Intestinal flora transplant bacteria mud capsule biological preparation and preparation method thereof

Abstract

The application discloses an intestinal flora transplant bacteria mud capsule biological preparation and a preparation method thereof. The biological preparation components comprise inulin, apple fruit powder, freeze-dried bergamot powder and freeze-dried daylily powder, and the proportions are inulin 75 parts, apple fruit powder 10 parts, freeze-dried bergamot powder 12 parts and freeze-dried daylily powder 3 parts. In the application, the two homologous medicinal and edible raw materials of bergamot and daylily are compounded according to a proportion of 12:3 and applied to the intestinal flora transplant bacteria mud capsule protective agent. The two raw materials have no gastrointestinal stimulation and no inhibition on the intestinal flora activity, and realize the synergistic medicinal value. Secondly, the bergamot and daylily have natural aromatic odors. After being compounded according to the core + synergistic proportion, a strong synergistic taste masking effect is formed, the fishy odor of the intestinal flora bacteria mud itself can be effectively masked, the flavor blending effect of the apple fruit powder is combined, the palatability of the bacteria mud capsule preparation is optimized, the aromatic substances are stable and not easy to volatilize during storage, the taste masking effect is durable, and the flora activity is not affected.

Description

Technical Field

[0001] This invention relates to the field of intestinal microecological preparation technology, and in particular to an intestinal flora transplantation bacterial mud capsule biological preparation and its preparation method. Background Technology

[0002] Gut microbiota transplantation (GMT) is an important treatment for diseases related to gut microbiota imbalance (such as inflammatory bowel disease and irritable bowel syndrome). Its core principle is to transfer the gut microbiota of a healthy donor to the patient's gut, restoring the patient's gut microbiota balance. The key to successful GMT formulation lies in maintaining the activity of the microbiota during preparation, storage, and transport, while simultaneously optimizing the palatability of the formulation to improve patient compliance.

[0003] Microbial sludge capsules are one of the mainstream dosage forms of intestinal flora transplantation preparations. The corresponding protective agents mostly use single ingredients such as trehalose, mannitol, glycerol, and sucrose or simple compound systems. Their main function is to maintain the activity of the flora, but they have obvious functional limitations and cannot meet the diversified needs of clinical applications.

[0004] Traditional preservatives can only protect the activity of intestinal flora without considering their medicinal value. They cannot provide additional support for patients' intestinal health and have a low degree of alignment with clinical needs. Intestinal flora sludge itself has a distinct fishy or putrid odor. Traditional preservatives often use a single odor-masking ingredient, which has a limited odor-masking effect. Moreover, the odor-masking substances are volatile and their effectiveness decreases significantly during storage, resulting in low patient compliance and affecting treatment efficacy. Summary of the Invention

[0005] In view of the shortcomings of the prior art, the purpose of this invention is to provide a biological agent for intestinal flora transplantation sludge capsules and its preparation method, which aims to solve the problems that traditional protectants mostly use a single taste-masking component, which has a limited taste-masking effect, and the taste-masking substances are volatile and their effects decay significantly during storage, resulting in low patient compliance and affecting the therapeutic effect.

[0006] To achieve the above objectives, the present invention adopts the following technical solution: Firstly, a biological agent for intestinal flora transplantation mud capsules, the biological agent components of which include inulin, apple fruit powder, freeze-dried bergamot powder, freeze-dried bitter orange powder, and intestinal flora mud.

[0007] Furthermore, the biological agent components, by mass parts, are in the following proportions: inulin 72-78 parts, apple fruit powder 9-11 parts, bergamot freeze-dried powder 11-13 parts, and bitter orange freeze-dried powder 2.5-3.5 parts.

[0008] Secondly, a method for preparing a gut microbiota transplantation microbial sludge capsule biological agent includes: Step 1: Prepare freeze-dried Buddha's Hand citron powder and freeze-dried bitter orange flower powder separately, and mix inulin and apple fruit powder with freeze-dried Buddha's Hand citron powder and freeze-dried bitter orange flower powder in proportion to prepare a protective agent; Step 2: Take fresh intestinal microbiota slurry, mix the preservative and fresh intestinal microbiota slurry in proportion, and stir at a stirring rate of 30-50 r / min for 3-5 minutes in a sterile environment at 2-8℃ to obtain mixed microbiota slurry; fill the mixed microbiota slurry into enteric-coated capsule shells, seal and package to obtain intestinal microbiota transplantation microbiota slurry capsules.

[0009] Furthermore, in step 1, the bergamot freeze-dried powder is prepared using a low-temperature freeze-drying process, with a freeze-drying temperature of 40℃-50℃ and a freeze-drying time of 12-24h, preserving its natural aromatic substances and active ingredients, and a particle size of 100-200 mesh.

[0010] Furthermore, in step 1, the freeze-dried powder of the substitute flower is prepared by low-temperature freeze-drying process, with a particle size of 100-200 mesh, retaining natural volatile oil components, and the volatile oil content is above 0.1%.

[0011] Furthermore, in step 1, the mass ratio of freeze-dried Buddha's Hand citron powder and freeze-dried bitter orange powder is 11:3.5-13:2.5, while the other two types of inulin and apple powder are added and mixed according to the required ratio of 7.5:1.

[0012] Furthermore, in step 2, the ratio of the protectant to the fresh intestinal flora sludge is 10:1, and the moisture content of the fresh intestinal flora sludge is 60-70%.

[0013] Furthermore, in step 2, the aseptic environment humidity is below 35%, and the finished intestinal flora transplantation sludge capsules are sealed and stored in a refrigerated environment at 4-8℃.

[0014] The technical solution adopted in this invention has the following beneficial effects: In this invention, two medicinal and edible raw materials, Buddha's Hand and Bitter Orange Flower, are compounded in a 12:3 ratio and applied to the preservative agent of intestinal flora transplantation sludge capsules. Neither raw material causes gastrointestinal irritation or inhibits intestinal flora activity, and both achieve synergistic medicinal value, overcoming the limitations of traditional preservatives that only focus on activity protection while neglecting auxiliary medicinal value. Secondly, Buddha's Hand and Bitter Orange Flower possess natural aromatic scents. When compounded in a core + synergistic ratio, they form a powerful synergistic taste-masking effect, effectively masking the fishy or putrid odor of the intestinal flora sludge itself. Combined with the flavor-harmonizing effect of apple powder, this further optimizes the palatability of the sludge capsule formulation, addressing the industry pain point of low patient compliance due to unpleasant odor in intestinal flora transplantation sludge capsules. Furthermore, the aromatic substances are stable and do not easily volatilize during storage, ensuring a long-lasting taste-masking effect without affecting flora activity. This achieves "activity protection + flavor harmonization + medicinal assistance + synergistic taste masking." Four-fold benefits, highly adapted to the characteristics and usage scenarios of bacterial mud capsule formulations; specifically adapted to bacterial mud capsule formulations, the selection of apple fruit powder can improve the stability of the bacterial mud system, avoid bacterial mud clumping, and perfectly adapt to capsule filling process; the fresh aroma of bergamot freeze-dried powder and bitter orange freeze-dried powder work together to accurately mask the fishy and putrid smell of bacterial mud, and the formula is adapted to the characteristics of enteric-coated capsules, without affecting the colonization of bacteria in the intestine. Detailed Implementation

[0015] To make the objectives, technical solutions, and effects of this invention clearer and more explicit, the following embodiments are provided to further illustrate the invention in detail. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the scope of the invention.

[0016] A gut microbiota transplantation microbial mud capsule biological preparation, the microbial mud capsule biological preparation components include a protectant prepared from inulin, apple fruit powder, freeze-dried bergamot powder, freeze-dried bitter orange flower powder and gut microbiota microbiota mud, the above components are mixed in any proportion.

[0017] The preferred proportions of the above components are as follows: the protective agent components are in the following proportions by mass: 72-78 parts inulin, 9-11 parts apple fruit powder, 11-13 parts freeze-dried Buddha's hand citron powder, and 2.5-3.5 parts freeze-dried bitter orange flower powder. The ratio of protective agent to fresh intestinal flora sludge is 10:1. All components meet food-grade or pharmaceutical-grade standards.

[0018] Secondly, a method for preparing a gut microbiota transplantation microbial sludge capsule biological agent includes: Step 1: Prepare freeze-dried Buddha's Hand citron powder and freeze-dried bitter orange flower powder separately, and mix inulin and apple fruit powder with freeze-dried Buddha's Hand citron powder and freeze-dried bitter orange flower powder in proportion to prepare a protective agent; The Buddha's Hand freeze-dried powder is prepared using a low-temperature freeze-drying process at 40℃-50℃ for 12-24 hours, preserving its natural aromatic substances and active ingredients. The particle size is 100-200 mesh, ensuring uniform mixing with the bacterial slurry and not affecting capsule formation. The Bitter Orange freeze-dried powder is also prepared using a low-temperature freeze-drying process, with a particle size of 100-200 mesh, preserving natural volatile oil components with a volatile oil content of over 0.1%. It features synergistic innovative components, is both medicinal and edible, causes no gastrointestinal irritation, enhances the aroma masking effect, and also possesses medicinal value. The mass ratio of Buddha's Hand freeze-dried powder to Bitter Orange freeze-dried powder is 11:3.5-13:2.5, while the other two, inulin and apple fruit powder, are added and mixed according to a 7.5:1 ratio. Inulin, as a core protective substrate, maintains the activity of intestinal flora in the bacterial slurry and also has prebiotic effects. Food-grade inulin with a purity ≥95% is selected. (2) Apple fruit powder: as an auxiliary protective agent, it enhances the stability of the fungal mud system and harmonizes the sweet flavor. It is made from natural freeze-dried apple fruit powder and contains no added sucrose or preservatives.

[0019] Step 2: Take fresh intestinal flora slurry, mix the preservative and fresh intestinal flora slurry in the specified ratio, and stir at a stirring rate of 30-50 r / min for 3-5 minutes in a sterile environment at 2-8℃ to obtain mixed slurry; fill the mixed slurry into enteric-coated capsule shells, seal and package to obtain intestinal flora transplantation slurry capsules; the ratio of preservative to fresh intestinal flora slurry is 10:1, and the moisture content of the fresh intestinal flora slurry is 60-70%, the humidity of the sterile environment is below 35%, and the finished intestinal flora transplantation slurry capsules are sealed and stored in a refrigerated environment at 4-8℃, with a shelf life of more than 6 months.

[0020] Example 1: Intestinal flora transplantation microbial sludge capsule protectant: Take 75g of inulin, 10g of apple fruit powder, 12g of freeze-dried bergamot powder, and 3g of freeze-dried bitter orange flower powder. Both the bergamot and bitter orange flower freeze-dried powders are prepared using a -40℃, 24h low-temperature freeze-drying process, with a particle size of 120 mesh. Mix the above components evenly under sterile conditions to obtain the preservative for the bacterial sludge capsules of this invention. Take 100g of fresh intestinal flora sludge with a moisture content of 65%, add 10g of this preservative, and stir for 4 minutes at 4℃, 30% humidity, and 40r / min to obtain a mixed bacterial sludge. Fill the mixed bacterial sludge into enteric-coated capsule shells using a fully automatic capsule filling machine, seal and package. Store the finished product in a refrigerated environment at 4-8℃; the shelf life can reach more than 6 months. To verify the technical effectiveness of the protective agent of this invention, multiple sets of comparative experiments were conducted, strictly following the "Microbial Limit Test Method" and the "Food Safety Toxicology Evaluation Procedure". The core test data are as follows (all are the average values ​​of 3 parallel tests, RSD≤3.5%, to ensure data reliability): Microbial activity test Test subjects: the mycelium sludge capsule protectant of this invention, the traditional protectant (trehalose + mannitol compound), and the single Buddha's Hand / Citrus aurantium protectant.

[0021] Experimental method: Each protective agent was mixed with the bacterial sludge and stored under specified conditions. The survival rate of the bacterial community was tested after 1 month, 3 months and 6 months of storage.

[0022] Experimental results: The bacterial sludge preparation treated with the protective agent of this invention achieved a bacterial survival rate of 86.7% after 6 months of storage, far exceeding the industrial storage requirements (the industry standard requirement is ≥75%). The bacterial sludge treatment group treated with traditional protective agents (trehalose:mannitol = 1:1) had a survival rate of only 73.8% after 6 months, the group treated with a single bergamot freeze-dried powder protective agent had a survival rate of 79.1%, and the group treated with a single dwarf citron freeze-dried powder protective agent had a survival rate of 77.8%, all significantly lower than the protective agent of this invention. Moreover, the survival rate of the protective agent of this invention perfectly matches the description of the 6-month shelf life of the bacterial sludge capsules, demonstrating a significant activity protection effect.

[0023] Gastrointestinal irritation test Test subjects: the mycelial mud capsule protectant of this invention (diluted according to the usage ratio) and physiological saline (blank control).

[0024] Experimental methods: Animal experiments (mice) were conducted. The drugs were administered by gavage for 7 consecutive days. The condition of the gastrointestinal mucosa, feeding and defecation of mice were observed, and the pathological sections of gastrointestinal tissue were examined.

[0025] Experimental Results: Forty SPF-grade Kunming mice were randomly divided into four groups (n=10 per group, half male and half female, weighing 20±2g). Each group was administered the following by gavage: a diluted protective agent of this invention (10-fold diluted to the clinical concentration), physiological saline (blank control), or a positive control (0.1mol / L hydrochloric acid). The administration was repeated for 7 consecutive days, with a daily gavage volume of 0.2mL / 10g body weight. After gavage, the gastric and intestinal mucosa of the mice were dissected and observed. The gastric and intestinal mucosa of the blank control group and the protective agent group were smooth, without congestion or edema, with a gastric mucosal damage index of 0.0±0.0. The gastric mucosa of the positive control group showed significant congestion and edema, with a damage index of 3.8±0.5. Histopathological examination of the gastrointestinal tissue showed that the gastrointestinal cells of the protective agent group were neatly arranged, without inflammatory cell infiltration, and there was no significant difference compared to the blank control group (P>0.05). This is consistent with the description in the component instructions that "Buddha's Hand and Bitter Orange have no gastrointestinal irritation," proving that the protective agent of this invention has no gastrointestinal irritation and good safety.

[0026] Odor evaluation test Test subjects: bacterial mud capsules treated with the protective agent of this invention, bacterial mud capsules treated with traditional protective agents, and bacterial mud preparations without added protective agents.

[0027] Experimental method: A sensory evaluation group of 10 people was formed to evaluate the odor of each preparation using a 1-10 scoring method (1 point: extremely poor odor, strong fishy smell; 10 points: good odor, no unpleasant odor, faint fragrance). The average value was taken.

[0028] Experimental Results: The initial odor score of the mycelial mud capsule formulation treated with the preservative of this invention was 9.2 points, and after 6 months of storage, the score remained at 8.8 points, decreasing by only 0.4 points. The initial score of the traditional preservative-treated group was 5.0 points, which decreased to 4.2 points after 6 months of storage. The initial score of the group without preservative was 2.0 points, with no odor-masking effect. The experimental results are consistent with the description in the component description that "Buddha's Hand and Bitter Orange have their own natural fragrance, and the aromatic substances are stable," proving that the synergistic odor-masking effect of the combination of two medicinal and edible raw materials is significant and long-lasting, while not affecting the activity of the microbial community.

[0029] Adaptability testing Test subject: The system after mixing the mycelium sludge capsule protectant of this invention with mycelium sludge; Test methods: The bacterial sludge forming rate, clumping and adhesion, capsule filling qualification rate, filling weight difference, capsule breakage rate and powder leakage rate were tested, with reference to the quality standards and industrial production requirements of enteric-coated capsules.

[0030] Experimental results: After the protective agent of this invention is mixed with the bacterial sludge, the bacterial sludge forming rate reaches 99.5%, with no clumping or adhesion (the number of clumping particles ≥500μm is 0); the capsule filling qualification rate reaches 99.8%, and the filling weight difference is ±2.1%, which meets the quality standard of enteric-coated capsules (weight difference ≤±3.0%); after the capsules are formed, there is no damage or powder leakage, and the damage rate and powder leakage rate are both 0.0%, which is fully compatible with industrial production processes (the industry standard requires a qualification rate ≥99.0%). This is consistent with the description in the preparation method that "the bacterial sludge has good formability and is suitable for capsule filling", and the dosage form compatibility is excellent.

[0031] The above test parameters are configured as follows: Basic test parameters Number of parallel experimental groups: 3 groups, mean RSD ≤ 3.5% Mouse experiment: 40 SPF-grade Kunming mice were used, with 10 mice in each group (half male and half female), weighing 20±2g. The gavage cycle was 7 days, and the daily gavage volume was 0.2mL / 10g body weight.

[0032] Odor evaluation: A sensory evaluation panel of 10 people, using a 1-10 scoring method.

[0033] Industry standard reference: microbial survival rate ≥75% (industrialization requirement); capsule filling weight difference ≤±3.0% (quality standard); capsule filling qualification rate ≥99.0% (industrialization requirement).

[0034] The values ​​for bacterial community activity testing (core verification items) are shown in Table 1: Table 1 Gastrointestinal irritation test values Positive control (0.1 mol / L hydrochloric acid): gastric mucosal damage index 3.8±0.5, with obvious congestion and edema.

[0035] Blank control (physiological saline): gastric mucosal damage index 0.0±0.0, mucosa smooth and without abnormalities.

[0036] The protective agent group of this invention: gastric mucosal damage index 0.0±0.0, no congestion and edema, and no inflammatory infiltration in pathological sections (P>0.05).

[0037] Related description: Consistent with the ingredient description that "Buddha's Hand and Bitter Orange have no gastrointestinal irritation".

[0038] The odor evaluation test values ​​are shown in Table 2: Table 2 Compatibility test values ​​(compatibility of microbial sludge capsules) The formula of this invention has the following characteristics: 99.5% mushroom mud forming rate, 99.8% filling qualification rate, weight difference ±2.1% (≤±3.0%), 0.0% breakage rate, and 0.0% powder leakage rate.

[0039] Key findings: No clumping or adhesion (0 particles with a clumping diameter ≥ 500 μm), meeting the quality standards for enteric-coated capsules and fully suitable for industrial production.

[0040] The novel formula for the intestinal flora transplantation sludge capsule protectant provided by this invention has the following significant beneficial effects compared to existing technologies: Active protection and stability: Through the core protective effect of inulin, combined with the synergistic assistance of the two medicinal and edible components of Buddha's Hand and Bitter Orange, the survival rate of the microbial community in the bacterial mud preparation still reaches 86.7% after 6 months of storage. It can effectively maintain the activity of the intestinal flora in the bacterial mud during the preparation, storage and transportation process, and extend the shelf life of the bacterial mud capsule preparation to more than 6 months, meeting the needs of industrialization and clinical application.

[0041] High safety: All components are food-grade or pharmaceutical-grade. The combination of Buddha's Hand and Bitter Orange, two medicinal and edible ingredients, causes no gastrointestinal irritation. Animal experiments have verified its good safety profile, making it suitable for long-term use.

[0042] Synergistic medicinal value: The combination of Buddha's Hand and Bitter Orange, two medicinal and edible raw materials, in a 12:3 ratio, achieves complementary medicinal value, which can help improve intestinal function, provide additional support for patients' intestinal health, and enhance the clinical value of the bacterial mud capsule preparation.

[0043] Significant and long-lasting odor masking effect: The synergistic odor masking system formed by Buddha's Hand and Bitter Orange can completely mask the fishy and rotten odor of the mushroom mud. The initial odor score is 9.2 points, and it still maintains 8.8 points after 6 months of storage. It optimizes the palatability of mushroom mud capsule formulation, significantly improves patient compliance, and solves industry pain points.

[0044] Highly adaptable: Specifically designed for bacterial sludge capsule formulations, it has excellent compatibility with bacterial sludge systems, achieving a bacterial sludge formation rate of 99.5% and a capsule filling qualification rate of 99.8%. It is perfectly adapted to industrial production processes such as capsule filling, requiring no adjustment to existing production equipment, making it highly practical and easy for large-scale production.

Claims

1. A gut microbiota transplantation microbial sludge capsule biological preparation, characterized in that, The biological agent components include inulin, apple fruit powder, freeze-dried bergamot powder, freeze-dried bitter orange flower powder, and intestinal flora sludge.

2. The intestinal flora transplantation sludge capsule biological preparation according to claim 1, characterized in that, The biological agent components, by mass parts, are in the following proportions: inulin 72-78 parts, apple fruit powder 9-11 parts, bergamot freeze-dried powder 11-13 parts, and bitter orange freeze-dried powder 2.5-3.5 parts.

3. A method for preparing the intestinal flora transplantation sludge capsule biological agent as described in claim 1, characterized in that, include: Step 1: Prepare freeze-dried Buddha's Hand citron powder and freeze-dried bitter orange flower powder separately, and mix inulin and apple fruit powder with freeze-dried Buddha's Hand citron powder and freeze-dried bitter orange flower powder in proportion to prepare a protective agent; Step 2: Take fresh intestinal microbiota slurry, mix the preservative and fresh intestinal microbiota slurry in proportion, and stir at a stirring rate of 30-50 r / min for 3-5 minutes in a sterile environment at 2-8℃ to obtain mixed microbiota slurry; fill the mixed microbiota slurry into enteric-coated capsule shells, seal and package to obtain intestinal microbiota transplantation microbiota slurry capsules.

4. The method for preparing the intestinal flora transplantation sludge capsule biological agent according to claim 3, characterized in that, In step 1, the bergamot freeze-dried powder is prepared using a low-temperature freeze-drying process. The freeze-drying temperature is 40℃-50℃, and the freeze-drying time is 12-24h, which preserves its natural aromatic substances and active ingredients. The particle size is 100-200 mesh.

5. The method for preparing the intestinal flora transplantation sludge capsule biological agent according to claim 3, characterized in that, In step 1, the freeze-dried powder of the substitute flower is prepared by low-temperature freeze-drying process, with a particle size of 100-200 mesh, retaining natural volatile oil components, and the volatile oil content is above 0.1%.

6. The method for preparing the intestinal flora transplantation sludge capsule biological agent according to claim 3, characterized in that, In step 1, the mass ratio of freeze-dried Buddha's Hand citron powder to freeze-dried bitter orange flower powder is 11:3.5-13:2.5, and inulin and apple powder are added and mixed according to the required ratio of 7.5:

1.

7. The method for preparing the intestinal flora transplantation sludge capsule biological agent according to claim 3, characterized in that, In step 2, the ratio of the preservative to the fresh intestinal flora sludge is 10:1, and the moisture content of the fresh intestinal flora sludge is 60-70%.

8. The method for preparing the intestinal flora transplantation sludge capsule biological agent according to claim 3, characterized in that, In step 2, the aseptic environment humidity is below 35%, and the finished intestinal flora transplantation sludge capsules are sealed and stored in a refrigerated environment at 4-8℃.

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