Herbal tea residue-containing Shiqi young pigeon functional feed and preparation method thereof

Abstract

The invention relates to the technical field of feed science and biology, and discloses a Shiqi young pigeon functional feed containing herbal tea residues and a preparation method of the Shiqi young pigeon functional feed, and the Shiqi young pigeon functional feed takes a biological modified herbal tea residue fiber carrier, anchored plant active matter and metabiotics, dormant state functional remodeling probiotics and galactooligosaccharide as main components. The preparation method comprises the following steps: carrying out synergistic solid-state fermentation on the herbal tea residue matrix by adopting lactobacillus plantarum and bacillus subtilis; adding a compound enzyme preparation into the material to perform targeted enzymolysis activation in a specific chemical microenvironment at a fermentation end point; and carrying out probiotic function remodeling treatment and low-temperature stabilization on the materials subjected to enzymolysis. Through multi-stage biological coupling treatment, cell walls of the herbal tea residues are effectively decomposed, antinutritional factors are degraded, and functional components such as probiotics, metabiotics and plant active matters are organically integrated into the same carrier. The product provided by the invention can significantly improve the growth performance, feed utilization rate and immune index of young pigeons.

Description

Technical Field

[0001] The invention relates to the fields of feed science and biotechnology, and in particular to a functional feed for Shiqi squab containing herbal tea residue and a preparation method thereof. Background Art

[0002] As a high-quality meat-producing poultry, the Shiqi squab industry places high demands on feed quality. Squab pigeons have a short growth cycle and a high metabolism. Furthermore, their digestive systems, particularly their intestinal microecological environments, are relatively fragile, making them highly sensitive to the nutritional value, safety, and digestibility of feed. Therefore, the development of functional feed additives that can improve feed utilization efficiency, enhance intestinal health, and enhance immunity is a current research hotspot in the pigeon farming industry.

[0003] Herbal tea is a traditional herbal beverage in South China. Its production process generates a large amount of plant residue, known as herbal tea slag. Rich in plant fiber and incompletely extracted active ingredients such as polyphenols and flavonoids, these residues hold great potential as a feed resource. However, current methods of utilizing herbal tea slag are relatively crude, typically simply drying and pulverizing them before adding them directly to feed as a filler. This approach presents significant technical drawbacks. The plant-based active ingredients in herbal tea slag are encapsulated within a dense cell wall structure composed of cellulose, hemicellulose, and lignin. Conventional physical pulverization is difficult to effectively break down these walls, resulting in extremely low dissolution and absorption efficiency of these functional ingredients in the pigeon's digestive tract, and low bioavailability. Furthermore, herbal tea slag naturally contains anti-nutritional factors such as tannins and phytic acid. These substances can impair the pigeon's absorption of key nutrients in feed, such as protein, and may reduce the feed's palatability, negatively impacting the pigeon's growth and development.

[0004] In order to overcome the above-mentioned defects, attempts have been made in the prior art to adopt biotechnology methods, such as separate microbial fermentation or enzymatic hydrolysis. Although these methods can improve the nutritional value of herbal tea residue to a certain extent, their treatment effects are still unsatisfactory. A simple solid-state fermentation process is difficult to achieve deep and controllable degradation of the complex plant cell wall structure, and the release of active substances is still insufficient. In contrast, if there is no effective pre-treatment to destroy the physical barrier of the cell wall, the enzyme action efficiency of the enzyme is low, the cost is high, and it is difficult to achieve effective degradation of antagonistic nutritional factors. In addition, even if the treated herbal tea residue is simply physically mixed with functional components such as probiotics, it cannot form an organic whole. There is a lack of effective synergistic mechanism between the components, and the survival rate of probiotics is low during storage and transportation and after entering the animal intestine. It is difficult to achieve synchronous release and functional complementarity with plant active substances at the site of action, thereby limiting the final application effect of the product. Summary of the Invention

[0005] In response to the shortcomings of the existing technology, the present invention provides a functional feed for Shiqi pigeons containing herbal tea residue and a preparation method thereof, which solves the problems of low bioavailability of active ingredients, difficulty in removing anti-nutritional factors, and inability to organically integrate and synergize functional components caused by traditional herbal tea residue processing methods.

[0006] To achieve the above objectives, the present invention is implemented through the following technical solutions: A functional feed for Shiqi pigeons containing herbal tea residues, comprising the following components: Bio-modified herbal tea residue fiber carrier: 80.0% to 95.0%; Anchored plant actives and postbiotics: 2.0% to 8.0%; Dormant function-remodeling probiotics: 1.0% to 5.0%; Galacto-oligosaccharides: 0.1% to 0.5%; Residual glycerin: 0.4% to 1.5%; Moisture: less than 8.0%.

[0007] Preferably, the bio-modified herbal tea residue fiber carrier is a carrier with a microporous structure derived from herbal tea residue and formed by microbial fermentation and enzymatic hydrolysis, and its surface is covered with in-situ generated lipopeptide biosurfactant.

[0008] Preferably, the dormant function-remodeling probiotics include Lactobacillus plantarum and Bacillus subtilis.

[0009] Preferably, the anchored plant actives and postbiotics comprise plant polyphenols and flavonoids dissociated from herbal tea residues, and organic acids and short-chain fatty acids produced by fermentation.

[0010] Preferably, the dormant functionally remodeled probiotics are probiotics that have been functionally induced with prebiotics.

[0011] A method for preparing a functional feed for Shiqi pigeons containing herbal tea residues comprises the following steps: S1. Collaborative solid-state fermentation step: fermenting the herbal tea residue using Lactobacillus plantarum and Bacillus subtilis capable of producing lipopeptide biosurfactants; S2. Targeted enzymatic activation step: After the fermentation is completed, a complex enzyme preparation is added to the material for enzymatic hydrolysis; The endpoint of the synergistic solid-state fermentation step is controlled by the ratio of organic acids to free polyphenols in the material reaching a specific threshold, so as to construct a chemical microenvironment that can activate the activity of the subsequent complex enzyme preparation; S3. Stabilization step: drying the enzymatically hydrolyzed material at low temperature to obtain the composition.

[0012] Preferably, the specific threshold value is a molar concentration ratio of organic acid to free polyphenols in the range of 2:1 to 5:1.

[0013] Preferably, in the targeted enzymatic hydrolysis activation step, an aqueous solution containing glycerol is added to the material, and the lipopeptide biosurfactant generated in situ in the fermentation step and glycerol are used to form an in situ emulsification-wetting system to improve the enzymatic hydrolysis efficiency.

[0014] Preferably, before the stabilization step, a probiotic function remodeling step is further included: adding galacto-oligosaccharides to the material and incubating at a specific temperature.

[0015] Preferably, the temperature of the collaborative solid-state fermentation is 35°C to 40°C; the temperature of the targeted enzymatic activation is 45°C to 50°C; and the incubation temperature of the probiotic function remodeling is 28°C to 32°C.

[0016] The present invention provides a functional feed for Shiqi pigeons containing herbal tea residue and a preparation method thereof. It has the following beneficial effects: 1. The present invention significantly improves the release rate of plant active substances from herbal tea residues by coupling synergistic solid-state fermentation with a subsequent targeted enzymatic activation step. Synergistic fermentation initially loosens the dense structure of plant cell walls, while the specific "acid-phenol" chemical microenvironment produced by fermentation creates highly active conditions for subsequent complex enzyme preparations, achieving efficient and deep degradation of the cellulose network, thereby fully releasing the encapsulated plant polyphenols, flavonoids, and other functional ingredients. This solves the problem of low bioavailability in the prior art due to the inability of physical pulverization to effectively break down the cell walls.

[0017] 2. The preparation method of the present invention can effectively degrade the anti-nutritional factors inherent in herbal tea residue and improve its palatability. During the synergistic solid-state fermentation step, the metabolic activities of the selected Lactobacillus plantarum and Bacillus subtilis can decompose and transform substances such as tannins and phytic acid that affect pigeon feeding and protein absorption. The organic acids such as lactic acid produced can also bring good flavor, thereby fundamentally improving the nutritional safety and application value of this raw material as a feed additive.

[0018] 3. This invention enhances the stability and functional efficiency of probiotics during feed storage and in the animal intestinal environment. On one hand, the bio-modified porous fiber carrier provides physical encapsulation and protection for the probiotics, improving their survival rate during storage and transportation. On the other hand, the unique probiotic functional remodeling step, through the use of galacto-oligosaccharides, pre-induces the probiotics' functionalities, shortening their adaptation period after entering the pigeon's intestines, allowing them to colonize more quickly and exert their effects on regulating the intestinal microecology.

[0019] 4. This invention achieves the organic integration and synergistic effect of multiple functional components through an integrated preparation process. The resulting composition is not a simple physical mixture of components, but rather integrates probiotics, postbiotics (fermentation metabolites), plant active ingredients, and prebiotics (galacto-oligosaccharides) into a single biological carrier. This structural coexistence enables the simultaneous release of each functional component at the same site of action in the animal intestine, promoting mutual synergy and achieving a comprehensive technical effect superior to that of each component added independently.

[0020] 5. The method design of the present invention improves the overall efficiency and control precision of the process. By utilizing the lipopeptide biosurfactant generated in situ during the fermentation step and exogenously added glycerol to form an emulsifying and wetting system, the mass transfer efficiency of the enzymatic hydrolysis reaction is greatly improved. Furthermore, by using the metabolites of the pre-fermentation to control and activate the subsequent enzymatic hydrolysis reaction, a close logical connection is established between the steps, making the automated control of the entire preparation process more convenient and effectively ensuring the quality uniformity between product batches. DETAILED DESCRIPTION

[0021] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the embodiments described are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of the present invention.

[0022] The sources and specifications of the main raw materials and reagents used in the following examples, comparative examples and test examples are as follows. Reagents not specifically specified are all commercially available products of analytical grade or higher.

[0023] Herbal tea residue: It is the mixed residue after water extraction of plant raw materials such as honeysuckle, licorice, and chrysanthemum, and then industrially dried.

[0024] Lactobacillus plantarum: strain number CICC6035, purchased from China Industrial Culture Collection Center (CICC).

[0025] Bacillus subtilis: strain number CICC20689, purchased from China Industrial Microbiology Culture Collection (CICC), has the ability to produce lipopeptide biosurfactants.

[0026] MRS liquid culture medium: purchased from Guangdong Huankai Microbiology Technology Co., Ltd., product number 023315.

[0027] LB liquid culture medium: purchased from Guangdong Huankai Microbiology Technology Co., Ltd., product number 028330.

[0028] Complex enzyme preparation: purchased from Novozymes (China) Investment Co., Ltd., it is a complex enzyme preparation containing cellulase, β-glucosidase and hemicellulase.

[0029] Glycerol: CAS number: 56-81-5, analytical grade, purchased from Sinopharm Chemical Reagent Co., Ltd.

[0030] Galacto-oligosaccharide: CAS number: 6587-31-1, food grade, purity ≥95%, purchased from Quantum Hi-Tech (China) Biotechnology Co., Ltd.

[0031] Gallic acid: CAS number: 149-91-7, standard product, purity ≥99%, purchased from Sigma-Aldrich Company, used for drawing the standard curve for the determination of total polyphenol content.

[0032] Folin-Ciocalteureagent: purchased from Merck, catalog number 1.09001.

[0033] Sodium carbonate: CAS number: 497-19-8, analytical grade, purchased from Sinopharm Chemical Reagent Co., Ltd.

[0034] Hydrochloric acid: CAS number: 7647-01-0, analytical grade, purchased from Sinopharm Chemical Reagent Co., Ltd.

[0035] Vanillin: CAS number: 121-33-5, analytical grade, purchased from Aladdin Reagent (Shanghai) Co., Ltd.

[0036] Basic daily diet: Prepared according to the recommended formula for meat pigeon production period in the agricultural industry standard of the People's Republic of China NY / T2119-2012 "Pigeon Feed". The main raw materials include corn, soybean meal, wheat, sorghum, etc.

[0037] Shiqi squab pigeons were purchased from Shiqi pigeon farm in Zhongshan City, Guangdong Province, 25 days old, weighing (350±20) g.

[0038] Animal ELISA kits: Pigeon immunoglobulin G (IgG), immunoglobulin A (IgA), and secretory immunoglobulin A (sIgA) ELISA kits were purchased from Nanjing Jiancheng Bioengineering Institute.

[0039] Example 1: This embodiment provides a specific preparation process of a functional feed core composition.

[0040] S1. Raw material pretreatment and bacterial activation: Take 1200g of herbal tea residue and dry it in a 65°C forced air drying oven for 12 hours to reduce the moisture content to less than 10%. Grind the dried herbal tea residue using an ultrafine grinder and pass it through a 100-mesh sieve. Inoculate the preserved Lactobacillus plantarum (CICC6035) into MRS liquid medium and incubate anaerobically at 37°C for 20 hours. Inoculate the preserved Bacillus subtilis (CICC20689) into LB liquid medium and incubate aerobically at 37°C at 160 rpm for 14 hours. Mix the two activated bacterial suspensions in a 1:1 volume ratio to prepare a composite bacterial suspension with a total viable count of no less than 1 × 109 × 109 CFU / mL.

[0041] S2. Collaborative solid-state fermentation: Take 1000g of pretreated herbal tea residue powder, add sterile water to adjust the moisture content of the material to 55% (w / w), and adjust the pH value to 6.2 with 1mol / LNaOH solution. Add 80mL of the composite bacterial suspension prepared above and mix evenly. Place the mixed material in a solid-state fermentation tank and perform anaerobic fermentation at 38°C. Starting from the 36th hour of fermentation, samples were taken every 4 hours, and the molar concentrations of total organic acids (lactic acid + acetic acid) and free polyphenols in the material were determined by high-performance liquid chromatography and Folin-phenol method. When the molar concentration ratio of the two reached 2.5:1, the fermentation was terminated.

[0042] S3. Targeted Enzymatic Activation: Add a sterile aqueous solution containing 1.0% (w / w) glycerol to the fermentation mixture, adjusting the total moisture content to 72% (w / w). Mix thoroughly. Subsequently, add 1.0% (w / w) of a complex enzyme preparation (Cellic® CTec2), based on the dry weight of the fermentation mixture. Raise the temperature of the mixture to 48°C and stir at 40 rpm / min for 7 hours.

[0043] S4. Probiotic Function Remodeling: After enzymatic hydrolysis, cool the material to 30°C. Add 0.3% (w / w) galacto-oligosaccharide (GOS) powder to the material, based on the current dry weight of the material. Gently stir until evenly combined, and incubate at this temperature for 1.5 hours.

[0044] S5. Low-temperature stabilization: Spread the remodeled material flat on a tray and prefreeze at -40°C for 4 hours. Then transfer it to a vacuum freeze dryer and dry it at a vacuum of less than 10 Pa. During the drying process, the plate temperature is gradually increased from -20°C to 25°C. At the end of drying, the moisture content of the material is less than 8%.

[0045] S6. Finished product: The dried material was crushed and passed through a 100-mesh sieve to obtain a light yellow, fluid powder product, designated as Product A.

[0046] Example 2: This embodiment provides another specific preparation process of a functional feed core composition.

[0047] Raw material pretreatment and bacterial activation: same as in Example 1.

[0048] Synergistic solid-state fermentation: Take 1000g of pretreated herbal tea residue powder, add sterile water to adjust the moisture content to 60% (w / w), and adjust the pH to 6.5 with 1 mol / L NaOH solution. Add 100mL of the composite bacterial suspension and mix thoroughly. Place the mixture in a solid-state fermentation tank and perform anaerobic fermentation at 40°C. Starting from the 36th hour of fermentation, samples are taken every 4 hours to determine the molar concentrations of total organic acids and free polyphenols in the material. Fermentation is terminated when the measured molar concentration ratio of the two reaches 4.5:1.

[0049] Targeted Enzymatic Activation: Add a sterile aqueous solution containing 1.2% (w / w) glycerol to the fermentation mixture, adjusting the total moisture content to 75% (w / w). Mix thoroughly. Then, add 1.2% (w / w) of a complex enzyme preparation (Cellic® CTec2), based on the dry weight of the fermentation mixture. Raise the temperature of the mixture to 50°C and stir at 40 rpm / min for 6 hours.

[0050] Probiotic Function Remodeling: After enzymatic hydrolysis, cool the material to 32°C. Add 0.4% (w / w) galacto-oligosaccharide (GOS) powder to the material based on the current dry weight of the material. After gentle mixing, incubate at this temperature for 1 hour.

[0051] Low temperature stabilization: same as Example 1.

[0052] Finished product: The dried material was crushed and passed through a 100-mesh sieve to obtain a light yellow powder with good fluidity, which was recorded as Product B.

[0053] Comparative Example 1 This comparative example prepared a powder of herbal tea residue without any biological treatment. Specifically, the herbal tea residue was subjected to the same drying and pulverization treatment as in step 1 of Example 1 to obtain a powdered product, designated as Product C.

[0054] Comparative Example 2 This comparative example prepared herbal tea residues subjected solely to solid-state fermentation. The specific procedures were as follows: After completing the "synergistic solid-state fermentation" step, the "targeted enzymatic activation" and "probiotic function remodeling" steps were omitted, and the fermented material was directly subjected to the "low-temperature stabilization" step to obtain a powdered product, designated Product D. The remaining steps and parameters were the same as in Example 1.

[0055] Comparative Example 3 This comparative example adopts the conventional "fermentation + enzymolysis" combined process to prepare the product. The specific operation is as follows: Compared with Example 1, the difference is: In the “cooperative solid-state fermentation” step, the endpoint is not controlled based on the ratio of organic acids to polyphenols, but the fermentation is directly fixed for 72 hours.

[0056] In the "targeted enzymatic activation" step, an equal volume of pure water (without glycerol) was used as the enzymatic medium instead of the glycerol aqueous solution. The remaining steps and parameters were the same as in Example 1. The resulting powdered product was designated Product E.

[0057] Comparative Example 4 This comparative example prepared a composition that was not subjected to the functional remodeling step. The specific operation was as follows: After completing the "targeted enzymatic activation" step, the "probiotic functional remodeling" step was omitted, and the enzymatically hydrolyzed material was directly subjected to the "low-temperature stabilization" step to obtain a powdered product, designated Product F. The remaining steps and parameters were the same as in Example 1.

[0058] Test Example 1: Determination of the dissolution rate of plant active ingredients This test example is intended to determine the total polyphenol dissolution rate of different products prepared in the examples and comparative examples under a simulated intestinal environment.

[0059] 1. Preparation of a standard curve: Accurately weigh the gallic acid standard and prepare a 1 mg / mL stock solution in deionized water. Aspirate appropriate amounts of the stock solution to prepare a series of standard working solutions with concentrations of 0, 10, 20, 40, 60, 80, and 100 μg / mL. Add 1.0 mL of each standard working solution to 5.0 mL of 10% (v / v) Folin-phenol reagent, mix thoroughly, and let stand for 5 minutes. Then, add 4.0 mL of 7.5% (w / v) sodium carbonate solution, mix thoroughly, and incubate at room temperature in the dark for 60 minutes. Measure the absorbance of each standard solution using a spectrophotometer at 765 nm. Plot a standard curve with gallic acid concentration as the horizontal axis and absorbance as the vertical axis to obtain a regression equation.

[0060] 2. Determination of Total Polyphenol Content in Samples: Accurately weigh 1.000 g each of Product A prepared in Example 1, Product B prepared in Example 2, Product C prepared in Comparative Example 1, Product D prepared in Comparative Example 2, and Product E prepared in Comparative Example 3 and place them in separate 100 mL Erlenmeyer flasks. Add 50 mL of pH 6.8 phosphate buffered saline (PBS) and shake in a constant-temperature water bath at 37°C and 150 rpm for 2 hours. After extraction, centrifuge the extract at 4000 rpm for 10 minutes, and collect the supernatant.

[0061] 3. Color Reaction and Calculation: Pipette 1.0 mL of the supernatant from each sample and perform the folin-phenol color reaction as described in step 1. Measure the absorbance. Substitute the measured absorbance into the standard curve regression equation to calculate the total polyphenol concentration in the supernatant. The total polyphenol dissolution rate is calculated using the following formula: Total polyphenol dissolution rate (%) = (total polyphenol concentration in supernatant × extract volume) / (sample mass) × 100% Each sample was tested three times and the average value was taken. The test results are recorded in Table 1.

[0062] Table 1 Comparison of total polyphenol dissolution rates of products under different treatment methods As can be seen from the test results in Table 1, the total polyphenol dissolution rates of products A and B prepared in Examples 1 and 2 were significantly higher than those of products C, D, and E prepared in Comparative Examples 1, 2, and 3. Product C is a herbal tea residue that has only been physically crushed. Its dense plant cell wall structure hinders the dissolution of internal polyphenols, resulting in the lowest dissolution rate. Product D has only been subjected to solid-state fermentation treatment. Although the initial decomposition of microorganisms has improved its dissolution rate compared to Product C, the effect is limited. Product E uses a conventional combination of fermentation and enzymolysis. Its dissolution rate is higher than that of Product D, but still lower than that of the products in the examples.

[0063] The preparation method provided by the present invention controls the fermentation endpoint during the coordinated solid-state fermentation step to achieve a specific "organic acid-free polyphenol" ratio, creating a favorable chemical reaction environment for the subsequent targeted enzymatic hydrolysis step. This specific chemical microenvironment can alter the molecular conformation of the subsequently added complex enzyme preparation, making its catalytic active sites more accessible to and effective on the cellulose substrate. This, compared to conventional enzymatic hydrolysis conditions, achieves a more thorough breakdown of the cell wall structure, allowing the release of a large number of physically embedded plant active substances.

[0064] Furthermore, during the targeted enzymatic hydrolysis step, this method adds glycerol to the medium and utilizes lipopeptide biosurfactants generated in situ during the fermentation process to form a highly efficient emulsifying and wetting system at the solid-liquid interface. This system reduces the surface tension of the enzymatic hydrolyzate, enhances its penetration into the microscopic pores within the fiber matrix, and improves the mass transfer efficiency between the enzyme and substrate. This dual mechanism, based on metabolite regulation and in situ mass transfer enhancement, works together to achieve a high level of total polyphenol dissolution rate in Products A and B, validating the effectiveness of the present method in enhancing the release of plant active ingredients.

[0065] Test Example 2: Determination of anti-nutritional factor (tannin) content This test example aims to determine the tannin content of different products prepared in the examples and comparative examples, in order to evaluate the degradation effect of the method of the present invention on the anti-nutritional factors in the herbal tea residue.

[0066] 1. Sample Preparation and Extraction: Accurately weigh 0.200 g each of Product A prepared in Example 1 and Product C prepared in Comparative Example 1 and place them in separate 50 mL centrifuge tubes. Add 20 mL of a 70% (volume fraction) methanol solution to each tube. Place the tubes in a 50°C water bath and shake at 150 rpm / min for 2 hours. After extraction, centrifuge the extract at 4000 rpm / min for 15 minutes, and collect the supernatant as the test solution.

[0067] 2. Color Reaction and Determination: Pipette 1.0 mL of each sample solution into a 10 mL stoppered test tube. Add 5.0 mL of vanillin-hydrochloric acid reagent (prepared freshly just before use by mixing equal volumes of 8% hydrochloric acid in methanol and 4% vanillin in methanol). Mix thoroughly and allow to react in the dark at 25°C for 20 minutes. After the reaction, immediately measure the absorbance at 500 nm using a spectrophotometer. Simultaneously, perform the same procedure using 70% methanol instead of the sample extract as a blank control.

[0068] 3. Content Calculation: Tannin content in the samples is measured by differences in absorbance. Higher absorbance values ​​indicate higher tannin content. Each sample was tested three times, and the average value was calculated. The test results are reported in Table 2.

[0069] Table 2 Comparison of tannin content of products under different treatment methods The test results in Table 2 show that the absorbance at 500 nm of Product A prepared in Example 1 is significantly lower than that of Product C prepared in Comparative Example 1. Since the absorbance is positively correlated with the tannin content in the sample, this result indicates that the tannin content in the herbal tea residue is significantly reduced after treatment with the method of the present invention.

[0070] Product C, made from untreated herbal tea waste, retains naturally occurring anti-nutritional factors such as tannins, resulting in a higher absorbance. High levels of tannins bind to protein in feed, reducing its nutritional value and affecting animal consumption due to their astringent taste.

[0071] In the preparation method provided by the present invention, collaborative solid-state fermentation is a key step in achieving tannin degradation. During this step, the selected Lactobacillus plantarum and Bacillus subtilis are able to grow and multiply on the herbal tea residue substrate and secrete various related degrading enzymes, such as tannase. The enzymes secreted by these microorganisms can hydrolyze the ester and glycosidic bonds in the tannin molecules, breaking them down into small molecules such as gallic acid and glucose, thereby degrading their anti-nutritional activity. Through the continuous fermentation process, the tannins in Product A are effectively degraded, verifying the effectiveness of the present method in improving the nutritional quality of raw materials and reducing anti-nutritional factors.

[0072] Test Example 3: In vitro simulation test of the efficiency of probiotics in utilizing prebiotics The purpose of this test case is to compare the growth ability of probiotics that have undergone a functional remodeling step and those that have not been treated with this step in a culture medium with galacto-oligosaccharides (GOS) as the sole carbon source through in vitro culture experiments.

[0073] 1. Preparation of Bacterial Suspensions: Accurately weigh 1.000 g each of Product A prepared in Example 1 and Product F prepared in Comparative Example 4 and place them in test tubes containing 9 mL of sterile saline. Vortex thoroughly to reconstitute the suspensions. Determine the total viable bacterial counts in both samples by serial dilution and plate count. Adjust the concentration of both suspensions to 1 × 107 to 1 × 107 CFU / mL using sterile saline.

[0074] 2. Culture and Assay: Prepare a basal liquid culture medium (composition: 10 g / L peptone, 5 g / L yeast extract, 5 g / L sodium chloride, 10 g / L GOS, pH 7.0) using GOS as the sole carbon source. Inoculate the culture medium with the adjusted concentrations of Product A and Product F at a 1% (v / v) inoculum. Incubate the culture in a 37°C incubator at 150 rpm / min. Starting at 0 hours, samples were taken every 2 hours and the optical density (OD600) of the culture medium was measured at 600 nm using a spectrophotometer to monitor bacterial growth. Each sample was assayed in triplicate, and the results were averaged. The test results are recorded in Table 3.

[0075] Table 3 Growth of probiotics in product A and product F in GOS medium The test results in Table 3 show that, in a culture environment with GOS as the sole carbon source, the OD600 value of the culture medium inoculated with Product A began to increase significantly after 2 hours, indicating a shorter growth hysteresis period. In contrast, the OD600 value of the culture medium inoculated with Product F increased slowly within the first 4 hours of culture, indicating a longer growth hysteresis period.

[0076] The probiotics in Product F were not treated with specific functional induction during the production process. When these dormant bacteria are placed in a new nutritional environment, their enzyme systems related to GOS metabolism (such as β-galactosidase) remain at basal expression levels. The bacteria need time to sense GOS in the environment, initiate transcription and translation of related genes, and synthesize sufficient enzymes to begin effectively utilizing this carbon source for proliferation. This process manifests itself as a prolonged period of growth stagnation on a macro scale.

[0077] The preparation method provided by the present invention includes a probiotic function remodeling step. In this step, by incubating the enzymatic hydrolysis material with GOS at a specific temperature before low-temperature stabilization, an environment is provided for the probiotics to pre-contact and adapt to GOS. This process allows the probiotics to synthesize and accumulate the enzymes necessary for utilizing GOS within their cells before entering dormancy. Therefore, when the probiotics in Product A encounter the GOS environment again, they can skip or significantly shorten the adaptation process of de novo enzyme synthesis, thereby achieving rapid proliferation, verifying the effectiveness of this functional remodeling step in improving the functional efficiency of probiotics.

[0078] Test Example 4: Animal feeding test This test example aims to evaluate the actual effects of the functional feed core composition prepared by the present invention on the growth performance and immune indicators of Shiqi pigeons through feeding experiments.

[0079] 1. Experimental design and grouping: 180 healthy, uniformly weighted (350 ± 20 g) 25-day-old Shiqi squabs were selected and randomly divided into three treatment groups, with 6 replicates in each group and 10 squabs in each replicate.

[0080] Control group: fed with basal diet.

[0081] Product C group: fed a diet containing 0.5% (w / w) of Product C (from Comparative Example 1) added to the basal diet.

[0082] Product A group: fed a diet supplemented with 0.5% (w / w) of Product A (from Example 1) in the basal diet.

[0083] 2. Feeding Management and Data Collection: The experiment was conducted under the same environmental and management conditions, with a pre-trial period of 3 days and a formal trial period of 28 days. During the trial, the pigeons had free access to food and water.

[0084] Growth performance measurement: At the beginning (day 1) and end (day 28) of the experiment, the fasting weight of each replicate was measured and the feed consumption was recorded to calculate the average daily gain (ADG), average daily feed intake (ADFI) and feed-to-gain ratio (FCR).

[0085] Immune Parameter Assays: On day 28 of the experiment, two squabs were randomly selected from each replicate. Blood was collected from the subwing vein, and serum was isolated. Serum immunoglobulin G (IgG) and immunoglobulin A (IgA) ELISA kits were used to measure the concentrations of the corresponding antibodies. After blood collection, the squabs were humanely sacrificed, and mid-jejunal tissue was obtained and homogenized. The intestinal mucosal secretory immunoglobulin A (sIgA) content was measured using a pigeon ELISA kit.

[0086] 3. Data processing: The test data are expressed as average values. The test results are recorded in Table 4.

[0087] Table 4 Effects of different additives on growth performance and immune indicators of Shiqi pigeons The test results in Table 4 show that compared with the control group and Product C group, feeding the diet supplemented with Product A significantly increased the average daily gain of Shiqi squabs and reduced the feed-to-gain ratio. Furthermore, the serum IgG and IgA concentrations, as well as the jejunal mucosal sIgA content, in the Product A group were higher than those in the control group and Product C group. There were no significant differences in any of the indicators in the Product C group compared with the control group, with some indicators even showing slight decreases.

[0088] The observed improvements in growth performance and feed utilization are directly related to the physicochemical properties of Product A. The preparation method provided by the present invention, through multi-stage biological treatment, achieves a higher dissolution rate and bioavailability of nutrients and active ingredients encapsulated by plant cell walls in herbal tea residue. Simultaneously, this method effectively degrades anti-nutritional factors, such as tannins, inherent in the raw materials. These two combined effects enable animals to more efficiently digest and absorb nutrients from the diet, resulting in higher weight gain and lower feed consumption.

[0089] The improved immune indicators in Group A are attributed to the multiple functional components incorporated within it. This combination of remodeled probiotics, prebiotic galacto-oligosaccharides, and postbiotics such as organic acids produced during fermentation, creates a synergistic effect within the animal's intestines. This synergistic effect regulates the balance of the intestinal microbiome, promotes the maturation of gut-associated lymphoid tissue, and enhances the intestinal mucosal barrier function and local immune response (as evidenced by increased sIgA levels), thereby influencing and enhancing the body's systemic humoral immunity (as evidenced by increased serum IgG and IgA concentrations).

[0090] While embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions, and variations may be made to these embodiments without departing from the principles and spirit of the invention, and that the scope of the invention is defined by the appended claims and their equivalents.

Claims

1. A functional feed for Shiqi pigeon containing herbal tea residue, characterized in that: Contains the following components: Bio-modified herbal tea residue fiber carrier: 80.0% to 95.0%; Anchored plant actives and postbiotics: 2.0% to 8.0%; Dormant function-remodeling probiotics: 1.0% to 5.0%; Galacto-oligosaccharides: 0.1% to 0.5%; Residual glycerin: 0.4% to 1.5%; Moisture: less than 8.0%.

2. The functional feed for Shiqi pigeons containing herbal tea residue according to claim 1, characterized in that: The bio-modified herbal tea residue fiber carrier is a carrier with a microporous structure derived from herbal tea residue and formed by microbial fermentation and enzymatic hydrolysis, and its surface is covered with in-situ generated lipopeptide biosurfactant.

3. The functional feed for Shiqi pigeons containing herbal tea residue according to claim 1, characterized in that: The dormant function remodeling probiotics include Lactobacillus plantarum and Bacillus subtilis.

4. The functional feed for Shiqi pigeons containing herbal tea residue according to claim 1, characterized in that: The anchored plant actives and postbiotics include plant polyphenols and flavonoids dissociated from herbal tea residues, and organic acids and short-chain fatty acids produced by fermentation.

5. The functional feed for Shiqi pigeons containing herbal tea residue according to claim 1, characterized in that: The dormant function-remodeling probiotics are probiotics that have undergone function-induction treatment with prebiotics.

6. A method for preparing a functional feed for Shiqi pigeons containing herbal tea residues, characterized in that: The functional feed for Shiqi pigeons containing herbal tea residues according to any one of claims 1 to 5 comprises the following steps: S1. Collaborative solid-state fermentation step: fermenting the herbal tea residue using Lactobacillus plantarum and Bacillus subtilis capable of producing lipopeptide biosurfactants; S2. Targeted enzymatic activation step: After the fermentation is completed, a complex enzyme preparation is added to the material for enzymatic hydrolysis; The endpoint of the synergistic solid-state fermentation step is controlled by the ratio of organic acids to free polyphenols in the material reaching a specific threshold, so as to construct a chemical microenvironment that can activate the activity of the subsequent complex enzyme preparation; S3. Stabilization step: drying the enzymatically hydrolyzed material at low temperature to obtain a composition.

7. The method for preparing a functional feed for Shiqi pigeons containing herbal tea residue according to claim 6, characterized in that: The specific threshold value is that the molar concentration ratio of organic acid to free polyphenols is in the range of 2:1 to 5:

1.

8. The method for preparing a functional feed for Shiqi pigeons containing herbal tea residue according to claim 6, characterized in that: In the targeted enzymatic hydrolysis activation step, an aqueous solution containing glycerol is added to the material, and the lipopeptide biosurfactant generated in situ in the fermentation step and glycerol are used to form an in situ emulsification-wetting system to improve the enzymatic hydrolysis efficiency.

9. The method for preparing a functional feed for Shiqi pigeons containing herbal tea residue according to claim 6, characterized in that: Before the stabilization step, a probiotic function remodeling step is also included: adding oligosaccharides to the material and incubating at a specific temperature.

10. The method for preparing the functional feed for Shiqi pigeons containing herbal tea residue according to claim 9, characterized in that: The temperature of the collaborative solid-state fermentation is 35°C to 40°C; the temperature of the targeted enzymatic activation is 45°C to 50°C; and the incubation temperature of the probiotic function remodeling is 28°C to 32°C.

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