Biological feed containing microbial microcapsule preparation and preparation method of biological feed

Abstract

The invention belongs to the field of feeds, and discloses a biological feed containing a microbial microcapsule preparation and a preparation method of the biological feed. The biological feed is prepared from the following components in parts by weight: 90 to 93 parts of wall-broken rhodopseudomonas palustris powder, 2 to 3 parts of lysine hydrochloride, 0.1 to 0.5 part of methionine, 0.1 to 0.5 part of threonine, 0.3 to 1 part of arginine, 2 to 3 parts of calcium hydrophosphate, 1 to 2 parts of calcium carbonate, 0.5 to 1.2 parts of an antioxidant additive, 0.3 to 0.8 part of modified nano montmorillonite and 1 to 5 parts of a microbial microcapsule preparation. According to the biological feed, the wall-broken rhodopseudomonas palustris serves as basic nutrition and a main microorganism source of the feed and is matched with the compound probiotics of the microorganism micro-capsule preparation, the intestinal nutrition utilization rate is effectively increased, meanwhile, the antioxidant additive is added, and the problems that a single additive is narrow in function and prone to oxidation are solved; the modified nano montmorillonite is added to solve the problem of antagonism of toxin adsorption and mineral absorption.

Description

Technical Field

[0001] This invention belongs to the field of feed technology, specifically relating to a biological feed containing microbial microcapsule preparations and its preparation method. Background Technology

[0002] As the aquaculture industry transforms towards "green, large-scale, and efficient" practices, the market demand for bio-feed, as a core product to replace antibiotics and improve aquaculture efficiency, continues to grow. Functional bio-feeds containing probiotics, enzymes, and microbial cells are mainly used in niche markets such as piglets, broiler chickens, and high-value aquatic products (e.g., whiteleg shrimp and California bass). Current research and development of bio-feed focuses on three main directions: first, optimizing microbial preparations, such as using microencapsulation technology to improve the survival rate of probiotics; second, synergistic effects of nutritional components, improving digestibility and utilization through adjustments to the ratio of amino acids, minerals, and functional bacteria; and third, enhancing environmental compatibility, developing products that can reduce nitrogen and phosphorus emissions and lower water pollution levels.

[0003] In existing products, probiotics and minerals are prone to antagonistic reactions; calcium ions can damage the cell membranes of probiotics, leading to reduced probiotic activity. Furthermore, it is difficult to simultaneously achieve synergistic effects of nutrient absorption, disease resistance, and stress reduction. For example, products focused on growth promotion (high amino acid content) often experience rapid oxidative rancidity; products focused on environmental protection (low-nitrogen formula) often exhibit poor growth performance. In addition, processing and storage stability are poor. The high temperature and humidity during feed pelleting lead to insufficient probiotic survival rates. After six months of storage at room temperature, the peroxide value and acid value of the products generally exceed standards, requiring the addition of high doses of chemical antioxidants, which does not meet the requirements of green farming. Summary of the Invention

[0004] To address the shortcomings mentioned in the background art, the present invention aims to provide a biological feed containing microbial microcapsule preparations and its preparation method. The feed uses broken-cell wall Rhodopseudomonas palustris as the basic nutrient and main microbial source, combined with a compound probiotic microbial microcapsule preparation, effectively improving intestinal nutrient utilization. Simultaneously, the addition of antioxidant additives addresses the issues of narrow function and easy oxidation associated with single additives, while the addition of modified nano-montmorillonite addresses the antagonistic relationship between toxin adsorption and mineral absorption.

[0005] The objective of this invention can be achieved through the following technical solutions: A biological feed containing microbial microcapsule preparation comprises the following components in parts by weight: 90-93 parts of broken-cell wall Rhodopseudomonas palustris powder, 2-3 parts of lysine hydrochloride, 0.1-0.5 parts of methionine, 0.1-0.5 parts of threonine, 0.3-1 parts of arginine, 2-3 parts of dicalcium phosphate, 1-2 parts of calcium carbonate, 0.5-1.2 parts of antioxidant additive, 0.3-0.8 parts of modified nano-montmorillonite, and 1-5 parts of microbial microcapsule preparation; The antioxidant additive is chitosan, which is enzymatically hydrolyzed and then combined with plant polyphenols via ultrasound-assisted hydrogen bonding. The plant polyphenols are green tea polyphenols and rosemary polyphenols mixed in a mass ratio of 3:1. The modified nano-montmorillonite is produced by introducing sodium ions through a hydrothermal method to expand the interlayer spacing of montmorillonite and then loading probiotic activators in situ. The probiotic activators are oligosaccharides, vitamin B and citric acid in a ratio of 5:3:2. The microbial microcapsule preparation is a core-shell structure formed by coating chitosan on the surface of probiotic sodium alginate composite microspheres. The probiotic sodium alginate composite microspheres are prepared by emulsion solidification method of composite probiotics and sodium alginate. The composite probiotics are Bacillus subtilis, Lactobacillus plantarum and Lactobacillus acidophilus mixed in a mass ratio of 2:1:1.

[0006] Preferably, the preparation method of the antioxidant additive includes the following steps: A1. Add chitosan to deionized water and stir to dissolve, obtaining a chitosan solution. Adjust the pH to 4.5 with hydrochloric acid, add the complex enzyme, and react in a 50°C constant temperature water bath for 3-5 hours. Raise the temperature to 90°C and keep it for 10-20 minutes to inactivate the complex enzyme. After cooling, adjust the pH to 6.5 with sodium hydroxide, centrifuge to remove insoluble matter, and concentrate the supernatant by rotary evaporation to obtain an enzymatically hydrolyzed chitosan solution with a solid content of 20%. A2. Add plant polyphenols to the enzymatically hydrolyzed chitosan solution, add deionized water to adjust the solid content to 15%, stir until the plant polyphenols are dissolved, sonicate for 15-25 minutes, and spray dry the reaction solution to obtain an antioxidant additive.

[0007] Preferably, the compound enzyme is a mixture of cellulase and papain in a 2:1 mass ratio, and the amount of compound enzyme added is 1.5 to 2.5% of the mass of chitosan.

[0008] Preferably, the mass ratio of plant polyphenols to enzymatically hydrolyzed chitosan is 1 to 10.

[0009] Preferably, the preparation method of modified nano-montmorillonite includes the following steps: B1. Add montmorillonite to deionized water and disperse it by ultrasonication to obtain a montmorillonite suspension. Add hydrochloric acid solution to adjust the pH to 3, stir the reaction at 60℃ for 0.5~1.5h, centrifuge and filter, wash the precipitate with deionized water until neutral, and dry to obtain activated montmorillonite. B2. Activated montmorillonite was added to deionized water and ultrasonically dispersed to obtain an activated montmorillonite suspension. Sodium chloride was added and stirred to dissolve the suspension. The suspension was then added to a high-pressure reactor and stirred at 120°C and 0.2 MPa for 4-6 hours. After centrifugation, filtration, and drying, nano-montmorillonite was obtained. B3. Add nano-montmorillonite to deionized water and ultrasonically disperse to obtain nano-montmorillonite dispersion. Add oligofructose, vitamin B and citric acid to deionized water and stir to dissolve to obtain probiotic activator solution. Slowly add probiotic activator solution to nano-montmorillonite dispersion. Stir and react in a 50℃ constant temperature water bath for 2-4 hours. After the reaction is completed, freeze dry to obtain modified nano-montmorillonite.

[0010] Preferably, the mass ratio of activated montmorillonite to sodium chloride is 8:1, the mass ratio of nano-montmorillonite to probiotic activator is 10:1, and vitamin B is a mixture of vitamin B1, vitamin B2, vitamin B6, and vitamin B12 in a mass ratio of 2:1:1:0.5.

[0011] Preferably, the preparation method of the microbial microcapsule formulation includes the following steps: C1. Add the compound probiotic powder to physiological saline to prepare 1×10 9 CFU / mL bacterial suspension was mixed with glycerol and maltodextrin and stirred for 20-30 min. Sodium alginate was added to deionized water and stirred in a 50℃ water bath for 0.5-1.5 h to obtain a 2wt% sodium alginate solution. The compound probiotic suspension was mixed with the sodium alginate solution, and liquid paraffin and Span 80 were slowly added. The mixture was emulsified by high-speed shearing for 10-20 min to form a uniform W / O type emulsion. C2. Slowly add calcium chloride solution to the emulsion, stir and solidify at 4°C for 20-40 min, let stand for 8-12 min after the reaction is complete, collect the lower layer of microspheres, wash with deionized water 3-5 times to obtain sodium alginate probiotic microspheres. C3. Add chitosan powder to acetic acid solution, stir in a 50°C water bath for 0.5~1.5h to prepare chitosan solution, adjust pH to 5.5 with NaOH, add sodium alginate probiotic microspheres to chitosan solution, and stir magnetically at 4°C for 30~50min. C4. Slowly add glutaraldehyde solution to the system, continue stirring at 4°C for 10-30 min, let stand for 10-20 min, collect the microcapsules, wash with deionized water 3-5 times, and freeze-dry to obtain the microbial microcapsule preparation.

[0012] Preferably, the compound probiotics and sodium alginate solution are mixed at a volume ratio of 1:4, and the volume ratio of liquid paraffin to the mixed solution of compound probiotic suspension and sodium alginate solution is 3:1.

[0013] Preferably, the mass ratio of sodium alginate probiotic microspheres to chitosan is 10:1.

[0014] A method for preparing a bio-feed containing microbial microcapsule formulation includes the following steps: S1. First, add dicalcium phosphate and calcium carbonate to the twin-shaft paddle mixer and stir at low speed for 4-6 minutes. Then, add lysine hydrochloride, methionine, threonine, and arginine to the mixer at one time. Stir for 4-6 minutes after each amino acid is added. S2. Slowly add the broken cell wall Rhodopseudomonas sphaeroides powder while stirring. After adding all the powder, continue stirring for 8-12 minutes. Add the antioxidant additive and stir for 6-10 minutes. Then add the modified nano-montmorillonite and continue stirring for 6-10 minutes. Finally, add the microbial microcapsule preparation and stir at low speed for 3-5 minutes to obtain the mixed powder of all components. S3. Feed the mixed powder of all components into a low-temperature ring die pellet mill, introduce saturated steam at a pressure of 0.2~0.3MPa and a temperature of 85~90℃, condition for 30~40s, control the pellet mill speed at 300~350r / min for extrusion granulation, and air-cool dry to obtain biological feed containing microbial microcapsule preparation.

[0015] The beneficial effects of this invention are: This invention's biological feed uses broken-cell wall Rhodopseudomonas palustris as the basic nutrient and main microbial source, providing high concentrations of easily absorbed protein, vitamins, coenzyme Q10, etc. Combined with microbial microencapsulation preparations of compound probiotics, it can improve the intestinal flora structure, reduce nutrient loss, and effectively improve intestinal nutrient utilization. At the same time, the addition of antioxidant additives and modified nano-montmorillonite creates a stable external environment for all active ingredients, helping them to grow and function better.

[0016] This invention uses chitosan as a raw material and replaces traditional chemical degradation with a complex enzymatic hydrolysis (cellulase + papain) to reduce the molecular weight of chitosan to 5000~8000 Da (small molecules are easily absorbed) while retaining amino activity. Green tea polyphenols and rosemary polyphenols are selected and compounded, and ultrasonic-assisted hydrogen bonding is used to avoid chemical cross-linking that destroys activity, forming a complex structure of chitosan backbone-polyphenol active sites, which solves the problems of easy oxidation of single polyphenols and narrow antibacterial spectrum of chitosan.

[0017] This invention employs a hydrothermal method to modify natural montmorillonite, introducing sodium-based ions to widen the interlayer spacing and enhance the adsorption capacity for intestinal toxins (such as aflatoxin and heavy metal Pb²⁺); and adds a probiotic activator. The activator is a compound of oligosaccharides, vitamin B, and citric acid in a 5:3:2 ratio, and is embedded into the montmorillonite nanolayers using in-situ loading technology, avoiding antagonistic reactions with minerals in the feed and achieving synergistic adsorption and activation.

[0018] This invention focuses on high activity retention, strong stress resistance, and suitability for feed applications. It uses the chitosan-sodium alginate coagulation method and optimizes the encapsulation process parameters to construct microcapsules with a core-shell structure, thereby achieving the activity protection of probiotics in feed processing, storage, and animal digestive tract.

[0019] Of course, any product implementing this invention does not necessarily need to achieve all of the advantages described above at the same time. Detailed Implementation

[0020] The technical solutions of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of the present invention.

[0021] Example 1 An antioxidant additive is prepared by combining chitosan with plant polyphenols via enzymatic hydrolysis and ultrasound-assisted hydrogen bonding. The plant polyphenols are green tea polyphenols and rosemary polyphenols mixed in a mass ratio of 3:1. The preparation method includes the following steps: A1. Add 10g of chitosan to 490mL of deionized water and stir to dissolve, obtaining a 2wt% chitosan solution. Adjust the pH to 4.5 with hydrochloric acid, add 0.13g of cellulase and 0.07g of papain, react in a 50℃ constant temperature water bath for 4h, raise the temperature to 90℃ and keep warm for 15min to inactivate cellulase and papain, cool and adjust the pH to 6.5 with sodium hydroxide, centrifuge to remove insoluble matter, and concentrate the supernatant by rotary evaporation to obtain an enzymatically hydrolyzed chitosan solution with a solid content of 20%. A2. Add 1.5g of green tea polyphenols and 0.5g of rosemary polyphenols to 100mL of enzymatically hydrolyzed chitosan solution, add deionized water to adjust the solid content to 15%, stir until the green tea polyphenols and rosemary polyphenols are dissolved, sonicate for 20min, and spray dry the reaction solution to obtain the antioxidant additive.

[0022] Example 2 A modified nano-montmorillonite is prepared by introducing sodium ions via a hydrothermal method to expand the interlayer spacing of montmorillonite, followed by in-situ loading of a probiotic activator. The probiotic activator is a compound of oligosaccharides, vitamin B, and citric acid in a 5:3:2 ratio. The citric acid is prepared by the following steps: B1. Add 50g of montmorillonite to 450mL of deionized water and disperse it by ultrasonication to obtain a 10wt% montmorillonite suspension. Add hydrochloric acid solution to adjust the pH to 3, stir and react at 60℃ for 1h, centrifuge and filter, wash the precipitate with deionized water until neutral, and dry to obtain activated montmorillonite. B2. Add 40g of activated montmorillonite to 460g of deionized water and disperse by ultrasonication to obtain an 8wt% activated montmorillonite suspension. Add 6g of sodium chloride, stir to dissolve, and then add to a high-pressure reactor. Stir and react at 120℃ and 0.2MPa for 5h. After centrifugation, filtration, and drying, nano-montmorillonite is obtained. B3. Add 45g of nano-montmorillonite to 255mL of deionized water and ultrasonically disperse to obtain a 15wt% nano-montmorillonite dispersion. Add 2.25g of fructooligosaccharide, 1.35g of vitamin B (vitamin B1:B2:B6:B12=2:1:1:0.5) and 0.9g of citric acid to 11.5mL of deionized water and stir to dissolve to obtain a probiotic activator solution. Slowly add the probiotic activator solution to the nano-montmorillonite dispersion, and stir in a 50℃ constant temperature water bath for 3h. After the reaction is completed, freeze dry to obtain the modified nano-montmorillonite.

[0023] Example 3 A microbial microcapsule formulation comprises a core-shell structure formed by coating chitosan onto the surface of probiotic sodium alginate composite microspheres. The probiotic sodium alginate composite microspheres are prepared by emulsion solidification of composite probiotics and sodium alginate. The composite probiotics are a mixture of Bacillus subtilis, Lactobacillus plantarum, and Lactobacillus acidophilus in a 2:1:1 mass ratio. The preparation method includes the following steps: C1. A compound probiotic powder consisting of 6.67g Bacillus subtilis, 3.33g Lactobacillus plantarum, and 3.33g Lactobacillus acidophilus was added to 100mL of physiological saline to prepare a 1×10⁻⁶ solution. 9 CFU / mL bacterial suspension was mixed with 0.25g glycerol and 0.25g maltodextrin and stirred for 25min. 8g sodium alginate was added to 392ml deionized water and stirred in a 50℃ water bath for 1h to obtain a 2wt% sodium alginate solution. The compound probiotic suspension and sodium alginate solution were mixed at a volume ratio of 1:4. 1200ml liquid paraffin and 20g Span 80 were slowly added and emulsified at high speed for 15min to form a uniform W / O type emulsion. C2. Slowly add calcium chloride solution to the emulsion, stir and solidify at 4°C for 30 min, let stand for 10 min after the reaction is complete, collect the lower layer of microspheres, wash with deionized water 3-5 times to obtain sodium alginate probiotic microspheres. C3. Add 1g of chitosan powder to 100mL of 1wt% acetic acid solution, stir in a water bath at 50℃ for 1h to prepare a chitosan solution, adjust the pH to 5.5 with NaOH, add 10g of sodium alginate probiotic microspheres to 100mL of chitosan solution, and stir magnetically at 4℃ for 40min. C4. Slowly add 0.1g of glutaraldehyde solution to the system, continue stirring at 4℃ for 10-30min, let stand for 10-20min, collect the microcapsules, wash with deionized water 3-5 times, and freeze-dry to obtain the microbial microcapsule preparation.

[0024] Example 4 A biological feed containing microbial microcapsule preparation comprises the following components in parts by weight: 90 parts of broken-cell wall Rhodopseudomonas palustris powder, 3 parts of lysine hydrochloride, 0.1 parts of methionine, 0.5 parts of threonine, 0.3 parts of arginine, 3 parts of dicalcium phosphate, 1 part of calcium carbonate, 1.2 parts of antioxidant additive, 0.3 parts of modified nano-montmorillonite, and 5 parts of microbial microcapsule preparation; the antioxidant additive was prepared in Example 1, the modified nano-montmorillonite was prepared in Example 2, and the microbial microcapsule preparation was prepared in Example 3.

[0025] The preparation method of the above-mentioned biological feed containing microbial microcapsule preparations includes the following steps: S1. First, add dicalcium phosphate and calcium carbonate to the twin-shaft paddle mixer and stir at low speed for 4 minutes. Then, add lysine hydrochloride, methionine, threonine, and arginine to the mixer at one time. Stir for 6 minutes after each amino acid is added. S2. Slowly add the broken cell wall Rhodopseudomonas sphaeroides powder while stirring. After adding all the powder, continue stirring for 12 minutes. Add the antioxidant additive and stir for 10 minutes. Then add the modified nano-montmorillonite and continue stirring for 6 minutes. Finally, add the microbial microcapsule preparation and stir at low speed for 5 minutes to obtain the mixed powder of all components. S3. Feed the mixed powder of all components into a low-temperature ring die pellet mill, introduce saturated steam at a pressure of 0.2 MPa and a temperature of 90°C, condition for 30 seconds, control the pellet mill speed at 350 r / min for extrusion granulation, and air-cool and dry to obtain biological feed containing microbial microcapsule preparation.

[0026] Example 5 A biological feed containing microbial microcapsule preparation comprises the following components in parts by weight: 93 parts of broken-cell wall Rhodopseudomonas palustris powder, 2 parts of lysine hydrochloride, 0.5 parts of methionine, 0.1 parts of threonine, 1 part of arginine, 2 parts of dicalcium phosphate, 2 parts of calcium carbonate, 0.5 parts of antioxidant additive, 0.8 parts of modified nano-montmorillonite, and 1 part of microbial microcapsule preparation; the antioxidant additive was prepared in Example 1, the modified nano-montmorillonite was prepared in Example 2, and the microbial microcapsule preparation was prepared in Example 3.

[0027] The preparation method of the above-mentioned biological feed containing microbial microcapsule preparations includes the following steps: S1. First, add dicalcium phosphate and calcium carbonate to the twin-shaft paddle mixer and stir at low speed for 6 minutes. Then, add lysine hydrochloride, methionine, threonine, and arginine to the mixer at one time, stirring for 4 minutes after each amino acid is added. S2. Slowly add the broken cell wall Rhodopseudomonas sphaeroides powder while stirring. After adding all the powder, continue stirring for 8 minutes. Add the antioxidant additive and stir for 6 minutes. Then add the modified nano-montmorillonite and continue stirring for 10 minutes. Finally, add the microbial microcapsule preparation and stir at low speed for 3 minutes to obtain the mixed powder of all components. S3. Feed the mixed powder of all components into a low-temperature ring die pellet mill, introduce saturated steam at a pressure of 0.3 MPa and a temperature of 85°C, condition for 40 seconds, control the pellet mill speed at 300 r / min for extrusion granulation, and air-cool dry to obtain biological feed containing microbial microcapsule preparation.

[0028] Example 6 A biological feed containing microbial microcapsule preparation comprises the following components in parts by weight: 91.5 parts of broken-cell wall Rhodopseudomonas palustris powder, 2.5 parts of lysine hydrochloride, 0.3 parts of methionine, 0.2 parts of threonine, 0.6 parts of arginine, 2.5 parts of dicalcium phosphate, 1.5 parts of calcium carbonate, 0.8 parts of antioxidant additive, 0.5 parts of modified nano-montmorillonite, and 2.5 parts of microbial microcapsule preparation; the antioxidant additive was prepared in Example 1, the modified nano-montmorillonite was prepared in Example 2, and the microbial microcapsule preparation was prepared in Example 3.

[0029] The preparation method of the above-mentioned biological feed containing microbial microcapsule preparations includes the following steps: S1. First, add dicalcium phosphate and calcium carbonate to the twin-shaft paddle mixer and stir at low speed for 5 minutes. Then, add lysine hydrochloride, methionine, threonine, and arginine to the mixer at one time, stirring for 5 minutes after each amino acid is added. S2. Slowly add the broken cell wall Rhodopseudomonas sphaeroides powder while stirring. After adding all the powder, continue stirring for 10 minutes. Add the antioxidant additive and stir for 8 minutes. Then add the modified nano-montmorillonite and continue stirring for 8 minutes. Finally, add the microbial microcapsule preparation and stir at low speed for 4 minutes to obtain the mixed powder of all components. S3. Feed the mixed powder of all components into a low-temperature ring die pellet mill, introduce saturated steam at a pressure of 0.25 MPa and a temperature of 88°C, condition for 35 seconds, control the pellet mill speed at 320 r / min for extrusion granulation, and air-cool dry to obtain biological feed containing microbial microcapsule preparation.

[0030] Comparative Example 1 A biological feed containing microbial microcapsule preparation comprises the following components in parts by weight: 91.5 parts of broken-cell wall Rhodopseudomonas palustris powder, 2.5 parts of lysine hydrochloride, 0.3 parts of methionine, 0.2 parts of threonine, 0.6 parts of arginine, 2.5 parts of dicalcium phosphate, 1.5 parts of calcium carbonate, 0.5 parts of modified nano-montmorillonite, and 2.5 parts of microbial microcapsule preparation; the modified nano-montmorillonite was prepared in Example 2, and the microbial microcapsule preparation was prepared in Example 3.

[0031] The preparation method of the above-mentioned biological feed containing microbial microcapsule preparations includes the following steps: S1. First, add dicalcium phosphate and calcium carbonate to the twin-shaft paddle mixer and stir at low speed for 5 minutes. Then, add lysine hydrochloride, methionine, threonine, and arginine to the mixer at one time, stirring for 5 minutes after each amino acid is added. S2. Slowly add the broken cell wall Rhodopseudomonas sphaeroides powder while stirring. After adding all the powder, continue stirring for 10 minutes. Then add the modified nano-montmorillonite and continue stirring for 8 minutes. Finally, add the microbial microcapsule preparation and stir at low speed for 4 minutes to obtain the mixed powder of all components. S3. Feed the mixed powder of all components into a low-temperature ring die pellet mill, introduce saturated steam at a pressure of 0.25 MPa and a temperature of 88°C, condition for 35 seconds, control the pellet mill speed at 320 r / min for extrusion granulation, and air-cool dry to obtain biological feed containing microbial microcapsule preparation.

[0032] Comparative Example 2 A biological feed containing microbial microcapsule preparation comprises the following components in parts by weight: 91.5 parts of broken-cell wall Rhodopseudomonas palustris powder, 2.5 parts of lysine hydrochloride, 0.3 parts of methionine, 0.2 parts of threonine, 0.6 parts of arginine, 2.5 parts of dicalcium phosphate, 1.5 parts of calcium carbonate, 0.8 parts of antioxidant additive, 0.5 parts of modified nano-montmorillonite, and 2.5 parts of microbial microcapsule preparation; the antioxidant additive was prepared in Example 1, and the microbial microcapsule preparation was prepared in Example 3.

[0033] The preparation method of the above-mentioned biological feed containing microbial microcapsule preparations includes the following steps: S1. First, add dicalcium phosphate and calcium carbonate to the twin-shaft paddle mixer and stir at low speed for 5 minutes. Then, add lysine hydrochloride, methionine, threonine, and arginine to the mixer at one time, stirring for 5 minutes after each amino acid is added. S2. Slowly add the broken cell wall Rhodopseudomonas sphaeroides powder while stirring. After adding all the powder, continue stirring for 10 minutes. Add the antioxidant additive and stir for 8 minutes. Finally, add the microbial microcapsule preparation and stir at low speed for 4 minutes to obtain the mixed powder of all components. S3. Feed the mixed powder of all components into a low-temperature ring die pellet mill, introduce saturated steam at a pressure of 0.25 MPa and a temperature of 88°C, condition for 35 seconds, control the pellet mill speed at 320 r / min for extrusion granulation, and air-cool dry to obtain biological feed containing microbial microcapsule preparation.

[0034] Comparative Example 3 A biological feed containing microbial microcapsule preparation comprises the following components in parts by weight: 91.5 parts of broken-cell wall Rhodopseudomonas palustris powder, 2.5 parts of lysine hydrochloride, 0.3 parts of methionine, 0.2 parts of threonine, 0.6 parts of arginine, 2.5 parts of dicalcium phosphate, 1.5 parts of calcium carbonate, 0.8 parts of antioxidant additive, 0.5 parts of modified nano-montmorillonite, and 2.5 parts of microbial microcapsule preparation; the antioxidant additive was prepared in Example 1, and the modified nano-montmorillonite was prepared in Example 2.

[0035] The preparation method of the above-mentioned biological feed containing microbial microcapsule preparations includes the following steps: S1. First, add dicalcium phosphate and calcium carbonate to the twin-shaft paddle mixer and stir at low speed for 5 minutes. Then, add lysine hydrochloride, methionine, threonine, and arginine to the mixer at one time, stirring for 5 minutes after each amino acid is added. S2. Slowly add the broken cell wall Rhodopseudomonas sphaeroides powder while stirring. After adding all the powder, continue stirring for 10 minutes. Add the antioxidant additive and stir for 8 minutes. Then add the modified nano-montmorillonite and continue stirring for 8 minutes to obtain the mixed powder of all components. S3. Feed the mixed powder of all components into a low-temperature ring die pellet mill, introduce saturated steam at a pressure of 0.25 MPa and a temperature of 88°C, condition for 35 seconds, control the pellet mill speed at 320 r / min for extrusion granulation, and air-cool dry to obtain biological feed containing microbial microcapsule preparation.

[0036] Performance testing I. Nutrient Utilization Efficiency Testing 1. Testing of nutrient utilization efficiency and stress and disease resistance in piglets (1) Nutritional indicators for piglets Protein digestibility: Using the total manure collection method, daily manure was collected from each group of piglets. After drying, the crude protein content in the manure was determined by the Kjeldahl method. Combined with the crude protein content in the feed, the protein digestibility was calculated using the formula: Protein digestibility (%) = (crude protein intake in feed - crude protein excretion in feces) / crude protein intake in feed × 100%; Feed conversion ratio: Record the initial weight, final weight and total feed intake of each group of piglets, and calculate according to the formula: Feed conversion ratio = Total feed intake / (Final total weight - Initial total weight).

[0037] (2) Indicators of piglet resistance to stress and disease Diarrhea rate: Observe the fecal morphology of piglets daily (loose feces are considered diarrhea), and calculate according to the formula: Diarrhea rate (%) = (Number of piglets with diarrhea × Number of days with diarrhea) / (Total number of piglets × Total number of days of rearing) × 100%; Immune organ index: After the breeding period, three piglets were randomly selected from each group for slaughter, and the spleen and thymus were separated. After weighing, the immune organ index (mg / g) was calculated according to the formula: Immune organ index (mg / g) = weight of immune organs / weight of piglet.

[0038] The obtained data is shown in Table 1 below: Table 1. Detection results of nutrient utilization efficiency and stress and disease resistance performance of piglets fed with the biological feed of this invention. ; As shown in Table 1, the protein digestibility of piglets in Example 6 was higher than that of the three comparative examples. This is because the small-molecule chitosan in the antioxidant additive promoted intestinal mucosal absorption, and polyphenols assisted in the decomposition of feed protein. The activator of modified nano-montmorillonite provided a carbon source for probiotics, enhancing their ability to decompose nutrients. The microbial microcapsule preparation directly supplemented beneficial bacteria in the intestines, improving the intestinal flora structure and reducing nutrient loss. Comparative Example 1 lacked antioxidant additives, resulting in the lowest protein digestibility due to the absence of polyphenols to aid digestion and chitosan to promote absorption. Comparative Example 3 lacked microbial microcapsule preparations, leading to the highest feed conversion ratio due to intestinal flora imbalance. This demonstrates that probiotics can improve intestinal nutrient utilization. The diarrhea rate in pigs in Example 6 was only 25.6% of that in Comparative Example 1 and 20.5% of that in Comparative Example 3. This is because the polyphenols in the antioxidant additive inhibited Escherichia coli, and the probiotics maintained the intestinal barrier. The spleen index and thymus index of piglets in Example 6 were significantly higher than those in the comparative examples. This is because the chitosan in the antioxidant additive activated immune cells, and the probiotics stimulated the development of immune organs.

[0039] 2. Testing of nutrient utilization efficiency and stress and disease resistance of grass carp (1) Nutritional indicators of grass carp Feed conversion ratio: Record the initial weight, final weight and total feed intake of each group of grass carp, and calculate according to the formula: Feed conversion ratio = Total feed intake / (Final total weight - Initial total weight). Crude fat deposition rate: After the aquaculture was completed, three grass carp were randomly selected from each group, and muscle tissue was dissected and extracted. The crude fat content of the muscle was detected by Soxhlet extraction. Combined with the initial crude fat content of the muscle (initial sampling test), the crude fat deposition rate (%) was calculated according to the formula: Crude fat deposition rate (%) = (final crude fat content of muscle - initial crude fat content of muscle) / initial crude fat content of muscle × 100%.

[0040] (2) Indicators of stress and disease resistance in grass carp Ammonia nitrogen stress survival rate: On the 30th day of rearing, each group of grass carp was transferred to water with an ammonia nitrogen concentration of 5 mg / L (normal ≤0.5 mg / L) for 48 hours, and the number of surviving fish was recorded: survival rate (%) = number of surviving grass carp / total number of fish × 100%; Vibrio infection incidence: After a stress test, Vibrio parahaemolyticus was inoculated into the water (concentration 1×10⁻⁶). 6CFU / mL), observe for 7 days, and record the number of diseased fish (gill rot and enteritis are considered as diseased): Incidence rate (%) = number of diseased grass carp / total number of fish × 100%.

[0041] The results are shown in Table 2 below: Table 2. Detection results of nutrient utilization efficiency and stress and disease resistance performance of grass carp fed with the biological feed of the present invention. ; The data in Table 2 show that the feed conversion ratio of grass carp in Example 6 was lower than that in Comparative Examples 1-3, for the same reason as the above-mentioned nutrient utilization efficiency test for piglets. The survival rate of grass carp under ammonia nitrogen stress in Example 6 was higher than that in Comparative Example 2 because the modified nano-montmorillonite adsorbed ammonia nitrogen in the water, reducing toxin damage, while the survival promoter enhanced the activity of grass carp stress-reducing kinase. The incidence of vibriosis in grass carp in Example 6 was lower than that in Comparative Example 1, verifying the inhibitory effect of the polyphenols in the antioxidant additive on pathogenic bacteria. Comparative Example 2, lacking nano-montmorillonite adsorption of toxins, still had a higher incidence rate.

[0042] 3. Storage stability testing (1) Probiotic activity retention rate At storage time of 0 days (initial), 3 months, and 6 months, 10g of feed was sampled from each group. The probiotic colony count was detected using the serial dilution plate count method (MRS medium). The activity retention rate (%) was calculated as follows: (number of colonies after storage / number of initial colonies) × 100%.

[0043] (2) Oxidative rancidity index Peroxide value (POV): The peroxide content in feed was determined by iodometric titration (unit: meq / kg); Acid value (AV): The free fatty acid content in feed was determined by potentiometric titration (unit: mgKOH / g), referring to GB5009.227-2016 standard.

[0044] The obtained data is shown in Table 3 below: Table 3. Results of storage stability test of the biological feed of the present invention ; As can be seen from the data in Table 3, after 6 months of storage at room temperature, the probiotic activity retention rate of Example 6 was 34.6% higher than that of Comparative Example 2. This is because the nano-montmorillonite interlayer structure of the modified nano-montmorillonite isolates oxygen and moisture, reducing the inactivation of probiotics. Comparative Example 1 was slightly lower than that of Example 6 because although the polyphenols of the antioxidant additive can resist oxidation, they have no physical protective effect, and the probiotics are still affected by the environment.

[0045] Example 6: The peroxide value (7.2 meq / kg) and acid value (3.8 mg KOH / g) after 6 months were lower than those of Comparative Examples 1-3, and far below the national standard limits (POV≤10 meq / kg, AV≤6 mg KOH / g). The key is that the polyphenols in the antioxidant additive replaced the chemical antioxidants, inhibiting the oxidation of fat in the feed. In contrast, Comparative Example 3 had the most severe rancidity because it lacked probiotic metabolites to assist in antioxidant activity.

[0046] In the description of this specification, references to terms such as "an embodiment," "example," "specific example," etc., indicate that a specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of the invention. In this specification, illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples.

[0047] The foregoing has shown and described the basic principles, main features, and advantages of the present invention. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The embodiments and descriptions in the specification are merely illustrative of the principles of the invention. Various changes and modifications can be made to the invention without departing from its spirit and scope, and all such changes and modifications fall within the scope of the claimed invention.

Claims

1. A biological feed containing microbial microcapsule preparations, characterized in that, It comprises the following components in parts by weight: 90-93 parts of broken-cell wall Rhodopseudomonas palustris powder, 2-3 parts of lysine hydrochloride, 0.1-0.5 parts of methionine, 0.1-0.5 parts of threonine, 0.3-1 parts of arginine, 2-3 parts of dicalcium phosphate, 1-2 parts of calcium carbonate, 0.5-1.2 parts of antioxidant additive, 0.3-0.8 parts of modified nano-montmorillonite, and 1-5 parts of microbial microcapsule preparation; The antioxidant additive is chitosan that has been enzymatically hydrolyzed and then combined with plant polyphenols via ultrasound-assisted hydrogen bonding. The plant polyphenols are green tea polyphenols and rosemary polyphenols mixed in a mass ratio of 3:

1. The modified nano-montmorillonite is prepared by introducing sodium ions through a hydrothermal method to expand the interlayer spacing of montmorillonite and then loading probiotic activators in situ. The probiotic activators are oligosaccharides, vitamin B and citric acid in a ratio of 5:3:

2. The microbial microcapsule preparation is a core-shell structure formed by coating chitosan on the surface of probiotic sodium alginate composite microspheres. The probiotic sodium alginate composite microspheres are prepared by emulsion solidification method of composite probiotics and sodium alginate. The composite probiotics are Bacillus subtilis, Lactobacillus plantarum and Lactobacillus acidophilus mixed in a mass ratio of 2:1:

1.

2. The biological feed containing microbial microcapsule preparation according to claim 1, characterized in that, The preparation method of the antioxidant additive includes the following steps: A1. Add chitosan to deionized water and stir to dissolve, obtaining a chitosan solution. Adjust the pH to 4.5 with hydrochloric acid, add the complex enzyme, and react in a 50°C constant temperature water bath for 3-5 hours. Raise the temperature to 90°C and keep it for 10-20 minutes to inactivate the complex enzyme. After cooling, adjust the pH to 6.5 with sodium hydroxide, centrifuge to remove insoluble matter, and concentrate the supernatant by rotary evaporation to obtain an enzymatically hydrolyzed chitosan solution with a solid content of 20%. A2. Add plant polyphenols to the enzymatically hydrolyzed chitosan solution, add deionized water to adjust the solid content to 15%, stir until the plant polyphenols are dissolved, sonicate for 15-25 minutes, and spray dry the reaction solution to obtain the antioxidant additive.

3. The biological feed containing microbial microcapsule preparation according to claim 2, characterized in that, The compound enzyme is a mixture of cellulase and papain in a 2:1 mass ratio, and the amount of compound enzyme added is 1.5 to 2.5% of the mass of chitosan.

4. The biological feed containing microbial microcapsule preparation according to claim 2, characterized in that, The mass ratio of plant polyphenols to enzymatically hydrolyzed chitosan is 1 to 10.

5. The biological feed containing microbial microcapsule preparation according to claim 1, characterized in that, The preparation method of the modified nano-montmorillonite includes the following steps: B1. Add montmorillonite to deionized water and disperse it by ultrasonication to obtain a montmorillonite suspension. Add hydrochloric acid solution to adjust the pH to 3, stir the reaction at 60℃ for 0.5~1.5h, centrifuge and filter, wash the precipitate with deionized water until neutral, and dry to obtain activated montmorillonite. B2. Activated montmorillonite was added to deionized water and ultrasonically dispersed to obtain an activated montmorillonite suspension. Sodium chloride was added and stirred to dissolve the suspension. The suspension was then added to a high-pressure reactor and stirred at 120°C and 0.2 MPa for 4-6 hours. After centrifugation, filtration, and drying, nano-montmorillonite was obtained. B3. Add nano-montmorillonite to deionized water and ultrasonically disperse to obtain nano-montmorillonite dispersion. Add oligofructose, vitamin B and citric acid to deionized water and stir to dissolve to obtain probiotic activator solution. Slowly add probiotic activator solution to nano-montmorillonite dispersion, and stir in a 50°C constant temperature water bath for 2-4 hours. After the reaction is completed, freeze dry to obtain the modified nano-montmorillonite.

6. The biological feed containing microbial microcapsule preparation according to claim 1, characterized in that, The activated montmorillonite and sodium chloride have a mass ratio of 8:1, the nano-montmorillonite and probiotic activator have a mass ratio of 10:1, and the vitamin B is a mixture of vitamin B1, vitamin B2, vitamin B6 and vitamin B12 in a mass ratio of 2:1:1:0.

5.

7. The biological feed containing microbial microcapsule preparation according to claim 1, characterized in that, The preparation method of the microbial microcapsule formulation includes the following steps: C1. Add the compound probiotic powder to physiological saline to prepare 1×10 9 CFU / mL bacterial suspension was mixed with glycerol and maltodextrin and stirred for 20-30 min. Sodium alginate was added to deionized water and stirred in a 50℃ water bath for 0.5-1.5 h to obtain a 2wt% sodium alginate solution. The compound probiotic suspension was mixed with the sodium alginate solution, and liquid paraffin and Span 80 were slowly added. The mixture was emulsified by high-speed shearing for 10-20 min to form a uniform W / O type emulsion. C2. Slowly add calcium chloride solution to the emulsion, stir and solidify at 4°C for 20-40 min, let stand for 8-12 min after the reaction is complete, collect the lower layer of microspheres, wash with deionized water 3-5 times to obtain sodium alginate probiotic microspheres. C3. Add chitosan powder to acetic acid solution, stir in a 50°C water bath for 0.5~1.5h to prepare chitosan solution, adjust pH to 5.5 with NaOH, add sodium alginate probiotic microspheres to chitosan solution, and stir magnetically at 4°C for 30~50min. C4. Slowly add glutaraldehyde solution to the system, continue stirring at 4°C for 10-30 min, let stand for 10-20 min, collect the microcapsules, wash with deionized water 3-5 times, and freeze-dry to obtain the microbial microcapsule preparation.

8. The biological feed containing microbial microcapsule preparation according to claim 7, characterized in that, The compound probiotics and sodium alginate solution are mixed at a volume ratio of 1:4, and the volume ratio of liquid paraffin to the mixed solution of compound probiotic suspension and sodium alginate solution is 3:

1.

9. The biological feed containing microbial microcapsule preparation according to claim 7, characterized in that, The mass ratio of sodium alginate probiotic microspheres to chitosan is 10:

1.

10. The method for preparing biological feed containing microbial microcapsule formulation according to claim 1, characterized in that, Includes the following steps: S1. First, add dicalcium phosphate and calcium carbonate to the twin-shaft paddle mixer and stir at low speed for 4-6 minutes. Then, add lysine hydrochloride, methionine, threonine, and arginine to the mixer at one time. Stir for 4-6 minutes after each amino acid is added. S2. Slowly add the broken cell wall Rhodopseudomonas sphaeroides powder while stirring. After adding all the powder, continue stirring for 8-12 minutes. Add the antioxidant additive and stir for 6-10 minutes. Then add the modified nano-montmorillonite and continue stirring for 6-10 minutes. Finally, add the microbial microcapsule preparation and stir at low speed for 3-5 minutes to obtain the mixed powder of all components. S3. Feed the mixed powder of all components into a low-temperature ring die pellet mill, introduce saturated steam at a pressure of 0.2~0.3MPa and a temperature of 85~90℃, condition for 30~40s, control the pellet mill speed at 300~350r / min for extrusion granulation, and air-cool dry to obtain biological feed containing microbial microcapsule preparation.