] The difference from Example 3 is that the Bacillus subtilis solution and the yeast solution are replaced with equal amounts of live bacteria with a concentration of 1×10 8 CFU / mL of Bacillus subtilis liquid, and the rest is the same as Example 3.

[0070] Comparative Example 4

[0071] The difference from Example 3 is that the Bacillus subtilis solution and the yeast solution are replaced with equal amounts of live bacteria with a concentration of 1×10 8 CFU / mL of yeast solution, and the rest is the same as in Example 3.

[0072] Comparative Example 5

[0073] The difference from Example 3 is that the yeast solution is replaced with an equal amount of live bacteria with a concentration of 1×10 8 CFU / mL of plant acid lactobacillus liquid, and the rest is the same as in Example 3.

[0074] Comparative Example 6

[0075] The difference from Example 3 is that the Bacillus subtilis solution is replaced with an equal amount of live bacteria with a concentration of 1×10 8 CFU / mL of plant acid lactobacillus liquid, and the rest is the same as in Example 3.

[0076] Comparative Example 7

[0077] The difference from Example 3 is that the concentration of live bacteria equal to that of the Bacillus subtilis solution is increased to 1×10 8 CFU / mL of plant acid lactobacillus liquid, and the rest is the same as in Example 3.

[0078] Comparative Example 8

[0079] The difference from Example 3 is that the temperature is adjusted to 20°C, specifically: the liver protection tablet residues are dried and sieved through a 60-mesh sieve, the sieve is taken to obtain the liver protection tablet residues, which are placed in a fermentation bag, and then the Bacillus subtilis solution and the yeast solution are inoculated into the liver protection tablet residues to ensure that the dosage ratio of the liver protection tablet residues, the Bacillus subtilis solution and the yeast solution is 20g:9mL:15mL, the mass ratio of Bacillus subtilis and the yeast is 3:5, and the fermentation is carried out under anaerobic conditions, temperature 20°C, moisture content 60wt%, C / N of 25 for 72h (3d) to obtain a fermented liver protection tablet residue feed additive.

[0080] Effect verification:

[0081] 1. The contents of the main bioactive substances (polysaccharides, polyphenols, flavonoids, saponins) in the fermented liver-protecting tablets residue feed additives obtained in Examples 1 to 3 and Comparative Examples 1 to 8 were tested, with 3 replicates for each sample. The results are as follows: Figure 1 And as shown in Table 1.

[0082] Figure 1 The contents of the main bioactive substances in the fermented liver protection tablets residue feed additives obtained in Example 3 and Comparative Examples 1 to 7, wherein a: polysaccharide content, b: polyphenol content, c: flavonoid content, d: saponin content; Figure 1 In the horizontal axis, A is comparative example 1, B is comparative example 2, C is comparative example 3, D is comparative example 4, E is comparative example 5, F is comparative example 6, G is embodiment 3, and H is comparative example 7. Figure 1It can be seen that the polysaccharide, polyphenol and flavonoid contents in the fermented liver-protecting tablets dregs feed additives obtained in other embodiments and comparative examples of adding strains compared to comparative example 1 (blank control group) have increased significantly (P<0.05), among which the effect of the Bacillus subtilis + yeast combination of Example 3 is the best, because there is mutualism and synergy between yeast and Bacillus subtilis. Yeast decomposes the dregs to produce soluble oligosaccharides and other metabolites, providing energy and substrates for Bacillus subtilis, providing suitable environmental conditions and metabolites, and promoting the growth and cellulose degradation of Bacillus subtilis. Bacillus subtilis decomposes complex polysaccharides and cellulose, produces organic acids and enzyme substances, and organic acids reduce pH values ​​to provide suitable acidity and alkalinity conditions for yeast, and Bacillus subtilis regulates oxygen concentration by consuming oxygen, provides a lower oxygen environment for yeast, and promotes the generation of products such as acid production and alcohol production. Yeast and Bacillus subtilis provide nutrients, enzymes and other auxiliary factors to each other, jointly promote growth and metabolic activity, and enhance the overall fermentation effect.

[0083] Table 1 Contents of main biologically active substances in the fermented liver protection tablets residue feed additives obtained in Examples 1 to 3, Comparative Example 1 and Comparative Example 8

[0084]

[0085] Table 1 shows the contents of the main bioactive substances in the fermented liver protection tablets residue feed additives obtained in Examples 1 to 3, Comparative Example 1 and Comparative Example 8. As can be seen from Table 1, the fermented liver protection tablets residue feed additives obtained in Examples 1 to 3 have increased contents of bioactive substances such as polysaccharides, polyphenols, and flavonoids compared to Comparative Example 1 and Comparative Example 8. Suitable temperature will promote bacterial metabolism and increase the catalytic efficiency and reaction rate of enzymes, promote the process of converting substrates into bioactive substances, and the content of bioactive substances will increase accordingly. This is because the cellulase produced by Bacillus subtilis can destroy plant cell walls and release compounds such as polysaccharides, polyphenols, flavonoids, and saponins. At the same time, enzymes such as amylase and protease can also hydrolyze various nutrients, further promoting biological transformation. These processes increase the content of polysaccharides, polyphenols, flavonoids, and saponins.

[0086] 2. The content of nutrients in the fermented liver protection tablet residue feed additives obtained in Examples 1 to 3 and Comparative Example 1 was tested, with 3 replicates for each sample. The results are shown in Table 2.

[0087] Table 2 Content of nutrients in the fermented liver protection tablets residue feed additives obtained in Examples 1 to 3 and Comparative Example 1

[0088]

[0089]

[0090] As shown in Table 2, the fermented liver-protecting tablets dregs feed additives obtained in Examples 1 to 3 have lower contents of pre-crude fiber, acid detergent fiber, and neutral detergent fiber than those in Comparative Example 1. This is because Bacillus subtilis decomposes part of the cellulose to generate sugars, thereby reducing the crude fiber content, which helps to reduce the fiber content of the raw material and makes it easier to digest and absorb. The crude fat, crude protein, and ash contents increased significantly. Crude fat, crude protein, and ash are the nutrients in the raw materials. Their increase indicates that the fermented liver-protecting tablets dregs feed additives obtained in Examples 1 to 3 can provide more nutrients.

[0091] 3. The heavy metal content and pesticide residue in the fermented liver protection tablets obtained in Example 2 were tested for 3 times. The results are shown in Tables 3 to 4 and Figures 4 to 6 shown.

[0092] The standards for heavy metal content and pesticide residues are based on the Feed Hygiene Standard GB 13078-2017.

[0093] Table 3 Heavy metal content in the fermented liver protection tablets residue feed additive obtained in Example 2

[0094] type unit standard result arsenic mg / kg ≤2.0 1.23±0.0510 lead mg / kg ≤10.0 2.09±0.0500 mercury mg / kg ≤0.1 0.01±0.0003 cadmium mg / kg ≤0.5 0.19±0.0040

[0095] It can be seen from Table 3 that the heavy metal contents in the fermented liver protection tablet residue feed additive obtained in Example 2 are all within the qualified range and meet the feed hygiene standard GB 13078-2017.

[0096] Table 4 Pesticide residues in the fermented liver protection tablets residue feed additives obtained in Example 2

[0097]

[0098]

[0099] Figure 4 The gas chromatography analysis chart of the organochlorine pesticide content in the fermented liver protection tablets residue feed additive obtained in Example 2; Figure 5 The gas chromatography analysis chart of the organophosphorus pesticide content in the fermented liver protection tablets residue feed additive obtained in Example 2; Figure 6 This is an analytical graph of the pyrethroid insecticide content in the fermented liver protection tablet residue feed additive obtained in Example 2 detected by gas chromatography.

[0100] From Table 4 and Figures 4 to 6 It can be seen that the content of the above-mentioned pesticides was not detected in the fermented liver protection tablet residue feed additive obtained in Example 2, which meets the requirements of the feed hygiene standard GB 13078-2017.

[0101] 4. The fermented liver protection tablet residue feed additive obtained in Example 2 and Comparative Example 1 was subjected to in vitro antibacterial test, in vivo broiler test and slaughter performance test.

[0102] (1) In vitro antibacterial experiment: Accurately weigh 100 mg of fermented liver protection tablets residue feed additive and add it to an EP tube containing 1 mL of ddH2O, mix well and set aside. Pour 20 mL of sterilized MH agar medium into a sterile culture dish, and inoculate 100 μL of 1×10 8 CFU / mL of bacterial solution (the bacterial species involved in the bacterial solution are Staphylococcus aureus ATCC29213, Escherichia coli ATCC25922, Salmonella xy202310 and Pseudomonas aeruginosa ATCC27853), spread evenly with a sterilized coating rod. Gently press the pre-sterilized Oxford cup onto the surface of the agar plate to ensure good contact with the agar. Add the fermented liver protection tablets residue feed additive mixture into the Oxford cup. Culture at 37°C for 24 hours and record the diameter (mm) of the circular inhibition zone formed around each well. The results are shown in Table 5.

[0103] Table 5 Antibacterial properties of fermented liver protection tablets residue feed additives obtained in Example 2 and Comparative Example 1 detected by Oxford cup method

[0104]

[0105]

[0106] As shown in Table 5, the fermented liver protection tablets residue feed additive obtained in Example 2 has a better inhibitory effect on Staphylococcus aureus, Escherichia coli, Salmonella and Pseudomonas aeruginosa than Comparative Example 1. The fermented liver protection tablets residue feed additive obtained in the present invention has a good inhibitory effect on Staphylococcus aureus, Escherichia coli, Salmonella and Pseudomonas aeruginosa.

[0107] (2) In vivo broiler experiment: 160 1-day-old AA white-feathered broilers were selected and adaptively raised for 3 days. They were divided into 4 groups according to the principle of average body weight (40±2g), namely the control group (fed with commercially available basic feed), the 1% fermented drug residue group (fed with commercially available basic feed and supplemented with 1% of the dry weight of the commercially available basic feed obtained in Example 2 as the fermented liver-protecting tablet drug residue feed additive), the 3% fermented drug residue group (fed with 3% of the dry weight of the commercially available basic feed added with 3% of the dry weight of the commercially available basic feed obtained in Example 2 as the fermented liver-protecting tablet drug residue feed additive), and the 5% fermented drug residue group (fed with 5% of the dry weight of the commercially available basic feed added with 5% of the dry weight of the commercially available basic feed obtained in Example 2 as the fermented liver-protecting tablet drug residue feed additive). Each group had 4 replicates, each replicate had 10 broilers, and the experimental period was 42 days. The experiment was divided into two stages, namely the chick stage (1st to 21st day) and the growth stage (22nd to 42nd day).

[0108] Determination of growth performance: At the end of the experiment, the broilers in each group were weighed and the feed intake of the broilers was recorded every day. The average daily weight gain, average daily feed intake and feed-to-weight ratio were calculated according to the following formula;

[0109] Average daily feed intake = (total feed intake / number of experimental broilers) / number of experimental days;

[0110] Average daily weight gain = (total weight gain / number of experimental broilers) / number of experimental days;

[0111] Feed-to-weight ratio = feed intake during the test period / weight gain during the test period.

[0112] The results are shown in Table 6.

[0113] Table 6 Effect of adding fermented liver protection tablets residue feed additive to feed on the growth performance of white-feathered broilers

[0114]

[0115] It can be seen from Table 6 that the fermented liver protection tablet residue feed additive obtained in Example 2 of the present invention can reduce the feed intake when added to broiler feed, but it also promotes weight gain. This is because the fermentation process can increase the bioavailability of the nutrients in the liver protection tablet residue and improve nutrient absorption. The fermented liver protection tablet residue contains probiotics and beneficial substances, which help to improve intestinal microecology, promote digestion and absorption, reduce intestinal diseases, and improve feed utilization.

[0116] (3) Slaughter performance test: On the 42nd day of the test, 2 broilers were randomly selected from 10 broilers in each replicate and their live weight was recorded. The broilers were killed by bleeding from the neck, feathers were plucked, and water was drained. The carcass weight, half-eviscerated weight, full-eviscerated weight, breast muscle weight, and leg muscle weight were weighed respectively. The slaughter rate, half-eviscerated rate, full-eviscerated rate, breast muscle rate, and leg muscle rate were calculated in accordance with "NY / T 823-2004 Terminology and Statistical Methods for Poultry Production Performance". The calculation formula is as follows:

[0117] Slaughter rate (%) = carcass weight (g) / live weight (g) × 100%;

[0118] Eviscerated rate (%) = Eviscerated weight (g) / carcass weight (g) × 100%;

[0119] Eviscerated rate (%) = eviscerated weight (g) / carcass weight (g) × 100%;

[0120] Breast muscle rate (%) = breast muscle weight (g) / eviscerated weight (g) × 100%;

[0121] Leg muscle rate (%) = leg muscle weight (g) / whole carcass weight (g) × 100%.

[0122] The results are shown in Table 7.

[0123] Table 7 Effect of adding fermented liver protection tablets residue feed additive to feed on slaughter performance of white-feathered broilers

[0124]

[0125] It can be seen from Table 7 that the fermented liver protection tablet residue feed additive obtained in Example 2 of the present invention is added to broiler feed to increase the live weight, carcass weight, whole carcass weight, half-clean carcass weight, breast muscle weight, whole leg weight and leg muscle weight of broilers, indicating that adding the fermented liver protection tablet residue feed additive obtained in the present invention to broiler feed can significantly increase the meat production performance of broilers.

[0126] (4) After the experiment, the liver tissue sections and kidney tissue sections of AA white-feathered broiler chickens in the control group (fed with a commercially available basic feed), the 1% fermented drug residue group (fed with a commercially available basic feed supplemented with 1% of the dry weight of the commercially available basic feed obtained in Example 2), the 3% fermented drug residue group (fed with a commercially available basic feed supplemented with 3% of the dry weight of the commercially available basic feed obtained in Example 2), and the 5% fermented drug residue group (fed with a commercially available basic feed supplemented with 5% of the dry weight of the commercially available basic feed obtained in Example 2) were observed. The results are as follows: Figure 2 and Figure 3 shown.

[0127] Figure 2 This is a graph showing the effect of the fermented liver protection tablet residue feed additive obtained in Example 2 on the liver tissue structure of broiler chickens; Figure 3 This is a graph showing the effect of the fermented liver-protecting tablet residue feed additive obtained in Example 2 on the kidney tissue structure of broiler chickens. Figure 2 It can be seen that the liver tissue structure of the control group and the group supplemented with the fermented liver protection tablet residue feed additive obtained in Example 2 is intact, the liver cords are distributed in a ray-like manner, and the liver cells are not edematous and degenerated, and are arranged neatly. Figure 3 It can be seen that the glomeruli, tubules and renal cells of the control group and the group adding the fermented liver protection tablets residue feed additive obtained in Example 2 were normal and had no morphological changes. The above results show that the fermented liver protection tablets residue feed additive obtained by the present invention has no obvious toxic reaction to the liver and kidney tissues of AA white-feathered broilers.

[0128] The various embodiments in this specification are described in a progressive manner, and each embodiment focuses on the differences from other embodiments. The same or similar parts between the various embodiments can be referenced to each other.

[0129] The above description of the disclosed embodiments enables one skilled in the art to implement or use the present invention. Various modifications to these embodiments will be apparent to one skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention will not be limited to the embodiments shown herein, but rather to the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A method for preparing a fermented liver protection tablet residue feed additive, characterized in that: The steps include: The liver protection tablet residues, Bacillus subtilis liquid and yeast liquid are mixed, inoculated and then cultured to obtain the fermented liver protection tablet residue feed additive.

2. The preparation method according to claim 1, characterized in that: The liver protection tablet residues are further dried and sieved before inoculation; and / or the mesh number of the liver protection tablet residues is ≥60 meshes.

3. The preparation method according to claim 1, characterized in that: The liver protection tablet residues are the residues remaining after effective components are extracted from Chinese medicinal materials during the production process of the liver protection tablets.

4. The preparation method according to claim 1, characterized in that: The dosage ratio of the liver protection tablet residue, Bacillus subtilis liquid and yeast liquid is 20g:3-15mL:3-15mL; and / or, the mass ratio of Bacillus subtilis in the Bacillus subtilis liquid and yeast in the yeast liquid is 1-5:1-5; and / or, the live bacteria concentration of the Bacillus subtilis liquid and the yeast liquid is independently 1.0×10 8 ~9.0×10 9 CFU / mL.

5. The preparation method according to claim 1, characterized in that: The culture conditions are set as follows: anaerobic conditions, a moisture content of 50-60 wt%, a temperature of 25-40° C., a C / N ratio of 25, and a time of 3-6 days.

6. The preparation method according to claim 5, characterized in that: The C / N value is regulated by glucose.

7. The fermented liver protection tablet residue feed additive prepared by the preparation method according to any one of claims 1 to 6.

8. Use of the fermented liver protection tablet residue feed additive according to claim 7 in preparing feed for poultry animals.

9. The use according to claim 8, characterized in that: The poultry animals include chickens and / or ducks; and / or the feed is a brooding feed or a growing feed.

10. The use according to claim 8, characterized in that: The amount of the fermented liver protection tablet residue feed additive added to the feed is 1-5wt%.

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