Breast perfusate for treating cow mastitis and preparation method thereof

Abstract

The invention discloses a breast perfusate for treating cow mastitis and a preparation method thereof, and relates to the technical field of veterinary drugs, the efficient viscosity reducer comprises the following components: herbaceous plant flavonoid extracts, lactobacillus fermentation metabolites, a suspending aid, a stabilizer, an osmotic pressure regulator and purified water; according to the invention, the herbaceous plant flavonoid extract and the lactobacillus fermentation metabolite are combined, the two components both have obvious antibacterial and anti-inflammatory effects, and the herbaceous plant flavonoid extract is rich in active components such as chlorogenic acid and dandelion flavones, so that the growth of pathogenic microorganisms can be effectively inhibited; lactic acid and bacteriocin in the lactobacillus fermentation metabolite can further enhance the antibacterial effect, regulate the microbial environment of the milk area and prevent infection recurrence, so that the combination not only improves the treatment effect, but also realizes the combination of treatment and prevention, and is beneficial to long-term management of the cow mastitis.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of veterinary medicine, in particular to a kind of treatment of cow mastitis breast perfusion fluid and preparation method thereof. BACKGROUND

[0002] In the dairy industry, mastitis is one of the main diseases affecting the health and production performance of dairy cows, which has brought huge economic losses to the industry. Mastitis not only leads to a decrease in milk production and quality, but also can cause systemic infection in dairy cows, and even death in severe cases.

[0003] Traditional treatment methods for cow mastitis mainly rely on antibiotic drugs. Although these drugs can control the disease to some extent, they have many shortcomings. First, long-term use of antibiotics can lead to drug resistance in pathogenic microorganisms, reducing the effectiveness of treatment and even rendering it ineffective. Second, antibiotic residues in the body of dairy cows can enter the human body through milk, posing a potential risk to consumer health. In addition, traditional antibiotic treatment can only control existing infections and is difficult to effectively regulate the microbial environment of the milk area to prevent the recurrence of infection.

[0004] In view of the shortcomings of traditional treatment methods for cow mastitis, such as drug residues, drug resistance, and insufficient ability to prevent the recurrence of infection, the present application has developed a breast perfusion fluid for treating cow mastitis and a preparation method thereof, which is particularly important. SUMMARY

[0005] The present application aims to overcome the shortcomings of the prior art and provides a breast perfusion fluid for treating cow mastitis and a preparation method thereof. By utilizing active ingredients from herbal plants and metabolic products of lactic acid bacteria, the present application achieves a comprehensive effect of antibacterial, anti-inflammatory, and regulation of the microbial environment of the milk area, thereby improving the treatment effect and effectively preventing the recurrence of infection. Moreover, the perfusion fluid uses natural ingredients, avoiding drug residues and drug resistance, and providing a safe and effective new approach for the treatment of cow mastitis.

[0006] To solve the above technical problems, the present application provides the following technical solution: a breast perfusion fluid for treating cow mastitis, prepared from the following raw materials in parts by weight: Herbal plant flavonoid extract, lactic acid bacteria fermentation metabolite, suspending agent, stabilizer, osmotic pressure regulator, and purified water; The herbal plant flavonoid extract is a mixture of honeysuckle extract and dandelion extract; The lactic acid bacteria fermentation metabolite is the supernatant after centrifugation and ultrafiltration of a mixed fermentation broth of Lactobacillus plantarum and Lactobacillus acidophilus.

[0007] Further, the herbal flavonoids extract 5-12 parts, lactic acid bacteria fermentation metabolites 8-15 parts, suspending agent 2-5 parts, stabilizer 1-3 parts, osmotic pressure regulator 3-8 parts, and purified water to 100 parts.

[0008] Further, in the herbal flavonoids extract, the mass ratio of honeysuckle extract to dandelion extract is 1:1.5-2.5; and the content of chlorogenic acid in the honeysuckle extract is ≥8%, and the content of dandelion flavone in the dandelion extract is ≥10%.

[0009] Further, in the lactic acid bacteria fermentation metabolites, the content of lactic acid is 15-20 g / L, and the content of bacteriocin is 0.8-1.2 g / L, and the ultrafiltration process uses an ultrafiltration membrane with a molecular weight cut-off of 5000 Da.

[0010] Further, the suspending agent is hydroxypropyl methyl cellulose K4M, the stabilizer is vitamin E acetate, and the osmotic pressure regulator is glycerol; and the osmotic pressure of the perfusion solution is controlled at 280-320 mOsm / kg, and the pH value is adjusted to 6.5-7.0.

[0011] On the other hand, a preparation method of a mammary perfusion solution for treating dairy cow mastitis, characterized in that the preparation method comprises the following steps: Preparation of herbal flavonoids extract: the herbal flavonoids extract is prepared by low-temperature ultrasonic extraction-membrane separation purification process, specifically including: mixing honeysuckle and dandelion and crushing, then ultrasonic extraction with 50% ethanol aqueous solution at 35-40℃, power 300-400W, 2 times, 30-45 minutes each time, the extract is microfiltered with 0.22μm organic filter membrane to remove impurities, then purified by ultrafiltration membrane with a molecular weight cut-off of 3000 Da, finally concentrated under reduced pressure at 50-55℃, vacuum degree-0.08 to-0.09 MPa, to solid content 20-25%, to prepare honeysuckle-dandelion mixed flavonoids extract, ready for use; Preparation of lactic acid bacteria fermentation metabolites: inoculate Lactobacillus plantarum and Lactobacillus acidophilus into MRS medium respectively, anaerobic culture at 37℃ for 18-24 hours to obtain single bacterial fermentation broth, mix according to volume ratio 1:1, then transfer to culture medium containing 10% sucrose, 5% soybean peptone and 2% yeast extract, pH 6.5, anaerobic fermentation at 37℃ for 36-48 hours, centrifuge the fermentation broth at 4℃, 8000r / min for 15 minutes, filter the supernatant through an ultrafiltration membrane with a molecular weight cut-off of 5000 Da to obtain the metabolites, ready for use; Mixing preparation: take purified water and heat to 40-45℃, add osmotic pressure regulator and stir to dissolve, add suspending agent and stabilizer in sequence, stir at a rotation speed of 300-500 r / min for 30-40 min to form a base solution; add flavonoids extract and metabolites to the base solution, continue to stir for 60-90 min until uniform and no particles, adjust the pH value to 6.5-7.0, add purified water to 100 parts, filter through a 0.22 mu sterile filter membrane to sterilize, and then pack into sterile filling devices and seal.

[0012] Further, in the preparation of the herbal plant flavonoids extract, the particle size of the mixture of honeysuckle and dandelion after crushing is controlled to be 40-60 mesh, and the amount of 50% ethanol aqueous solution added during ultrasonic extraction is 10-15 times the total mass of the raw material.

[0013] Further, in the mixing preparation, the whole mixing preparation process is carried out in a sterile environment, the sterile filling device used for packing is a soft silica gel nozzle, and the capacity of the filling device can be adjusted to 20-50 mL according to the size of the cow teat.

[0014] Compared with the prior art, the mammary gland perfusion solution for treating cow mastitis and the preparation method thereof have the following beneficial effects: I. The herbal plant flavonoids extract and the lactic acid bacteria fermentation metabolite have significant antibacterial and anti-inflammatory effects, the herbal plant flavonoids extract is rich in active ingredients such as chlorogenic acid and dandelion flavonoids, and can effectively inhibit the growth of pathogenic microorganisms; the lactic acid and bacteriocin in the lactic acid bacteria fermentation metabolite can further enhance the antibacterial effect, regulate the microbial environment of the milk area, and prevent the recurrence of infection, so that the combination not only improves the treatment effect, but also realizes the combination of treatment and prevention, which is helpful for the long-term management of cow mastitis.

[0015] II. The present application uses natural herbal plant extract and lactic acid bacteria fermentation product as the main component, which avoids the drug residue and drug resistance problems that may be caused by traditional antibiotics, at the same time, by accurately controlling the osmotic pressure and pH value of the perfusion solution, the mildness of the product to the cow mammary gland tissue is ensured, the irritation and discomfort are reduced, in addition, the sterile filling device used for packing adopts a soft silica gel nozzle, and the capacity can be adjusted according to the size of the cow teat, which further improves the comfort and accuracy of use, thereby effectively improving the welfare level of the cow.

[0016] Other advantages, objects, and features of the present application will be in part apparent and in part pointed out hereinafter in the specification, and in part will be learned from the practice of the application. BRIEF DESCRIPTION OF DRAWINGS

[0017] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the drawings needed to be used in the embodiments or prior art description will be briefly introduced. Obviously, the drawings in the following description are only some embodiments of the present application, and other drawings can be obtained by those skilled in the art without creative effort on the basis of these drawings.

[0018] Figure 1 A kind of treatment of bovine mastitis breast infusion liquid and its preparation method flow chart. DETAILED DESCRIPTION

[0019] In order to further illustrate the technical means and effects adopted by the present application to achieve the predetermined object of the application, the specific embodiments, structures, features and effects according to the present application will be described in detail below in combination with the drawings and preferred embodiments.

[0020] Examples one to three only change the content of herbal flavonoids extract Example one

[0021] The content of herbal flavonoids extract is 5 parts, and the mass ratio of honeysuckle extract to dandelion extract is 1:2. It is detected that the content of chlorogenic acid in honeysuckle extract is 9.2%, and the content of dandelion flavonoids in dandelion extract is 11.5%, which meets the quality requirements.

[0022] Lactic acid bacteria fermentation metabolites: 12 parts, the content of lactic acid is 18 g / L, and the content of bacteriocin is 1.0 g / L, which meets the technical index.

[0023] Suspending agent: 3.5 parts, hydroxypropyl methyl cellulose K4M is selected.

[0024] Stabilizer: 2 parts, vitamin E acetate is used.

[0025] Osmotic pressure regulator: 5.5 parts, glycerol is used.

[0026] Purified water: supplement to 100 parts, to ensure that the water quality meets the standard of sterile injection water.

[0027] Raw material pretreatment: mix honeysuckle and dandelion according to the mass ratio of 1:2, put them into the pulverizer for crushing, control the particle size through 40-60 mesh standard sieve, and get the mixed powder raw material.

[0028] Ultrasonic extraction: take the mixed powder raw material above, add 12 times of the total mass of 50% ethanol aqueous solution, place it in an ultrasonic extraction tank, set the extraction temperature to 38℃, the ultrasonic power to 350W, and perform ultrasonic extraction twice, each time for 40 minutes. Collect the two extraction liquids and combine them. Membrane separation and purification: the combined extraction liquid is micro-filtered through a 0.22μm organic filter membrane to remove impurities, raw material residues and macromolecular impurities; then the micro-filtered filtrate is purified through an ultrafiltration membrane with a molecular weight cut-off of 3000Da to cut off ineffective macromolecular substances, and the ultrafiltration permeate is collected. Reduced pressure concentration: transfer the ultrafiltration permeate into a reduced pressure concentration tank, set the concentration temperature to 52℃, the vacuum degree to -0.085MPa, and perform reduced pressure concentration until the solid content in the concentrated liquid reaches 23%, then stop the concentration to obtain the herbal plant flavonoid extract, which is sealed and refrigerated for standby.

[0029] Single-bacterium culture: respectively inoculate the Lactobacillus plantarum strain and the Lactobacillus acidophilus strain into sterilized MRS medium, and place it in a 37℃ anaerobic incubator for 22 hours to obtain Lactobacillus plantarum single-bacterium fermentation liquid and Lactobacillus acidophilus single-bacterium fermentation liquid.

[0030] Mixed fermentation: mix the two single-bacterium fermentation liquids uniformly at a volume ratio of 1:1, and transfer the mixed fermentation liquid into sterilized special medium, the medium composition is 10% sucrose, 5% soybean peptone, 2% yeast extract, and the pH value is adjusted to 6.5 in advance, and the mixed fermentation liquid is fermented in an anaerobic environment at 37℃ for 42 hours to obtain the mixed fermentation liquid.

[0031] Centrifugal separation: transfer the mixed fermentation liquid into a high-speed centrifuge, set the temperature to 4℃, the rotation speed to 8000r / min, and centrifuge for 15 minutes to remove the bacterial bodies and precipitate impurities, and collect the supernatant. Ultrafiltration purification: filter the supernatant after centrifugation through an ultrafiltration membrane with a molecular weight cut-off of 5000Da to further remove macromolecular impurities, and obtain the lactic acid bacteria fermentation metabolite, which is sealed and refrigerated for standby.

[0032] Preparation of base liquid: take an appropriate amount of purified water, heat it to 42℃, add 5.5 parts of glycerol, stir until completely dissolved; then add 3.5 parts of hydroxypropyl methylcellulose K4M and 2 parts of vitamin E acetate in sequence, adjust the stirring equipment speed to 400r / min, and continue stirring for 35 minutes to form a uniform and transparent base liquid.

[0033] Raw material mixing: slowly add 8 parts of herbal plant flavonoid extract and 12 parts of lactic acid bacteria fermentation metabolite to the base liquid under stirring, continue stirring at a speed of 400r / min for 75 minutes until the mixed liquid is uniform and particle-free to form the preliminary perfusion liquid.

[0034] Parameter adjustment and feeding: the pH value of the initial perfusion solution is adjusted to 6.8 using a pH adjuster; then purified water is added to a total mass of 100 parts, and stirring is performed for 5 minutes to ensure uniform mixing.

[0035] Sterilization and sub-packaging: the adjusted perfusion solution is filtered and sterilized through a 0.22 μm sterile filter, and the entire process is operated in a Class 100 sterile environment; after sterilization, the perfusion solution is sub-packaged into sterile perfusion containers, the perfusion containers use soft silicone nozzles, the capacity is selected to be 30 mL, and after sub-packaging is completed, the containers are immediately sealed, labeled, and stored in a cold storage at 2-8℃, the final product has an osmotic pressure of 302 mOsm / kg and a pH of 6.7. Effects: test subjects: 30 clinically mastitis Holstein cows, treatment regimen: 30 mL of perfusion per day for 5 consecutive days. Improvement of clinical symptoms: after 7 days of treatment, the clinical cure rate was 56.7% and the symptom relief rate was 100%; after 14 days of treatment, the final clinical cure rate was 80%, 4 cows needed to extend the treatment for 3 days, and 2 cows still had mild swelling. Pathogen clearance: after 7 days of treatment, the total clearance rate was 66.7%, 75% for streptococcus, 60% for E. coli, and 62.5% for Staphylococcus aureus; after 14 days of treatment, the total clearance rate was 83.3%, 91.7% for streptococcus, 80% for E. coli, and 75% for Staphylococcus aureus, and 3 cows were detected with a small amount of pathogenic bacteria.

[0036] Milk quality recovery: after 14 days of treatment, the average somatic cell count was 258 million cells / mL, the recovery rate was 70%, the milk protein was 2.9%, the recovery rate was 80%, and the milk fat was 3.4%, the recovery rate was 90%. Production performance: after 14 days of treatment, the daily milk yield of the affected side was 8.9 kg, and the recovery to the healthy milk area was 71.2%; after 30 days of treatment, the daily milk yield of the affected side was 11.2 kg, and the recovery to the healthy milk area was 90.4%.

[0037] Safety: no adverse reactions, and the safety compliance rate was 100%. Example Two

[0038] The herbal flavonoid extract was 10 parts, and the mass ratio of honeysuckle extract to dandelion extract was 1:2. The content of chlorogenic acid in the honeysuckle extract was 9.2%, and the content of dandelion flavonoids in the dandelion extract was 11.5%, both of which met the quality requirements.

[0039] Lactic acid bacteria fermentation metabolites: 12 parts, the lactic acid content was 18 g / L and the bacteriocin content was 1.0 g / L, which met the technical indicators.

[0040] Suspending agent: 3.5 parts, hydroxypropyl methylcellulose K4M was selected.

[0041] Stabilizer: 2 parts, vitamin E acetate was used.

[0042] Osmotic pressure regulator: 5.5 parts, using glycerol.

[0043] Purified water: make up to 100 parts, ensure that the water quality meets the standard of sterile water for injection.

[0044] Raw material pretreatment: mix honeysuckle and dandelion at a mass ratio of 1:2, put them into a pulverizer for crushing, control the particle size through a 40-60 mesh standard sieve to obtain mixed powder raw materials.

[0045] Ultrasonic extraction: take the above-mentioned mixed powder raw materials, add 12 times the total mass of 50% ethanol aqueous solution, place them in an ultrasonic extraction tank, set the extraction temperature to 38°C and the ultrasonic power to 350W, perform ultrasonic extraction for 2 times, each time for 40 minutes, collect and combine the two extraction liquids. Membrane separation and purification: filter the combined extraction liquid through a 0.22μm organic filter membrane for microfiltration to remove raw material residues and macromolecular impurities; then purify the microfiltration filtrate through an ultrafiltration membrane with a molecular weight cut-off of 3000Da to cut off ineffective macromolecular substances and collect the ultrafiltration permeate. Reduced pressure concentration: transfer the ultrafiltration permeate into a reduced pressure concentration tank, set the concentration temperature to 52°C and the vacuum degree to -0.085MPa, perform reduced pressure concentration until the solid content in the concentrated liquid reaches 23%, stop the concentration, and obtain herbal plant flavonoid extract, which is sealed and refrigerated for standby use.

[0046] Single strain culture: respectively inoculate Lactobacillus plantarum and Lactobacillus acidophilus into sterilized MRS medium, and culture them in a 37°C anaerobic incubator for 22 hours to obtain Lactobacillus plantarum single strain fermentation liquid and Lactobacillus acidophilus single strain fermentation liquid.

[0047] Mixed fermentation: uniformly mix the two single strain fermentation liquids at a volume ratio of 1:1, transfer the mixed fermentation liquid into sterilized special medium, the medium composition is 10% sucrose, 5% soybean peptone, and 2% yeast extract, adjust the pH value to 6.5 in advance, and ferment the mixed fermentation liquid in a 37°C anaerobic environment for 42 hours to obtain the mixed fermentation liquid.

[0048] Centrifugal separation: transfer the mixed fermentation liquid into a high-speed centrifuge, set the temperature to 4°C and the rotation speed to 8000r / min, centrifuge for 15 minutes to remove bacterial bodies and precipitate impurities, and collect the supernatant. Ultrafiltration purification: filter the supernatant after centrifugation through an ultrafiltration membrane with a molecular weight cut-off of 5000Da to further remove macromolecular impurities, and obtain lactic acid bacteria fermentation metabolites, which are sealed and refrigerated for standby use.

[0049] Base solution preparation: take a certain amount of purified water, heat to 42℃, add weighed 5.5 parts of glycerol, stir until completely dissolved; then add 3.5 parts of hydroxypropyl methyl cellulose K4M and 2 parts of vitamin E acetate in turn, adjust the stirring equipment speed to 400 r / min, continue stirring for 35 minutes, form a uniform transparent base solution.

[0050] Raw material mixing: under stirring, slowly add 8 parts of herbal flavonoids extract and 12 parts of lactic acid bacteria fermentation metabolites to the base solution, continue stirring at 400 r / min for 75 minutes until the mixture is uniform and particle-free, forming a preliminary perfusion solution.

[0051] Parameter adjustment and feeding: use pH adjuster to adjust the pH of the preliminary perfusion solution to 6.8; then add purified water to a total mass of 100 parts, stir for 5 minutes to ensure uniform mixing.

[0052] Sterilization and dispensing: filter the adjusted perfusion solution through a 0.22 μm sterile filter membrane, the whole process is operated in a hundred-level sterile environment; after sterilization, the perfusion solution is dispensed into sterile perfusion devices, the perfusion devices use soft silicone nozzles, the capacity is selected as 30 mL, after dispensing, immediately seal, label, and store in a 2-8℃ refrigerator, the final product has an osmotic pressure of 306 mOsm / kg and a pH of 6.8.

[0053] Effect: test subjects: 30 clinically diagnosed mastitis Holstein cows, treatment plan: perfusion of 30 mL daily for 5 consecutive days. Improvement of clinical symptoms: after 7 days of treatment, the clinical cure rate was 80% and the symptom relief rate was 100%; after 14 days of treatment, the final clinical cure rate was 96.7% and only 1 cow needed an additional 2 days of treatment. Pathogen clearance: after 7 days of treatment, the total clearance rate was 86.7%, 91.7% for streptococcus, 85% for E. coli, and 83.3% for Staphylococcus aureus; after 14 days of treatment, the total clearance rate was 98.3%, 100% for streptococcus, 100% for E. coli, and 95.8% for Staphylococcus aureus, only 1 cow was detected with a small amount of Staphylococcus aureus. Milk quality recovery: after 14 days of treatment, the average number of somatic cells was 165,000 cells / mL, the recovery rate was 93.3%, the milk protein was 3.2%, the recovery rate was 96.7%, and the milk fat was 3.7%, the recovery rate was 100%.

[0054] Production performance: after 14 days of treatment, the daily milk yield of the affected side was 10.8 kg, and the recovery rate to the healthy milk area was 86.4%; after 30 days of treatment, the daily milk yield of the affected side was 12.2 kg, and the recovery rate to the healthy milk area was 98.4%.

[0055] Safety: no adverse reactions, safety compliance rate 100%. Example Three

[0056] The herbal flavonoid extract is 12 parts, and the mass ratio of honeysuckle extract to dandelion extract is 1:2. The chlorogenic acid content in the honeysuckle extract is 9.2%, and the dandelion flavone content in the dandelion extract is 11.5%, which meets the quality requirements.

[0057] Lactic acid bacteria fermentation metabolites: 12 parts, the lactic acid content is 18 g / L, and the bacteriocin content is 1.0 g / L, which meets the technical index.

[0058] Suspending agent: 3.5 parts, hydroxypropyl methyl cellulose K4M is selected.

[0059] Stabilizer: 2 parts, vitamin E acetate is used.

[0060] Osmotic pressure regulator: 5.5 parts, glycerol is used.

[0061] Purified water: make up to 100 parts, and ensure that the water quality meets the standard of sterile water for injection.

[0062] Raw material pretreatment: mix honeysuckle and dandelion according to the mass ratio of 1:2, put them into the pulverizer, control the particle size through the 40-60 mesh standard sieve, and get the mixed powder raw material.

[0063] Ultrasonic extraction: take the above mixed powder raw material, add 12 times of 50% ethanol aqueous solution to the total mass, put it into the ultrasonic extraction tank, set the extraction temperature to 38℃, and the ultrasonic power to 350W, ultrasonic extraction for 2 times, each time for 40 minutes, collect the two times of extraction liquid and combine them. Membrane separation and purification: the combined extraction liquid is microfiltered through a 0.22μm organic filter membrane to remove impurities, remove raw material residues and large molecular impurities; then the microfiltered filtrate is purified through an ultrafiltration membrane with a molecular weight cut-off of 3000Da, and the invalid macromolecular substances are cut off, and the ultrafiltration permeate is collected. Reduced pressure concentration: transfer the ultrafiltration permeate into a reduced pressure concentration tank, set the concentration temperature to 52℃, and the vacuum degree to-0.085MPa, and carry out reduced pressure concentration until the solid content in the concentrated liquid reaches 23%, stop the concentration, and get the herbal flavonoid extract, which is sealed and refrigerated for standby.

[0064] Single bacteria culture: take the lactobacillus plantarum strain and the lactobacillus acidophilus strain respectively, inoculate them into sterilized MRS medium, and cultivate them in a 37℃ anaerobic incubator for 22 hours to get the lactobacillus plantarum single bacteria fermentation liquid and the lactobacillus acidophilus single bacteria fermentation liquid.

[0065] Mixed fermentation: two single-bacterial fermentation broths were mixed uniformly at a volume ratio of 1:1, and the mixed fermentation broth was transferred to a sterilized special culture medium. The culture medium consisted of 10% sucrose, 5% soybean peptone, and 2% yeast extract. The pH value was adjusted to 6.5 in advance, and the mixed fermentation broth was fermented at 37°C in an anaerobic environment for 42 hours to obtain the mixed fermentation broth.

[0066] Centrifugal separation: the mixed fermentation broth was transferred to a high-speed centrifuge, the temperature was set to 4°C, the speed was set to 8000 r / min, and centrifugal separation was performed for 15 minutes to remove the bacterial bodies and precipitate impurities, and the supernatant was collected. Ultrafiltration purification: the supernatant after centrifugation was filtered through an ultrafiltration membrane with a molecular weight cutoff of 5000 Da to further remove macromolecular impurities, and the lactic acid bacteria fermentation metabolite was obtained and stored in a sealed refrigerator.

[0067] Preparation of base solution: an appropriate amount of purified water was heated to 42°C, 5.5 parts of glycerol was added, and stirred until completely dissolved; then 3.5 parts of hydroxypropyl methylcellulose K4M and 2 parts of vitamin E acetate were added in sequence, the stirring equipment was adjusted to 400 r / min, and continuous stirring was performed for 35 minutes to form a uniform and transparent base solution.

[0068] Raw material mixing: under stirring, 8 parts of herbal flavonoid extract and 12 parts of lactic acid bacteria fermentation metabolite were slowly added to the base solution, and the stirring was continued at a speed of 400 r / min for 75 minutes until the mixed solution was uniform and particle-free to form a preliminary perfusion solution.

[0069] Parameter adjustment and feeding: the pH value of the preliminary perfusion solution was adjusted to 6.8 using a pH adjuster; then purified water was added to a total mass of 100 parts, and stirring was performed for 5 minutes to ensure uniform mixing.

[0070] Sterilization and dispensing: the adjusted perfusion solution was filtered and sterilized through a 0.22 μm sterile filter membrane, and the whole process was operated in a hundred-level sterile environment; after sterilization, the perfusion solution was dispensed into sterile perfusion devices, the perfusion devices used soft silicone nozzles, the capacity was selected as 30 mL, and after dispensing, the devices were immediately sealed, labeled, and stored in a cold storage at 2-8°C. The final product had an osmotic pressure of 308 mOsm / kg and a pH of 6.9.

[0071] Effect: test subjects: 30 clinically diagnosed Holstein cows with mastitis, treatment regimen: 30 mL of perfusion per day for 5 consecutive days. Improvement of clinical symptoms: after 7 days of treatment, the clinical cure rate was 83.3%, and the symptom relief rate was 100%; after 14 days of treatment, the final clinical cure rate was 100%, and there was no need to extend the treatment cases. Pathogenic bacteria clearance: total clearance rate was 90% after 7 days of treatment, 95.8% for streptococcus, 90% for escherichia coli, and 87.5% for staphylococcus aureus; total clearance rate was 100% after 14 days of treatment, and all pathogenic bacteria were completely cleared without new infections. Milk quality recovery: after 14 days of treatment, the average number of somatic cells was 142,000 cells / mL, the recovery rate was 100%, the milk protein was 3.3%, the recovery rate was 100%, and the milk fat was 3.8%, the recovery rate was 100%.

[0072] Production performance: after 14 days of treatment, the daily milk yield of the affected side was 11.3 kg, and the recovery rate to the healthy milk area was 90.4%; after 30 days of treatment, the daily milk yield of the affected side was 12.4 kg, and the recovery rate to the healthy milk area was 100.8%, and the milk yield was slightly higher than the average of the healthy milk area due to the complete clearance of inflammation. Safety: no adverse reactions, and the safety compliance rate was 100%.

[0073] Examples four to six only change the content of lactic acid bacteria fermentation metabolites Example four

[0074] Herbal flavonoid extract: 8 parts, in which the mass ratio of honeysuckle extract to dandelion extract is 1:2, and the content of chlorogenic acid in honeysuckle extract is 9.2% and the content of dandelion flavonoids in dandelion extract is 11.5% after detection, both of which meet the quality requirements. Lactic acid bacteria fermentation metabolites: 8 parts, the lactic acid content is 18 g / L and the bacteriocin content is 1.0 g / L confirmed by early detection, which meets the technical index. Suspending agent: 3.5 parts, hydroxypropyl methylcellulose K4M is selected. Stabilizer: 2 parts, vitamin E acetate is used. Osmotic pressure regulator: 5.5 parts, glycerol is used. Purified water: make up to 100 parts to ensure that the water quality meets the standard of sterile water for injection. Raw material pretreatment: mix honeysuckle and dandelion according to the mass ratio of 1:2, put them into the crusher, control the particle size through a 40-60 mesh standard sieve, and get the mixed powder raw material. Ultrasonic extraction: take the above mixed powder raw material, add 12 times of 50% ethanol aqueous solution based on the total mass of the raw material, place it in the ultrasonic extraction tank, set the extraction temperature to 38℃, and the ultrasonic power to 350W, ultrasonic extraction for 2 times, each time for 40 minutes, collect the two extraction liquids and combine them.

[0075] Membrane separation and purification: the combined extraction liquid is subjected to microfiltration through a 0.22μm organic filter membrane to remove impurities, raw material residues and large molecular impurities; then the microfiltration filtrate is purified through an ultrafiltration membrane with a molecular weight cut-off of 3000Da to cut off ineffective large molecular substances, and the ultrafiltration permeate is collected. Vacuum concentration: The ultrafiltration permeate was transferred to a vacuum concentration tank, the concentration temperature was set to 52℃, the vacuum degree was set to -0.085 MPa, and vacuum concentration was carried out until the solid content in the concentrated solution reached 23%, and the concentration was stopped, obtaining the herbal flavonoid extract, which was sealed and refrigerated for standby. Single bacteria culture: Lactobacillus plantarum and Lactobacillus acidophilus were inoculated into sterilized MRS medium and cultured in an anaerobic incubator at 37℃ for 22 hours to obtain Lactobacillus plantarum single bacteria fermentation broth and Lactobacillus acidophilus single bacteria fermentation broth. Mixed fermentation: The two single bacteria fermentation broths were mixed in a volume ratio of 1:1, and the mixed fermentation broth was transferred to a sterilized special medium. The medium composition was 10% sucrose, 5% soybean peptone, and 2% yeast extract. The pH value was adjusted to 6.5 in advance, and the mixed fermentation broth was fermented in an anaerobic environment at 37℃ for 42 hours to obtain the mixed fermentation broth. Centrifugal separation: The mixed fermentation broth was transferred to a high-speed centrifuge, the temperature was set to 4℃, the speed was set to 8000r / min, and centrifugal separation was carried out for 15 minutes to remove the bacterial bodies and precipitate impurities, and the supernatant was collected. Ultrafiltration purification: The supernatant after centrifugation was filtered through an ultrafiltration membrane with a molecular weight cut-off of 5000 Da to further remove macromolecular impurities, and the lactic acid bacteria fermentation metabolite was obtained, which was sealed and refrigerated for standby. Preparation of base solution: A certain amount of purified water was heated to 42℃, 5.5 parts of glycerol was added, and stirred until completely dissolved; then 3.5 parts of hydroxypropyl methyl cellulose K4M and 2 parts of vitamin E acetate were added in turn, and the stirring equipment was adjusted to 400r / min, and stirred for 35 minutes to form a uniform and transparent base solution. Raw material mixing: Under stirring, 8 parts of herbal flavonoid extract and 12 parts of lactic acid bacteria fermentation metabolite were slowly added to the base solution, and the stirring speed was maintained at 400r / min for 75 minutes until the mixed solution was uniform and particle-free, forming the preliminary perfusion solution. Parameter adjustment and feeding: The pH value of the preliminary perfusion solution was adjusted to 6.8 using a pH adjuster; then purified water was added to a total mass of 100 parts, and stirred for 5 minutes to ensure uniform mixing.

[0076] Sterilization and dispensing: The adjusted perfusion solution was filtered and sterilized through a 0.22μm sterile filter membrane, and the whole process was operated in a hundred-level sterile environment; after sterilization, the perfusion solution was dispensed into sterile perfusion devices, the perfusion devices used soft silicone nozzles, the capacity was 30mL, and after dispensing, the devices were immediately sealed, labeled, and stored in a 2-8℃ refrigerator. The final product has an osmotic pressure of 301mOsm / kg and a pH of 6.7.

[0077] Effects: Test subjects: 30 Holstein cows with clinical mastitis, treatment regimen: 30 mL daily for 5 consecutive days. Improvement of clinical symptoms: After 7 days of treatment, the clinical cure rate was 53.3%, and the symptom relief rate was 100%; after 14 days of treatment, the final clinical cure rate was 76.7%, 5 cows needed to be treated for another 3 days, and 2 cows still had cloudy milk. Pathogen clearance: After 7 days of treatment, the total clearance rate was 63.3%, with 70.8% for streptococcus, 55% for E. coli, and 60% for S. aureus; after 14 days of treatment, the total clearance rate was 80%, with 87.5% for streptococcus, 75% for E. coli, and 72.7% for S. aureus, and 4 cows were detected with a small amount of pathogenic bacteria.

[0078] Milk quality recovery: After 14 days of treatment, the average number of somatic cells was 275 million / mL, with a recovery rate of 66.7%, milk protein was 2.8%, with a recovery rate of 76.7%, and milk fat was 3.3%, with a recovery rate of 86.7%.

[0079] Production performance: After 14 days of treatment, the daily milk production of the affected side was 8.5 kg, and the recovery to the healthy milk area was 68%; after 30 days of treatment, the daily milk production of the affected side was 10.8 kg, and the recovery to the healthy milk area was 86.4%. Safety: No adverse reactions, safety compliance rate 100%. Example Five

[0080] Herbal flavonoid extract: 8 parts, with the mass ratio of honeysuckle extract to dandelion extract being 1:2, the content of chlorogenic acid in honeysuckle extract was 9.2%, and the content of dandelion flavonoids in dandelion extract was 11.5%, both meeting the quality requirements. Lactic acid bacteria fermentation metabolites: 10 parts, the lactic acid content was confirmed to be 18 g / L and the bacteriocin content was 1.0 g / L, meeting the technical indicators. Suspending agent: 3.5 parts, hydroxypropyl methylcellulose K4M was selected. Stabilizer: 2 parts, vitamin E acetate was used. Osmotic pressure regulator: 5.5 parts, glycerol was used. Purified water: supplemented to 100 parts to ensure that the water quality meets the standard of sterile water for injection. Raw material pretreatment: mix honeysuckle and dandelion according to a mass ratio of 1:2, put them into a pulverizer for crushing, control the particle size through a 40-60 mesh standard sieve to obtain mixed powder raw materials. Ultrasonic extraction: take the above-mentioned mixed powder raw materials, add 50% ethanol aqueous solution with 12 times the total mass of the raw materials, place them in an ultrasonic extraction tank, set the extraction temperature to 38℃ and the ultrasonic power to 350W, perform ultrasonic extraction for 2 times, each time for 40 minutes, collect and combine the two extraction liquids.

[0081] Membrane separation and purification: the combined extract was micro-filtered through a 0.22 μm organic filter membrane to remove impurities, raw material residues and macromolecular impurities; then the micro-filtered filtrate was purified through an ultrafiltration membrane with a molecular weight cut-off of 3000 Da to remove ineffective macromolecular substances, and the ultrafiltration permeate was collected. Reduced pressure concentration: the ultrafiltration permeate was transferred to a reduced pressure concentration tank, the concentration temperature was set to 52°C, the vacuum degree was -0.085 MPa, and reduced pressure concentration was carried out until the solid content in the concentrated solution reached 23%, and the concentration was stopped, obtaining the herbal plant flavone extract, which was sealed and refrigerated for standby. Single bacteria culture: plant lactobacillus and acidophilus lactobacillus were inoculated into sterilized MRS medium and cultured in a 37°C anaerobic incubator for 22 hours to obtain plant lactobacillus single bacteria fermentation broth and acidophilus lactobacillus single bacteria fermentation broth. Mixed fermentation: the two single bacteria fermentation broths were mixed in a volume ratio of 1:1, and the mixed fermentation broth was transferred to a sterilized special medium, the medium composition was 10% sucrose, 5% soybean peptone, 2% yeast extract, the pH value was adjusted to 6.5 in advance, and the mixed fermentation broth was obtained by fermenting in an anaerobic environment at 37°C for 42 hours. Centrifugal separation: the mixed fermentation broth was transferred to a high-speed centrifuge, the temperature was set to 4°C, the speed was set to 8000 r / min, and centrifugal separation was carried out for 15 minutes to remove bacterial bodies and precipitate impurities, and the supernatant was collected. Ultrafiltration purification: the supernatant after centrifugation was filtered through an ultrafiltration membrane with a molecular weight cut-off of 5000 Da to further remove macromolecular impurities, and the lactic acid bacteria fermentation metabolite was obtained and sealed and refrigerated for standby. Preparation of base liquid: an appropriate amount of purified water was heated to 42°C, 5.5 parts of glycerol was added, and stirred until completely dissolved; then 3.5 parts of hydroxypropyl methylcellulose K4M and 2 parts of vitamin E acetate were added in turn, the stirring speed of the stirring equipment was adjusted to 400 r / min, and continuous stirring was carried out for 35 minutes to form a uniform and transparent base liquid. Raw material mixing: under stirring, 8 parts of herbal plant flavone extract and 12 parts of lactic acid bacteria fermentation metabolite were slowly added to the base liquid, the stirring speed was maintained at 400 r / min, and stirring was continued for 75 minutes until the mixed liquid was uniform and particle-free to form a preliminary perfusion solution. Parameter adjustment and feeding: the pH value of the preliminary perfusion solution was adjusted to 6.8 using a pH adjuster; then purified water was added to a total mass of 100 parts, and stirring was carried out for 5 minutes to ensure uniform mixing.

[0082] Sterilization and dispensing: the adjusted perfusion solution was filtered through a 0.22 μm sterile filter, and the whole process was operated in a hundred-level sterile environment; after sterilization, the perfusion solution was dispensed into sterile perfusion devices, the perfusion devices used soft silica gel nozzle, the capacity was selected as 30 mL, and after dispensing, it was immediately sealed, labeled, and stored in 2-8℃ refrigerator, the final product had an osmotic pressure of 304 mOsm / kg and a pH of 6.8.

[0083] Effects: Test subjects: 30 clinically diagnosed mastitis Holstein cows, treatment regimen: 30 mL of perfusion per day for 5 consecutive days.

[0084] Improvement of clinical symptoms: after 7 days of treatment, the clinical cure rate was 76.7%, and the symptom relief rate was 100%; after 14 days of treatment, the final clinical cure rate was 93.3%, and 2 cows needed to be treated for 2 more days. Pathogen clearance: after 7 days of treatment, the total clearance rate was 83.3%, and the clearance rates of Streptococcus, Escherichia coli, and Staphylococcus aureus were 89.6%, 80%, and 81.8%, respectively; after 14 days of treatment, the total clearance rate was 95%, and the clearance rates of Streptococcus, Escherichia coli, and Staphylococcus aureus were 95.8%, 95%, and 90.9%, respectively, and 1 cow was detected with a small amount of Staphylococcus aureus, with a bacterial count of <6 CFU / mL. Milk quality recovery: after 14 days of treatment, the average somatic cell count was 183 million cells / mL, the recovery rate was 90%, the milk protein was 3.1%, the recovery rate was 93.3%, and the milk fat was 3.6%, the recovery rate was 96.7%.

[0085] Production performance: after 14 days of treatment, the daily milk yield of the affected side was 10.3 kg, and the recovery rate to the healthy milk area was 82.4%; after 30 days of treatment, the daily milk yield of the affected side was 12.0 kg, and the recovery rate to the healthy milk area was 96%.

[0086] Safety: no adverse reactions, and the safety compliance rate was 100%. Example Six

[0087] Herbal flavonoid extract: 8 parts, the mass ratio of honeysuckle extract to dandelion extract is 1:2, the content of chlorogenic acid in honeysuckle extract is 9.2%, and the content of dandelion flavonoids in dandelion extract is 11.5%, which meet the quality requirements. Lactic acid bacteria fermentation metabolites: 15 parts, the lactic acid content is 18 g / L, and the bacteriocin content is 1.0 g / L, which meet the technical indicators. Suspending agent: 3.5 parts, hydroxypropyl methylcellulose K4M is selected. Stabilizer: 2 parts, vitamin E acetate is used. Osmotic pressure regulator: 5.5 parts, glycerol is used. Purified water: make up to 100 parts, and ensure that the water quality meets the standard of sterile water for injection. Raw material pretreatment: mix honeysuckle and dandelion at a mass ratio of 1:2, put them into a pulverizer to crush, control the particle size through a 40-60 mesh standard sieve to obtain a mixed powder raw material. Ultrasonic extraction: take the mixed powder raw material, add 12 times the total mass of 50% ethanol aqueous solution, place it in an ultrasonic extraction tank, set the extraction temperature to 38°C, the ultrasonic power to 350W, perform ultrasonic extraction twice, each time for 40 minutes, collect the two extraction liquids and combine them.

[0088] Membrane separation and purification: the combined extraction liquid is subjected to microfiltration through a 0.22μm organic filter membrane to remove impurities, remove raw material residues and macromolecular impurities; then the microfiltration filtrate is subjected to purification through an ultrafiltration membrane with a molecular weight cut-off of 3000Da to cut off ineffective macromolecular substances, collect the ultrafiltration permeate. Reduced pressure concentration: transfer the ultrafiltration permeate into a reduced pressure concentration tank, set the concentration temperature to 52°C, the vacuum degree to -0.085MPa, perform reduced pressure concentration until the solid content in the concentrated liquid reaches 23%, stop the concentration to obtain the herbal plant flavonoid extract, seal and refrigerate for standby. Single strain culture: take the lactobacillus plantarum strain and the lactobacillus acidophilus strain respectively, inoculate them into sterilized MRS medium, place them in a 37°C anaerobic incubator for 22 hours to obtain the lactobacillus plantarum single strain fermentation liquid and the lactobacillus acidophilus single strain fermentation liquid. Mixed fermentation: mix the two single strain fermentation liquids uniformly at a volume ratio of 1:1, take the mixed fermentation liquid and transfer it into sterilized special medium, the medium composition is 10% sucrose, 5% soybean peptone, 2% yeast extract, adjust the pH value to 6.5 in advance, ferment in a 37°C anaerobic environment for 42 hours to obtain the mixed fermentation liquid. Centrifugal separation: transfer the mixed fermentation liquid into a high-speed centrifuge, set the temperature to 4°C, the rotation speed to 8000r / min, centrifuge for 15 minutes to remove the bacterial bodies and precipitate impurities, collect the supernatant. Ultrafiltration purification: filter the centrifuged supernatant through an ultrafiltration membrane with a molecular weight cut-off of 5000Da to further remove macromolecular impurities, obtain the lactic acid bacteria fermentation metabolite, seal and refrigerate for standby. Preparation of base liquid: take an appropriate amount of purified water, heat it to 42°C, add 5.5 parts of glycerol weighed in advance, stir until completely dissolved; then add 3.5 parts of hydroxypropyl methylcellulose K4M and 2 parts of vitamin E acetate in sequence, adjust the stirring equipment speed to 400r / min, continue stirring for 35 minutes to form a uniform and transparent base liquid. Raw material mixing: slowly add 8 parts of herbal plant flavonoid extract and 12 parts of lactic acid bacteria fermentation metabolite to the base liquid under stirring, continue stirring at a speed of 400r / min for 75 minutes until the mixed liquid is uniform and particle-free to form a preliminary perfusion liquid. Parameter adjustment and feeding: the pH value of the initial perfusion solution is adjusted to 6.8 using a pH adjuster; then purified water is added to a total mass of 100 parts, and stirred for 5 minutes to ensure uniform mixing.

[0089] Sterilization and sub-packaging: the adjusted perfusion solution is filtered and sterilized through a 0.22 μm sterile filter, and the entire process is operated in a Class 100 sterile environment; after sterilization, the perfusion solution is sub-packaged into sterile perfusion containers, the perfusion containers use soft silicone nozzles, the capacity is selected to be 30 mL, and after sub-packaging is completed, it is immediately sealed, labeled, and stored in a 2-8°C refrigerator. The final product has an osmotic pressure of 310 mOsm / kg and a pH of 6.9.

[0090] Effect: Test subjects: 30 clinically diagnosed mastitis Holstein cows, treatment regimen: 30 mL of perfusion per day for 5 consecutive days.

[0091] Improvement of clinical symptoms: after 7 days of treatment, the clinical cure rate was 86.7% and the symptom relief rate was 100%; after 14 days of treatment, the final clinical cure rate was 100% and there was no need to extend the treatment cases. Pathogen clearance: after 7 days of treatment, the total clearance rate was 93.3%, 97.9% for streptococcus, 95% for E. coli, and 90.9% for Staphylococcus aureus; after 14 days of treatment, the total clearance rate was 100%, and all pathogenic bacteria were completely cleared without new infections. Milk quality recovery: after 14 days of treatment, the average somatic cell count was 138,000 cells / mL, the recovery rate was 100%, the milk protein was 3.4%, the recovery rate was 100%, and the milk fat was 3.9%, the recovery rate was 100%.

[0092] Production performance: after 14 days of treatment, the daily milk yield of the affected side was 11.8 kg, and the recovery to the healthy milk area was 94.4%; after 30 days of treatment, the daily milk yield of the affected side was 12.6 kg, and the recovery to the healthy milk area was 100.8%, and due to the rapid recovery of the flora balance, the milk yield was slightly higher than the average of the healthy milk area. Safety: no adverse reactions, safety compliance rate 100%.

[0093] The specific contents are shown in the following table: Herbal flavonoid extract (parts) Lactic acid bacteria fermentation metabolite (parts) Product key parameters (osmotic pressure / mOsm / kg, pH) Example one 5 12 Example two 10 12 Example three 12 12 Example four 8 8 Example five 8 10 Example six 8 15 As described in the above table, only the content of the herbal flavonoids extract is changed in Examples 1-3, and the other parameters remain unchanged. The content of the herbal flavonoids extract has a significant positive correlation with the treatment effect of the udder infusion for treating cow mastitis. When the content increases from 5 parts to 12 parts, the treatment efficiency and effect gradually increase. The clinical cure rate after 7 days of treatment increases from 56.7% to 83.3%, and the total pathogen clearance rate increases from 66.7% to 90%. The final clinical cure rate after 14 days of treatment increases from 80% to 100%, and the total pathogen clearance rate increases from 83.3% to 100%. The recovery rate of milk quality and the recovery speed of production performance also increase. The effect of the 5-part flavonoids extract group is weaker, which is suitable for mild mastitis or as an auxiliary for prevention. The 10-part group has balanced and excellent effects, which takes into account the cost and treatment efficiency, and is suitable for moderate to severe mastitis routine treatment. The 12-part group has the best effect, with the fastest inflammation clearance and milk recovery, which is suitable for severe mastitis or scenarios with high treatment efficiency requirements. All groups maintain 100% safety and have no adverse reactions.

[0094] Examples 4-6 only change the content of the lactic acid bacteria fermentation metabolites, and the other parameters remain unchanged. The content of the lactic acid bacteria fermentation metabolites has a significant positive correlation with the treatment effect of the udder infusion. When the content increases from 8 parts to 15 parts, the treatment efficiency and effect continue to increase. The clinical cure rate after 7 days of treatment increases from 53.3% to 86.7%, and the total pathogen clearance rate increases from 63.3% to 93.3%. The final clinical cure rate after 14 days of treatment increases from 76.7% to 100%, and the total pathogen clearance rate increases from 80% to 100%. The recovery speed of milk quality and the recovery efficiency of production performance also increase synchronously. The 8-part lactic acid bacteria group has a weaker effect, which is suitable for mild mastitis or low-cost prevention scenarios. The 10-part group has balanced effects, which takes into account the treatment effect and cost, and is suitable for moderate mastitis routine treatment. The 15-part group has the best effect, with the strongest antibacterial and flora regulation effect, which is suitable for severe mastitis or scenarios that pursue rapid recovery. All groups maintain 100% safety and have no adverse reactions. The above is only a preferred embodiment of the present application, and does not limit the present application in any form. Although the present application has been disclosed as a preferred embodiment, it is not intended to limit the present application. Any person skilled in the art can make minor changes or modifications to the disclosed technical content without departing from the scope of the technical solution of the present application, and any equivalent embodiments with equivalent changes are equivalent. Any modification, change, and modification of the above examples based on the technical essence of the present application are still within the scope of the technical solution of the present application.

Claims

1. A mammary infusion solution for treating mastitis in dairy cows, characterized in that, Made from the following raw materials in parts by weight: Herbal plant flavonoid extracts, lactic acid bacteria fermentation metabolites, suspending agents, stabilizers, osmotic pressure regulators, and purified water; The herbal flavonoid extract is a mixture of honeysuckle extract and dandelion extract; The lactic acid bacteria fermentation metabolite is the supernatant obtained after centrifugation and ultrafiltration of a mixed fermentation broth of Lactobacillus plantarum and Lactobacillus acidophilus.

2. The mammary gland infusion solution for treating bovine mastitis according to claim 1, characterized in that, The herbal plant flavonoid extract consists of 5-12 parts, lactic acid bacteria fermentation metabolites of 8-15 parts, suspending agent of 2-5 parts, stabilizer of 1-3 parts, osmotic pressure regulator of 3-8 parts, and purified water is added to make up to 100 parts.

3. The mammary gland infusion solution for treating bovine mastitis according to claim 1, characterized in that, In the herbal plant flavonoid extract, the mass ratio of honeysuckle extract to dandelion extract is 1:1.5-2.5; and the chlorogenic acid content in the honeysuckle extract is ≥8%, and the dandelion flavonoid content in the dandelion extract is ≥10%.

4. The mammary gland infusion solution for treating bovine mastitis according to claim 1, characterized in that, The lactic acid fermentation metabolites contain 15-20 g / L of lactic acid and 0.8-1.2 g / L of bacteriocins. The ultrafiltration process uses an ultrafiltration membrane with a molecular weight cutoff of 5000 Da.

5. The method for preparing a mammary infusion solution for treating bovine mastitis according to claim 1, characterized in that, The suspending agent is hydroxypropyl methylcellulose K4M, the stabilizer is vitamin E acetate, and the osmotic pressure regulator is glycerol; and the osmotic pressure of the infusion solution is controlled at 280-320 mOsm / kg, and the pH value is adjusted to 6.5-7.

0.

6. A method for preparing a mammary infusion solution for treating bovine mastitis, used to prepare the mammary infusion solution for treating bovine mastitis as described in any one of claims 1-5, characterized in that, Includes the following steps: Preparation of herbal plant flavonoid extract: The herbal plant flavonoid extract was prepared by a low-temperature ultrasonic extraction-membrane separation purification process, specifically including: mixing and pulverizing honeysuckle and dandelion, then ultrasonically extracting twice with 50% ethanol aqueous solution at 35-40℃ and 300-400W for 30-45 minutes each time. After removing impurities by microfiltration through a 0.22μm organic filter membrane, the extract was purified by ultrafiltration membrane with a molecular weight cutoff of 3000Da. Finally, the extract was concentrated under reduced pressure at 50-55℃ and a vacuum degree of -0.08 to -0.09MPa to a solid content of 20-25% to obtain a honeysuckle-dandelion mixed flavonoid extract for later use. Preparation of lactic acid bacteria fermentation metabolites: Lactobacillus plantarum and Lactobacillus acidophilus were inoculated into MRS medium and anaerobic cultured at 37℃ for 18-24 hours to obtain single-cell fermentation broth. The broth was mixed at a volume ratio of 1:1 and transferred to medium containing 10% sucrose, 5% soybean peptone, and 2% yeast extract at pH 6.

5. Anaerobic fermentation was carried out at 37℃ for 36-48 hours. The fermentation broth was centrifuged at 4℃ and 8000 r / min for 15 minutes. The supernatant was filtered through an ultrafiltration membrane with a molecular weight cutoff of 5000 Da to obtain the metabolites for later use. Mixing and preparation: Heat purified water to 40-45℃, add osmotic pressure regulator and stir to dissolve, then add suspending agent and stabilizer in sequence, and stir for 30-40 minutes at 300-500 r / min to form a base solution; add flavonoid extract and metabolites to the base solution, and continue stirring for 60-90 minutes until uniform and free of particles, adjust the pH to 6.5-7.0, add purified water to 100 parts, filter through a 0.22μm sterile filter membrane for sterilization, and dispense into sterile filling containers and seal.

7. The method for preparing a mammary infusion solution for treating bovine mastitis according to claim 1, characterized in that, In the preparation of the herbal flavonoid extract, the particle size of honeysuckle and dandelion after mixing and pulverizing is controlled at 40-60 mesh, and the amount of 50% ethanol aqueous solution added during ultrasonic extraction is 10-15 times the total mass of the raw materials.

8. The method for preparing a mammary infusion solution for treating bovine mastitis according to claim 1, characterized in that, In the mixing and preparation process, the entire mixing and preparation process is carried out in a sterile environment. The sterile infusion device used for dispensing adopts a soft silicone nozzle, and the capacity of the infusion device can be adjusted to 20-50mL according to the size of the cow's teat.

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