A traditional Chinese medicine composition for preventing and treating eperythrozoons disease of animals and application thereof

Abstract

The present application belongs to the technical field of pharmaceutical preparation, and discloses a traditional Chinese medicine composition for preventing and treating eperythrozoonosis of animals, which is composed of the following raw medicinal materials in parts by weight: 25-35 parts of Corydalis decumbens, 12-25 parts of Hypericum japonicum, 12-25 parts of Hemerocallis fulva, 10-20 parts of Jatropha curcas leaves, and 8-16 parts of Smilax china. The raw medicinal materials are Chinese medicinal materials or Chinese medicine extracts equivalent to the crude drug amount of the above-mentioned Chinese medicinal materials. The traditional Chinese medicine composition can greatly improve the cure rate of sick pigs and effectively reduce breeding losses. Moreover, the traditional Chinese medicine composition is green, safe, non-toxic, and has no side effects, and has the advantages of prevention and treatment, and good market application value.

Description

Technical Field

[0001] This invention relates to the field of veterinary drug technology, specifically to a traditional Chinese medicine composition for the prevention and treatment of hemotropic mycoplasmosis in animals and its application. Background Technology

[0002] Eperythrozoonosis is a zoonotic infectious disease caused by *Eperythrozoon* (EH), which parasitizes the surface of red blood cells, plasma, tissue fluid, and bone marrow in humans and animals. Its main clinical symptoms include anemia, jaundice, and fever. Since Schilling and Dinger discovered *E. coccoides* in rodents in 1928, cases of eperythrozoonosis have been reported in various countries. In 1934, Adler et al. discovered a microorganism with a morphology similar to *E. coccoides* in cattle and named it *E. wenyoni*. Subsequently, infections with *E. coccoides* have been confirmed in pigs, cattle, sheep, goats, horses, rabbits, dogs, cats, monkeys, camels, foxes, donkeys, deer, wildebeest, llamas, birds, and humans. To date, dozens of species of *E. coccoides* have been discovered and named.

[0003] Porcine eperythrozoonosis is an acute, febrile infectious disease of pigs caused by *Eperythrozoon suis*. The main clinical symptoms are lethargy and fever, along with a tendency to sleep and huddle together. As the disease spreads, infected pigs may develop anemia and jaundice, and petechiae of varying degrees may appear under the skin pores, especially on the surface of the skin, abdomen, and inner sides of the limbs. In severe cases, the skin may turn purplish-red, making it easily noticeable. Some infected pigs may develop varying degrees of conjunctivitis, causing eyelid adhesions and affecting vision. Blood physiological and biochemical indicators will change accordingly in mild and severe cases of porcine eperythrozoon infection.

[0004] Mycoplasma eperythrozoonosis is an opportunistic pathogen that is prevalent during the hot summer months. It typically affects pigs with weakened immune systems, and outbreaks are easily triggered by farrowing, overcrowding, long-distance transport, inclement weather, poor husbandry practices, or changes in pen or feed. The disease has many transmission routes, the most important being through the bites of blood-sucking insects (flies, mosquitoes, scabies mites). Transmission of Mycoplasma eperythrozoonosis has been confirmed through contact, bloodborne, vertical transmission, or through blood-sucking insects such as mosquitoes, flies, and ectoparasites. Pigs with scabies mites can transmit the disease through broken skin.

[0005] While eperythrozoonosis rarely causes death when it occurs alone, it is very easy to cause mixed infections with other diseases, leading to high fever and difficulty in treatment. Clinically, this disease is often mixed with diseases such as Toxoplasma gondii, classical swine fever, and Streptococcus suis, which is often overlooked. Later, it can trigger diseases such as porcine circovirus and paraswine parasitic infection, which are very difficult to treat.

[0006] Currently, the treatment of porcine eperythrozoonosis mainly relies on antibiotics, such as oxytetracycline. However, with the increasing drug resistance of pathogens, the clinical efficacy is not ideal. Furthermore, the overuse of antibiotics in animal husbandry not only leads to drug resistance in pathogens but also causes drug residues in livestock products, endangering human food safety.

[0007] Given the potential harm that eperythrozoonosis poses to actual aquaculture production, and the lack of highly effective, green, and environmentally friendly treatment drugs, this invention aims to find a traditional Chinese medicine composition that is effective and inexpensive in preventing and treating eperythrozoonosis. Summary of the Invention

[0008] In view of the problems and shortcomings of the existing technology, the purpose of this invention is to provide a traditional Chinese medicine composition for the prevention and treatment of animal eperythrozoonosis and its application.

[0009] To achieve the objectives of this invention, the technical solution adopted is as follows:

[0010] The first aspect of this invention provides a traditional Chinese medicine composition for preventing and treating animal eperythrozoonosis. The traditional Chinese medicine composition consists of the following raw materials in parts by weight: 25-35 parts of Corydalis yanhusuo, 12-25 parts of Lysimachia christinae, 12-25 parts of Hemerocallis fulva, 10-20 parts of Melia toosendan leaves, and 8-16 parts of Gynostemma pentaphyllum. The raw materials are derived from traditional Chinese medicinal materials or traditional Chinese medicine extracts equivalent to the raw amount of the above-mentioned traditional Chinese medicinal materials.

[0011] According to the above-mentioned traditional Chinese medicine composition, preferably, the traditional Chinese medicine composition is composed of the following raw materials in parts by weight: 28-31 parts of Corydalis yanhusuo, 18-22 parts of Lysimachia christinae, 18-22 parts of Hemerocallis fulva, 14-17 parts of Melia toosendan leaves, and 10-12 parts of Gynostemma pentaphyllum.

[0012] According to the above-mentioned traditional Chinese medicine composition, preferably, the traditional Chinese medicine composition is composed of the following raw materials in parts by weight: 30 parts of Corydalis yanhusuo, 20 parts of Lysimachia christinae, 20 parts of Hemerocallis fulva, 15 parts of Melia toosendan leaves, and 10 parts of Gynostemma pentaphyllum.

[0013] According to the above-mentioned traditional Chinese medicine composition, preferably, the animal includes pigs, cattle, sheep, horses, rabbits, dogs, cats, monkeys, camels, foxes, donkeys, deer, wildebeests, llamas, etc. More preferably, the animal is a pig.

[0014] The second aspect of the present invention provides a method for preparing the traditional Chinese medicine composition described in the first aspect above. The method is as follows: Corydalis yanhusuo, Lysimachia christinae, Hemerocallis fulva, Melia toosendan leaves, and Gynostemma pentaphyllum are mixed in proportion and then extracted with a solvent. After extraction, the mixture is filtered and the filtrate is collected to obtain the traditional Chinese medicine composition.

[0015] According to the preparation method described above, preferably, the solvent is a water or ethanol solution. More preferably, the solvent is water.

[0016] According to the above preparation method, preferably, the mass ratio of the solvent to the total weight of the active pharmaceutical ingredient is 6 to 10:1.

[0017] According to the above preparation method, preferably, the extraction method is flash extraction.

[0018] According to the above preparation method, preferably, the flash extraction voltage is 150V to 200V, the extraction speed is 7000r / min to 9000r / min, and the extraction time is 10 to 20min.

[0019] According to the above preparation method, preferably, the raw material is crushed and sieved before extraction; more preferably, the crushing temperature is 5-10℃ and the sieve mesh size is 10-20 mesh.

[0020] The third aspect of this invention provides the use of the traditional Chinese medicine composition described in the first aspect above in the preparation of a medicament for the prevention and / or treatment of hemotropic mycoplasmosis in animals.

[0021] According to the above application, preferably, the animal includes pigs, cattle, sheep, horses, rabbits, dogs, cats, monkeys, camels, foxes, donkeys, deer, wildebeest, llamas, etc. More preferably, the animal is a pig.

[0022] The fourth aspect of the present invention provides a medicament for the prevention and / or treatment of hemotropic mycoplasmosis in animals, said medicament being made from the traditional Chinese medicine composition described in the first aspect above and pharmaceutically acceptable excipients.

[0023] According to the above-described drug, preferably, the drug is an oral dosage form. More preferably, the drug is an oral liquid, granules, tablets, or powder. Most preferably, the drug is a granule, and the pharmaceutical excipients include, but are not limited to, maltodextrin, sucrose, glucose, etc.

[0024] According to the above-mentioned drugs, preferably, the method of using granule drugs is to mix 1 kg of granule drugs with 500-1000 kg of feed and use it continuously for 5-7 days.

[0025] According to the aforementioned drug, preferably, the animal includes pigs, cattle, sheep, horses, rabbits, dogs, cats, monkeys, camels, foxes, donkeys, deer, wildebeest, llamas, etc. More preferably, the animal is a pig.

[0026] The fifth aspect of the present invention provides a method for preparing a granular drug for the prevention and / or treatment of porcine eperythrozoonosis, comprising concentrating the liquid of the traditional Chinese medicine composition described in the first aspect above and performing a paste-forming process, then adding excipients and granulating to obtain the granular drug.

[0027] According to the above preparation method, preferably, the concentration process is carried out using a double-effect concentrator until a clear paste with a relative density of 1.15 to 1.18 (50 to 60°C) is formed.

[0028] According to the above preparation method, preferably, the granulation process is carried out using a fluidized bed granulation device, with a feed rate of 12-14 r / min and an air volume of 84-86 m³ / min. 3 / h.

[0029] All components in the traditional Chinese medicine composition of this invention can be purchased commercially available products. The pharmacological properties of each component in the traditional Chinese medicine composition are as follows:

[0030] Corydalis fulva: The whole herb of Corydalis fulva, a plant in the Papaveraceae family, is bitter and cold in nature. It enters the lung, liver, and gallbladder meridians, clears heat and toxins, benefits the liver and gallbladder, cools the blood and stops bleeding; it is mainly used for high fever due to febrile diseases, damp-heat jaundice, etc.

[0031] Tianjihuang; the whole herb of *Gynostemma pentaphyllum*, a plant in the Clusiaceae family. It has a sweet and bitter taste and is cool in nature. It enters the lung, liver, and stomach meridians, and clears heat and dampness, detoxifies, disperses blood stasis and reduces swelling. It is mainly used for damp-heat jaundice, diarrhea, dysentery, intestinal abscesses, etc.

[0032] Daylily flower: The dried flower buds of the daylily (Hemerocallis fulva) or goldenrod (Hemerocallis fulva) plant, belonging to the lily family. It has a sweet taste and cool properties. It promotes urination and eliminates dampness, clears heat and quenches thirst, relieves depression and soothes the chest; used for symptoms such as painful urination, fever, and thirst.

[0033] The leaves of the Chinaberry tree (Melia azedarach), a plant in the Myrtaceae family, are pungent and bitter in taste, and neutral in nature. They are used to regulate qi, resolve dampness, detoxify, and kill parasites. They are primarily used for abdominal distension and pain due to dampness, and dysentery.

[0034] Hejincao: The whole herb of *Gnaphalium affine*, a plant in the Caryophyllaceae family. It has a bland taste and neutral properties; it promotes urination, relieves strangury, regulates qi, relieves pain, and helps heal bones and injuries. It is mainly used for urinary tract infections caused by heat and urinary stones.

[0035] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0036] (1) For symptoms of jaundice such as lethargy, fever, anemia, jaundice, and varying degrees of petechiae and conjunctivitis caused by porcine eperythrozoonosis, this invention uses Corydalis yanhusuo as the principal herb, which is bitter and cold in nature and can clear heat from the lung, liver, and gallbladder meridians; it uses Lysimachia christinae and Hemerocallis fulva as assistant herbs, both of which are sweet and promote body fluid production, and are mainly used to clear heat, promote bile secretion, remove dampness, disperse blood stasis and reduce swelling, and assist the principal herb in clearing the liver and promoting bile secretion; it uses Melia toosendan leaves and Gynostemma pentaphyllum as adjuvant herbs, Melia toosendan leaves are pungent and bitter, and can regulate qi and resolve dampness, and when combined with Gynostemma pentaphyllum, which is bland and neutral in nature, can promote urination, relieve strangury, regulate qi and relieve pain, and assist the principal herb in regulating qi and resolving dampness; the combined use of these herbs can achieve the effects of clearing the liver and promoting bile secretion, and removing dampness and jaundice.

[0037] (2) The efficacy experiment of the traditional Chinese medicine composition of the present invention on porcine eperythrozoonosis shows that the traditional Chinese medicine composition of the present invention has good preventive and therapeutic effects on porcine eperythrozoonosis, can significantly improve the clinical symptoms of porcine eperythrozoonosis, increase the cure rate of diseased pigs, and effectively reduce breeding losses; moreover, the traditional Chinese medicine composition of the present invention has the advantages of being green and safe, having no toxic side effects, and combining prevention and treatment, filling the gap in the lack of traditional Chinese medicine products for porcine eperythrozoonosis in the existing national veterinary drug products, which can reduce the use of antibiotics for such diseases in clinical practice, reduce drug resistance and veterinary drug residues, and ensure food safety and human health.

[0038] (3) The drug of the present invention for the prevention and / or treatment of animal hemotropic mycoplasmosis can be mixed with feed or drinking water, is easy to use, is suitable for large-scale pig farms, and has broad development space and good market prospects. Attached Figure Description

[0039] Figure 1 Photos showing the clinical symptoms of pigs infected with porcine eperythrozoonosis. Detailed Implementation

[0040] To enable those skilled in the art to better understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below with reference to specific embodiments.

[0041] The following detailed description is exemplary and intended to provide further illustration of the invention. Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. It should be noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the scope of exemplary embodiments of the invention.

[0042] Unless otherwise specified, the experimental methods in the following examples all employ conventional techniques in this technical field or follow the conditions recommended by the manufacturer; reagents or instruments whose manufacturers are not specified are all commercially available products.

[0043] Example 1:

[0044] A drug for preventing and treating eperythrozoonosis in animals, the drug being made from a traditional Chinese medicine composition and pharmaceutically acceptable excipients. By weight, the traditional Chinese medicine composition comprises the following raw materials: 30 parts of Corydalis yanhusuo, 20 parts of Lysimachia christinae, 20 parts of Hemerocallis fulva, 15 parts of Melia toosendan leaves, and 10 parts of Gynostemma pentaphyllum.

[0045] The preparation method of the above-mentioned drug is as follows: Weigh out the following raw materials according to their respective weight proportions in the above-mentioned traditional Chinese medicine composition: Corydalis yanhusuo, Lysimachia christinae, Hemerocallis fulva, Melia toosendan leaves, and Gynostemma pentaphyllum. Mix the above raw materials in proportion, pulverize them, pass them through a 20-mesh sieve, and add them to a flash extractor. Add water equal to 10 times the total weight of the raw materials to the flash extractor for extraction. The extraction voltage is 150V, the rotation speed is 9000r / min, and the extraction time is 15min, repeated twice. After extraction, the liquid and residue are transferred together to a filter press. The pressure is controlled at 0.12MPa, and the filter cloth pore size is 200 mesh. After filtration, the collected liquid is transferred to a double-effect concentrator for concentration until a clear extract with a relative density of 1.15 (50℃) is formed. Then, add the excipient maltodextrin and further granulate using a fluidized bed granulator. During granulation, the feed rate is 12r / min and the air volume is 85m³ / min. 3 / h, after granulation, the granules are sieved and packaged to obtain the final product.

[0046] Example 2:

[0047] A drug for preventing and treating eperythrozoonosis in animals, the drug being made from a traditional Chinese medicine composition and pharmaceutically acceptable excipients. By weight, the traditional Chinese medicine composition comprises the following raw materials: 35 parts of Corydalis yanhusuo, 15 parts of Lysimachia christinae, 18 parts of Hemerocallis fulva, 15 parts of Melia toosendan leaves, and 12 parts of Gynostemma pentaphyllum.

[0048] The preparation method of the above-mentioned drug is the same as that in Example 1.

[0049] Example 3:

[0050] A drug for preventing and treating eperythrozoonosis in animals, the drug being made from a traditional Chinese medicine composition and pharmaceutically acceptable excipients. By weight, the traditional Chinese medicine composition comprises the following raw materials: 25 parts of Corydalis yanhusuo, 15 parts of Lysimachia christinae, 15 parts of Hemerocallis fulva, 10 parts of Melia toosendan leaves, and 8 parts of Gynostemma pentaphyllum.

[0051] The preparation method of the above-mentioned drug is the same as that in Example 1.

[0052] Example 4:

[0053] A drug for preventing and treating eperythrozoonosis in animals, the drug being made from a traditional Chinese medicine composition and pharmaceutically acceptable excipients. By weight, the traditional Chinese medicine composition comprises the following raw materials: 25 parts of Corydalis yanhusuo, 12 parts of Lysimachia christinae, 12 parts of Hemerocallis fulva, 10 parts of Melia toosendan leaves, and 8 parts of Lysimachia christinae.

[0054] The preparation method of the above-mentioned drug is the same as that in Example 1.

[0055] Example 5:

[0056] A drug for preventing and treating eperythrozoonosis in animals, the drug being made from a traditional Chinese medicine composition and pharmaceutically acceptable excipients. By weight, the traditional Chinese medicine composition comprises the following raw materials: 26 parts of Corydalis yanhusuo, 16 parts of Lysimachia christinae, 16 parts of Hemerocallis fulva, 14 parts of Melia toosendan leaves, and 10 parts of Gynostemma pentaphyllum.

[0057] The preparation method of the above-mentioned drug is the same as that in Example 1.

[0058] Example 6:

[0059] A drug for preventing and treating eperythrozoonosis in animals, the drug being made from a traditional Chinese medicine composition and pharmaceutically acceptable excipients. By weight, the traditional Chinese medicine composition comprises the following raw materials: 30 parts of Corydalis yanhusuo, 20 parts of Lysimachia christinae, 18 parts of Hemerocallis fulva, 18 parts of Melia toosendan leaves, and 12 parts of Gynostemma pentaphyllum.

[0060] The preparation method of the above-mentioned drug is the same as that in Example 1.

[0061] Example 7:

[0062] A drug for preventing and treating eperythrozoonosis in animals, the drug being made from a traditional Chinese medicine composition and pharmaceutically acceptable excipients. By weight, the traditional Chinese medicine composition comprises the following raw materials: 35 parts of Corydalis yanhusuo, 20 parts of Lysimachia christinae, 25 parts of Hemerocallis fulva, 20 parts of Melia toosendan leaves, and 16 parts of Gynostemma pentaphyllum.

[0063] The preparation method of the above-mentioned drug is the same as that in Example 1.

[0064] Example 8:

[0065] A drug for preventing and treating eperythrozoonosis in animals, the drug being made from a traditional Chinese medicine composition and pharmaceutically acceptable excipients. By weight, the traditional Chinese medicine composition comprises the following raw materials: 35 parts of Corydalis yanhusuo, 12 parts of Lysimachia christinae, 12 parts of Hemerocallis fulva, 10 parts of Melia toosendan leaves, and 8 parts of Gynostemma pentaphyllum.

[0066] The preparation method of the above-mentioned drug is the same as that in Example 1.

[0067] Example 9:

[0068] A drug for preventing and treating eperythrozoonosis in animals, the drug being made from a traditional Chinese medicine composition and pharmaceutically acceptable excipients. By weight, the traditional Chinese medicine composition comprises the following raw materials: 25 parts of Corydalis yanhusuo, 25 parts of Lysimachia christinae, 20 parts of Hemerocallis fulva, 15 parts of Melia toosendan leaves, and 15 parts of Gynostemma pentaphyllum.

[0069] The preparation method of the above-mentioned drug is the same as that in Example 1.

[0070] Example 10:

[0071] A drug for preventing and treating eperythrozoonosis in animals, the drug being made from a traditional Chinese medicine composition and pharmaceutically acceptable excipients. By weight, the traditional Chinese medicine composition comprises the following raw materials: 28 parts of Corydalis yanhusuo, 20 parts of Lysimachia christinae, 20 parts of Hemerocallis fulva, 18 parts of Melia toosendan leaves, and 11 parts of Gynostemma pentaphyllum.

[0072] The preparation method of the above-mentioned drug is the same as that in Example 1.

[0073] Example 11:

[0074] A drug for preventing and treating eperythrozoonosis in animals, the drug being made from a traditional Chinese medicine composition and pharmaceutically acceptable excipients. By weight, the traditional Chinese medicine composition comprises the following raw materials: 31 parts of Corydalis yanhusuo, 18 parts of Lysimachia christinae, 18 parts of Hemerocallis fulva, 15 parts of Melia toosendan leaves, and 12 parts of Gynostemma pentaphyllum.

[0075] The preparation method of the above-mentioned drug is the same as that in Example 1.

[0076] Efficacy verification test of the drug for preventing and treating eperythrozoonosis in animals according to this invention:

[0077] (I) Efficacy verification against hemotropic mycoplasmosis in rats

[0078] To verify the efficacy of the drug for preventing and treating animal eperythrozoonosis according to the present invention, efficacy verification experiments were conducted using the drugs prepared in Examples 1, 2, and 3 of the present invention as examples. Simultaneously, to compare with the drug for preventing and treating animal eperythrozoonosis according to the present invention, the present invention also set up the following comparative experiments, namely Comparative Examples 1 to 5. The drugs in Comparative Examples 1 to 5 are basically the same as those in Example 1 of the present invention, except that the raw materials used in the traditional Chinese medicine composition in the formula are different. See Table 1 for details of the raw materials used in the traditional Chinese medicine compositions of Comparative Examples 1 to 5.

[0079] Table 1. Raw materials of the traditional Chinese medicine compositions used in each experimental group.

[0080] Example 1 30 parts of Corydalis yanhusuo, 20 parts of Lysimachia christinae, 20 parts of Hemerocallis fulva, 15 parts of Melia toosendan leaves, and 10 parts of Lysimachia christinae. Example 2 35 parts of Corydalis yanhusuo, 15 parts of Lysimachia christinae, 18 parts of Hemerocallis fulva, 15 parts of Melia toosendan leaves, and 12 parts of Lysimachia christinae. Example 3 25 parts of Corydalis yanhusuo, 15 parts of Lysimachia christinae, 15 parts of Hemerocallis fulva, 10 parts of Melia toosendan leaves, and 8 parts of Lysimachia christinae. Comparative Example 1 30 portions of Corydalis yanhusuo Comparative Example 2 20 portions of *Lysimachia christinae* and 20 portions of daylily. Comparative Example 3 15 parts of chinaberry leaves and 10 parts of sedge Comparative Example 4 30 parts of Corydalis yanhusuo, 15 parts of Melia toosendan leaves, and 10 parts of Clematis armandii. Comparative Example 5 20 parts of *Lysimachia christinae*, 20 parts of daylily flowers, 15 parts of *Melia azedarach* leaves, and 10 parts of *Gynostemma pentaphyllum*.

[0081] 1. Laboratory animals:

[0082] The SPF-grade SD rats used in this experiment were purchased from the Henan Provincial Experimental Animal Center. They were 4-5 weeks old, with half males and half females. Blood was collected from the tail vein for microscopic examination of fresh blood pressure smears to confirm that the rats were not infected with Mycoplasma suis.

[0083] 2. Test methods

[0084] (1) Experimental drugs:

[0085] The experimental drugs were the drugs prepared in Examples 1 to 3 of this invention and the drugs prepared in Comparative Examples 1 to 5.

[0086] (2) Experimental grouping and treatment:

[0087] After all experimental animals completed their quarantine period, they were randomly grouped using Excel software to ensure that each animal had an equal chance of being assigned to an experimental group, minimizing the influence of human factors on the experiment. Before each drug administration experiment, rats were randomly divided into 10 groups of 10 each, with half males and half females: control group (hereinafter referred to as "Group A"), model group (hereinafter referred to as "Group B"), Example 1 group (hereinafter referred to as "Group C"), Example 2 group (hereinafter referred to as "Group D"), Example 3 group (hereinafter referred to as "Group E"), comparative example 1 group (hereinafter referred to as "Group F"), comparative example 2 group (hereinafter referred to as "Group G"), comparative example 3 group (hereinafter referred to as "Group H"), comparative example 4 group (hereinafter referred to as "Group I"), and comparative example 5 group (hereinafter referred to as "Group J").

[0088] Blood samples from pigs infected with *Eperythrozoon suis* were washed three times with sterile PBS and then intraperitoneally inoculated into rats in all groups except group A, 0.5 ml per rat. Five days after artificial infection, blood was collected from the tail vein and observed under a 1000x oil immersion microscope. A large number of irregularly edged dentate erythrocytes were found in the field of view. The erythrocytes were degenerated and fragmented, or had spherical, disc-shaped, ring-shaped, comma-shaped, and rod-shaped appendages attached around them. This indicated that a successful *Eperythrozoon suis* infection model had been obtained.

[0089] (3) Dosing regimen

[0090] Route of administration: The intended route of clinical use for this test product is oral administration; therefore, gavage administration was adopted.

[0091] Administration method: Add 5g of the drugs prepared in Examples 1-3 and Comparative Examples 1-5 to physiological saline to a final volume of 100ml and mix well. The administration volume is 20ml / kg·bw, administered 5 days after infection with Mycoplasma suis. Using a 12-gauge gavage needle, insert the tip into the rat's mouth and slowly advance it downwards along the right corner of the mouth, allowing the tip of the tube to slide past the maxilla and into the pharynx. When resistance is felt, slightly tilt the tip of the tube ventrally to allow it to enter the esophagus. Once it has reached the stomach, insert it slightly further forward and then begin injecting the drug.

[0092] Control group (Group A): Administered normal saline 20 mL / kg bw by gavage once a day for 7 consecutive days;

[0093] Model group (Group B): Administered 20 mL / kg bw of normal saline by gavage once a day for 7 consecutive days;

[0094] Example 1 Group (Group C): The drug prepared in Example 1 was administered by gavage at a dose of 20 mL / kg·bw once a day for 7 consecutive days;

[0095] Example 2 group (Group D): The drug prepared in Example 2 was administered by gavage at a rate of 20 mL / kg·bw once a day for 7 consecutive days;

[0096] Example 3 group (Group E): The drug prepared in Example 3 was administered by gavage at a rate of 20 mL / kg·bw once a day for 7 consecutive days;

[0097] Comparative Example 1 (F Group): The drug prepared in Comparative Example 1 was administered by gavage at a rate of 20 mL / kg·bw once a day for 7 consecutive days.

[0098] Comparative Example 2 (Group G): The drug prepared in Comparative Example 2 was administered by gavage at a rate of 20 mL / kg·bw once a day for 7 consecutive days.

[0099] Comparative Example 3 (H Group): The drug prepared in Comparative Example 3 was administered by gavage at a dose of 20 mL / kg·bw once a day for 7 consecutive days.

[0100] Comparative Example 4 (Group I): The drug prepared in Comparative Example 4 was administered by gavage at a dose of 20 mL / kg·bw once a day for 7 consecutive days.

[0101] Comparative Example 5 (J Group): The drug prepared in Comparative Example 5 was administered by gavage at a dose of 20 mL / kg·bw once a day for 7 consecutive days.

[0102] (4) Observation and detection indicators:

[0103] During the experiment, the clinical status of rats in each group was observed from day 1 after inoculation with Mycoplasma eperythrocytes throughout the entire experimental period. Weight changes in rats were recorded before inoculation with Mycoplasma eperythrocyte-infected pig blood samples, 5 days after inoculation, 7 days after drug administration, and 14 days after the end of drug administration. Complete blood count and Mycoplasma eperythrocyte infection rate in erythrocytes were measured 14 days after drug withdrawal.

[0104] 3. Experimental Results:

[0105] (1) Clinical symptoms

[0106] Throughout the experiment, the animals in the control group (Group A) were healthy, had normal diets, and showed no adverse clinical symptoms. Five days after inoculation, compared with the control group, rats in all groups exhibited clinical symptoms such as lethargy, huddling together, yellow urine, and pale mucous membranes, indicating successful model establishment. In the treatment group C (Example 1), after 7 days of drug administration and 14 days of observation, the clinical symptoms of eperythrozoonosis in rats disappeared, and they returned to normal. In the other treatment groups, some or all animals still had symptoms such as lethargy and pale mucous membranes that were not completely relieved.

[0107] (2) Weight measurement

[0108] The statistical results of weight changes in rats in each group before inoculation with porcine blood samples infected with Mycoplasma suis, 5 days after inoculation, 7 days after administration, and 14 days after the end of administration are shown in Table 2-3.

[0109] Table 2. Weight changes (g) of male rats in different groups

[0110]

[0111] Note: Data with the same letter in the same column indicates no significant difference (P>0.05); data with different letters in the same column indicates significant difference (P<0.05). The same applies below.

[0112] Table 3. Weight changes (g) of female rats in different groups

[0113]

[0114] Tables 2 and 3 show that the body weight of rats in each group was uniform before inoculation, with no significant difference. After inoculation with hemotropic mycoplasma, all groups of rats showed a significant decrease in body weight compared to the control group. Except for the control group and the model group, the other groups were treated with different drugs. After 7 days of drug administration, the body weight of rats in group B (model group) was lower, while the body weight of rats in groups C to J recovered to varying degrees. Among them, the body weight of rats in group C increased more rapidly, showing a significant difference compared with the other treatment groups (P < 0.05). After 14 days of observation after drug withdrawal, the body weight of rats in group C basically returned to normal levels, with no significant difference compared with the control group (P > 0.05); the body weight of rats in groups D and E was significantly better than that of rats in groups F to J; the body weight of animals in the model group remained lower, showing a significant difference compared with the other groups (P < 0.05).

[0115] (3) Blood routine test:

[0116] After 14 days of observation, blood samples were collected from rats in each group for routine blood tests. The measured parameters were red blood cell count (RBC) and hemoglobin (HGB), and the results are shown in Table 4.

[0117] Table 4. Results of routine blood tests in rats of each group

[0118]

[0119] Table 4 shows that after the administration was completed and observed for 14 days, compared with the blank group (group A), the number of red blood cells and the content of hemoglobin in rats in each group were significantly reduced (P < 0.05). Mycoplasma suis infection caused a decrease in the number of red blood cells and the content of hemoglobin in rats. After treatment, the RBC and HGB results of rats in group C were basically similar to those in group A, with no significant difference (P > 0.05), and were significantly higher than those in groups F to J.

[0120] (4) Erythrocyte erythrozoon infection rate

[0121] On the first day of drug administration, blood samples were taken from 10 fields of view of each rat in each group for microscopic examination. The average number of red blood cells in each field of view was recorded, and the infection rate of erythropoietin was calculated (number of red blood cells infected with erythropoietin in a single field of view / number of red blood cells * 100%). Within 7 days of drug administration and 14 days after the end of drug administration, blood samples were collected from the tail vein of each group of rats at regular intervals every day for microscopic examination to observe the changes in the infection rate of erythropoietin. The results are shown in Table 5.

[0122] Table 5. Results of Detection of Erythrocyte and Mycoplasma Infection Rate in Each Group of Rats

[0123]

[0124] Table 5 shows that in group B (model group), without drug treatment, the erythrocyte infection rate and self-healing rate were very low, and the erythrocyte infection rate remained high after 7 days of drug administration and 14 days of observation. After treatment, the infection rate in each drug-treated group decreased significantly. In group C, the erythrocytes of the experimental animals were completely cleared of hemotropic mycoplasmosis, and the erythrocyte hemotropic mycoplasmosis infection rate was 0 after 14 days of observation. The results indicate that the drug for preventing and treating hemotropic mycoplasmosis in animals according to this invention has a good therapeutic effect on porcine hemotropic mycoplasmosis, and the therapeutic effect is significantly better than that prepared in comparative examples 1-5. This demonstrates that the combination of *Corydalis yanhusuo*, *Lysimachia christinae*, *Hemerocallis fulva*, *Melia azedarach* leaves, and *Gynostemma pentaphyllum* in this invention achieves a synergistic effect, greatly improving the therapeutic effect on hemotropic mycoplasmosis infection in experimental animals.

[0125] (II) Verification of the efficacy of the drug of this invention on pigs with porcine hemotropic mycoplasmosis

[0126] To verify the efficacy of the drug for preventing and treating animal eperythrozoonosis according to the present invention, efficacy verification experiments were conducted using the drugs prepared in Examples 1, 2, and 3 of the present invention as examples. Simultaneously, to compare with the drug for preventing and treating animal eperythrozoonosis according to the present invention, the following comparative experiments were also set up: Comparative Examples 1 to 5 and Comparative Example 6. The drugs in Comparative Examples 1 to 5 are basically the same as those in Example 1 of the present invention, except that the raw materials used in the traditional Chinese medicine composition in the formula are different. See Table 1 for details of the raw materials used in the traditional Chinese medicine compositions of Comparative Examples 1 to 5. The experimental drug used in Comparative Example 6 was doxycycline hydrochloride soluble powder (1000g / bag, production batch number: 2022040301), produced by Henan Muxiang Animal Pharmaceutical Co., Ltd.

[0127] 1. Laboratory animals:

[0128] The affected pig herd was provided by Xindi Livestock Farm in Yiyang County, Henan Province. Specifically, it consisted of pigs suspected of having porcine eperythrozoonosis, exhibiting obvious symptoms such as anemia, jaundice, and fever, appearing naturally on the farm approximately 3 days prior. 280 pigs of similar weight, sex, and growth stage, diagnosed with porcine eperythrozoonosis, were selected for treatment trials. Clinical symptoms of the affected pigs are shown in the attached photos. Figure 1 As shown.

[0129] 2. Test methods

[0130] (1) Experimental drugs:

[0131] The experimental drugs were the drugs prepared in Examples 1 to 3 of this invention, the drugs prepared in Comparative Examples 1 to 5, and doxycycline hydrochloride soluble powder (1000g / bag, production batch number: 2022040301) in Comparative Example 6, which were produced by Henan Muxiang Animal Pharmaceutical Co., Ltd.

[0132] (2) Experimental Groups:

[0133] Two hundred and eighty pigs diagnosed with porcine parvovirus infection were divided into ten groups (Experimental Group 1 to Experimental Group 10). Experimental Group 10 served as the untreated control group (i.e., no drug intervention was used). To reduce economic losses for the farm, Experimental Group 10 had 10 infected pigs, while the other groups had 30 infected pigs each. These pigs were isolated and kept in a controlled environment with free access to water.

[0134] (3) Dosing regimen

[0135] The dosing regimens for Experiment 1 through Experiment 10 are shown in Table 6.

[0136] Table 6. Dosing regimens for Experimental Groups 1 through 10

[0137]

[0138]

[0139] (4) Observation and detection indicators:

[0140] The body temperature of pigs in each group was measured and recorded using the conventional rectal method 1 day before the experiment, 7 days after medication, and 14 days after observation. The infection rate of erythropoietin in sick pigs in each group before and after medication was measured, and the efficacy was evaluated.

[0141] 3. Experimental Results

[0142] (1) Changes in body temperature

[0143] The body temperature of pigs in each group was measured and recorded using the conventional rectal method 1 day before the experiment, 7 days after medication, and 14 days after observation. The results are shown in Table 7.

[0144] Table 7 shows the body temperature detection results of the diseased pigs in each experimental group.

[0145]

[0146] Table 7 shows that the body temperature of animals in all groups was high before drug administration, with no significant difference (P>0.05). In the untreated control group, the body temperature of the experimental animals remained high after the experiment. In the drug-treated groups, the body temperature decreased after 7 days of drug administration. The temperature decrease in experimental groups 1-3 was significantly different from that in experimental groups 4-8 (P<0.05). Animals in experimental group 1 recovered to normal body temperature, showing the best therapeutic effect, comparable to that of experimental group 9. Furthermore, after 14 days of observation, the body temperature of animals in experimental groups 1-3 did not rebound, indicating a good therapeutic effect.

[0147] (2) Erythrocyte eperythrozoon infection rate

[0148] Blood samples were collected from the ear vein in the untreated control group, the treated group 1 day before medication, 7 days after medication, and 14 days after the end of observation. The blood samples were smeared, stained with Wright's stain, and examined under a microscope (oil immersion, 1000×) for 20 fields of view. The total number of red blood cells and the number of red blood cells infected with mycoplasma were recorded. The infection rate of mycoplasma erythrocytes was calculated. The results are shown in Table 8.

[0149] Erythrocyte eperythrozoon infection rate = (Number of erythrocytes infected with eperythrozoon / Total number of erythrocytes) × 100%

[0150] Table 8 shows the results of the detection of erythrocyte-epidermosome infection rate in diseased pigs in each experimental group.

[0151]

[0152] Table 8 shows that in experimental groups 10, without drug treatment, the infection rate of erythropoietin in the experimental animals showed an increasing trend. After drug treatment, the infection rate of erythropoietin in the experimental animals decreased significantly, and the number of parasites decreased. Specifically, the infection rate of erythropoietin in experimental groups 1-3 was significantly lower than that in experimental groups 4-8 after 7 days of treatment and 14 days of observation. Furthermore, experimental group 1 showed the best treatment effect; after 7 days of treatment, the infection rate of erythropoietin in the experimental animals decreased to approximately 5.3%, and after 14 days of observation following the end of treatment, the infection rate of erythropoietin in the experimental animals decreased to 0%, with no recurrence, comparable to the treatment effect of doxycycline hydrochloride in experimental group 9. This indicates that the erythropoietin in the experimental animals was completely eradicated without recurrence. The above results demonstrate that the present invention, by combining *Corydalis yanhusuo*, *Lysimachia christinae*, *Hemerocallis fulva*, *Melia azedarach* leaves, and *Gynostemma pentaphyllum*, achieves a synergistic effect, greatly improving the treatment effect on porcine erythropoietinosis.

[0153] (3) Clinical efficacy observation and efficacy evaluation criteria:

[0154] Observation period: Observation begins at the start of the first dose of medication and continues for 14 days after discontinuation of medication. After 14 days of observation, blood is collected from the ear vein, smears are prepared, Wright staining is performed, and microscopic examination is conducted. The total number of red blood cells and the number of red blood cells infected with Mycoplasma erythrozoon are recorded, and the Mycoplasma erythrozoon infection rate is calculated.

[0155] The efficacy evaluation criteria are as follows:

[0156] Cure: The pigs' body temperature returned to normal after medication, and the infection rate of hemozoonosis under microscopic examination was 0.

[0157] Effective: Pigs' body temperature returned to normal after medication, and microscopic examination showed that the infection rate of hemozoonosis was less than 10%.

[0158] Ineffective: Pigs' body temperature did not return to normal after medication, and microscopic examination showed an infection rate of more than 10% for Hemozoella epidermosome.

[0159] Cure rate = (Number of cured animals / Number of animals in the group) × 100%.

[0160] Effective rate = (number of cured animals + number of effective animals) / number of animals in the group × 100%.

[0161] Inefficiency = Number of invalid animals / Number of animals in the group × 100%.

[0162] The statistical results of the treatment effects of each experimental group are shown in Table 9.

[0163] Table 9 shows the statistical results of the treatment effects in each experimental group.

[0164]

[0165] Table 9 shows that the clinical symptoms of the untreated control group animals did not improve significantly, and symptoms such as fever remained severe. The efficacy rates of experimental groups 1-3 were high, exceeding 90%, comparable to the control group treated with doxycycline hydrochloride soluble form. Experimental groups 4-8 showed some therapeutic effect on porcine eperythrozoonosis, but the effect was inferior to that of experimental groups 1-3. These results further confirm that the combination of *Corydalis yanhusuo*, *Lysimachia christinae*, *Hemerocallis fulva*, *Melia azedarach* leaves, and *Gynostemma pentaphyllum* in this invention has a synergistic effect, providing good prevention and treatment of eperythrozoonosis, significantly improving the clinical symptoms of porcine eperythrozoonosis, increasing the cure rate of affected pigs, and effectively reducing breeding losses.

[0166] In summary, the traditional Chinese medicine composition of this invention has good preventive and therapeutic effects on porcine eperythrozoonosis, significantly improving the clinical symptoms of the disease, increasing the cure rate of affected pigs, and effectively reducing breeding losses. Furthermore, this traditional Chinese medicine composition fills the gap in existing national veterinary drug products for porcine eperythrozoonosis, reducing the clinical use of antibiotics for this disease, lowering drug resistance and veterinary drug residues, and ensuring food safety and human health. This invention can be mixed with feed or drinking water, is convenient to use, suitable for large-scale pig farms, and has broad development potential and a promising market prospect.

[0167] In summary, the above description is merely a preferred embodiment of the present invention and is not intended to limit the invention in any other way. Any person skilled in the art may use the above technical content as inspiration to make changes or modifications. These are equivalent embodiments with variations. However, any simple modifications, equivalent changes, and modifications made to the above embodiments based on the technical essence of the present invention without departing from the inventive concept are still within the protection scope of the claims of the present invention.

Claims

1. A traditional Chinese medicine composition for preventing and treating hemotropic mycoplasmosis in animals, characterized in that, The raw materials are composed of the following parts by weight: 25-35 parts of Corydalis yanhusuo, 12-25 parts of Lysimachia christinae, 12-25 parts of Hemerocallis fulva, 10-20 parts of Melia toosendan leaves, and 8-16 parts of Gynostemma pentaphyllum. The raw materials are derived from Chinese medicinal materials or Chinese medicinal extracts equivalent to the amount of the above-mentioned raw Chinese medicinal materials.

2. The traditional Chinese medicine composition according to claim 1, characterized in that, It is made from the following raw materials in parts by weight: 28-31 parts of Corydalis yanhusuo, 18-22 parts of Lysimachia christinae, 18-22 parts of Hemerocallis fulva, 14-17 parts of Melia toosendan leaves, and 10-12 parts of Gynostemma pentaphyllum.

3. The method for preparing the traditional Chinese medicine composition according to claim 1 or 2, characterized in that, The herbs Corydalis yanhusuo, Lysimachia christinae, Hemerocallis fulva, Melia toosendan leaves, and Gynostemma pentaphyllum were mixed in a certain proportion and then extracted with a solvent. After extraction, the mixture was filtered and the filtrate was collected to obtain the traditional Chinese medicine composition.

4. The preparation method according to claim 3, characterized in that, The solvent is a water or ethanol solution.

5. The preparation method according to claim 4, characterized in that, The extraction method is flash extraction.

6. The preparation method according to claim 5, characterized in that, The flash extraction voltage is 150V to 200V, the extraction speed is 7000r / min to 9000r / min, and the extraction time is 10 to 20min.

7. The use of the traditional Chinese medicine composition according to claim 1 or 2 in the preparation of a medicament for the prevention and / or treatment of hemotropic mycoplasmosis in animals.

8. A drug for the prevention and / or treatment of hemotropic mycoplasmosis in animals, characterized in that, The drug is made from the traditional Chinese medicine composition as described in claim 1 or 2 and pharmaceutically acceptable excipients.

9. The medicament according to claim 8, characterized in that, The drug is an oral dosage form.

10. The medicament according to claim 9, characterized in that, The drug is an oral liquid, granule, tablet, or powder.

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