Traditional Chinese medicine composition for preventing and treating exogenous wind-heat syndrome of livestock and poultry and application of traditional Chinese medicine composition

Abstract

The invention discloses a traditional Chinese medicine composition for preventing and treating exogenous wind-heat syndrome of livestock and poultry and application thereof, and belongs to the technical field of traditional Chinese veterinary medicines. The traditional Chinese medicine composition is prepared from the following components in parts by weight: 60 to 100 parts of radix isatidis, 20 to 60 parts of gypsum, 20 to 60 parts of rhizoma phragmitis, 10 to 40 parts of radix rehmanniae, 10 to 30 parts of radix curcumae, 10 to 30 parts of rhizoma anemarrhenae, 10 to 30 parts of rhizoma acori graminei, 10 to 30 parts of herba pogostemonis and 15 to 50 parts of fructus forsythiae. The composition disclosed by the invention can be used for effectively treating exogenous wind-heat syndrome of livestock or poultry caused by infection of viruses such as chicken infectious bronchitis virus, avian influenza virus and swine influenza virus and respiratory tract symptoms caused by the exogenous wind-heat syndrome, and the resistance reducing and replacing effects are obvious.

Description

Technical Field

[0001] This invention belongs to the field of traditional Chinese veterinary medicine technology, specifically relating to a traditional Chinese medicine composition for preventing and treating exogenous wind-heat syndrome in livestock and poultry and its application. Background Technology

[0002] Chickens suffering from exogenous wind-heat syndrome are a common and frequently occurring disease throughout the year. They are particularly prevalent due to poor husbandry practices, hot seasons, or sudden climate changes in late spring and early autumn with drastic temperature fluctuations. Wind-heat is a yang evil, easily transforming body fluids into phlegm. Therefore, affected chickens often exhibit difficulty breathing due to increased phlegm in the lungs, obstructing the flow of qi. They frequently shake their heads, easily expelling thick phlegm, and exhibit difficulty breathing, often stretching their necks upwards and breathing with their mouths open. If left untreated or mistreated, this syndrome can easily develop into lung-heat cough and wheezing syndrome. During seasonal transitions and periods of drastic climate change, such as when chicken coops are damaged, chickens are highly susceptible to respiratory infectious diseases such as colds, bronchitis (pneumonia), and pneumonia, including infectious bronchitis, infectious laryngotracheitis, infectious coryza, Newcastle disease, and avian influenza (H9N2). These diseases often present with symptoms of exogenous wind-heat syndrome in their early stages.

[0003] Swine exogenous wind-heat syndrome is a common disease in livestock farming. It is often caused by sudden changes in climate and large temperature fluctuations during hot seasons, resulting in wind-heat. Affected pigs often exhibit high fever, slight chills with mild shivering, or even heat intolerance instead of chills. They prefer to lie in cool or damp places, are lethargic, with drooping heads and ears, and have poor appetite. Their ears and nose are warm, their body temperature is elevated, and they may sweat slightly. Initially, they have a clear nasal discharge, with intermittent nasal sweating. Later, their snout becomes dry, and they develop a thick, yellow or white, viscous purulent nasal discharge, cough, and a persistent cough. They also experience dry mouth, a slightly red tongue with a thin white or yellow coating, and a floating, rapid pulse. The wind-heat pathogen invades through the nose and throat, damaging these organs and causing nasal congestion and frequent sneezing. The throat is swollen, painful, and sensitive; affected pigs will flinch when their throat is touched, and this easily triggers coughing. The lungs are a delicate organ; when attacked by wind-heat pathogens, lung qi rises abnormally, easily causing coughing and rapid breathing. When lung heat descends, it easily leads to dryness and heat in the large intestine, resulting in constipation and dark yellow urine. Wind-heat, being a yang evil, easily transforms body fluids into phlegm, hence the common appearance of vomiting and neck stretching in sick pigs after coughing. If this condition is left untreated or mistreated, it can easily develop into lung-heat cough and wheezing syndrome. During seasonal changes and climate shifts, respiratory infections in pigs, such as colds, bronchitis (pneumonia), early-stage pneumonia, and those resulting from regrouping or long-distance transport, often exhibit wind-heat syndrome symptoms in their pathological process.

[0004] Currently, wind-heat colds are widespread in livestock and poultry, posing a significant challenge to the livestock industry. High stocking densities and improper feeding management lead to decreased immune levels in farmed animals, making them more susceptible to invasion by bacteria, viruses, and other microorganisms, resulting in a range of diseases. Clinically, respiratory diseases are often treated with antiviral drugs and antibiotics; however, the irrational use of antibiotics can easily lead to drug-resistant bacteria and other side effects. Commonly used antiviral and antibacterial drugs, such as ribavirin and amantadine, often have transient or even serious adverse reactions, such as allergic reactions, liver and kidney damage, aplastic anemia, and hemolytic anemia, as well as gastrointestinal adverse reactions like nausea, vomiting, and constipation. Antiviral and antibacterial drugs primarily target pathogenic microorganisms and have limited effectiveness in relieving symptoms such as fever, severe inflammation, and cough caused by these pathogens.

[0005] Therefore, clinically, there is a need for a class of drugs that can cover a variety of symptoms caused by wind-heat colds, and that are effective, fast-acting, and have few side effects. Traditional Chinese medicine (TCM) has significant advantages in this regard. Research in TCM shows that Chinese herbs are abundant, inexpensive, and have few side effects. Furthermore, TCM treatments involve multiple pathways, multiple stages, and multiple targets. TCM preparations with pungent and cooling properties that relieve exterior symptoms and clear heat and detoxify have good antipyretic, anti-inflammatory, antibacterial, and antimicrobial effects, and have few side effects, effectively relieving cold symptoms and quickly achieving therapeutic goals. Summary of the Invention

[0006] The present invention aims to solve the technical problems in the prior art and provide a traditional Chinese medicine composition for preventing and treating exogenous wind-heat syndrome in livestock and poultry and its application.

[0007] To achieve the above objectives, the specific technical solution of the present invention is as follows:

[0008] In a first aspect, the present invention provides a traditional Chinese medicine composition for preventing and treating exogenous wind-heat syndrome in livestock and poultry, wherein the raw material components of the traditional Chinese medicine composition include Isatis root, gypsum, reed rhizome, Rehmannia root, turmeric, anemarrhena rhizome, acorus tatarinowii rhizome, patchouli, and forsythia.

[0009] The raw material components in the traditional Chinese medicine composition are, by weight, 60-100 parts of Isatis root, 20-60 parts of gypsum, 20-60 parts of reed rhizome, 10-40 parts of Rehmannia root, 10-30 parts of turmeric, 10-30 parts of anemarrhena rhizome, 10-30 parts of acorus tatarinowii rhizome, 10-30 parts of patchouli rhizome, and 15-50 parts of forsythia fruit.

[0010] Preferably, the raw material components in the traditional Chinese medicine composition are, by weight, 80-85 parts of Isatis root, 35-40 parts of gypsum, 35-40 parts of reed rhizome, 20-25 parts of Rehmannia root, 15-20 parts of turmeric, 15-20 parts of anemarrhena rhizome, 15-20 parts of acorus tatarinowii rhizome, 15-20 parts of patchouli rhizome, and 30-35 parts of forsythia fruit.

[0011] In a second aspect, the present invention provides a method for preparing a traditional Chinese medicine composition for preventing and treating exogenous wind-heat syndrome in livestock and poultry, comprising the following steps:

[0012] (1) Weigh each of the above raw material components separately, add 10-14 times the amount of purified water to the raw material components and decoct 3 times, each time for 1-2 hours, while collecting the volatile oil and volatile oil emulsion; combine the decoctions, filter and set aside.

[0013] (2) Concentrate the decoction under reduced pressure to an appropriate volume, add the antibacterial agent and stir until dissolved, then add the volatile oil and volatile oil emulsion and mix well.

[0014] (3) Adjust the pH value with 10% sodium hydroxide solution, add purified water to make up the volume, mix well, filter, and fill into a container to obtain the product.

[0015] The vacuum concentration temperature in step (2) is 50–70°C.

[0016] The vacuum degree of the reduced pressure concentration in step (2) is 0.08 to 0.09 MPa.

[0017] The volume of vacuum concentration in step (2) is 60% to 70% of the total volume of the finished product.

[0018] The antibacterial agent mentioned in step (2) is potassium sorbate, and the addition amount is 0.2% to 0.3%.

[0019] The pH value in step (3) is adjusted to 4.0 to 6.0.

[0020] The pharmaceutical composition can be prepared into various formulations for use by livestock and poultry according to pharmaceutical methods, including but not limited to oral liquid formulations, powders, granules, and soluble powders.

[0021] In a third aspect, the present invention provides the application of a traditional Chinese medicine composition for preventing and treating exogenous wind-heat syndrome in livestock and poultry in the preparation of a medicine for relieving exogenous wind-heat syndrome caused by livestock and poultry viruses and the respiratory symptoms caused thereto.

[0022] The livestock and poultry viruses mentioned are chicken infectious bronchitis virus, avian influenza virus, and swine influenza virus.

[0023] In the above technical solution, the functions of each component are as follows:

[0024] Isatis root is the dried root of the cruciferous plant Isatis indigotica, containing polysaccharides, alkaloids, phenylpropanoids, organic acids, triterpenoids, nucleosides, and other components. Among them, the polysaccharides in Isatis root have pharmacological effects such as antiviral, antibacterial, free radical scavenging, and immunomodulatory effects.

[0025] Gypsum is a commonly used herb in Traditional Chinese Medicine (TCM) for clearing heat and purging fire, belonging to the category of pungent and cold herbs. It has the effects of clearing heat and purging fire, relieving irritability and quenching thirst, and is used for exogenous febrile diseases, high fever with thirst, and cough and wheezing due to lung heat. It is often paired with other Chinese herbs to exert synergistic effects or reduce toxicity. The combination of gypsum and anemarrhena asphodeloides (Zhimu) is a classic combination for treating febrile diseases of the Yangming channel in typhoid fever, as it has the effects of clearing heat, generating fluids, moistening dryness, and quenching thirst.

[0026] Reed rhizome is the fresh or dried rhizome of the reed plant (Phragmites communis), a member of the Poaceae family. It has the effects of clearing heat, relieving irritability, promoting body fluid production, stopping vomiting, and promoting diuresis. Reed rhizome also possesses antioxidant, anti-aging, liver-protective, antibacterial, litholytic, and blood sugar-lowering properties.

[0027] Rehmannia glutinosa is a perennial herb belonging to the genus Rehmannia in the family Scrophulariaceae. It contains various types of compounds, including iridoids and their glycosides, phenolic compounds, sugars, amino acids, and organic acids, and possesses a variety of pharmacological effects such as anti-aging, hypoglycemic, anti-inflammatory, and immune system enhancement.

[0028] Turmeric is the dried tuberous root of *Curcuma longa*, *Curcuma zedoaria*, *Curcuma fangchi*, or *Curcuma zedoaria*, all belonging to the ginger family. It has the effects of regulating qi and relieving depression, cooling the blood and breaking up blood stasis. When used with *Acorus tatarinowii*, *Forsythia suspensa*, *Gardenia jasminoides*, *Rheum palmatum*, alum, Baijin pill, Gleditsia sinensis, and centipede, it can clear the heart and open the orifices; when used with *Rehmannia glutinosa*, *Paeonia suffruticosa*, *Gardenia jasminoides*, talc, *Rehmannia glutinosa*, and *Plantago asiatica*, it can clear heat and cool the blood.

[0029] Anemarrhena asphodeloides is the rhizome of the plant Anemarrhena asphodeloides, which contains mangiferin, anemarrhena saponins and anemarrhena polysaccharides, among other components; it has antipyretic, anti-inflammatory, diuretic and analgesic effects.

[0030] As one of the few medicines that can open the mind and stimulate consciousness, Acorus tatarinowii has a long history of medicinal use and possesses a variety of medicinal effects, making it widely used in clinical practice. Its volatile oil, the effective component of Acorus tatarinowii, has pharmacological activities such as stimulating awakening, sedation, improving cognitive function, anticoagulation, and enhancing hypoxia tolerance.

[0031] Patchouli is a perennial herbaceous plant of the Lamiaceae family. It has pharmacological effects such as antibacterial, anti-spirochete, digestive promotion, relief of gastrointestinal damage, and inhibition of respiratory pathogens.

[0032] Forsythia is the dried fruit of the Forsythia suspensa plant, belonging to the Oleaceae family. Its chemical composition is complex, mainly containing glycosides, phenylethyl glycosides, lignans, flavonoids, natural alcohols, coumarins, and volatile components. It has effects such as inhibiting pathogenic microorganisms, antipyretic and anti-inflammatory properties, liver protection, diuresis, and antiemetic effects.

[0033] Beneficial effects:

[0034] (1) The traditional Chinese medicine composition of the present invention can be used to treat the exogenous wind-heat syndrome and respiratory symptoms caused by infection with chicken infectious bronchitis virus, avian influenza virus, swine influenza virus, etc. in livestock or poultry.

[0035] (2) The traditional Chinese medicine composition of the present invention can be administered by drinking water, which is convenient to use and has low cost. Detailed Implementation

[0036] Unless otherwise specified, the experimental methods used in the following examples are conventional methods. The raw materials used in the following examples are all commercially available products that can be purchased from the market, unless otherwise specified.

[0037] The above content will be further described in detail below through specific embodiments. However, this should not be construed as limiting the scope of the above subject matter to the following embodiments. All technologies implemented based on the content of this invention fall within the scope.

[0038] Example 1

[0039] A traditional Chinese medicine oral liquid preparation for preventing and treating exogenous wind-heat syndrome in livestock and poultry, wherein the main components and their weights in the oral liquid are as follows: Isatis root 100g, gypsum 60g, reed rhizome 60g, Rehmannia root 40g, turmeric 30g, anemarrhena rhizome 30g, acorus tatarinowii rhizome 30g, patchouli 30g, and forsythia fruit 50g. Excipients and their dosage: potassium sorbate 1g. pH value: adjusted to 4.0.

[0040] Its preparation process is as follows:

[0041] (1) Weigh all the above-mentioned Chinese herbal components separately. First, put the Chinese herbal components into a round-bottom flask. Add water 10 times the total amount of raw materials to the round-bottom flask and decoct 3 times, 2 hours each time (collect volatile oil and volatile oil emulsion at the same time). Combine the decoctions, filter, and set aside.

[0042] (2) Concentration: Concentrate the decoction at 50℃ and 0.09Mpa under reduced pressure to 60% of the total volume of the finished product. Add 1g of potassium sorbate and stir until dissolved. Then add volatile oil and volatile oil emulsion and mix well. Adjust the pH value to 4.0 with 10% sodium hydroxide solution, add water to make up to 500ml, mix well, filter, and fill into bottles to obtain the final product.

[0043] Example 2

[0044] A traditional Chinese medicine oral liquid preparation for preventing and treating exogenous wind-heat syndrome in livestock and poultry, wherein the main components and their weights in the oral liquid are as follows: Isatis root 85g, gypsum 40g, reed rhizome 40g, Rehmannia root 25g, turmeric 20g, anemarrhena rhizome 20g, acorus tatarinowii rhizome 20g, patchouli 20g, and forsythia fruit 35g. Excipients and their dosage: potassium sorbate 1.5g. pH value: adjusted to 5.0.

[0045] Its preparation process is as follows:

[0046] (1) Weigh all the above-mentioned Chinese herbal components separately. First, put the Chinese herbal components into a round-bottom flask. Add water 12 times the total amount of raw materials to the round-bottom flask and decoct 3 times, 1.5 hours each time (collect volatile oil and volatile oil emulsion at the same time). Combine the decoctions, filter and set aside.

[0047] (2) Concentration: Concentrate the decoction at 60℃ and 0.085Mpa under reduced pressure to 70% of the total volume of the finished product. Add 1.5g of potassium sorbate and stir until dissolved. Then add volatile oil and volatile oil emulsion and mix well. Adjust the pH value to 5.0 with 10% sodium hydroxide solution, add water to make up to 500ml, mix well, filter, and fill into bottles to obtain the final product.

[0048] Example 3

[0049] A traditional Chinese medicine oral liquid preparation for preventing and treating exogenous wind-heat syndrome in livestock and poultry, wherein the main components and their weights in the oral liquid are as follows: Isatis root 82g, gypsum 38g, reed rhizome 37g, rehmannia root 22g, turmeric 17g, anemarrhena rhizome 17g, acorus tatarinowii rhizome 17g, patchouli 18g, and forsythia fruit 32g. Excipients and their dosage: potassium sorbate 1g. pH value: adjusted to 6.0.

[0050] Its preparation process is as follows:

[0051] (1) Weigh all the above-mentioned Chinese herbal components separately. First, put the Chinese herbal components into a round-bottom flask. Add water 14 times the total amount of raw materials to the round-bottom flask and decoct 3 times, 1 hour each time (collect volatile oil and volatile oil emulsion at the same time). Combine the decoctions, filter, and set aside.

[0052] (2) Concentration: Concentrate the decoction at 70℃ and 0.08Mpa under reduced pressure to 60% of the total volume of the finished product. Add 1g of potassium sorbate and stir until dissolved. Then add volatile oil and volatile oil emulsion and mix well. Adjust the pH value to 6.0 with 10% sodium hydroxide solution, add water to make up to 500ml, mix well, filter, and fill into bottles to obtain the final product.

[0053] Example 4

[0054] A traditional Chinese medicine oral liquid preparation for preventing and treating exogenous wind-heat syndrome in livestock and poultry, wherein the main components and their weights in the oral liquid are as follows: Isatis root 80g, gypsum 35g, reed rhizome 35g, Rehmannia root 20g, turmeric 15g, anemarrhena rhizome 15g, acorus tatarinowii rhizome 15g, patchouli 15g, and forsythia fruit 30g. Excipients and their dosage: potassium sorbate 1.5g. pH value: adjusted to 4.0.

[0055] Its preparation process is as follows:

[0056] (1) Weigh all the above-mentioned Chinese herbal components separately. First, put the Chinese herbal components into a round-bottom flask. Add water 12 times the total amount of raw materials to the round-bottom flask and decoct 3 times, 1 hour each time (collect volatile oil and volatile oil emulsion at the same time). Combine the decoctions, filter and set aside.

[0057] (2) Concentration: Concentrate the decoction at 50℃ and 0.09Mpa under reduced pressure to 70% of the total volume of the finished product. Add 1.5g of potassium sorbate and stir until dissolved. Then add volatile oil and volatile oil emulsion and mix well. Adjust the pH value to 4.0 with 10% sodium hydroxide solution, add water to make up to 500ml, mix well, filter, and fill into bottles to obtain the final product.

[0058] Example 5

[0059] A traditional Chinese medicine oral liquid preparation for preventing and treating exogenous wind-heat syndrome in livestock and poultry, wherein the main components and their weights in the oral liquid are as follows: Isatis root 60g, gypsum 20g, reed rhizome 20g, Rehmannia root 10g, turmeric 10g, anemarrhena rhizome 10g, acorus tatarinowii rhizome 10g, patchouli 10g, and forsythia fruit 15g. Excipients and their dosage: potassium sorbate 1g. pH value: adjusted to 5.0.

[0060] Its preparation process is as follows:

[0061] (1) Weigh all the above-mentioned Chinese herbal components separately. First, put the Chinese herbal components into a round-bottom flask. Add water 10 times the total amount of raw materials to the round-bottom flask and decoct 3 times, 1.5 hours each time (collect volatile oil and volatile oil emulsion at the same time). Combine the decoctions, filter and set aside.

[0062] (2) Concentration: Concentrate the decoction at 60°C and 0.085 MPa under reduced pressure to 60% of the total volume of the finished product. Add 1g of potassium sorbate and stir until dissolved. Then add volatile oil and volatile oil emulsion and mix well. Adjust the pH value to 5.0 with 10% sodium hydroxide solution, add water to make up to 500ml, mix well, filter, and fill into bottles to obtain the final product.

[0063] Comparative Example 1

[0064] For example, in Example 12 of patent CN100431596C, the components and their weights in the drug composition for treating avian influenza are as follows: 600g of Isatis root, 400g of Forsythia, 300g of Gypsum, 600g of Phragmites rhizome, 100g of Rehmannia root, 300g of Patchouli, 200g of Acorus tatarinowii, 100g of Curcuma longa, and 100g of Anemarrhena asphodeloides.

[0065] The preparation process is as follows: as in Example 12 of patent CN100431596C.

[0066] Example 6: Clinical efficacy trial against chicken infectious bronchitis virus

[0067] 1. Purpose of the experiment: To conduct a comparative experiment on the efficacy of IBV in 12-day-old broiler chickens artificially infected with IBV.

[0068] 2. Experimental design: 160 12-day-old white-feathered broiler chickens were randomly divided into groups of 20 each, namely blank control group, model control group, Example 1 group, Example 2 group, Example 3 group, Example 4 group, Example 5 group, and comparative example 1 group.

[0069] (1) Modeling method: On the first day, each chicken in the experimental group was given 0.6 ml of IBDV via nasal drops or eye drops. The medication was administered 48 hours after infection when symptoms such as fever, chills, huddling together, nasal or respiratory secretions, coughing, wheezing, and nasal discharge appeared.

[0070] (2) Dosing regimen: Blank control group: No modeling, administered blank solvent without drug by gavage; Model control group: After modeling, administered blank solvent without drug by gavage; Examples 1-5: After modeling, administered via drinking water (2ml of drug added per 1L of water); Comparative Example 1: After modeling, the powder was added to a 0.25%-0.5% sodium carboxymethyl cellulose solution to prepare a suspension for gavage administration, with a dosage of 1g / kg.

[0071] 3. Detection indicators and test results:

[0072] (1) Detection indicators: After 5 days of drug administration, observe for 24 hours and conduct clinical symptom observation and necropsy symptom scoring. See Table 1 and Table 2 for specific scores.

[0073] Table 1 Clinical Scoring Criteria

[0074]

[0075] (2) Results of the experiment: The clinical symptom scores are shown in Table 3, and the autopsy scores are shown in Table 4.

[0076] Table 3 Clinical symptom scoring results

[0077] Experimental Groups mental state Respiratory condition Blank group 5 4.9 Model group 2.4 1.9 Example 1 4.5 4.6 Example 2 4.8 4.8 Example 3 4.7 4.6 Example 4 4.5 4.3 Example 5 4.2 3.6 Comparative Example 1 3.9 3.9

[0078] Table 4. Autopsy Symptom Scoring Results

[0079] Experimental Groups trachea Intestines kidney Blank group 1.9 1.9 1.9 Model group 0.6 0.8 0.5 Example 1 1.7 1.5 1.4 Example 2 1.8 1.8 1.5 Example 3 1.8 1.7 1.7 Example 4 1.7 1.6 1.5 Example 5 1.4 1.3 1.3 Comparative Example 1 1.5 1.3 1.2

[0080] 4. Conclusion

[0081] Except for the control group, the other groups developed symptoms of exogenous wind-heat syndrome, such as fever, chills, nasal or respiratory secretions, cough, wheezing, and nasal discharge, 48 hours after infection with the infectious bronchitis virus. Therefore, the model of exogenous wind-heat syndrome caused by infectious bronchitis virus was successfully constructed.

[0082] Based on the results of each group of experiments, in the clinical symptom score, compared with the model group, Examples 1-5 significantly improved the mental state, respiratory tract, and diseased organs of the sick chickens, while Comparative Example 1 showed some improvement in the mental state, respiratory tract, and diseased organs of the sick chickens. In the necropsy score, Examples 1-5 and Comparative Example 1 all showed certain therapeutic effects on the trachea, intestines, and kidneys of the sick chickens. Among them, Examples 1-4 showed significant improvement in tracheal and kidney lesions, while Comparative Example 1 showed some effect in improving tracheal lesions. Therefore, Examples 1-5 can alleviate respiratory symptoms caused by infection with infectious bronchitis virus, and can also improve the lesions of organs such as the trachea and kidneys. Considering both the cost and therapeutic effect, Examples 2-4 are preferred.

[0083] Example 7: In vitro test against H9 subtype avian influenza virus

[0084] 1. Implementation Method

[0085] (1) Preparation of 1% chicken blood erythrocytes: Collect 3-5 mL of blood from the heart of a healthy chicken and place it in a 15 mL centrifuge tube containing sodium heparin. Gently invert the tube to mix, avoiding red blood cell breakage. Centrifuge at 3000 r / min for 5 min. Aspirate the supernatant, add 8-10 mL of PBS, mix well, and centrifuge again. Wash the red blood cells, repeating 1-2 times until the supernatant is no longer red. After the last centrifugation, discard the supernatant, measure the volume of red blood cells, and prepare a 1% chicken blood erythrocyte suspension with PBS. It can be stored at 4℃ for 1-2 days.

[0086] (2) HA (Hemagglutination assay): Take a clean 96-well V-type reaction plate and add 25 μL of PBS per well. Add 25 μL of allantoic fluid containing the virus to well 1, mix well, and then aspirate 25 μL to well 2. Repeat this serial dilution up to well 11, discarding 25 μL of the mixture from well 11. Well 12 serves as the red blood cell control group (negative control). Add 25 μL of 1% chicken blood red blood cell suspension to each of the above wells, mix well, and incubate at 37°C for 20-30 min.

[0087] (3) Result Interpretation: Stand the reaction plate upright and read the results immediately within a few seconds. A negative result indicates red blood cells flowing in a tear-like pattern. If red blood cells are scattered at the bottom of the well, forming a uniform thin layer, and do not flow when tilted, it indicates that the red blood cells have been agglutinated by the virus, which is a positive result. The highest dilution factor of the agglutinated antigen, i.e., the HA titer, is calculated when HA ≥ 4(-log2). M When the virus infection chicken embryo model is successfully constructed, it is determined that the model is successfully constructed.

[0088] (4) Experimental results:

[0089] Table 5 Results of in vitro anti-avian influenza virus assay

[0090]

[0091]

[0092] Note: Blood coagulation (HA): When HA ≥ 4 (-log2) M The viral infection model was considered successfully constructed at the specified time. Compared to the model group, a lower HA titer in the drug group indicates a better antiviral effect; compared to the model group, the drug group... ## The difference was highly significant (P<0.01). # The difference was statistically significant (P<0.05).

[0093] 2. Conclusion

[0094] Based on the experimental results, in terms of therapeutic effect, Examples 1-5 and Comparative Example 1 showed significant differences compared to the model group; in terms of preventive effect, Examples 2, 3, and 4 showed extremely significant differences compared to the model group, while Examples 1, 5, and the Comparative Example showed significant differences. Therefore, Examples 1-4 have a certain preventive and therapeutic effect against avian influenza virus, and considering both cost and therapeutic effect, Examples 2-4 are preferred.

[0095] Example 8: In vitro anti-swine influenza virus test

[0096] 1. Implementation Method

[0097] (1) Administration method

[0098] ① Blank control group: 96-well plates with confluent monolayer cells were cultured in a 37℃, 5% CO2 incubator for 72-120 h.

[0099] ② Virus model group: 96-well plates that have grown a confluent monolayer of cells were first inoculated with 1000 TCID. 50 Add 100 μL of virus solution per mL, discard after 2 h, then add 100 μL of cell maintenance medium (2% fetal bovine serum) per well, and incubate at 37°C in a 5% CO2 incubator for 72-120 h.

[0100] ③Inhibitory effect: 1000 TCID⁻¹ is first seeded into a 96-well plate that has already grown a monolayer of cells. 50 Add 100 μL of virus solution per mL, discard after 2 hours, then add 100 μL of drug-containing culture medium per well. Incubate at 37°C and 5% CO2 for 4 hours, then discard the drug-containing culture medium. Add 100 μL of cell maintenance medium (2% fetal bovine serum) to each well and continue culturing for 72-120 hours.

[0101] ④ Blocking effect: Add 100 μL of drug-containing medium to each well of a 96-well plate that has already grown a monolayer of cells. Incubate at 37°C and 5% CO2 for 4 hours, then discard the medium containing the test sample and add 1000 TCID50. 50Add 100 μL of virus solution per mL, discard the virus solution after 2 hours, add 100 μL of cell maintenance medium (2% fetal bovine serum) to each well, and continue culturing for 72-120 hours.

[0102] ⑤ Direct inactivation: Add 100 μL of the drug and virus mixture to each well of a 96-well plate that has been confluent with a monolayer of cells. Incubate at 37°C for 4 hours, then discard the liquid. Add 100 μL of cell maintenance medium (2% fetal bovine serum) to each well and continue culturing for 72-120 hours.

[0103] (2) Detection methods and result interpretation:

[0104] ①CCK8 method: After cells have been treated with drugs / viruses for 72-120 hours, the cell maintenance medium in the 96-well plate is discarded and replaced with fresh basal medium, 100 μL / well, and 10 μL of CCK8 solution is added to each well. After incubation in a cell culture incubator for 2 hours, the absorbance value at 450 nm is detected using an ELISA reader to obtain the OD value of each well.

[0105] ②Result Interpretation: The protective rate of the drug against virus-infected cells = [OD] (药物组) -OD (病毒模型组) ] / [OD (不加药空白组) -OD (病毒模型组) ]×100%

[0106] (3) Test Results

[0107] Table 6. Results of in vitro anti-swine influenza virus assay

[0108]

[0109] Note: 1. The protective rate of a drug against virus-infected cells = [OD] (药物组) -OD (病毒模型组) ] / [OD (不加药空白组) -OD (病毒模型组) ×100%; Cell protection rate: <0 no protection and damages cells, 10%-20% low protection rate, 20%-30% relatively high protection rate, >30% high protection rate.

[0110] 2. Conclusion

[0111] Based on the experimental results, Examples 1-5 showed high protection rates against cells infected with swine influenza virus in terms of inhibition, blocking, and direct inactivation. Examples 2, 3, and 4 demonstrated good inhibition of swine influenza virus replication, blocking of virus adsorption, and direct inactivation. Comparative Example 1 showed some protection against swine influenza virus-infected cells in terms of inhibition and blocking, but low protection in terms of direct inactivation. Therefore, Examples 2, 3, and 4 exhibited significant anti-swine influenza virus effects.

[0112] Example 9: Clinical efficacy trial of exogenous wind-heat syndrome in broilers

[0113] 1. Experimental Background

[0114] A poultry farm in Changzhi City, Shanxi Province, raised 270,000 17-day-old Lifeng chickens. 30-40% of them showed respiratory symptoms such as wheezing, coughing, nasal discharge, respiratory rales, and open-mouth breathing. The samples were sent to a disease testing center and identified as a mixed infection of infectious bronchitis and H9 subtype avian influenza.

[0115] 2. Experimental Objective

[0116] The clinical use of the formulation of this invention verifies whether it can be used to relieve exogenous wind-heat syndrome and respiratory symptoms caused by mixed infection of infectious bronchitis virus and H9 avian influenza virus.

[0117] 3. Test Plan

[0118] 3.1 Experimental Subjects

[0119] The experiment selected four chicken houses (40,000 chickens in each house) with similar ages and medication procedures. Experiment 1 used Example 2 of the present invention; Experiment 2 used Example 3 of the present invention; Experiment 3 used Example 4 of the present invention; and Experiment 4 used Comparative Example 1.

[0120] 3.2 Test Materials

[0121] The samples used in this experiment were provided by Ruipu Research Institute. Experiment 1 was the traditional Chinese medicine preparation of Example 2 of this invention, 500ml / bottle, batch number 20230801; Experiment 2 was the traditional Chinese medicine preparation of Example 3 of this invention, 500ml / bottle, batch number 20230802; Experiment 3 was the traditional Chinese medicine preparation of Example 4 of this invention, 500ml / bottle, batch number 20230803; Comparative Example 1 was the traditional Chinese medicine preparation of Comparative Example 1, 500ml / bag, batch number 20230804.

[0122] 3.3 Test Methods

[0123] The experimental group received the medication via drinking water twice daily for 5 days; the control group received the medication mixed with feed twice daily for 5 days. Mortality rates and clinical symptoms were recorded for each group.

[0124] Criteria for evaluating therapeutic efficacy:

[0125] ①Cure: Clinical respiratory symptoms disappear, mortality rate, mental state, and stool condition return to normal.

[0126] ②Effective: Clinical respiratory symptoms improved, mortality rate decreased, mental state improved, and stool condition improved significantly.

[0127] ③ Ineffective: No improvement was observed in clinical respiratory symptoms, mortality rate, mental status, or stool condition.

[0128] 4. Test Results

[0129] (1) Death rate

[0130] Table 7 Mortality rate (‰) before and after medication

[0131] experimental group Experimental Group 1 (‰) Experimental Group 2 (‰) Experimental group 3 (‰) Comparative Example 1 (‰) 3 days before medication 9.4 9.7 9.3 9.5 Day 1 of medication 8.2 8.3 7.8 8.7 Day 2 of medication 7.4 7.1 6.3 7.6 Day 3 of medication 6.6 6.3 5.2 6.4 Day 4 of medication 5.4 5.2 4.5 5.2 Day 5 of medication 4.7 4.4 4.2 5.0 Day 1 after medication ends 4.4 4.1 3.9 4.7 Day 2 after medication ended 3.9 3.7 3.5 5.6 Day 3 after medication ended 3.5 3.2 2.8 5.8

[0132] (2) Cure rate

[0133] Table 8. Cure rate (%) after medication completion

[0134] Experimental Groups Experiment 1 group Experimental Group 2 Experiment 3 groups Comparative Example 1 Cure rate (%) 96.4％ 95.2％ 94.7％ 86.3％

[0135] 5. Conclusion

[0136] In terms of mortality rate, the mortality rates of Examples 2, 3, and 4 and Comparative Example 1 of this invention can be significantly reduced by approximately 10% to 30%; however, the mortality rate of Comparative Example 1 increased slightly 2 and 3 days after administration. In terms of cure rate, the cure rates of Examples 2, 3, and 4 are significantly higher than those of Comparative Example 1. Therefore, this invention can alleviate exogenous wind-heat syndrome and respiratory symptoms caused by mixed infection of infectious bronchitis virus and H9 subtype avian influenza virus.

[0137] Example 10: Clinical efficacy trial of exogenous wind-heat syndrome in pigs

[0138] 1. Experimental Background

[0139] A pig farm in Hebei Province discovered fattening pigs suspected of having swine influenza. The sick pigs had elevated body temperature, lethargy, loss of appetite, and mucus in their eyes and nose. Some pigs also showed respiratory symptoms such as difficulty breathing, wheezing, and coughing. The samples were sent to a disease testing center, where the virus was identified as swine influenza virus infection.

[0140] 2. Experimental Objective

[0141] The clinical use of the formulation of this invention verifies whether this invention can be used to alleviate exogenous wind-heat syndrome caused by swine influenza infection and the resulting respiratory symptoms.

[0142] 3. Test Plan

[0143] 3.1 Experimental Subjects

[0144] Eighty sick pigs with obvious clinical respiratory symptoms were selected for the experiment. Experiment 1 used Example 2 of the present invention; Experiment 2 used Example 3 of the present invention; Experiment 3 used Example 4 of the present invention; and Experiment 4 used Comparative Example 1.

[0145] 3.2 Test Materials

[0146] The samples used in this experiment were provided by Ruipu Research Institute. Experiment 1 was the traditional Chinese medicine preparation of Example 2 of this invention, 500ml / bottle, batch number 20230801; Experiment 2 was the traditional Chinese medicine preparation of Example 3 of this invention, 500ml / bottle, batch number 20230802; Experiment 3 was the traditional Chinese medicine preparation of Example 4 of this invention, 500ml / bottle, batch number 20230803; Comparative Example 1 was the traditional Chinese medicine preparation of Comparative Example 1, 500ml / bag, batch number 20230804.

[0147] 3.3 Test Methods

[0148] All experimental groups were administered the medication orally once a day for 7 consecutive days; control group 1 was administered the medication by mixing it into feed once a day for 7 consecutive days.

[0149] Criteria for evaluating therapeutic efficacy:

[0150] ① Cured: Clinical symptoms such as fever, difficulty breathing, and runny nose completely disappear, body temperature returns to normal, and mental state and appetite return to normal.

[0151] ②Effective: Clinical symptoms such as fever, difficulty breathing, and runny nose are significantly reduced, and mental state and appetite gradually recover.

[0152] ③ Ineffective: Symptoms do not improve significantly or worsen.

[0153] ④ Overall effective rate = (number of cured + number of effective cases) / total number of cases in each group.

[0154] 4. Test Results

[0155] Table 9. Cure status after medication completion

[0156] Experimental Groups Animal number cure rate efficiency inefficiency Overall effectiveness (%) Experiment 1 group 20 12 7 1 95％ Experimental Group 2 20 10 8 2 90％ Experiment 3 groups 20 7 10 3 85％ Comparative Example 1 20 5 8 7 65％

[0157] 5. Conclusion

[0158] As can be seen from Table 9, the total effective rate of the two groups of Examples 2 and 3 of the present invention can reach more than 90%, and the total effective rate of the four groups of Examples 4 can also reach 85%, which are all higher than the comparative example 1. It can be seen that the present invention can alleviate the exogenous wind-heat syndrome caused by swine influenza virus and the respiratory symptoms it causes.

[0159] The above embodiments are only used to illustrate the technical solutions of the present invention, and are not intended to limit it. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A traditional Chinese medicine composition for preventing and treating exogenous wind-heat syndrome in livestock and poultry, characterized in that, The raw materials in the traditional Chinese medicine composition include Isatis root, gypsum, reed rhizome, Rehmannia root, turmeric, anemarrhena rhizome, acorus tatarinowii rhizome, patchouli, and forsythia.

2. The traditional Chinese medicine composition according to claim 1, characterized in that, The raw material components in the traditional Chinese medicine composition are, by weight, 60-100 parts of Isatis root, 20-60 parts of gypsum, 20-60 parts of reed rhizome, 10-40 parts of Rehmannia root, 10-30 parts of turmeric, 10-30 parts of anemarrhena rhizome, 10-30 parts of acorus tatarinowii rhizome, 10-30 parts of patchouli rhizome, and 15-50 parts of forsythia fruit.

3. The traditional Chinese medicine composition according to claim 2, characterized in that, The raw material components in the traditional Chinese medicine composition are, by weight, 80-85 parts of Isatis root, 35-40 parts of gypsum, 35-40 parts of reed rhizome, 20-25 parts of Rehmannia root, 15-20 parts of turmeric, 15-20 parts of anemarrhena rhizome, 15-20 parts of acorus tatarinowii rhizome, 15-20 parts of patchouli rhizome, and 30-35 parts of forsythia fruit.

4. A method for preparing a traditional Chinese medicine composition for preventing and treating exogenous wind-heat syndrome in livestock and poultry as described in any one of claims 1-3, comprising the following steps: (1) Weigh the above raw material components according to the prescription amount, add 10-14 times the amount of purified water to the raw material components and decoct 3 times, 1-2 hours each time, while collecting the volatile oil and volatile oil emulsion; combine the decoctions, filter and set aside. (2) Concentrate the decoction under reduced pressure to a certain volume, add the antibacterial agent and stir until dissolved, add the volatile oil and volatile oil emulsion, and mix well; (3) Adjust the pH value with 10% sodium hydroxide solution, add purified water to make up the volume, mix well, filter, and fill into a container to obtain the product.

5. The preparation method according to claim 4, characterized in that, The temperature of the vacuum concentration in step (2) is 50-70℃; the vacuum degree of the vacuum concentration is 0.08-0.09 MPa; and the volume of the vacuum concentration is 60%-70% of the total volume of the decoction.

6. The preparation method according to claim 4, characterized in that, The antibacterial agent mentioned in step (2) is potassium sorbate, and the addition amount is 0.2% to 0.3%.

7. The preparation method according to claim 4, characterized in that, The pH value in step (3) is 4.0 to 6.

0.

8. The use of the traditional Chinese medicine composition for preventing and treating exogenous wind-heat syndrome in livestock and poultry as described in any one of claims 1-3 in the preparation of a medicine for treating exogenous wind-heat syndrome caused by livestock and poultry viruses and the respiratory symptoms caused thereto.

9. The application according to claim 8, characterized in that, The livestock and poultry viruses mentioned are chicken infectious bronchitis virus, avian influenza virus, and swine influenza virus.

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