Poultry postbiotic preparation prepared by solid-state fermentation of traditional Chinese medicine by-products and preparation method of poultry postbiotic preparation

Abstract

The invention relates to the technical field of feed additives, and particularly discloses a poultry postbiotic preparation prepared by solid-state fermentation of traditional Chinese medicine byproducts and a preparation method of the poultry postbiotic preparation. The preparation method of the postbiotic preparation for poultry prepared by solid-state fermentation of the traditional Chinese medicine byproducts comprises the following steps: mixing the radix astragali byproducts, the honeysuckle leaf byproducts, soybean meal powder and sodium alginate; cooling the hydrothermal modified material, and adding composite cellulase, glucose oxidase and composite strains; performing three-stage solid-state fermentation on the inoculated material; and mixing the fermentation material with the modified montmorillonite, and carrying out pulsed electric field treatment inactivation. Sodium alginate is adopted to form a gel network in hydrothermal pre-modification to protect heat-sensitive active ingredients, compound cellulase and glucose oxidase are synchronously added to relieve catabolic repression, and modified montmorillonite is adopted for in-situ adsorption and pulsed electric field non-thermal inactivation, so that the heat-sensitive active ingredients are effectively protected. The stability of the functional components in the intestinal environment and the storage stability of the product are enhanced.

Description

Technical Field

[0001] This application relates to the field of feed additive technology, and more specifically, to poultry postbiotic preparations prepared by solid-state fermentation of traditional Chinese medicine by-products and their preparation methods. Background Technology

[0002] With the large-scale and intensive development of livestock and poultry farming, the intestinal health of poultry has received increasing attention. Metabiotics, as a general term for inactivated probiotics and their metabolites, have the functions of regulating intestinal flora, enhancing immune function, and improving the integrity of the intestinal barrier. Compared with live bacteria preparations, they have advantages such as good storage stability, resistance to high temperatures in feed pelleting, and no ecological risks of live bacteria release. They have become a research hotspot in the field of feed additives. Solid-state fermentation technology, with its characteristics of low energy consumption, low wastewater discharge, and wide availability of culture medium, is widely used in the preparation of feed additives.

[0003] Existing poultry postbiotic preparations are mostly made by mixing wheat bran, soybean meal and other auxiliary materials, then steaming them, inoculating them with microorganisms such as Bacillus subtilis, and then fermenting them in a solid state under natural ventilation conditions. Finally, they are dried and pulverized by hot air. However, it is difficult to meet the metabolic needs of aerobic and anaerobic bacteria during the fermentation process, which can easily lead to metabolic conflicts, affect the targeted enrichment of functional components, and result in a decline in the product's functional effects. Summary of the Invention

[0004] To address the problem that existing poultry postbiotic preparations are prone to metabolic conflicts, which affect the targeted enrichment of functional components and lead to a decline in product efficacy, this application provides a poultry postbiotic preparation prepared by solid-state fermentation of traditional Chinese medicine by-products and its preparation method.

[0005] The poultry post-biotic preparation prepared by solid-state fermentation of traditional Chinese medicine by-products and its preparation method provided in this application adopt the following technical solution:

[0006] In the first aspect, this application provides a method for preparing poultry post-biotic preparations by solid-state fermentation of traditional Chinese medicine by-products, using the following technical solution:

[0007] A method for preparing poultry post-biotic preparations from solid-state fermentation of traditional Chinese medicine by-products includes the following steps:

[0008] Astragalus by-products, honeysuckle leaf by-products, soybean meal powder and sodium alginate were mixed, and water was added for hydrothermal pre-modification to obtain hydrothermally modified materials.

[0009] After cooling the hydrothermally modified material, add compound cellulase, glucose oxidase and compound bacterial strain, mix them to obtain the inoculum material;

[0010] The inoculum material is subjected to three-stage solid-state fermentation to obtain fermented material;

[0011] The fermentation material was mixed with modified montmorillonite and then inactivated by pulsed electric field treatment to obtain inactivated material;

[0012] The inactivated material is dried and pulverized at low temperature to obtain a poultry postbiotic preparation.

[0013] By adopting the above technical solution, the use of sodium alginate in hydrothermal pre-modification to form a gel network to protect heat-sensitive active ingredients, the simultaneous addition of compound cellulase and glucose oxidase to relieve catabolism inhibition, the use of a three-stage solid-state fermentation incorporating anaerobic pulses to achieve a smooth transition in microbial metabolism, and the use of modified montmorillonite for in-situ adsorption and non-thermal inactivation by pulsed electric fields, can increase the enrichment of short-chain fatty acids and extracellular polysaccharides, inhibit the formation of biogenic amines, enhance the stability of functional ingredients in the intestinal environment, and improve the storage stability of the product. This solves the problem that existing poultry postbiotic preparations are prone to metabolic conflicts, affecting the targeted enrichment of functional ingredients and leading to a decline in product efficacy.

[0014] Preferably, the solid-liquid ratio for hydrothermal pre-modification after adding water is 1:0.7~0.9, the temperature is 120~140℃, the pressure is 0.2~0.3MPa, and the holding time is 30~45min.

[0015] By adopting the above technical solution and utilizing hydrothermal pre-modification, the gel network formed by sodium alginate can encapsulate heat-sensitive components such as astragalus polysaccharide and chlorogenic acid, reducing high-temperature degradation losses. At the same time, hydrothermal conditions enable selective depolymerization of lignin and improve cellulose accessibility, thereby achieving a balance between protecting active ingredients and improving the fermentability of the matrix.

[0016] Preferably, the temperature after cooling is 25-35℃, the amount of compound cellulase added is 15-25 FPU / g dry basis, the amount of glucose oxidase added is 5-10 U / g dry basis, and the compound strain is composed of Bacillus subtilis, Lactobacillus acidophilus, and Aspergillus niger mixed at a live count ratio of 2-4:1:1-3, with a total inoculum of 5×10⁻⁶. 7 The mixing speed is 20~50 r / min and the mixing time is 5~15 min.

[0017] By adopting the above technical solution, and simultaneously adding compound cellulase and glucose oxidase, the cellulase directly hydrolyzes the exposed cellulose to produce fermentable sugars, and the glucose oxidase immediately converts glucose into gluconic acid, thereby relieving the metabolic inhibition and lowering the pH of the system. At the same time, the strain ratio ensures the metabolic synergy between cellulose-degrading bacteria and acid-producing bacteria.

[0018] Preferably, the three-stage solid-state fermentation specifically includes the following steps:

[0019] The inoculum was subjected to forced ventilation at a temperature of 30-32℃ and a ventilation ratio of 0.3-0.7vvm, with a relative humidity of 85%-90%, for 24 hours.

[0020] Then, at a temperature of 35-37℃, an anaerobic pulse was performed every 3-5 hours, during which CO2 was introduced at a flow rate of 0.1-0.3 vvm for 3-8 minutes. The compost was turned over during the time outside of the pulse period, and the treatment lasted for 24 hours.

[0021] Then, at a temperature of 35~42℃, in a completely sealed environment, CO2 or N2 is continuously introduced at a flow rate of 0.05~0.15 vvm for 24 hours to obtain fermented material.

[0022] By adopting the above technical solution, the three-stage fermentation promotes the rapid proliferation and enzyme production of Aspergillus niger and Bacillus subtilis in the aerobic stage, achieves a smooth transition of microbial metabolism in the anaerobic pulse stage so that Lactobacillus acidophilus can adapt to low oxygen conditions in advance, and completes the targeted enrichment of short-chain fatty acids in the strict anaerobic stage. This avoids metabolic conflicts between aerobic and anaerobic bacteria and achieves efficient accumulation of functional components.

[0023] Preferably, the field strength for the pulsed electric field treatment inactivation is 15~25kV / cm, the pulse width is 20~50μs, the pulse frequency is 10~30Hz, the treatment time is 5~15min, the material thickness is ≤2cm, and the material temperature during the treatment is ≤45℃.

[0024] By adopting the above technical solution, under the pulsed electric field conditions, the high-intensity electric field irreversibly destroys the microbial cell membrane structure to achieve complete inactivation. At the same time, since the treatment process does not produce a significant temperature rise, the degradation of heat-sensitive metabolites such as short-chain fatty acids and extracellular polysaccharides is avoided, thus ensuring the inactivation effect while preserving the functional components of the product to the greatest extent.

[0025] Preferably, the low-temperature drying temperature is 50~60℃, the vacuum degree is -0.08~-0.09MPa, and the drying is carried out until the moisture content is ≤10%, and the particle size after pulverization is 60~100 mesh.

[0026] By adopting the above technical solution, drying under low temperature vacuum conditions can effectively remove moisture without causing Maillard reaction or thermal degradation. Combined with appropriate particle size, the product has good flowability and dispersibility, which is conducive to uniform addition to feed and consumption by poultry.

[0027] Preferably, the Astragalus by-product is obtained by ethanol extraction and drying of Astragalus, with a water content ≤15%, and the Honeysuckle leaf by-product is obtained by water extraction and drying of Honeysuckle leaves, with a water content ≤15%.

[0028] By adopting the above technical solution and using a specific extraction process, the traditional Chinese medicine by-products obtained not only retain the active ingredients such as astragalus polysaccharide and chlorogenic acid, but also avoid the influence of extraction solvent residue on the fermentation process. At the same time, controlling the water content is beneficial to adjusting the moisture content of the material during the hydrothermal pre-modification process.

[0029] Preferably, the viscosity of the sodium alginate is 200~400 mPa·s, and the modified montmorillonite is sodium-based activated montmorillonite with a cation exchange capacity ≥90 mmol / 100g and a particle size ≤200 mesh.

[0030] By adopting the above technical solution, sodium alginate in this viscosity range can form a uniform and stable gel network under hydrothermal conditions, achieving effective encapsulation of heat-sensitive components. Sodium-based activated montmorillonite with high cation exchange capacity can adsorb metabolites through electrostatic interaction to form intercalation complexes, thereby playing a sustained-release role while protecting functional components.

[0031] Preferably, the weight proportions of the mixed astragalus by-product, honeysuckle leaf by-product, soybean meal powder and sodium alginate are: 40-60 parts of astragalus by-product, 20-30 parts of honeysuckle leaf by-product, 10-15 parts of soybean meal powder, and 0.5-1.5 parts of sodium alginate, and the amount of modified montmorillonite added is 5-8 parts by weight.

[0032] By adopting the above technical solutions, the functional components of Astragalus membranaceus by-products and Lonicera japonica leaf by-products complement each other, soybean meal powder adjusts the carbon-nitrogen ratio to a suitable range for microbial growth, the amount of sodium alginate added is sufficient to form a protective gel network, the amount of modified montmorillonite added can achieve full adsorption of metabolites, and the overall formula ensures the synergistic effect of all elements in the fermentation system.

[0033] Secondly, the poultry post-biotic preparation prepared by solid-state fermentation of traditional Chinese medicine by-products in this application adopts the following technical solution:

[0034] Poultry post-biotic preparations prepared by solid-state fermentation of traditional Chinese medicine by-products are obtained through the above-mentioned preparation method.

[0035] By adopting the above technical solution, the poultry postbiotic preparation obtained by this preparation method contains high levels of short-chain fatty acids and extracellular polysaccharides, low levels of biogenic amines, and high stability of functional components in the poultry intestinal environment. The product has a high activity retention rate during storage and has good application prospects.

[0036] In summary, this application has the following beneficial effects:

[0037] 1. Because this application uses sodium alginate to form a gel network to protect heat-sensitive active ingredients during hydrothermal pre-modification, and simultaneously adds compound cellulase and glucose oxidase to relieve catabolism inhibition, and employs a three-stage solid-state fermentation process including anaerobic pulses to achieve a smooth transition of microbial metabolism, and uses modified montmorillonite for in-situ adsorption and non-thermal inactivation by pulsed electric field, it can increase the enrichment of short-chain fatty acids and extracellular polysaccharides, inhibit the formation of biogenic amines, enhance the stability of functional ingredients in the intestinal environment and the storage stability of the product, and solve the problem that existing poultry postbiotic preparations are prone to metabolic conflicts, affecting the targeted enrichment of functional ingredients and leading to a decline in product functional effects.

[0038] 2. This application incorporates sodium alginate into the hydrothermal pre-modification process, utilizing the gel network formed under hydrothermal conditions to encapsulate heat-sensitive active ingredients such as astragalus polysaccharide and chlorogenic acid, reducing high-temperature degradation losses. Simultaneously, the hydrothermal conditions facilitate selective depolymerization of lignin and improve cellulose accessibility, providing high-quality substrates for subsequent enzymatic hydrolysis and fermentation. During the simultaneous saccharification and fermentation stage, the synergistic action of compound cellulase and glucose oxidase achieves rapid hydrolysis of cellulose and immediate conversion of released glucose into gluconic acid, effectively relieving catabolism inhibition and avoiding metabolic conflicts between enzymatic hydrolysis and fermentation in traditional processes. Furthermore, the anaerobic pulse regulation in the three-stage solid-state fermentation ensures a smooth transition between the metabolic activities of aerobic and anaerobic bacteria, achieving targeted enrichment of short-chain fatty acids and extracellular polysaccharides while keeping the content of biogenic amines at a low level.

[0039] 3. This application utilizes modified montmorillonite after fermentation for in-situ adsorption. Its layered structure adsorbs short-chain fatty acids, extracellular polysaccharides, and other metabolites into the interlayer to form intercalation complexes. This not only protects the metabolites from external environmental influences but also endows the product with sustained-release function, extending the duration of action of functional components in the poultry intestine. At the same time, it employs pulsed electric field non-thermal inactivation technology to replace traditional high-temperature inactivation. It uses a high-intensity electric field to irreversibly destroy the microbial cell membrane structure to achieve complete inactivation. No significant temperature rise is generated during the process, avoiding the thermal degradation of heat-sensitive components such as short-chain fatty acids and extracellular polysaccharides. Attached Figure Description

[0040] Figure 1 This is a flowchart of the preparation method of poultry postbiotic preparation prepared by solid-state fermentation of traditional Chinese medicine by-products provided in this application. Detailed Implementation

[0041] The present application will be further described in detail below with reference to the accompanying drawings and embodiments.

[0042] Technical concept: Existing poultry postbiotic preparations are mostly made by solid-state fermentation of auxiliary materials such as wheat bran and soybean meal with microorganisms, followed by hot air drying. However, the dense complex formed by lignin and cellulose in the by-products of traditional Chinese medicine constitutes a physical barrier to the microbial enzyme system, resulting in low cellulose degradation efficiency and low yield of metabolites. At the same time, it is difficult to meet the metabolic needs of aerobic and anaerobic bacteria in the traditional solid-state fermentation process, which is prone to metabolic conflicts. Furthermore, the hot air drying after fermentation will destroy heat-sensitive functional components such as short-chain fatty acids and extracellular polysaccharides, which seriously affects the functional effect of the product.

[0043] Based on the above findings, this application utilizes sodium alginate to form a gel network during hydrothermal pre-modification, which can protect heat-sensitive active ingredients such as astragalus polysaccharide and chlorogenic acid. Simultaneously, hydrothermal conditions enable selective depolymerization of lignin and improve cellulose accessibility. Adding glucose oxidase during the simultaneous saccharification and fermentation stage allows for the immediate conversion of glucose released by the enzyme into gluconic acid, thereby relieving catabolism inhibition. The use of a three-stage solid-state fermentation incorporating anaerobic pulses enables a smooth transition between aerobic and anaerobic bacterial metabolism. The use of modified montmorillonite for in-situ adsorption and pulsed electric field non-thermal inactivation protects heat-sensitive metabolites and achieves complete inactivation, thus solving the problem that existing poultry postbiotic preparations are prone to metabolic conflicts, affecting the targeted enrichment of functional components and leading to a decline in product efficacy.

[0044] Unless otherwise specified, all experimental methods used below are conventional methods. All materials, reagents, methods, and instruments used, unless otherwise specified, are conventional materials, reagents, methods, and instruments in this field, which can be obtained commercially or prepared according to literature methods by those skilled in the art.

[0045] To better understand the above technical solutions, the technical solutions of the present invention will be clearly and completely described below in conjunction with embodiments.

[0046] The "byproducts of traditional Chinese medicine" mentioned in this application refer to solid residues, scraps, offcuts, tailings, or waste generated during the extraction, processing, or cutting of traditional Chinese medicinal materials. These include, but are not limited to: dried residue remaining after alcohol extraction of Astragalus membranaceus, dried leaf residue remaining after water extraction of Lonicera japonica, and fragments, powders, and irregularly shaped lumps generated during the production of prepared slices of traditional Chinese medicine. These byproducts may be processed without further extraction or only with simple drying and pulverization, retaining some of the active ingredients of the original medicinal materials, and can be used as a solid-state fermentation substrate.

[0047] The following is a further description with reference to the embodiments:

[0048] Example 1: Please refer to the appendix Figure 1 A method for preparing poultry postbiotic preparations by solid-state fermentation of traditional Chinese medicine by-products includes the following steps:

[0049] Astragalus by-products, honeysuckle leaf by-products, soybean meal powder and sodium alginate were mixed, and water was added for hydrothermal pre-modification to obtain hydrothermally modified materials.

[0050] The solid-liquid ratio for hydrothermal pre-modification after adding water was 1:0.8, the temperature was 130℃, the pressure was 0.25MPa, and the holding time was 37.5min. The Astragalus by-product was obtained by ethanol extraction and drying of Astragalus, with a water content ≤15%. The Honeysuckle leaf by-product was obtained by water extraction and drying of Honeysuckle leaves, with a water content ≤15%. The viscosity of sodium alginate was 300 mPa·s. Viscosity was measured using a 1% aqueous solution at 20℃. The weight proportions of the mixed Astragalus by-product, Honeysuckle leaf by-product, soybean meal powder, and sodium alginate were: 50 parts Astragalus by-product, 25 parts Honeysuckle leaf by-product, 12.5 parts soybean meal powder, and 1 part sodium alginate.

[0051] After cooling the hydrothermally modified material, add compound cellulase, glucose oxidase and compound bacterial strain, mix them to obtain the inoculum material;

[0052] The temperature after cooling was 30℃, the amount of compound cellulase added was 20 FPU / g dry basis, the amount of glucose oxidase added was 7.5 U / g dry basis, and the compound strain was composed of Bacillus subtilis, Lactobacillus acidophilus and Aspergillus niger mixed in a live count ratio of 3:1:2, with a total inoculum of 5×10⁻⁶ cells / g. 7 The mixture was prepared at a stirring speed of 35 r / min for 10 min, with a dry basis of CFU / g.

[0053] The inoculum material is subjected to three-stage solid-state fermentation to obtain fermented material;

[0054] The three-stage solid-state fermentation process includes the following steps:

[0055] The inoculum was treated at 31°C with forced ventilation at a ventilation ratio of 0.5vvm and a relative humidity of 87.5% for 24 hours.

[0056] Then, at a temperature of 36℃, an anaerobic pulse was performed every 4 hours, during which CO2 was introduced at a flow rate of 0.2 vvm for 5.5 minutes. The pile was turned over during the time outside of the pulse period, and the treatment lasted for 24 hours.

[0057] Then, at a temperature of 38.5℃, in a completely sealed environment, CO2 or N2 is continuously introduced at a flow rate of 0.1 vvm for 24 hours to obtain fermented material.

[0058] The fermentation material was mixed with modified montmorillonite and then inactivated by pulsed electric field treatment to obtain inactivated material;

[0059] The pulsed electric field inactivation treatment had an electric field strength of 20 kV / cm, a pulse width of 35 μs, a pulse frequency of 20 Hz, a treatment time of 10 min, a material thickness ≤ 2 cm, and a material temperature ≤ 45℃ during treatment. The modified montmorillonite was sodium-based activated montmorillonite with a cation exchange capacity ≥ 90 mmol / 100 g and a particle size ≤ 200 mesh. The amount of modified montmorillonite added was 6.5 parts by weight.

[0060] The inactivated material is dried and pulverized at low temperature to obtain a poultry postbiotic preparation.

[0061] The low-temperature drying process involves a temperature of 55℃, a vacuum degree of -0.085MPa, and drying until the moisture content is ≤10%, resulting in a particle size of 75 mesh after pulverization.

[0062] Poultry post-biotic preparations prepared by solid-state fermentation of traditional Chinese medicine by-products are obtained through the above-mentioned preparation method.

[0063] Example 2: This example differs from Example 1 above in that:

[0064] A method for preparing poultry post-biotic preparations from solid-state fermentation of traditional Chinese medicine by-products includes the following steps:

[0065] Astragalus by-products, honeysuckle leaf by-products, soybean meal powder and sodium alginate were mixed, and water was added for hydrothermal pre-modification to obtain hydrothermally modified materials.

[0066] The solid-liquid ratio for hydrothermal pre-modification after adding water was 1:0.7, the temperature was 120℃, the pressure was 0.2MPa, and the holding time was 30min. The Astragalus by-product was obtained by ethanol extraction and drying of Astragalus, with a water content ≤15%. The Honeysuckle leaf by-product was obtained by water extraction and drying of Honeysuckle leaves, with a water content ≤15%. The viscosity of sodium alginate was 200 mPa·s. Viscosity was measured using a 1% aqueous solution at 20℃. The weight proportions of the mixed Astragalus by-product, Honeysuckle leaf by-product, soybean meal powder, and sodium alginate were: 40 parts Astragalus by-product, 20 parts Honeysuckle leaf by-product, 10 parts soybean meal powder, and 0.5 parts sodium alginate.

[0067] After cooling the hydrothermally modified material, add compound cellulase, glucose oxidase and compound bacterial strain, mix them to obtain the inoculum material;

[0068] The temperature after cooling was 25℃, the amount of compound cellulase added was 15 FPU / g dry basis, the amount of glucose oxidase added was 5 U / g dry basis, and the compound strain was composed of Bacillus subtilis, Lactobacillus acidophilus and Aspergillus niger mixed at a live count ratio of 2:1:1, with a total inoculum of 5×10⁻⁶ cells / g. 7 The mixture was prepared at a stirring speed of 20 r / min for 5 min, with a mixing time of 5 min.

[0069] The inoculum material is subjected to three-stage solid-state fermentation to obtain fermented material;

[0070] The three-stage solid-state fermentation process includes the following steps:

[0071] The inoculum was treated at 30°C with forced ventilation at a ventilation ratio of 0.3vvm and a relative humidity of 85% for 24 hours.

[0072] Then, at a temperature of 35℃, an anaerobic pulse was performed every 3 hours, during which CO2 was introduced at a flow rate of 0.1 vvm for 3 minutes. The pile was turned over during the time outside of the pulse period, and the treatment lasted for 24 hours.

[0073] Then, at a temperature of 35°C, in a completely sealed environment, CO2 or N2 is continuously introduced at a flow rate of 0.05 vvm for 24 hours to obtain fermented material.

[0074] The fermentation material was mixed with modified montmorillonite and then inactivated by pulsed electric field treatment to obtain inactivated material;

[0075] The pulsed electric field inactivation treatment had an electric field strength of 15 kV / cm, a pulse width of 20 μs, a pulse frequency of 10 Hz, a treatment time of 5 min, a material thickness ≤ 2 cm, and a material temperature ≤ 45℃ during treatment. The modified montmorillonite was sodium-based activated montmorillonite with a cation exchange capacity ≥ 90 mmol / 100 g and a particle size ≤ 200 mesh. The amount of modified montmorillonite added was 5 parts by weight.

[0076] The inactivated material is dried and pulverized at low temperature to obtain a poultry postbiotic preparation.

[0077] The low-temperature drying process involves a temperature of 50℃, a vacuum degree of -0.08MPa, and drying until the moisture content is ≤10%, with a particle size of 60 mesh after pulverization.

[0078] Poultry post-biotic preparations prepared by solid-state fermentation of traditional Chinese medicine by-products are obtained through the above-mentioned preparation method.

[0079] Example 3: This example differs from Example 1 above in that:

[0080] A method for preparing poultry post-biotic preparations from solid-state fermentation of traditional Chinese medicine by-products includes the following steps:

[0081] Astragalus by-products, honeysuckle leaf by-products, soybean meal powder and sodium alginate were mixed, and water was added for hydrothermal pre-modification to obtain hydrothermally modified materials.

[0082] The solid-liquid ratio for hydrothermal pre-modification after adding water was 1:0.9, the temperature was 140℃, the pressure was 0.3MPa, and the holding time was 45min. The Astragalus by-product was obtained by ethanol extraction and drying of Astragalus, with a water content ≤15%. The Honeysuckle leaf by-product was obtained by water extraction and drying of Honeysuckle leaves, with a water content ≤15%. The viscosity of sodium alginate was 400 mPa·s. Viscosity was measured using a 1% aqueous solution at 20℃. The weight proportions of the mixed Astragalus by-product, Honeysuckle leaf by-product, soybean meal powder, and sodium alginate were: 60 parts Astragalus by-product, 30 parts Honeysuckle leaf by-product, 15 parts soybean meal powder, and 1.5 parts sodium alginate.

[0083] After cooling the hydrothermally modified material, add compound cellulase, glucose oxidase and compound bacterial strain, mix them to obtain the inoculum material;

[0084] The temperature after cooling was 35℃, the amount of compound cellulase added was 25 FPU / g dry basis, the amount of glucose oxidase added was 10 U / g dry basis, and the compound strain was composed of Bacillus subtilis, Lactobacillus acidophilus and Aspergillus niger mixed in a live count ratio of 4:1:3, with a total inoculum of 5×10⁻⁶ cells / g. 7 The mixture was prepared at a stirring speed of 50 r / min for 15 min, with a mixing time of CFU / g dry basis.

[0085] The inoculum material is subjected to three-stage solid-state fermentation to obtain fermented material;

[0086] The three-stage solid-state fermentation process includes the following steps:

[0087] The inoculum was treated at 32°C with forced ventilation at a ventilation ratio of 0.7vvm and a relative humidity of 90% for 24 hours.

[0088] Then, at a temperature of 37℃, an anaerobic pulse was performed every 5 hours, during which CO2 was introduced at a flow rate of 0.3 vvm for 8 minutes. The pile was turned over during the time outside of the pulse period, and the treatment lasted for 24 hours.

[0089] Then, at a temperature of 42℃, in a completely sealed environment, CO2 or N2 is continuously introduced at a flow rate of 0.15 vvm for 24 hours to obtain fermented material.

[0090] The fermentation material was mixed with modified montmorillonite and then inactivated by pulsed electric field treatment to obtain inactivated material;

[0091] The pulsed electric field inactivation treatment had an electric field strength of 25 kV / cm, a pulse width of 50 μs, a pulse frequency of 30 Hz, a treatment time of 15 min, a material thickness ≤ 2 cm, and a material temperature ≤ 45℃ during treatment. The modified montmorillonite was sodium-based activated montmorillonite with a cation exchange capacity ≥ 90 mmol / 100 g and a particle size ≤ 200 mesh. The amount of modified montmorillonite added was 8 parts by weight.

[0092] The inactivated material is dried and pulverized at low temperature to obtain a poultry postbiotic preparation.

[0093] The low-temperature drying process involves a temperature of 60℃, a vacuum degree of -0.09MPa, and drying until the moisture content is ≤10%, with a particle size of 100 mesh after pulverization.

[0094] Poultry post-biotic preparations prepared by solid-state fermentation of traditional Chinese medicine by-products are obtained through the above-mentioned preparation method.

[0095] Comparative Example 1: A method for preparing a poultry postbiotic preparation, comprising the following steps:

[0096] Wheat bran, soybean meal powder, and corn flour are mixed evenly in a weight ratio of 5:3:2 to obtain a mixed base material;

[0097] Add the mixed base material to deionized water at a solid-liquid ratio of 1:1.2, adjust the pH to 6.5, steam sterilize at 121℃ for 30 minutes, and cool to 30℃.

[0098] The activated Bacillus subtilis and lactic acid bacteria suspensions were inoculated into the sterilized substrate at an inoculation rate of 10% v / w, and stirred evenly. The viable count ratio of Bacillus subtilis to lactic acid bacteria suspensions was 1:1, and the total viable count was 1×10⁻⁶. 8 CFU / mL;

[0099] The inoculated material was placed in a solid fermentation tank, and the fermentation temperature was controlled at 32℃ and the relative humidity at 85%. Natural ventilation was used for fermentation for 72 hours, during which the material was turned over once every 12 hours.

[0100] After fermentation, the fermented material is dried with hot air at 60℃ until the moisture content is ≤10%, and then crushed through an 80-mesh sieve to obtain a post-biotic preparation for feed.

[0101] Comparative Example 2: This comparative example differs from Example 1 above in that:

[0102] Sodium alginate was not added during hydrothermal pre-modification. Everything else was the same as in Example 1.

[0103] Comparative Example 3: This comparative example differs from Example 1 above in that:

[0104] No glucose oxidase was added. Everything else was the same as in Example 1.

[0105] Comparative Example 4: This comparative example differs from Example 1 above in that:

[0106] In the three-stage solid-state fermentation, anaerobic pulse treatment is not performed; that is, continuous microaerobic conditions are used during the 24-48h stage, with natural turning of the compost, and no CO2 pulse treatment is applied. The rest is the same as in Example 1.

[0107] Comparative Example 5: This comparative example differs from Example 1 above in that:

[0108] The pulsed electric field inactivation was replaced with high-temperature inactivation, that is, after fermentation, the material was kept at 80°C for 30 minutes for thermal inactivation. The rest was the same as in Example 1.

[0109] Performance testing: The following performance tests were conducted on the poultry postbiotic preparations prepared in Examples 1-3 and Comparative Examples 1-5.

[0110] Determination of short-chain fatty acid content: The total content of short-chain fatty acids in fermentation products, including acetic acid, propionic acid, and butyric acid, was determined by gas chromatography. 2g of sample was weighed, 10mL of deionized water was added, and the mixture was ultrasonically extracted for 30min. The supernatant was collected by centrifugation, filtered through a 0.22μm filter membrane, and then injected for analysis. Chromatographic conditions: 30m × 0.25mm × 0.25μm FFAP capillary column; injection port temperature 250℃; detector temperature 250℃; temperature program: initial temperature 80℃ held for 1min, then increased to 200℃ at 10℃ / min and held for 5min. Quantification was performed using external standard method. The results are expressed as milligrams of short-chain fatty acids per gram of dry sample.

[0111] Extracellular polysaccharide content determination: The phenol-sulfuric acid method was used to determine the extracellular polysaccharide content in the fermentation product. 1 g of sample was weighed, 10 mL of deionized water was added, and the mixture was extracted in an 80 °C water bath for 2 h. The supernatant was collected by centrifugation, 3 times the volume of anhydrous ethanol was added, and the mixture was allowed to stand overnight at 4 °C. The precipitate was collected by centrifugation, and after redissolving in deionized water, the absorbance was measured at 490 nm using glucose as a standard. The extracellular polysaccharide content was calculated, and the results were expressed as milligrams of extracellular polysaccharide per gram of dry sample.

[0112] Determination of total biogenic amines: The total biogenic amines in the fermentation products, including putrescine, cadaverine, histamine, and tyramine, were determined by high performance liquid chromatography (HPLC). 2 g of sample was weighed, added to 10 mL of 0.1 mol / L hydrochloric acid solution, and extracted by ultrasonication for 30 min. The supernatant was collected by centrifugation, derivatized with dansyl chloride, and then injected for determination. Chromatographic conditions: 250 mm × 4.6 mm, 5 μm C18 column; mobile phase: 65:35 v / v methanol:water; flow rate: 1.0 mL / min; detection wavelength: 254 nm. The total biogenic amine content was calculated using the external standard method, and the results were expressed as milligrams of biogenic amines per kilogram of dry sample.

[0113] Poultry intestinal simulated digestion survival rate determination: The stability of functional components in postbiotic preparations in the poultry intestinal environment was determined by in vitro simulated digestion method; 1g of sample was weighed, 10mL of simulated gastric juice was added, and the mixture was shaken and digested at 37℃ for 2h, and the precipitate was collected by centrifugation; the precipitate was added to 10mL of simulated intestinal juice, and the mixture was shaken and digested at 37℃ for 4h, and the supernatant was collected by centrifugation. The residual amounts of short-chain fatty acids and extracellular polysaccharides were determined, and the activity retention rate was calculated; the simulated gastric juice contained 0.2% sodium chloride and 0.35% pepsin, and the pH was adjusted to 2.5; the simulated intestinal juice contained 0.68% potassium dihydrogen phosphate and 0.5% trypsin, and the pH was adjusted to 6.5.

[0114] Storage stability test: The samples prepared in Examples 1-3 and Comparative Examples 1-5 were packaged in aluminum foil composite bags and placed in a constant temperature and humidity chamber at 40°C and 75% relative humidity for accelerated storage for 6 months. The short chain fatty acid content was measured at 0 months and 6 months, and the activity retention rate was calculated.

[0115] The test results are shown in Table 1.

[0116] Table 1

[0117] Group Short-chain fatty acids mg / g extracellular polysaccharide mg / g Total biogenic amines mg / kg Intestinal mimic activity retention rate % % of activity retention after 6 months of storage Example 1 48.6 15.3 42 92.3 91.5 Example 2 45.2 14.1 48 90.1 89.8 Example 3 47.5 14.8 45 91.2 90.6 Comparative Example 1 18.3 6.2 256 68.5 62.3 Comparative Example 2 32.5 10.5 68 82.6 80.5 Comparative Example 3 28.6 9.8 58 78.3 75.2 Comparative Example 4 35.2 11.2 52 85.1 82.4 Comparative Example 5 42.1 13.5 48 75.6 68.2

[0118] As can be seen from Examples 1 to 3 and Comparative Examples 1 to 5, and Table 1, this application promotes the enrichment of short-chain fatty acids and extracellular polysaccharides through a combination of hydrothermal pre-modification, simultaneous glucose oxidase saccharification fermentation, anaerobic pulsed three-stage fermentation, and pulsed electric field inactivation. At the same time, it effectively inhibits the formation of biogenic amines and improves the stability and storage stability of functional components in the intestinal simulated environment.

[0119] As can be seen from Example 1 and Comparative Example 2, and Table 1, the addition of sodium alginate during the hydrothermal pre-modification process can protect the heat-sensitive active ingredients from high-temperature degradation. This protective agent forms a gel network under hydrothermal conditions, reducing the loss of functional polysaccharides and flavonoids, thereby providing a higher quality substrate for subsequent fermentation.

[0120] As can be seen from Example 1 and Comparative Example 3, and Table 1, the addition of glucose oxidase can convert the glucose released by the enzyme into gluconic acid in a timely manner, relieve the inhibition of catabolism on cellulase synthesis, and enable Aspergillus niger and Bacillus subtilis to continuously secrete cellulase, thereby improving the cellulose degradation efficiency and the yield of metabolites.

[0121] As can be seen from Example 1 and Comparative Example 4, and Table 1, the anaerobic pulse treatment in the three-stage solid-state fermentation can achieve a smooth transition of microbial metabolism. By creating a local anaerobic environment through intermittent CO2 replacement, Lactobacillus acidophilus can adapt to low oxygen conditions in advance, avoiding metabolic stress caused by the sudden switch in stage three, thereby ensuring the continuous and efficient acid production metabolism.

[0122] As can be seen from Example 1 and Comparative Example 5, and Table 1, pulsed electric field non-thermal inactivation can retain the functional components of fermentation products. Inactivation is achieved by irreversibly destroying the microbial cell membrane structure through the action of electric field, avoiding the damage of heat-sensitive short-chain fatty acids and extracellular polysaccharides caused by high temperature treatment, thereby maintaining the activity of the product during storage and application.

[0123] This specific embodiment is merely an explanation of this application and is not intended to limit it. After reading this specification, those skilled in the art can make modifications to this embodiment without contributing any inventive step, but such modifications are protected by patent law as long as they fall within the scope of the claims of this application.

Claims

1. A method for preparing poultry postbiotic preparations from solid-state fermentation of traditional Chinese medicine by-products, characterized in that, Includes the following steps: Astragalus by-products, honeysuckle leaf by-products, soybean meal powder and sodium alginate were mixed, and water was added for hydrothermal pre-modification to obtain hydrothermally modified materials. After cooling the hydrothermally modified material, add compound cellulase, glucose oxidase and compound bacterial strain, mix them to obtain the inoculum material; The inoculum material is subjected to three-stage solid-state fermentation to obtain fermented material; The fermentation material was mixed with modified montmorillonite and then inactivated by pulsed electric field treatment to obtain inactivated material; The inactivated material is dried and pulverized at low temperature to obtain a poultry postbiotic preparation.

2. The method for preparing poultry post-biotic preparations by solid-state fermentation of traditional Chinese medicine by-products according to claim 1, characterized in that: The solid-liquid ratio for hydrothermal pre-modification after adding water is 1:0.7~0.9, the temperature is 120~140℃, the pressure is 0.2~0.3MPa, and the holding time is 30~45min.

3. The method for preparing poultry post-biotic preparations by solid-state fermentation of traditional Chinese medicine by-products according to claim 1, characterized in that: The cooled temperature is 25-35℃, the amount of compound cellulase added is 15-25 FPU / g dry basis, the amount of glucose oxidase added is 5-10 U / g dry basis, and the compound strain is composed of Bacillus subtilis, Lactobacillus acidophilus, and Aspergillus niger mixed at a live count ratio of 2-4:1:1-3, with a total inoculum of 5×10⁻⁶. 7 The mixing speed is 20~50 r / min and the mixing time is 5~15 min.

4. The method for preparing poultry post-biotic preparations by solid-state fermentation of traditional Chinese medicine by-products according to claim 1, characterized in that: The three-stage solid-state fermentation process specifically includes the following steps: The inoculum was subjected to forced ventilation at a temperature of 30-32℃ and a ventilation ratio of 0.3-0.7vvm, with a relative humidity of 85%-90%, for 24 hours. Then, at a temperature of 35-37℃, an anaerobic pulse was performed every 3-5 hours, during which CO2 was introduced at a flow rate of 0.1-0.3 vvm for 3-8 minutes. The compost was turned over during the time outside of the pulse period, and the treatment lasted for 24 hours. Then, at a temperature of 35~42℃, in a completely sealed environment, CO2 or N2 is continuously introduced at a flow rate of 0.05~0.15 vvm for 24 hours to obtain fermented material.

5. The method for preparing poultry post-biotic preparations by solid-state fermentation of traditional Chinese medicine by-products according to claim 1, characterized in that: The pulsed electric field inactivation treatment has an electric field strength of 15~25kV / cm, a pulse width of 20~50μs, a pulse frequency of 10~30Hz, a treatment time of 5~15min, and a material thickness of ≤2cm, with the material temperature ≤45℃ during the treatment.

6. The method for preparing poultry post-biotic preparations by solid-state fermentation of traditional Chinese medicine by-products according to claim 1, characterized in that: The low-temperature drying temperature is 50~60℃, the vacuum degree is -0.08~-0.09MPa, and the drying is carried out until the moisture content is ≤10%. The particle size of the pulverized material is 60~100 mesh.

7. The method for preparing poultry post-biotic preparations by solid-state fermentation of traditional Chinese medicine by-products according to claim 1, characterized in that: The Astragalus by-product is obtained by ethanol extraction and drying of Astragalus, with a water content ≤15%. The Honeysuckle leaf by-product is obtained by water extraction and drying of Honeysuckle leaves, with a water content ≤15%.

8. The method for preparing poultry post-biotic preparations by solid-state fermentation of traditional Chinese medicine by-products according to claim 1, characterized in that: The viscosity of the sodium alginate is 200~400 mPa·s, and the modified montmorillonite is sodium-based activated montmorillonite with a cation exchange capacity ≥90mmol / 100g and a particle size ≤200 mesh.

9. The method for preparing poultry post-biotic preparations by solid-state fermentation of traditional Chinese medicine by-products according to claim 1, characterized in that: The weight proportions of the mixture of Astragalus membranaceus by-product, honeysuckle leaf by-product, soybean meal powder and sodium alginate are as follows: 40-60 parts of Astragalus membranaceus by-product, 20-30 parts of honeysuckle leaf by-product, 10-15 parts of soybean meal powder, and 0.5-1.5 parts of sodium alginate. The amount of modified montmorillonite added is 5-8 parts by weight.

10. A poultry post-biotic preparation prepared by solid-state fermentation of traditional Chinese medicine by-products, characterized in that: It is prepared by the preparation method according to any one of claims 1-9.

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