A probiotic biological microcapsule and its preparation method and application

Abstract

The present invention discloses a probiotic bio-microcapsule and its preparation method and application. The probiotic bio-microcapsule includes a probiotic core material and a wall material coating the probiotic core material; the probiotic core material is obtained by coating a composite bacterial agent with a coating agent; the composite bacterial agent includes Bacillus lentus and Bacillus velez; the coating agent is composed of chitosan, resistant starch and modified polyacrylamide. The probiotic bio-microcapsule provided by the present invention, based on the synergistic effect and dual protective effect of the coating and the wall material, can not only ensure that the composite bacterial agent safely reaches the colonization site, but also can further provide nutrition and protection for the colonization process, significantly improving the survival rate of probiotics during transportation in the body environment and the survival rate after colonization, and at the same time can be stored for a long time in an external environment at room temperature, reducing the cost of storage and transportation, and can be widely used in biological feed.

Description

Technical Field

[0001] The present invention relates to the technical field of microorganisms, and in particular to a probiotic biological microcapsule and a preparation method and application thereof. Background Art

[0002] As the livestock industry continues to expand and become increasingly intensive, the search for efficient, safe, and sustainable production methods has become a core issue for the industry. Against this backdrop, probiotics are moving from research laboratories to large-scale applications, becoming a highly anticipated engine of green growth. Their core value lies in their gradual replacement of antibiotics, once widely used but associated with serious resistance and residue issues, providing a safe, non-toxic, and highly effective solution for the livestock industry. This shift not only impacts economic benefits but also significantly contributes to food safety, public health, and ecological balance.

[0003] A large number of studies have confirmed that probiotics, as a type of living microorganisms that have a certain promoting effect on host health, have multiple physiological functions in treatment and health care, such as: enhancing the body's immune function, regulating the animal's intestinal microecological environment, inhibiting the reproduction of harmful pathogens, improving feed conversion rate, maintaining animal health and promoting growth, etc.

[0004] Common probiotics, such as lactic acid bacteria, are inherently fragile and susceptible to environmental factors. Therefore, probiotic products must be stored at low temperatures and refrigerated to maximize the number of active probiotics within them, as the colony count decreases over time. Furthermore, the highly acidic environment of gastric juice and pepsin can effectively kill a large number of unprotected bacteria, and the high concentration of bile salts in the front of the small intestine has a powerful dissolving and damaging effect on bacterial cell membranes. As a result, after oral administration of untreated probiotics, the proportion of live bacteria that ultimately reach and colonize target areas such as the large intestine is often less than 1%. This low survival rate means that the actual number of effective bacteria is far lower than the theoretical requirement, severely restricting the manifestation of probiotic effects.

[0005] Currently, probiotic products are generally applied in the pharmaceutical field in the form of tablets, capsules, powders, etc., and generally need to be stored in low-temperature refrigeration, which limits the storage environment and storage time. In addition, under the constraints of harsh environments such as gastric acid and pepsin, the survival rate of current probiotic products and the survival rate after colonization still cannot meet actual growth needs. At the same time, the probiotic products with microcapsule structures in the existing technology have insufficient bonding strength and biocompatibility between the core material and the wall material, resulting in their structural stability and probiotic survival rate still needing to be improved.

[0006] Patent CN201911378534.7 discloses a microbial composite agent, a microencapsulated composite probiotic preparation, a meat rabbit feed, and a preparation method and application. The microbial composite agent is used as a core material and directly mixed with a wall material to form a microcapsule preparation. However, its ability to improve the colonization survival rate still cannot meet the increasing demand.

[0007] Therefore, there is an urgent need for a probiotic bio-microcapsule that can significantly improve the survival rate of probiotics during delivery in the in vivo environment and the survival rate after colonization, and is easy to preserve for a long time. Summary of the Invention

[0008] Purpose of the invention: In view of the defects of the prior art, the purpose of the present invention is to provide a probiotic bio-microcapsule and its preparation method and application, which can significantly improve the survival rate of probiotics during transportation in the in vivo environment and the survival rate after colonization, and can be easily preserved for a long time.

[0009] Technical solution:

[0010] In one aspect, the present invention provides a probiotic biological microcapsule comprising a probiotic core material and a wall material coating the probiotic core material;

[0011] The probiotic core material is prepared by coating the composite bacterial agent with a coating agent;

[0012] The composite bacterial agent includes Bacillus lentus and Bacillus velez;

[0013] The coating agent consists of chitosan, resistant starch and modified polyacrylamide;

[0014] The modified polyacrylamide has a structure shown in the following formula A:

[0015] ,

[0016] Among them, a:b:c=20:5:3, and the molecular weight is 8000-15000.

[0017] Furthermore, the effective viable bacteria count in the composite bacterial agent is not less than 5×10 9 CFU / g; the effective viable bacterial count ratio of the Bacillus lentus and Bacillus velezii is (1-3): (1-3).

[0018] The probiotic microcapsules provided by the present invention have a combination of composite bacterial agents that can stably release amylase, lipase, cellulase, and protease, and can hydrolyze a variety of complex proteins in the animal intestine into small molecules such as polypeptides and amino acids, which is beneficial to the digestion and absorption of the animal intestine.

[0019] Furthermore, the mass ratio of chitosan, resistant starch and modified polyacrylamide in the coating agent is (5-8): (5-8): (12-15).

[0020] In the probiotic bio-microcapsules provided by the present invention, the coating agent is composed of chitosan, resistant starch and modified polyacrylamide. On the one hand, the three can cross-link with each other to form a stable coating structure, uniformly coating the composite bacterial agent to form a core material, which not only limits the shape of the bacterial agent to facilitate combination with the wall material to form microcapsules, but also further enhances the protective effect and improves the stress resistance of the probiotics. On the other hand, the three have excellent biocompatibility and nutritional properties for the probiotics, not only can they effectively coat the probiotics without affecting their activity, but the resistant starch and modified polyacrylamide can also provide nutrition for their growth, thereby improving their colonization survival rate.

[0021] Furthermore, the modified polyacrylamide is prepared by the following steps:

[0022] (1) Add acrylamide, dimethyl vinylphosphonate, 3-methacryloyldopamine and deionized water into the reactor, stir and mix them evenly, and then introduce nitrogen;

[0023] (2) Adding an initiator to the reactor of step (1), heating to 90-95°C under nitrogen protection and keeping the temperature to react for 18-24 hours, cooling, filtering, washing and drying to obtain the modified polyacrylamide.

[0024] In the probiotic bio-microcapsules provided by the present invention, catecholamine structures and phosphate groups are polymerized in the modified polyacrylamide, which can improve the binding ability and promote colonization. On the one hand, the catecholamine structure has excellent binding ability with the probiotic biofilm, which can improve the coating effect in the core material, thereby enhancing the survival rate and colonization effect of the composite bacterial agent; on the other hand, the phosphate group structure can not only provide a phosphorus source for the growth of probiotics in the early colonization process, thereby improving the survival rate of colonization, but also enhance the binding effect between the coating structure and the wall material, thereby improving the structural stability of the microcapsules.

[0025] Furthermore, in step (1), the molar ratio of acrylamide, dimethyl vinylphosphonate and 3-methacryloyldopamine is 20:5:3;

[0026] In the step (2), the initiator is selected from ammonium persulfate or potassium persulfate.

[0027] Furthermore, the wall material is selected from one of sodium alginate, chitosan or starch.

[0028] In the probiotic bio-microcapsules provided by the present invention, the phosphate ester structure and amide structure based on the modified polyacrylamide in the coating agent can be further cross-linked with structures such as carboxyl and hydroxyl groups in the wall material, thereby improving the bonding ability between the core material and the wall material, thereby enhancing the structural stability of the microcapsules, which can not only improve their stability in the organism, but also enhance their storage and transportation stability in the external environment at room temperature.

[0029] Another aspect of the present invention provides a method for preparing any of the above-mentioned probiotic microcapsules, comprising the following steps:

[0030] (1) Bacillus lentus and Bacillus velezii are mixed to prepare a composite bacterial agent, and chitosan, resistant starch and modified polyacrylamide are mixed to prepare a coating agent;

[0031] (2) After the composite bacterial agent and the coating agent are evenly mixed, they are sheared at high speed to form the probiotic core material;

[0032] (3) The probiotic core material and the wall material are mixed evenly and then spray-dried to obtain the probiotic bio-microcapsules.

[0033] Furthermore, in step (2), the mass ratio of the composite bacterial agent to the coating agent is (1-2): (3-4); the rotation speed of the high-speed shear is 8000-12000 rpm, and the time is 2-5 minutes.

[0034] Furthermore, in step (3), the mass ratio of the probiotic core material to the wall material is 1:2-2.5; the spray drying conditions are: inlet air temperature 120-140°C, fan frequency 40-50Hz, peristaltic speed 5-10rpm, and outlet air temperature 50-60°C.

[0035] Finally, the present invention also provides the use of any one of the above-mentioned probiotic microcapsules in biological feed.

[0036] Beneficial effects:

[0037] (1) The probiotic bio-microcapsules provided by the present invention are prepared by combining a core material made by coating a composite bacterial agent with a coating agent and a wall material. Based on the synergistic effect and dual protective effect of the coating and the wall material, it can not only ensure that the composite bacterial agent arrives safely at the colonization site, but also further provide nutrition and protection for the colonization process, significantly improve the survival rate of probiotics during transportation in the body environment and the survival rate after colonization, and at the same time can be stored for a long time in an external environment at room temperature, reducing the cost of storage and transportation, and can be widely used in biological feed.

[0038] (2) The probiotic bio-microcapsules provided by the present invention have a combination of composite bacterial agents that can stably release amylase, lipase, cellulase and protease, and can hydrolyze a variety of complex proteins in the animal intestine into small molecules such as polypeptides and amino acids, which is beneficial to the digestion and absorption of the animal intestine.

[0039] (3) The probiotic bio-microcapsules provided by the present invention have a coating agent composed of chitosan, resistant starch and modified polyacrylamide. On the one hand, the three can cross-link with each other to form a stable coating structure, uniformly coating the composite bacterial agent to form a core material, which can not only limit the shape of the bacterial agent to be easy to combine with the wall material to form a microcapsule, but also further improve the protective effect and enhance the resistance of probiotics to adversity; on the other hand, the three have excellent biocompatibility and nutritional properties for probiotics, not only can they effectively coat the probiotics without affecting their activity, but the resistant starch and modified polyacrylamide can also provide nutrition for their growth, thereby improving their colonization survival rate.

[0040] (4) The probiotic bio-microcapsules provided by the present invention have catecholamine structures and phosphate groups polymerized in the modified polyacrylamide, which can improve the binding ability and promote colonization. On the one hand, the catecholamine structure has excellent binding ability with the probiotic biofilm, which can improve the coating effect in the core material, thereby enhancing the survival rate and colonization effect of the composite bacterial agent; on the other hand, the phosphate group structure can not only provide a phosphorus source for the growth of probiotics in the early colonization process, thereby improving the survival rate of colonization, but also enhance the binding effect between the coating structure and the wall material, thereby improving the structural stability of the microcapsules.

[0041] (5) The probiotic bio-microcapsules provided by the present invention can further cross-link with the carboxyl, hydroxyl and other structures in the wall material based on the phosphate ester structure and amide structure of the modified polyacrylamide in the coating agent, thereby improving the bonding ability between the core material and the wall material, thereby enhancing the structural stability of the microcapsules, which can not only improve their stability in the organism, but also enhance their storage and transportation stability in the external environment at room temperature. DETAILED DESCRIPTION

[0042] The present invention will be described below in conjunction with specific embodiments. It should be noted that the following examples are illustrative of the present invention and are intended only to illustrate the present invention and are not intended to limit the present invention. Other combinations and various modifications within the scope of the present invention may be made without departing from the spirit or scope of the present invention.

[0043] Bacillus lentus can be Bacillus lentus MES857, purchased from China Industrial Microbiological Culture Collection Administration Center, the purchase address is Building 6, No. 24 Jiuxianqiao Middle Road, Chaoyang District, Beijing, the purchase date is July 23, 2018, and the deposit number is CICC 10365;

[0044] The Bacillus velezensis may be Bacillus velezensis MES861, purchased from the General Microbiology Center of China Culture Collection Administration, the purchase address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, the purchase date is December 3, 2018, and the deposit number is CGMCC No. 16857;

[0045] The CAS number of 3-methacryloyldopamine is 471915-89-6; the remaining reagents and equipment are conventional reagents and equipment in this technical field.

[0046] Preparation of modified polyacrylamide-1

[0047] (1) Add 0.2 mol of acrylamide, 0.05 mol of dimethyl vinylphosphonate, 0.03 mol of 3-methacryloyldopamine, and 100 mL of deionized water into the reactor, stir and mix until uniform, and then introduce nitrogen;

[0048] (2) Add 0.3 g of ammonium persulfate to the reactor of step (1), heat to 95° C. under nitrogen protection and keep warm for 24 hours, then cool, filter, wash and dry to obtain the modified polyacrylamide-1. The obtained modified polyacrylamide-1 has a molecular weight of 12,000.

[0049] Preparation of modified polyacrylamide-2

[0050] The preparation method is basically the same as that of modified polyacrylamide-1, except that dimethyl vinylphosphonate is replaced by an equal amount of acrylamide.

[0051] Preparation of modified polyacrylamide-3

[0052] The preparation method is basically the same as that of modified polyacrylamide-1, except that 3-methacryloyldopamine is replaced with an equal amount of acrylamide.

[0053] Example 1

[0054] Probiotic microcapsules were prepared by the following steps:

[0055] (1) Bacillus lentus and Bacillus velezii are mixed to prepare a composite bacterial agent, and chitosan, resistant starch and modified polyacrylamide-1 are mixed to prepare a coating agent;

[0056] (2) After uniformly mixing 1 part of the composite bacterial agent with 4 parts of the coating agent by weight, the mixture was subjected to high-speed shearing at a speed of 10,000 rpm for 2-5 minutes to prepare the probiotic core material;

[0057] (3) After 5 parts of probiotic core material and 10 parts of sodium alginate were evenly mixed, the probiotic bio-microcapsules were prepared by spray drying at an inlet air temperature of 130°C, a fan frequency of 50 Hz, a peristaltic speed of 10 rpm, and an outlet air temperature of 50°C;

[0058] The effective viable bacteria count of the composite bacterial agent in step (1) is 8×10 9 CFU / g; the effective viable count ratio of the Bacillus lentus to the Bacillus Velezii is 1:1;

[0059] In the step (1), the mass ratio of chitosan, resistant starch and modified polyacrylamide-1 is 1:1:3.

[0060] Example 2

[0061] The method is basically the same as Example 1, except that the mass ratio of chitosan, resistant starch and modified polyacrylamide-1 in step (1) is changed to 2:2:3.

[0062] Example 3

[0063] The method is basically the same as Example 1, except that the coating agent in step (2) is changed to 1.5 parts; and the 10 parts of sodium alginate in step (3) is changed to 12 parts of starch.

[0064] Comparative Example 1

[0065] Probiotic microcapsules were prepared by the following steps:

[0066] (1) Mixing Bacillus lentus and Bacillus velezii to prepare a composite bacterial agent;

[0067] (2) After 1 part of the composite bacterial agent and 10 parts of sodium alginate were evenly mixed, the probiotic microcapsules were prepared by spray drying at an inlet air temperature of 130°C, a fan frequency of 50 Hz, a peristaltic speed of 10 rpm, and an outlet air temperature of 50°C;

[0068] The effective viable bacteria count of the composite bacterial agent in step (1) is 8×10 9 CFU / g; the effective viable bacterial count ratio of the Bacillus lentus to the Bacillus Velez is 1:1.

[0069] Comparative Example 2

[0070] The process is basically the same as Example 1, except that the modified polyacrylamide-1 in step (1) is replaced by an equal amount of modified polyacrylamide-2.

[0071] Comparative Example 3

[0072] The process is basically the same as Example 1, except that the modified polyacrylamide-1 in step (1) is replaced by an equal amount of modified polyacrylamide-3.

[0073] Comparative Example 4

[0074] The process is basically the same as Example 1, except that the resistant starch in step (1) is replaced by an equal amount of chitosan.

[0075] Performance Testing

[0076] Detection of viable bacteria count in microcapsules: 1 g of the products of Examples 1 to 3 and Comparative Examples 1 to 4 were taken respectively, and the number of viable bacteria in the microcapsules was determined by plate count method. The experiment was repeated three times in parallel, and the number of viable bacteria was recorded as -0.

[0077] Storage capacity test: 1 g of the products of Examples 1 to 3 and Comparative Examples 1 to 4 were stored at room temperature for 120 days, and the number of viable bacteria in the microcapsules was determined by the plate count method. The experiment was repeated three times in parallel, and the number of viable bacteria was recorded as -1.

[0078] Gastric acid and bile salt resistance test:

[0079] Simulated gastric fluid: Take 100 mL of 0.3 mol / L NaCl solution, adjust the pH to 2.5 with hydrochloric acid solution, add 1 g of pepsin after sterilization, use a magnetic stirrer to fully dissolve it, and store at 4°C for later use.

[0080] Weigh 1 g of each product from Example 1 to Example 3 and Comparative Examples 1 to Comparative Examples 4 into 20 mL of simulated gastric fluid, and simulate the gastric digestion process under anaerobic conditions at 37°C and 90 rpm. Samples were taken after 2 hours, and the number of viable bacteria in the microcapsules was determined by the plate count method. The experiment was repeated three times, and the number of viable bacteria was recorded as -2.

[0081] To prepare simulated small intestinal digestive fluid: Dissolve 3.4g of KH2PO4 in an appropriate amount of ultrapure water and dilute to 500mL. Add 100mL of the prepared KH2PO4 solution and adjust the pH to 8.0 with 1mol / L NaOH. After sterilization, add 1g of trypsin and 0.3g of porcine bile salts. Dissolve thoroughly using a magnetic stirrer and store at 4°C until ready for use.

[0082] Weigh 1 g of each product from Example 1 to Example 3 and Comparative Example 1 to Comparative Example 4 into 40 mL of simulated small intestinal digestive fluid. Simulate the small intestinal digestion process under anaerobic conditions at 37°C and 90 rpm. Take samples after 4 hours and determine the number of viable bacteria in the microcapsules by the plate count method. Repeat the experiment three times and record the number of viable bacteria - 3.

[0083] The test results are shown in Table 1 below:

[0084] Table 1

[0085]

[0086] According to the comparison of the test results of Examples 1 to 3 with those of Comparative Example 1, the probiotic biological microcapsules provided by the present invention can significantly improve the storage capacity and gastric acid and bile salt resistance of the microcapsules by coating the composite bacterial agent with a coating agent, so that the microcapsules can effectively pass through the biological stomach and colonize in target areas such as the large intestine, and can be widely used in biological feed.

[0087] According to the comparison of the test results of Examples 1 to 3 with Comparative Examples 2 to 3, the probiotic bio-microcapsules provided by the present invention can significantly improve the binding ability between the composite bacterial agent, the coating and the wall material in the bio-microcapsules, as well as the colonization ability of the probiotics, by using modified polyacrylamide containing a catecholamine structure and a phosphate group structure as the main component of the coating agent.

[0088] According to the comparison of the test results of Examples 1 to 3 with Comparative Example 4, the probiotic bio-microcapsules provided by the present invention can be cross-linked with other components to form a stable coating structure to improve the stress resistance of the microcapsules by adding resistant starch to the coating agent.

[0089] Colonization ability test:

[0090] 1. Preparation of meat rabbit feed: 2 parts of the products of Examples 1 to 3 and Comparative Examples 1 to 4, respectively, were mixed with 15 parts of green hay, 20 parts of soybean meal powder, 8 parts of rapeseed cake, 20 parts of wheat bran, 25 parts of pea flour, 5 parts of methionine, 30 parts of alfalfa powder, 20 parts of carrots, and 0.5 parts of salt, and then transferred to a grinder for pulverization. The mixture was passed through a 60-mesh sieve and granulated into feed granules with a particle size of 3.0 mm to prepare different meat rabbit feeds containing probiotic bio-microcapsules.

[0091] 2. Healthy New Zealand rabbits of similar birth date and weight were selected and randomly divided into three groups, each containing four replicates, with 40 rabbits (half male and half female) in each group. The experimental setup consisted of: a control group fed standard feed; and experimental groups fed rabbit feed supplemented with the products of Examples 1-3 and Comparative Examples 1-4. The rabbits were fed once daily in the morning, noon, and evening, with free access to water, and maintained under conventional breeding practices for 30 days. When the rabbits reached three months of age, ten were randomly selected and slaughtered. Their weights were weighed on an empty stomach in the morning, and their protein content and average weight were determined.

[0092] The test results are shown in Table 2 below:

[0093] Table 2

[0094]

[0095] According to the comparison of the test results of Examples 1 to 3 with the control group, it can be seen that the composite bacterial agent of the probiotic bio-microcapsules provided by the present invention can effectively improve the absorption effect of the organism on the feed, and the form of the bio-microcapsules can significantly improve the colonization ability of the probiotics, further improving the growth effect of the organism.

[0096] According to the comparison of the test results of Examples 1 to 3 with those of Comparative Example 1, the probiotic bio-microcapsules provided by the present invention can significantly improve the colonization ability of probiotics by coating the composite bacterial agent with a coating agent, so that the probiotics can effectively improve the growth effect of biological edible feed, and can be widely used in biological feed.

[0097] According to the comparison of the test results of Examples 1 to 3 with Comparative Examples 2 to 3, the probiotic bio-microcapsules provided by the present invention can significantly improve the binding ability between the composite bacterial agent, the coating and the wall material in the bio-microcapsules, as well as the colonization ability of the probiotics, by using modified polyacrylamide containing a catecholamine structure and a phosphate group structure as the main component of the coating agent.

[0098] According to the comparison of the test results of Examples 1 to 3 with Comparative Example 4, the probiotic bio-microcapsules provided by the present invention can provide nutrition for the growth and preservation of probiotics by adding resistant starch in the coating agent, thereby improving the colonization survival rate of probiotics.

[0099] The above embodiments are only for illustrating the technical concept and features of the present invention. Its purpose is to enable people familiar with this technology to understand the content of the present invention and implement it. It cannot be used to limit the scope of protection of the present invention. Any equivalent changes or modifications made according to the spirit of the present invention should be included in the scope of protection of the present invention.

Claims

1. A probiotic bio-microcapsule, characterized in that: It comprises a probiotic core material and a wall material covering the probiotic core material; The probiotic core material is prepared by coating the composite bacterial agent with a coating agent; The composite bacterial agent includes Bacillus lentus and Bacillus velez; The coating agent consists of chitosan, resistant starch and modified polyacrylamide; The modified polyacrylamide has a structure shown in the following formula A: ， Where a: b:c=20:5:3, molecular weight is 8000-15000.

2. The probiotic microcapsule according to claim 1, characterized in that The effective viable bacteria count in the composite bacterial agent is not less than 5×10 9 CFU / g; the effective viable bacterial count ratio of the Bacillus lentus and Bacillus velezii is (1-3): (1-3).

3. The probiotic microcapsule according to claim 1, characterized in that The mass ratio of chitosan, resistant starch and modified polyacrylamide in the coating agent is (5-8): (5-8): (12-15).

4. The probiotic microcapsule according to claim 1, characterized in that The modified polyacrylamide is prepared by the following steps: (1) Add acrylamide, dimethyl vinylphosphonate, 3-methacryloyldopamine and deionized water into the reactor, stir and mix them evenly, and then introduce nitrogen; (2) Adding an initiator to the reactor of step (1), heating to 90-95°C under nitrogen protection and keeping the temperature to react for 18-24 hours, cooling, filtering, washing and drying to obtain the modified polyacrylamide.

5. The probiotic microcapsule according to claim 4, characterized in that: In step (1), the molar ratio of acrylamide, dimethyl vinylphosphonate and 3-methacryloyldopamine is 20:5:3; In the step (2), the initiator is selected from ammonium persulfate or potassium persulfate.

6. The probiotic microcapsule according to claim 1, characterized in that The wall material is selected from one of sodium alginate, chitosan or starch.

7. The method for preparing the probiotic microcapsules according to any one of claims 1 to 6, characterized in that: The following steps are involved: (1) Bacillus lentus and Bacillus velezii are mixed to prepare a composite bacterial agent, and chitosan, resistant starch and modified polyacrylamide are mixed to prepare a coating agent; (2) After the composite bacterial agent and the coating agent are evenly mixed, they are sheared at high speed to form the probiotic core material; (3) The probiotic core material and the wall material are mixed evenly and then spray-dried to obtain the probiotic bio-microcapsules.

8. The method for preparing probiotic microcapsules according to claim 7, characterized in that: The mass ratio of the composite bacterial agent to the coating agent in step (2) is (1-2): (3-4); the rotation speed of the high-speed shear is 8000-12000 rpm, and the time is 2-5 minutes.

9. The method for preparing probiotic microcapsules according to claim 7, characterized in that: The mass ratio of the probiotic core material to the wall material in step (3) is 1:2-2.5; the spray drying conditions are: inlet air temperature 120-140°C, fan frequency 40-50Hz, peristaltic speed 5-10rpm, and outlet air temperature 50-60°C.

10. Use of the probiotic microcapsule according to any one of claims 1 to 6 in biological feed.

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