Antibacterial diuretic traditional Chinese medicine compound preparation and preparation method thereof
By using composite enzymatic hydrolysis technology and low-temperature extraction process, combined with specific Chinese medicinal materials, an antibacterial and diuretic Chinese medicinal compound preparation was prepared, which solved the side effect problem of antibiotics and chemical diuretics in the existing technology and achieved significant antibacterial and diuretic effects.
Patent Information
- Application Number
- CN202510858408.0
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-06-25
- Publication Date
- 2025-09-16
AI Technical Summary
The use of antibiotics and chemical diuretics in existing livestock and pet feeds poses the risk of drug resistance and side effects, and traditional Chinese medicine extraction processes destroy the active ingredients, making it impossible to achieve significant antibacterial and diuretic effects.
By using composite enzymatic hydrolysis technology and low-temperature extraction process, combined with Chinese medicinal materials such as cat's whiskers grass, ground ivy, quassia, corn silk, stone vine, Centella asiatica and hemp seed meal, a Chinese herbal compound preparation with both antibacterial and diuretic functions was prepared.
It significantly improves the bioavailability of active ingredients, specifically inhibits urinary tract pathogens, promotes urine excretion, reduces the risk of electrolyte disorders, and has stable product quality and potential for industrial production.
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Abstract
Description
Technical Field
[0001] The present invention belongs to the technical field of feed additives, and specifically relates to a traditional Chinese medicine compound preparation with both antibacterial and diuretic functions. The compound preparation is suitable for addition to livestock, poultry and pet feed. The preparation method innovatively combines composite enzymatic hydrolysis technology and low-temperature extraction process, which can significantly improve the bioavailability of active ingredients. Background Art
[0002] At present, traditional livestock and pet feed and nutritional supplements on the market mostly add antibiotics to achieve antibacterial and diuretic functions. However, the indiscriminate addition of antibiotics can easily lead to drug resistance, thus limiting their application.
[0003] Chemical diuretics are also used on the market to enhance the antibacterial and diuretic functions, but the use of chemical diuretics also carries the risk of residues and side effects, which limits their further promotion and application.
[0004] In addition, conventional Chinese medicine extraction processes on the market (such as water decoction, alcohol precipitation, etc.) will cause the effective ingredients to be destroyed, the content of small molecule peptides is relatively low, and cannot achieve good antibacterial and diuretic effects.
[0005] Based on this, this application was developed.
[0006] In the present invention, the active ingredients of cat's whiskers grass include rosmarinic acid and phenolic acids, which have antibacterial, diuretic and anti-inflammatory effects.
[0007] Herba Euphorbiae Ichthyophthirii, whose active ingredients include ellagic acid and quercetin, has the effects of blocking adhesion to the urothelium, achieving targeted antibacterial effects, scavenging hydroxyl free radicals, and protecting renal tubules from oxidative damage. Quassin, whose active ingredients include quassin and alkaloids, has the effects of inhibiting Gram-negative bacteria, enhancing the phagocytic activity of macrophages against pathogens, and relieving painful urination.
[0008] Corn silk, whose active ingredients include corn silk polysaccharide and potassium ions, has the effects of balancing electrolytes, providing osmotic diuresis, and lowering uric acid.
[0009] Pyrrosia, whose active ingredients include chlorogenic acid and pyrrostrum polysaccharide, has anti-inflammatory, immune-enhancing, and stone-preventing effects. Centella asiatica, whose active ingredients include asiaticoside and madecassic acid, has the effects of repairing urinary tract epithelial cells, resisting fibrosis, and providing stress protection, alleviating urination anxiety in animals.
[0010] Hemp seed meal, whose active ingredients include hemp seed protein peptide and arginine, has the effects of inhibiting urease, blocking bacterial ammonia production, reducing intestinal colonization of pathogenic bacteria, supplementing nutrition and improving feed protein utilization. Summary of the Invention
[0011] The present invention aims to overcome the shortcomings of the prior art and provide a traditional Chinese medicine compound preparation with significant antibacterial and diuretic effects. The compound preparation is suitable for addition to livestock and pet feed and can significantly improve the bioavailability of the active ingredients.
[0012] Another object of the present invention is to provide a method for preparing the above-mentioned antibacterial and diuretic Chinese medicine compound preparation, which innovatively combines composite enzymatic hydrolysis technology and low-temperature extraction process, and can significantly improve the bioavailability of the active ingredients.
[0013] The third object of the present invention is to provide the use of the above-mentioned antibacterial and diuretic Chinese medicine compound preparation as a feed additive for livestock, poultry and pets.
[0014] In order to achieve the above-mentioned object of the invention, the present invention adopts the following technical solutions: An antibacterial and diuretic traditional Chinese medicine compound preparation is mainly prepared from the following raw materials in parts by weight: 15-25 parts of cat's whiskers grass, 10-20 parts of euphorbia pulex, 5-15 parts of quassia, 20-30 parts of corn silk, 10-15 parts of pyrrosia, 8-12 parts of centella asiatica, and 8-12 parts of hemp seed meal.
[0015] Furthermore, the Chinese medicine compound preparation is mainly made of the following raw materials in parts by weight: 20-25 parts of cat's whiskers grass, 10-15 parts of herba euphorbiae, 10-15 parts of quassia, 20-25 parts of corn silk, 12-15 parts of pyrrosia, 8-10 parts of centella asiatica, and 10-12 parts of hemp seed meal.
[0016] A method for preparing the above-mentioned antibacterial and diuretic Chinese medicine compound preparation comprises the following steps: 1. Raw material pretreatment 1) Cat's Whiskers: Wash and cut into sections, dry to a moisture content of ≤8%, crush and sieve; 2) Euphorbia pulegata: Degreasing to remove fat-soluble impurities; 3) Hemp seed meal: Soak the hemp seed meal in NaOH solution, wash with water until neutral, dry and crush for later use; 4) Inoculate Aspergillus niger onto PDA slant medium and culture at 30 ± 2°C for 48 ± 12 hours; 5) Inoculate Lactobacillus plantarum into MRS liquid medium and culture at 37±1°C for 24±6 hours; 2. Preparation process 1) Mixing Herba Miscanthi, Herba Euphorbiae, Herba Quassima, Herba Corni, Herba Pyrrosiae, and Herba Centellae, adding citric acid buffer for extraction, and performing solid-liquid separation to obtain the medicinal residue and the extract; 2) The extracted medicinal residue is mixed with hemp seed meal, and glucose and ammonium sulfate are added as carbon and nitrogen sources for staged fermentation; In the first stage, the Aspergillus niger obtained in step 4) was added and fermented at 30±2°C for 24±6 hours; In the second stage, the Lactobacillus plantarum obtained in step 5) was added and fermented at 37±1°C for 24±6 hours; 3) The fermentation product is adjusted to a pH of 7-8, and after adding complex enzymes, it is enzymatically hydrolyzed in a 40-60°C water bath for 3-5 hours; 4) Heat at 80-95°C for 8-20 minutes to inactivate the enzyme, centrifuge, and collect the supernatant; 5) The supernatant is mixed with the extract obtained in step 1) and spray-dried to obtain the product.
[0017] Specifically, in the raw material pretreatment, the Herba Euphorbiae Ichthyophthirius is degreased by CO2 supercritical extraction at a pressure of 20-30 MPa and a temperature of 30-50° C. to remove fat-soluble impurities; and the hemp seed meal is soaked in a 0.1-0.2 mol / L NaOH solution for 1-3 hours.
[0018] In the preparation method of the above-mentioned antibacterial and diuretic traditional Chinese medicine compound preparation, citric acid buffer solution can be added at a material-liquid ratio of 1 g: 10-20 mL.
[0019] Furthermore, glucose accounting for 1-2% of the sum of the mass of the medicinal residues and the hemp seed meal and ammonium sulfate accounting for 0.5-1% of the sum of the mass of the medicinal residues and the hemp seed meal can be added as carbon and nitrogen sources.
[0020] Furthermore, Aspergillus niger accounts for 4-6% of the mass of the mixture of the medicinal residue and the hemp seed meal; and Lactobacillus plantarum accounts for 4-6% of the mass of the mixture of the medicinal residue and the hemp seed meal.
[0021] Furthermore, a composite enzyme accounting for 1-2% of the sum of the weight of the medicinal residue and the hemp seed meal is added, wherein the composite enzyme is composed of a mixture of neutral protease and flavor protease in a mass ratio of 2-3:1.
[0022] The present invention also provides the use of the antibacterial and diuretic traditional Chinese medicine compound preparation as a feed additive for livestock, poultry and pets.
[0023] Compared with the prior art, the present invention has the following advantages and beneficial effects: (1) This formula specifically inhibits urinary tract pathogens, promotes urine excretion and reduces the risk of electrolyte imbalance; (2) The present invention preferably adopts high-voltage pulsed electric field (PEF) extraction technology to improve the extraction rate of heat-sensitive components; (3) Using hemp seed meal to replace animal-derived peptides not only reduces the cost of raw materials, but also eliminates any organic solvent residues throughout the entire process; (4) The method of the present invention uses staged solid-state fermentation and enzymatic hydrolysis to convert medicinal residues into functional oligosaccharides and small molecule peptides, which improves resource utilization, stabilizes product quality, and has the potential for industrial production. DETAILED DESCRIPTION
[0024] The technical solution of the present invention is further described in detail below in conjunction with the embodiments, but the protection scope of the present invention is not limited thereto.
[0025] In the following examples, the raw materials used are all commercially available products that can be directly purchased or prepared using conventional methods in the art. For example, hemp seed meal is a by-product of hemp seed oil extraction, and has a protein content of ≥35%.
[0026] Aspergillus niger (strain number: CICC 2462) was purchased from the Chinese Academy of Food Fermentation Industries. Lactobacillus plantarum (strain number: CICC 20265) was purchased from Shanghai Luwei Technology Co., Ltd.
[0027] Neutral protease (200,000 U / g) was purchased from Ningxia Xiasheng Industrial Group Co., Ltd.; flavor protease (450,000 U / g) was purchased from Guangdong Yidoli Biotechnology.
[0028] PDA slant culture medium: purchased from Qingdao Haibo Biotechnology (HKM-P001, pre-made dry powder culture medium).
[0029] MRS liquid culture medium: purchased from Beijing Luqiao Technology (HB0384-1, dehydrated culture medium).
[0030] Example 1 An antibacterial and diuretic traditional Chinese medicine compound preparation containing a small molecule peptide The composition and weight ratio of the invention are as follows: 20 parts of cat's whiskers grass, 15 parts of herba euphorbiae, 10 parts of quassia, 25 parts of corn silk, 12 parts of pyrrosia, 10 parts of centella asiatica and 10 parts of hemp seed meal.
[0031] Preparation method: 1. Raw material pretreatment 1. Cat's Whiskers: Wash and cut into 1-2 cm segments, dry with hot air at 50°C until the moisture content is ≤8%, and grind through a 60-mesh sieve. 2. Euphorbia pulex: Use CO2 supercritical extraction to remove fat (pressure 25MPa, temperature 40℃) to remove fat-soluble impurities; 3. Hemp seed meal: Soak hemp seed meal (protein content ≥35%) in 0.1mol / L NaOH solution for 2 hours, wash with water until neutral, dry and crush for later use; 4. Inoculate Aspergillus niger into PDA slant medium and culture at 30°C for 48 hours; 5. Inoculate Lactobacillus plantarum into MRS liquid culture medium and culture at 37°C for 24 hours.
[0032] 2. Preparation process 1. Mix Herba Miscanthi, Herba Euphorbiae, Herba Quassima, Herba Corni, Herba Pyrrosiae, and Herba Centellae, add citric acid buffer (pH 6.0), with a solid-liquid ratio of 1:15 (g / mL), and extract using a high-voltage pulsed electric field (electric field intensity 30 kV / cm, pulse frequency 100 Hz, the same below). After extraction, filter to obtain the medicinal residue and extract; 2. The extracted medicinal residues are mixed with the hemp seed meal, and glucose accounting for 1% of the total mass of the medicinal residues and the hemp seed meal and ammonium sulfate accounting for 0.5% of the total mass of the medicinal residues and the hemp seed meal are added as carbon and nitrogen sources to carry out staged fermentation; The first stage (Aspergillus niger accounts for 5% of the mass of the mixture of medicinal residues and hemp seed meal): add the Aspergillus niger obtained in step 4 and ferment at a temperature of 30°C for 24 hours; The second stage (Lactobacillus plantarum accounts for 5% of the mass of the mixture of medicinal residue and hemp seed meal): add the Lactobacillus plantarum obtained in step 5 and ferment at a temperature of 37°C for 24 hours; 3. The pH of the fermentation product was adjusted to 7.5, and a complex enzyme (the complex enzyme was composed of a mixture of neutral protease and flavor protease in a mass ratio of 2:1) accounting for 1.5% of the sum of the mass of the medicinal residue and the hemp seed meal was added, and enzymolysis was carried out in a 50°C water bath for 4 hours; 4. Heat at 90°C for 10 minutes to inactivate the enzyme, centrifuge (8000 rpm, 15 minutes) and collect the supernatant; 5. Take the supernatant and mix it with the extract in step 1 and spray dry it to obtain the product of the present invention.
[0033] Example 2 An antibacterial and diuretic traditional Chinese medicine compound preparation containing a small molecule peptide The composition and weight ratio of the invention are as follows: 25 parts of cat's whiskers grass, 10 parts of herba euphorbiae, 15 parts of quassia, 20 parts of corn silk, 15 parts of pyrrosia, 8 parts of centella asiatica and 12 parts of hemp seed meal.
[0034] Preparation method: 1. Raw material pretreatment 1. Cat's Whiskers: Wash and cut into 1-2 cm segments, dry with hot air at 50°C until the moisture content is ≤8%, and grind through a 60-mesh sieve. 2. Euphorbia pulex: Use CO2 supercritical extraction to remove fat (pressure 25MPa, temperature 40℃) to remove fat-soluble impurities; 3. Hemp seed meal: Soak hemp seed meal (protein content ≥35%) in 0.1mol / L NaOH solution for 2 hours, wash with water until neutral, dry and crush for later use; 4. Inoculate Aspergillus niger into PDA slant medium and culture at 30°C for 48 hours; 5. Inoculate Lactobacillus plantarum into MRS liquid culture medium and culture at 37°C for 24 hours.
[0035] 2. Preparation process 1. Mix the herbs of Herba Miscanthifolia, Herba Euphorbiae Herba, Herba Quassiae, Herba Corni Stigmae, Herba Pyrrosiae, and Herba Centellae, add citric acid buffer solution (pH 6.0), and the solid-liquid ratio is 1:15 (g / mL). Extract the herbs using a high-voltage pulsed electric field. After extraction, filter the mixture to obtain the medicinal residue and the extract. 2. The extracted medicinal residues were mixed with the pretreated hemp seed meal, and glucose accounting for 1% of the total mass of the medicinal residues and hemp seed meal and ammonium sulfate accounting for 0.5% of the total mass of the medicinal residues and hemp seed meal were added as carbon and nitrogen sources for staged fermentation: The first stage (Aspergillus niger accounts for 6% of the mass of the mixture of medicinal residues and hemp seed meal): add the Aspergillus niger obtained in step 4 and ferment at a temperature of 30°C for 24 hours; The second stage (Lactobacillus plantarum accounts for 4% of the mass of the mixture of medicinal residue and hemp seed meal): add the Lactobacillus plantarum obtained in step 5 and ferment at a temperature of 37°C for 24 hours; 3. The pH of the fermentation product was adjusted to 7.5, and a complex enzyme (the complex enzyme was composed of a mixture of neutral protease and flavor protease in a mass ratio of 2:1) accounting for 1.5% of the sum of the mass of the medicinal residue and the hemp seed meal was added, and enzymolysis was carried out in a 50°C water bath for 4 hours; 4. Heat at 90°C for 10 minutes to inactivate the enzyme, centrifuge (8000 rpm, 15 minutes) and collect the supernatant; 5. Take the supernatant and mix it with the extract in step 1 and spray dry it to obtain the product of the present invention.
[0036] Example 3 An antibacterial and diuretic traditional Chinese medicine compound preparation containing a small molecule peptide The composition and weight ratio of the preparation are as follows: 23 parts of cat's whiskers grass, 18 parts of herba euphorbiae, 13 parts of quassia, 22 parts of corn silk, 14 parts of pyrrosia, 9 parts of centella asiatica and 11 parts of hemp seed meal.
[0037] Preparation method: 1. Raw material pretreatment 1. Cat's Whiskers: Wash and cut into 1-2 cm segments, dry with hot air at 50°C until the moisture content is ≤8%, and grind through a 60-mesh sieve. 2. Euphorbia pulex: Use CO2 supercritical extraction to remove fat (pressure 25MPa, temperature 40℃) to remove fat-soluble impurities; 3. Hemp seed meal: Soak hemp seed meal (protein content ≥35%) in 0.1mol / L NaOH solution for 2 hours, wash with water until neutral, dry and crush for later use; 4. Inoculate Aspergillus niger into PDA slant medium and culture at 30°C for 48 hours; 5. Inoculate Lactobacillus plantarum into MRS liquid culture medium and culture at 37°C for 24 hours.
[0038] 2. Preparation process 1. Mix the herbs of Herba Miscanthifolia, Herba Euphorbiae Herba, Herba Quassiae, Herba Corni Stigmae, Herba Pyrrosiae, and Herba Centellae, add citric acid buffer solution (pH 6.0), and the solid-liquid ratio is 1:15 (g / mL). Extract the herbs using a high-voltage pulsed electric field. After extraction, filter the mixture to obtain the medicinal residue and the extract. 2. The extracted medicinal residues were mixed with the pretreated hemp seed meal, and glucose accounting for 1% of the total mass of the medicinal residues and hemp seed meal and ammonium sulfate accounting for 0.5% of the total mass of the medicinal residues and hemp seed meal were added as carbon and nitrogen sources for staged fermentation: The first stage (Aspergillus niger accounts for 5% of the mass of the mixture of medicinal residues and hemp seed meal): add the Aspergillus niger obtained in step 4 and ferment at a temperature of 30°C for 24 hours; The second stage (Lactobacillus plantarum accounts for 5% of the mass of the mixture of medicinal residues and hemp seed meal): add the Lactobacillus plantarum obtained in step 5 and ferment at a temperature of 37°C for 24 hours; 3. The pH of the fermentation product was adjusted to 7.5, and a complex enzyme (the complex enzyme was composed of a mixture of neutral protease and flavor protease in a mass ratio of 2:1) accounting for 1.5% of the sum of the mass of the medicinal residue and the hemp seed meal was added, and enzymolysis was carried out in a 50°C water bath for 4 hours; 4. Heat at 90°C for 10 minutes to inactivate the enzyme, centrifuge (8000 rpm, 15 minutes) and collect the supernatant; 5. Take the supernatant and mix it with the extract in step 1 and spray dry it to obtain the product of the present invention.
[0039] Comparative Example 1: No enzymatic treatment The composition and weight ratio of the invention are as follows: 20 parts of cat's whiskers grass, 15 parts of herba euphorbiae, 10 parts of quassia, 25 parts of corn silk, 12 parts of pyrrosia, 10 parts of centella asiatica and 10 parts of hemp seed meal.
[0040] Preparation method: 1. Raw material pretreatment 1. Cat's Whiskers: Wash and cut into 1-2 cm segments, dry with hot air at 50°C until the moisture content is ≤8%, and grind through a 60-mesh sieve. 2. Euphorbia pulex: Use CO2 supercritical extraction to remove fat (pressure 25MPa, temperature 40℃) to remove fat-soluble impurities; 3. Hemp seed meal: Soak hemp seed meal (protein content ≥35%) in 0.1mol / L NaOH solution for 2 hours, wash with water until neutral, dry and crush for later use; 4. Inoculate Aspergillus niger into PDA slant medium and culture at 30°C for 48 hours; 5. Inoculate Lactobacillus plantarum into MRS liquid culture medium and culture at 37°C for 24 hours.
[0041] 2. Preparation process 1. Mix the herbs of Herba Miscanthifolia, Herba Euphorbiae Herba, Herba Quassiae, Herba Corni Stigmae, Herba Pyrrosiae, and Herba Centellae, add citric acid buffer solution (pH 6.0), and the solid-liquid ratio is 1:15 (g / mL). Extract the herbs using a high-voltage pulsed electric field. After extraction, filter the mixture to obtain the medicinal residue and the extract. 2. The extracted medicinal residues were mixed with the pretreated hemp seed meal, and glucose accounting for 1% of the total mass of the medicinal residues and hemp seed meal and ammonium sulfate accounting for 0.5% of the total mass of the medicinal residues and hemp seed meal were added as carbon and nitrogen sources for staged fermentation: The first stage (Aspergillus niger accounts for 5% of the mass of the mixture of medicinal residues and hemp seed meal): add the Aspergillus niger obtained in step 4 and ferment at a temperature of 30°C for 24 hours; The second stage (Lactobacillus plantarum accounts for 5% of the mass of the mixture of medicinal residues and hemp seed meal): add the Lactobacillus plantarum obtained in step 5 and ferment at a temperature of 37°C for 24 hours; 3. After fermentation, centrifuge (8000 rpm, 15 minutes) and take the supernatant; 4. Take the supernatant and mix it with the extract in step 1 and spray dry it to obtain this product.
[0042] Comparative Example 2 Adjusting the ratio of raw materials The composition and proportion by weight are as follows: 35 parts of cat's whiskers grass, 10 parts of herba euphorbiae, 15 parts of quassia, 10 parts of corn silk, 15 parts of pyrrosia, 8 parts of centella asiatica and 12 parts of hemp seed meal.
[0043] Preparation method: 1. Raw material pretreatment 1. Cat's Whiskers: Wash and cut into 1-2 cm segments, dry with hot air at 50°C until the moisture content is ≤8%, and grind through a 60-mesh sieve. 2. Euphorbia pulex: Use CO2 supercritical extraction to remove fat (pressure 25MPa, temperature 40℃) to remove fat-soluble impurities; 3. Hemp seed meal: Soak hemp seed meal (protein content ≥35%) in 0.1mol / L NaOH solution for 2 hours, wash with water until neutral, dry and crush for later use; 4. Inoculate Aspergillus niger into PDA slant medium and culture at 30°C for 48 hours; 5. Inoculate Lactobacillus plantarum into MRS liquid culture medium and culture at 37°C for 24 hours.
[0044] 2. Preparation process 1. Mix the herbs of Herba Miscanthifolia, Herba Euphorbiae Herba, Herba Quassiae, Herba Corni Stigmae, Herba Pyrrosiae, and Herba Centellae, add citric acid buffer solution (pH 6.0), and the solid-liquid ratio is 1:15 (g / mL). Extract the herbs using a high-voltage pulsed electric field. After extraction, filter the mixture to obtain the medicinal residue and the extract. 2. The extracted medicinal residues are mixed with the hemp seed meal, and glucose accounting for 1% of the total mass of the medicinal residues and the hemp seed meal and ammonium sulfate accounting for 0.5% of the total mass of the medicinal residues and the hemp seed meal are added as carbon and nitrogen sources to carry out staged fermentation; The first stage (Aspergillus niger accounts for 6% of the mass of the mixture of medicinal residues and hemp seed meal): add the Aspergillus niger obtained in step 4 and ferment at a temperature of 30°C for 24 hours; The second stage (Lactobacillus plantarum accounts for 4% of the mass of the mixture of medicinal residue and hemp seed meal): add the Lactobacillus plantarum obtained in step 5 and ferment at a temperature of 37°C for 24 hours; 3. The pH of the fermentation product was adjusted to 7.5, and a complex enzyme (the complex enzyme was composed of a mixture of neutral protease and flavor protease in a mass ratio of 2:1) accounting for 1.5% of the sum of the mass of the medicinal residue and the hemp seed meal was added, and enzymolysis was carried out in a 50°C water bath for 4 hours; 4. Heat at 90°C for 10 minutes to inactivate the enzyme, centrifuge (8000 rpm, 15 minutes) and collect the supernatant; 5. Take the supernatant and mix it with the extract in step 1 and spray dry it to obtain this product.
[0045] Comparative Example 3 Single strain fermentation The composition and weight ratio of the invention are as follows: 20 parts of cat's whiskers grass, 15 parts of herba euphorbiae, 10 parts of quassia, 25 parts of corn silk, 12 parts of pyrrosia, 10 parts of centella asiatica and 10 parts of hemp seed protein peptide.
[0046] Preparation method: 1. Raw material pretreatment 1. Cat's Whiskers: Wash and cut into 1-2 cm segments, dry with hot air at 50°C until the moisture content is ≤8%, and grind through a 60-mesh sieve. 2. Euphorbia pulex: Use CO2 supercritical extraction to remove fat (pressure 25MPa, temperature 40℃) to remove fat-soluble impurities; 3. Hemp seed meal: Soak hemp seed meal (protein content ≥35%) in 0.1mol / L NaOH solution for 2 hours, wash with water until neutral, dry and crush for later use; 4. Inoculate Aspergillus niger into PDA slant medium and culture at 30℃ for 48 hours.
[0047] 2. Core Preparation Process 1. Mix the herbs of Herba Miscanthifolia, Herba Euphorbiae Herba, Herba Quassiae, Herba Corni Stigmae, Herba Pyrrosiae, and Herba Centellae, add citric acid buffer solution (pH 6.0), and the solid-liquid ratio is 1:15 (g / mL). Extract the herbs using a high-voltage pulsed electric field. After extraction, filter the mixture to obtain the medicinal residue and the extract. 2. The extracted medicinal residues were mixed with the pre-treated hemp seed dregs, and glucose accounting for 1% of the total mass of the medicinal residues and the hemp seed dregs, and ammonium sulfate accounting for 0.5% of the total mass of the medicinal residues and the hemp seed dregs, and the aspergillus niger obtained in step 4 (accounting for 5% of the mass of the medicinal residues and the hemp seed dregs mixture) was added and fermented at 30°C for 48 hours; 3. The pH of the fermentation product was adjusted to 7.5, and a complex enzyme (the complex enzyme was composed of a mixture of neutral protease and flavor protease in a mass ratio of 2:1) accounting for 1.5% of the sum of the mass of the medicinal residue and the hemp seed meal was added, and enzymolysis was carried out in a 50°C water bath for 4 hours; 4. Heat at 90°C for 10 minutes to inactivate the enzyme, centrifuge (8000 rpm, 15 minutes) and collect the supernatant; 5. Take the supernatant and mix it with the extract in step 1 and spray dry it to obtain this product.
[0048] Test Example 1: Detection of small molecule peptide content and antioxidant capacity 1. Test materials The products prepared in Example 1, Comparative Example 1, Comparative Example 2 and Comparative Example 3.
[0049] 2. Test methods 2.1 Detection indicators 2.2 Test results 2.2.1 Comparison of peptide molecular weight distribution As can be seen from the table above, the proportion of large molecular peptides >1000 Da in the non-enzymatic hydrolysis group and the single-bacteria fermentation group increased significantly (p < 0.01), while small molecular peptides (<500 Da) decreased by 67% and 43%, respectively. The proportion of small molecular peptides (<500 Da) in the product of the embodiment of the present invention was as high as 68.2%, significantly higher than the 22.5% in Comparative Example 1 and 38.7% in Comparative Example 3.
[0050] 2. Differences in functional activity DPPH IC50: The sample concentration required to achieve a 50% DPPH free radical scavenging rate. A lower IC50 indicates a stronger free radical scavenging ability per unit concentration of the sample.
[0051] As shown in the table above, compared with the product of Example 1 of the present invention, the DPPH IC50 of the non-enzymatic hydrolysis group of Comparative Example 1 increased by 232%, and the DPPH IC50 of the raw material ratio-adjusted group of Comparative Example 2 increased to 0.58, an increase of 132%. This shows that the antioxidant capacity of the product of Example 1 of the present invention is stronger.
[0052] From this, we can see that enzymatic hydrolysis can convert large molecular proteins into small molecular active peptides (<500 Da), and small molecular peptides are easier to penetrate cell membranes to exert antioxidant effects. At the same time, the proportion of the original formula can better exert a synergistic effect.
[0053] Experimental Example 2: Detection of Metabolites and ABTS Free Radical Scavenging Rate (IC50) 1. Test materials Products prepared in Example 1 and Comparative Example 3 2. Test methods 2.1 Detection indicators 3. Test results Metabolites and functional activity differences As can be seen from the above table, the anti-ABTS IC50 of Example 1 group was reduced by 43.18% compared with that of Comparative Example 2, and the diversity of metabolites doubled from 6 in Comparative Example 2 to 12 phenolic acid species, thereby enhancing the synergistic effect of antioxidant and anti-inflammatory functions.
[0054] Test Example 3 Comparison of antibacterial and diuretic effects 1. Test method 1.1 Experimental Design Subjects: 180 urinary tract infection model rats (half male and half female, weighing 200±20 g) were randomly divided into 4 groups: patent group (Example 1), non-enzymatic hydrolysis group (Comparative Example 1), raw material ratio adjustment group (Comparative Example 2), and single strain fermentation group (Comparative Example 3).
[0055] Usage: 0.5 g / kg·d was administered by gavage for 7 consecutive days to observe the antibacterial and diuretic effects.
[0056] 1.2 Detection methods and indicators 2. Test results 2.1 Comparison of antibacterial effects Note: ▲ indicates significant difference from the patent group (p<0.01) As can be seen from the above table, the antibacterial effect of the patent group of Example 1 is significantly better than that of the comparative groups, with the bacterial load reduced to 1 / 40-1 / 32, and the levels of inflammatory factors (IL-6, TNF-α) decreased by 50%-70%.
[0057] 2.2 Comparison of diuretic effects As can be seen from the table above, compared with the control groups, the 24-hour urine volume, urine sodium and urine potassium excretion of the patent group of Example 1 increased significantly, indicating that it can significantly enhance the diuretic effect of the test animals.
[0058] Conclusion: Comparative Example 1 systematically verified the decisive role of the composite enzymatic hydrolysis process on the functional activity of the product by eliminating the enzymatic hydrolysis step.
[0059] Comparative Example 2 verified the rationality of the original formula ratio by changing the raw material ratio.
[0060] Comparative Example 3, by eliminating the Lactobacillus plantarum fermentation stage, clearly reveals the decisive role of dual-bacteria synergy and staged parameter regulation (temperature, pH) on the functional activity of the product.
[0061] Clinical trial data confirms that the present invention, through the production of small molecule peptides, optimization of raw material ratios and dual-bacteria staged fermentation, is significantly superior to traditional processes in terms of antibacterial and diuretic function and safety, and has clear innovation and application value.
Claims
1. An antibacterial and diuretic Chinese medicine compound preparation, characterized in that: The traditional Chinese medicine compound preparation is mainly prepared from the following raw materials in parts by weight: 15-25 parts of cat's whiskers, 10-20 parts of herba euphorbiae, 5-15 parts of quassia, 20-30 parts of corn silk, 10-15 parts of pyrrosia, 8-12 parts of centella asiatica and 8-12 parts of hemp seed meal.
2. The antibacterial and diuretic Chinese medicine compound preparation according to claim 1, wherein: The traditional Chinese medicine compound preparation is mainly prepared from the following raw materials in parts by weight: 20-25 parts of cat's whiskers, 10-15 parts of euphorbia pulex, 10-15 parts of quassia, 20-25 parts of corn silk, 12-15 parts of pyrrosia, 8-10 parts of centella asiatica and 10-12 parts of hemp seed meal.
3. The method for preparing the antibacterial and diuretic Chinese medicinal compound preparation according to claim 1 or 2, characterized in that: The steps include:
1. Raw material pretreatment 1) Cat's Whiskers: Wash and cut into sections, dry to a moisture content of ≤8%, crush and sieve; 2) Euphorbia pulegata: Degreasing to remove fat-soluble impurities; 3) Hemp seed meal: Soak the hemp seed meal in NaOH solution, wash with water until neutral, dry and crush for later use; 4) Inoculate Aspergillus niger onto PDA slant medium and culture at 30 ± 2°C for 48 ± 12 hours; 5) Inoculate Lactobacillus plantarum into MRS liquid medium and culture at 37±1°C for 24±6 hours; 2. Preparation process 1) Mixing Herba Miscanthi, Herba Euphorbiae, Herba Quassima, Herba Corni, Herba Pyrrosiae, and Herba Centellae, adding citric acid buffer for extraction, and performing solid-liquid separation to obtain the medicinal residue and the extract; 2) The extracted medicinal residue is mixed with hemp seed meal, and glucose and ammonium sulfate are added as carbon and nitrogen sources for staged fermentation; In the first stage, the Aspergillus niger obtained in step 4) was added and fermented at 30±2°C for 24±6 hours; In the second stage, the Lactobacillus plantarum obtained in step 5) was added and fermented at 37±1°C for 24±6 hours; 3) The fermentation product is adjusted to a pH of 7-8, and after adding complex enzymes, it is enzymatically hydrolyzed in a 40-60°C water bath for 3-5 hours; 4) Heat at 80-95℃ to inactivate the enzyme, centrifuge, and collect the supernatant; 5) The supernatant is mixed with the extract obtained in step 1) and spray-dried to obtain the product.
4. The method for preparing the antibacterial and diuretic Chinese medicine compound preparation as claimed in claim 3, characterized in that: In the raw material pretreatment, the Herba Euphorbiae Ichthyophthirius is degreased by CO2 supercritical extraction at a pressure of 20-30 MPa and a temperature of 30-50°C; the hemp seed meal is soaked in a 0.1-0.2 mol / L NaOH solution for 1-3 hours.
5. The method for preparing the antibacterial and diuretic Chinese medicine compound preparation as claimed in claim 3, characterized in that: Add citric acid buffer at a material-liquid ratio of 1g:10-20mL.
6. The method for preparing the antibacterial and diuretic Chinese medicine compound preparation as claimed in claim 3, characterized in that: Glucose accounting for 1-2% of the sum of the mass of the medicinal residues and the hemp seed meal and ammonium sulfate accounting for 0.5-1% of the sum of the mass of the medicinal residues and the hemp seed meal are added as carbon and nitrogen sources.
7. The method for preparing the antibacterial and diuretic Chinese medicine compound preparation as claimed in claim 3, characterized in that: Aspergillus niger accounts for 4-6% of the mass of the mixture of medicinal residues and hemp seed meal; Lactobacillus plantarum accounts for 4-6% of the mass of the mixture of medicinal residues and hemp seed meal.
8. The method for preparing the antibacterial and diuretic Chinese medicine compound preparation as claimed in claim 3, characterized in that: A composite enzyme accounting for 1-2% of the total mass of the medicinal residues and the hemp seed meal is added, wherein the composite enzyme is composed of a mixture of neutral protease and flavor protease in a mass ratio of 2-3:
1.
9. Use of the antibacterial and diuretic Chinese medicine compound preparation according to claim 1 or 2 as a feed additive for livestock, poultry and pets.