Fermented Chinese medicine preparation for preventing and treating African swine fever and its preparation method and application

Abstract

The present invention provides a fermented Chinese medicine preparation that can effectively prevent or treat African swine fever, and relates to the technical field of Chinese medicine pharmaceutical manufacturing. The Chinese medicine preparation is mainly composed of 17 Chinese medicines, including wild grass incense, astragalus, echinacea, cooked rehmannia polysaccharide, tangerine peel, camellia, salvia miltiorrhiza, Poria, Atractylodes macrocephala, Platycodon grandiflorum, Cinnabarinia, Herba Cyperi, jujube, licorice, Acanthopanax senticosus, Ligustrum lucidum, and bilberry; the present invention uses the principle of Chinese medicine dialectical implementation, and the components complement each other. The Chinese medicine composition of the present invention has the effects of clearing away heat and detoxicating, and invigorating qi, and can achieve the purpose of strengthening the body without leaving evil, removing evil without hurting the body, and keeping the body healthy and the evil not being able to interfere. The fermented Chinese medicine preparation of the present invention can effectively inhibit the adsorption, internalization and replication stages of African swine fever virus on cells, fundamentally enhance the body's resistance to viruses such as African swine fever, reduce the incidence of the disease, conform to the concept of "preventive treatment" of Chinese medicine, and has significant preventive and therapeutic effects on African swine fever clinically.

Description

Technical Field

[0001] The present invention relates to the technical field of traditional Chinese medicine pharmaceutical preparation, and in particular to a fermented traditional Chinese medicine preparation for preventing and treating African swine fever, and a preparation method and application thereof. Background Art

[0002] African swine fever (ASF) is an acute, severe, highly contagious disease caused by the African swine fever virus (ASFV). The mortality rate of infected pigs is as high as 100%. ASFV was first discovered in Kenya in 1921 and has spread to many countries around the world, causing huge economic losses to the pig industry. ASF disease is acute and severe, with the main symptoms being high fever, ecchymoses on the body surface, and organ bleeding. ASFV is a DNA virus with a double-stranded DNA structure. The overall structure is relatively complex and not stable enough. It is easy to mutate and can evade the body's immune system to a certain extent, making it difficult for the body to be immune.

[0003] At present, there are many difficulties in developing safe and effective vaccine products, and there is no specific vaccine, and no antibiotics and chemical drugs that can effectively prevent ASF have been found. The prevention and control of ASF still mainly rely on detection and culling. Chinese veterinary medicine uses natural medicine, which has outstanding advantages such as no residue, improving the body's immunity, and regulating non-specific immunity, which has positive significance for the prevention and control of African swine fever. Chinese herbal medicine has many functions such as clearing away heat and detoxification, nourishing the stomach and spleen, removing dampness and phlegm, replenishing qi and nourishing blood, promoting blood circulation and removing blood stasis, consolidating the foundation and nourishing the essence, strengthening the body and eliminating evil, balancing yin and yang, etc. It can be targeted according to the cause and pathogenesis of the disease, and comprehensive use can play a comprehensive adjustment and treatment role. Therefore, in the comprehensive prevention and control of African swine fever, my country's traditional Chinese herbal medicine can improve the immunity of animal bodies, alleviate symptoms, and play a good role in prevention and treatment. Summary of the invention

[0004] 1. Technical issues to be resolved

[0005] In view of the deficiencies in the prior art, the present invention provides a fermented Chinese medicine preparation for preventing and treating African swine fever, as well as a preparation method and application thereof. The fermented Chinese medicine preparation can improve the immunity of pigs, enhance resistance to viruses such as African swine fever, have significant preventive and therapeutic effects on African swine fever, and reduce the incidence of African swine fever.

[0006] (II) Technical solution

[0007] To achieve the above objectives, the present invention is implemented through the following technical solutions:

[0008] On the one hand, the present invention provides a fermented Chinese medicine preparation for preventing and treating African swine fever, the Chinese medicine composition comprising the following Chinese medicine raw materials by weight: 30-50g of wild grass fragrance, 20-50g of astragalus, 5-20g of echinacea, 5-20g of cooked rehmannia polysaccharide, 5-20g of tangerine peel, 10-30g of camellia, 10-30g of salvia miltiorrhiza, 10-30g of Poria, 10-30g of Atractylodes macrocephala, 10-30g of Platycodon grandiflorum, 10-30g of Verniciflua cinnabarifolia, 10-30g of Cyperus rotundus, 10-30g of jujube, 5-20g of licorice, 5-20g of Acanthopanax senticosus, 5-20g of Ligustrum lucidum, and 5-20g of blueberry.

[0009] Furthermore, the Chinese medicine composition includes the following Chinese medicine raw materials by weight: 35-50g of wild grass fragrance, 25-40g of astragalus, 5-15g of echinacea, 5-15g of cooked rehmannia polysaccharide, 5-15g of tangerine peel, 10-20g of camellia, 10-20g of salvia miltiorrhiza, 10-20g of Poria, 10-20g of Atractylodes macrocephala, 10-20g of Platycodon grandiflorum, 10-20g of Verniciflua cinnabarifolia, 10-20g of Cyperus rotundus, 15-25g of jujube, 5-15g of licorice, 5-15g of Acanthopanax senticosus, 5-15g of Ligustrum lucidum, and 5-15g of blueberry.

[0010] Furthermore, the Chinese medicine composition includes the following Chinese medicine raw materials by weight: 40g of wild grass fragrance, 30g of astragalus, 9g of echinacea, 9g of cooked rehmannia polysaccharide, 9g of tangerine peel, 15g of camellia, 15g of salvia miltiorrhiza, 15g of Poria, 15g of Atractylodes macrocephala, 15g of Platycodon grandiflorum, 15g of Cinnabarinum, 15g of Cyperus rotundus, 20g of jujube, 9g of licorice, 9g of Acanthopanax senticosus, 9g of Ligustrum lucidum, and 9g of Vaccinium myrtles.

[0011] Furthermore, the Chinese medicine composition also includes: 0.02-0.1g vitamin C, 0.02-0.1g vitamin A, 0.5-2g Bacillus subtilis powder, 0.5-2g lactobacillus powder, 0.2-1g bifidobacterium powder, and 0.2-1g Clostridium butyricum powder.

[0012] Furthermore, the Chinese medicine composition also includes: 0.08g vitamin C, 0.06g vitamin A, 1.2g Bacillus subtilis powder, 1g Lactobacillus powder, 0.6g Bifidobacterium powder, and 0.4g Clostridium butyricum powder.

[0013] On the other hand, the present invention provides a preparation process of a fermented Chinese medicine preparation for preventing and treating African swine fever, which is prepared according to the following steps:

[0014] (1) Weigh wild grass, astragalus, echinacea, rehmannia polysaccharide, tangerine peel, camellia, salvia miltiorrhiza, tuckahoe, atractylodes, platycodon, cinnabarinum, scutellaria baicalensis, jujube, licorice, acanthopanax senticosus, glossy privet fruit, and bilberry respectively according to the above weights, wash, dry, and crush the above raw materials, pass through a 60-mesh sieve, and mix them evenly to obtain a mixed Chinese medicine powder for later use;

[0015] (2) adding 4-6 times the weight of deionized water to the mixed Chinese medicine powder obtained in step (1) and soaking it for more than 40 minutes, boiling it over high heat and then decocting it over low heat for more than 90 minutes, filtering the residue to collect the filtrate, repeatedly extracting the residue 2 to 3 times, and mixing the filtrate collected each time to obtain a Chinese medicine extract;

[0016] (3) distilling and concentrating the Chinese medicine extract obtained in step (2) to finally obtain a concentrated Chinese medicine extract with a concentration of 1-1.2 g / mL;

[0017] (4) Weigh vitamin C, vitamin A, Bacillus subtilis powder, Lactobacillus powder, Bifidobacterium powder, and Clostridium butyricum powder respectively according to the above weights, and add them to the concentrated acid Chinese medicine extract obtained in step (3) in sequence, mix well, and place in a sealed sterile container for fermentation at 30-37° C. for more than 72 hours;

[0018] (5) After fermentation is completed, the fermentation liquid is filtered to obtain a clear solution without solid impurities, and then bottled to obtain the fermented Chinese medicine preparation.

[0019] The fermented Chinese medicine preparation is used to prepare drugs for preventing and treating African swine fever; the Chinese medicine composition is used to prepare any drug dosage form acceptable to animals, including but not limited to powders, granules, decoctions, and oral liquids.

[0020] On the other hand, the fermented Chinese medicine preparation provided by the present invention is used to prevent and treat African swine fever, and the dosage is 20mL / 50kg for oral administration of each medicine, with preventive administration for 7-14 days and therapeutic administration for more than 14 days.

[0021] Pharmacological effects of each Chinese medicinal material in the fermented Chinese medicine preparation of the present invention:

[0022] Wild grass fragrance: It tastes spicy and is cool in nature. It has the effects of dispelling wind and clearing heat, promoting qi and blood circulation, detoxifying and reducing swelling. It can be used to treat colds and fever, abdominal pain caused by food accumulation, dysentery, malaria, carbuncle, urticaria and other symptoms.

[0023] Astragalus: sweet in taste, slightly warm in nature, enters the spleen and lung meridians; has the effects of replenishing qi and strengthening the exterior, expelling toxins and pus, promoting diuresis, and promoting tissue regeneration; used for symptoms such as qi deficiency and fatigue, long-term diarrhea and prolapse of the anus, spontaneous sweating, edema, uterine prolapse, proteinuria of chronic nephritis, diabetes, and non-healing wounds; Astragalus contains effective ingredients such as carbohydrates, multiple amino acids, proteins, choline, betaine, folic acid, vitamin P, amylase, and multiple trace elements, which have the effects of enhancing immunity and anti-virus.

[0024] Echinacea: It mainly contains caffeic acid and its derivatives, alkylamide compounds, polysaccharides and glycoproteins. Compounds such as volatile oils have the main pharmacological effects of immunomodulation, anti-inflammatory, antiviral, antibacterial and antioxidant.

[0025] Rehmannia glutinosa polysaccharide: It is an extract of Rehmannia glutinosa, which has immune and tumor transplantation activities, and has the effects of strengthening the heart, lowering blood pressure, protecting the myocardium, inhibiting thrombosis and lowering blood lipids on the cardiovascular system.

[0026] Tangerine peel: It tastes spicy and bitter, is warm in nature, and belongs to the spleen and lung meridians; it has the effects of regulating qi and strengthening the spleen, resolving phlegm and drying dampness; it is used for spleen and stomach disharmony, vomiting, cough, bloating and loss of appetite, etc. Tangerine peel mainly contains effective ingredients such as flavonoids, alkaloids, limonoids, volatile oils and trace elements; it has pharmacological effects such as inhibiting gastrointestinal smooth muscle, promoting digestive juice secretion, promoting choleretic effect, relaxing bronchial smooth muscle, and resisting shock.

[0027] Camellia: It is sweet, bitter, and pungent, and cool in nature. It enters the liver and lung meridians. It has the effects of cooling blood, stopping bleeding, dispersing blood stasis, and reducing swelling. It is used for symptoms such as vomiting blood, metrorrhagia, intestinal wind, bloody dysentery, hematuria, bruises, and burns.

[0028] Salvia miltiorrhiza: bitter in taste, slightly cold in nature, enters the heart and liver meridians; has immunoregulatory effects on cytokines, antibodies, immune complexes and immune cells, and has many functions such as improving microcirculation, protecting the liver and anti-inflammation, promoting blood circulation and removing blood stasis, calming the mind, relieving pain, dilating blood vessels, and lowering blood pressure.

[0029] Poria: sweet, light, neutral in nature, enters the heart, lung, spleen, and kidney meridians; has the effects of promoting diuresis and eliminating dampness, strengthening the spleen, and calming the mind; used to treat edema, oliguria, phlegm and fluid, dizziness, palpitations, spleen deficiency, poor appetite, and restlessness; Poria mainly contains triterpenes and polysaccharide compounds, which have pharmacological effects such as diuresis, anti-inflammatory, and liver protection.

[0030] Atractylodes: bitter, sweet, and warm in nature, it enters the spleen and stomach meridians; it has the effects of strengthening the spleen and replenishing Qi, drying dampness and promoting diuresis, stopping sweating, and calming the fetus; it is often used for symptoms such as spleen deficiency, poor appetite, diarrhea, loose stools, and fetal movement disorder; its main chemical components include volatile oils, polysaccharides, and lipids, which have anti-tumor, anti-inflammatory, and digestive system regulation effects.

[0031] Platycodon grandiflorum: It is mild in nature, bitter in taste, and belongs to the lung meridian. It has the effects of clearing the lungs and removing phlegm, relieving sore throat and expelling pus, benefiting the five internal organs, replenishing qi and blood, replenishing the five labors, and nourishing qi. It is used to treat cough with a lot of phlegm, chest fullness, sore throat and hoarseness, lung carbuncle and vomiting pus. Platycodon grandiflorum contains chemical substances such as platycodon saponins, polygala acid, platycodon inulin, glucose, and phytosterol. Pharmacological studies in recent years have shown that the roots, stems, leaves, flowers, and fruits of Platycodon grandiflorum have very significant expectorant, antitussive, anti-inflammatory, anti-ulcer, sedative and analgesic, hypothermic, anti-edema, and diuretic effects. The expectorant and antitussive effects of Platycodon grandiflorum are due to the fact that it can significantly increase the secretion of respiratory mucus. Its intensity is comparable to that of ammonium chloride, and it has a significant expectorant effect.

[0032] Cinnabar Lotus: It tastes bitter, pungent, and cold in nature. It enters the heart, lung, and liver meridians. It has the effects of clearing away heat and detoxifying, regulating qi and relieving pain. It is used for carbuncles, swellings, summer heat, diarrhea, dysentery, chest and abdominal pain, toothache, sore throat, vomiting blood, snake bites and other diseases. The main pharmacological effects are analgesia and antipyretic.

[0033] Herba Cynanchum: It is pungent, bitter and cold in nature, and enters the liver and gallbladder meridians; it has the effects of clearing the liver and purging fire, improving eyesight, dispersing nodules and reducing swelling; it is used for symptoms such as red and swollen eyes, night pain in the eyes, headaches and dizziness; the main chemical components include triterpenes, sterols, flavonoids, organic acids, coumarins and other effective ingredients, which have certain pharmacological activities in antibacterial, antiviral, anti-inflammatory, immune regulation, anti-oxidation, free radical scavenging, anti-tumor, antihypertensive, hypoglycemic, and lipid-regulating.

[0034] Jujube: sweet in taste, warm in nature, and enters the spleen and stomach meridians; it has the effects of tonifying the spleen and stomach, replenishing Qi and promoting body fluid, regulating the Ying and Wei, and detoxifying; it is used to treat stomach deficiency, poor appetite, weak spleen and loose stools, insufficient Qi and blood, disharmony between the Ying and Wei, palpitations and other symptoms; jujube is rich in nutrients, including amino acids, sugars, vitamins and various trace elements, and contains a large amount of nucleotide derivatives. Jujube has a good auxiliary therapeutic effect on diseases such as tumors, hypertension, hypercholesterolemia, and has the effects of delaying aging, anti-oxidation, improving immunity, protecting the liver, etc.

[0035] Licorice: sweet in taste, neutral in nature, enters the heart, spleen, lung, and stomach meridians; has the effects of tonifying the spleen and replenishing qi, clearing away heat and detoxifying, removing phlegm and relieving cough, relieving acute pain, and harmonizing various medicines; used for spleen and stomach weakness, fatigue, palpitations and shortness of breath, cough with sputum, carbuncles, sores and other symptoms. Licorice is also good at harmonizing the properties of medicines and alleviating the toxicity of drugs.

[0036] Acanthopanax: sweet, slightly bitter, and warm in nature; it enters the spleen, lung, heart, and kidney meridians; it has the effects of invigorating qi and strengthening the spleen, nourishing the kidneys and calming the mind, and can treat spleen and lung qi deficiency syndrome, kidney deficiency and low back and knee pain syndrome, heart and spleen deficiency, insomnia, etc.

[0037] Fructus Ligustri Lucidi: It is cool in nature, sweet and bitter in taste, and enters the liver and kidney meridians; it has the effects of nourishing the liver and kidney, improving eyesight and blackening hair; it is used to treat tinnitus, deafness, dizziness, weakness of waist and knees and other symptoms caused by liver and kidney yin deficiency; Fructus Ligustri Lucidi mainly contains effective ingredients such as stearic acid, palmitic acid, linolenic acid, oleic acid, oleanolic acid and D-mannitol. Modern pharmacological studies have shown that Fructus Ligustri Lucidi has the effects of lowering blood lipids, lowering blood sugar, antioxidant activity, resisting liver damage, and enhancing immunity.

[0038] Bilberry: Bilberry fruit is sour and sweet in taste, neutral in nature, and enters the stomach and large intestine meridians; it has the effects of clearing away heat and dampness, and stopping diarrhea; it is used for enteritis, dysentery and other diseases; it contains flavonoids, triterpenes, cyclopentane ether terpenes, organic acids, phenolic acids, sugars, amino acids, etc.; it has antioxidant, antibacterial, anti-inflammatory, anti-cancer, antiviral and other pharmacological effects.

[0039] (III) Beneficial effects

[0040] The present invention provides a fermented Chinese medicine preparation that can effectively prevent or treat African swine fever. The fermented Chinese medicine preparation is mainly composed of 17 Chinese medicines such as wild grass incense, astragalus, echinacea, cooked rehmannia polysaccharide, tangerine peel, camellia, salvia miltiorrhiza, Poria, Atractylodes macrocephala, Platycodon grandiflorum, Cinnabarinia, Herba Cynoglossi, jujube, licorice, Acanthopanax senticosus, Ligustrum lucidum, and bilberry. The present invention uses the principle of TCM dialectical implementation. In the prescription, wild grass incense dispels wind and heat, promotes qi and blood circulation, detoxifies and reduces swelling, and astragalus replenishes qi and consolidates the exterior, promotes diuresis and removes toxins. They are all monarch drugs. Atractylodes macrocephala can help astragalus strengthen the spleen and lungs, replenish qi and qi, and raise yang and lift the sunken, so as to nourish the source of qi and blood biochemical, stabilize the innate foundation, and strengthen the body and eliminate evil, so as to achieve the merit of "positive qi is inside, and evil cannot interfere". It replenishes qi and nourishes blood, and has the effect of removing rot and promoting muscle growth, and astringing sores, so it is matched with cooked rehmannia, and these two are ministerial drugs. Atractylodes macrocephala replenishes qi and strengthens the spleen to transport and transform water and dampness. When combined with Poria cocos, it can not only strengthen the spleen and control water, but also play the role of transporting body fluid to the four parts of the body, and is good at treating plague in all seasons. Rehmannia glutinosa combined with Ligustrum lucidum can replenish essence and fill the marrow, replenish blood and nourish yin. Acanthopanax senticosus and dried orange peel combined with Poria cocos can dry dampness and promote qi, strengthen the spleen and stomach. Pathogenic pathogens enter the body and cause blood heat to run wild, so it is combined with Salvia miltiorrhiza, which is cold in nature and cools blood, and can also activate blood circulation, and has the function of clearing blood stasis and heat to eliminate carbuncle and swelling. Vermilion lotus, camellia, and big head grass clear heat and detoxify, stop bleeding, Echinacea purpurea and bilberry are combined to play the role of detoxification, clearing heat and promoting diuresis, licorice combined with Platycodon grandiflorum can relieve throat and cough, jujube regulates the spleen and stomach internally, and harmonizes the Ying and Wei externally and can also harmonize all the medicines, which is the role of adjuvant. When used together, all the medicines can achieve the functions of clearing heat and detoxification, replenishing the middle and replenishing qi, strengthening the body, strengthening the body, avoiding plague and expelling epidemics; the monarch, minister, adjuvant and envoy complement each other and treat according to syndrome differentiation. The Chinese medicine composition of the present invention has the effects of clearing away heat, detoxifying, and invigorating qi, and can achieve the purpose of strengthening the body without leaving evil, removing evil without hurting the body, and keeping the body healthy and preventing evil from interfering. The fermented Chinese medicine preparation of the present invention can effectively inhibit the adsorption, internalization and replication stages of African swine fever virus on cells in vitro, fundamentally enhance the body's resistance to viruses such as African swine fever, and reduce the incidence of the disease, which is in line with the concept of "preventive treatment" of traditional Chinese medicine, and has significant preventive and therapeutic effects on African swine fever clinically, helping the development of the pig farming industry under the normalization of African swine fever. BRIEF DESCRIPTION OF THE DRAWINGS

[0041] Figure 1 The effect of different drugs on the viral activity in PAM cells; 1 represents the ribavirin + virus group; 2 represents the virus group; 3 represents the drug 1 + virus group; 4 represents the drug 2 + virus group; 5 represents the drug 3 + virus group; 6 represents the drug 4 + virus group; 7 represents the drug 5 + virus group; 8 represents the drug 6 + virus group.

[0042] Figure 2 A is the effect of different drugs on virus adsorption in PAM cells; Figure 2 B shows the effects of different drugs on viral penetration in PAM cells; Figure 2C (QPCR test) and 2D (HAD50 assay) are the effects of different drugs on viral replication in PAM cells. DETAILED DESCRIPTION

[0043] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution in the embodiments of the present invention will be clearly and completely described below in combination with the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0044] Example 1

[0045] A fermented Chinese medicine preparation for preventing and treating African swine fever, the fermented Chinese medicine preparation comprising the following raw materials by weight: 30 grams of wild grass incense, 20 grams of astragalus, 5 grams of echinacea, 5 grams of cooked rehmannia polysaccharide, 5 grams of tangerine peel, 10 grams of camellia, 10 grams of salvia miltiorrhiza, 10 grams of poria, 10 grams of atractylodes, 10 grams of platycodon, 10 grams of cinnabar lotus, 10 grams of big head grass, 10 grams of jujube, 5 grams of liquorice, 5 grams of acanthopanax, 5 grams of privet fruit, 5 grams of bilberry, 0.02 grams of vitamin C, 0.02 grams of vitamin A, 0.8 grams of bacillus subtilis powder, 0.6 grams of lactobacillus powder, 0.4 grams of bifidobacterium powder, and 0.2 grams of Clostridium butyricum powder.

[0046] The fermented Chinese medicine preparation is prepared according to the following steps:

[0047] (1) Weigh astragalus, echinacea, rehmannia polysaccharide, tangerine peel, camellia, salvia miltiorrhiza, tuckahoe, atractylodes, platycodon, cinnabarinum, scutellaria baicalensis, jujube, liquorice, acanthopanax senticosus, glossy privet fruit, and bilberry respectively according to the above weights, wash, dry, and crush the above raw materials, pass through a 60-mesh sieve, and mix them evenly to obtain a mixed Chinese medicine powder for later use;

[0048] (2) adding 4-6 times the weight of deionized water to the mixed Chinese medicine powder obtained in step (1) and soaking it for more than 40 minutes, boiling it over high heat and then decocting it over low heat for more than 90 minutes, filtering the residue to collect the filtrate, repeatedly extracting the residue 2 to 3 times, and mixing the filtrate collected each time to obtain a Chinese medicine extract;

[0049] (3) distilling and concentrating the Chinese medicine extract obtained in step (2) to finally obtain a concentrated Chinese medicine extract with a concentration of 1-1.2 g / mL;

[0050] (4) Weigh vitamin C, vitamin A, Bacillus subtilis powder, Lactobacillus powder, Bifidobacterium powder, and Clostridium butyricum powder respectively according to the above weights, and add them to the concentrated acid Chinese medicine extract obtained in step (3) in sequence, mix well, and place in a sealed sterile container for fermentation at 30-37° C. for more than 72 hours;

[0051] (5) After the fermentation is completed, the fermentation liquid is filtered to obtain a clear solution without solid impurities, and then spray-dried to obtain a powder or granular solution to obtain the fermented Chinese medicine preparation.

[0052] Example 2

[0053] The difference between this embodiment and embodiment 1 is that the weight of each raw material is:

[0054] 50g of wild grass fragrance, 50g of astragalus, 20g of echinacea, 20g of cooked rehmannia polysaccharide, 20g of tangerine peel, 30g of camellia, 30g of salvia miltiorrhiza, 30g of poria, 30g of atractylodes, 30g of platycodon, 30g of cinnabar lotus, 30g of big head grass, 30g of jujube, 20g of licorice, 20g of acanthopanax, 20g of glossy privet, 20g of bilberry, 0.1g of vitamin C, 0.1g of vitamin A, 2g of Bacillus subtilis powder, 2g of lactobacillus powder, 1g of bifidobacterium powder, and 1g of Clostridium butyricum powder.

[0055] Example 3

[0056] The difference between this embodiment and embodiment 1 is that the weight of each raw material is:

[0057] 35g of wild grass fragrance, 25g of astragalus, 7g of echinacea, 7g of cooked rehmannia polysaccharide, 7g of tangerine peel, 12g of camellia, 12g of salvia miltiorrhiza, 12g of poria, 12g of atractylodes, 12g of platycodon, 12g of cinnabar lotus, 12g of big head grass, 15g of jujube, 7g of licorice, 7g of acanthopanax, 7g of glossy privet, 7g of bilberry, 0.06g of vitamin C, 0.04g of vitamin A, 1g of Bacillus subtilis powder, 1g of lactobacillus powder, 0.6g of bifidobacterium powder, and 0.4g of Clostridium butyricum powder.

[0058] Example 4

[0059] The difference between this embodiment and embodiment 1 is that the weight of each raw material is:

[0060] 50g of wild grass fragrance, 40g of astragalus, 15g of echinacea, 15g of cooked rehmannia polysaccharide, 15g of tangerine peel, 20g of camellia, 20g of salvia miltiorrhiza, 20g of poria, 20g of atractylodes, 20g of platycodon, 20g of cinnabar lotus, 20g of big head grass, 25g of jujube, 15g of licorice, 15g of acanthopanax, 15g of glossy privet, 15g of bilberry, 0.08g of vitamin C, 0.06g of vitamin A, 1.2g of Bacillus subtilis powder, 0.8g of lactobacillus powder, 0.5g of bifidobacterium powder, and 0.5g of Clostridium butyricum powder.

[0061] Example 5

[0062] The difference between this embodiment and embodiment 1 is that the weight of each raw material is:

[0063] Wild grass fragrance 40g, astragalus 35g, echinacea 12g, cooked rehmannia polysaccharide 12g, tangerine peel 10g, camellia 18g, salvia miltiorrhiza 16g, Poria 14g, Atractylodes 14g, Platycodon 18g, Verniciflua 12g, Herba Cynanchifoliae 14g, jujube 16g, licorice 10g, Acanthopanax 8g, Ligustrum lucidum 8g, Bacillus vaccinium 8g, vitamin C 0.07g, vitamin A 0.05g, Bacillus subtilis powder 1g, Lactobacillus powder 1g, Bifidobacterium powder 0.6g, Clostridium butyricum powder 0.4g.

[0064] Example 6

[0065] The difference between this embodiment and embodiment 1 is that the weight of each raw material is:

[0066] Wild grass fragrance 45g, astragalus 42g, echinacea 18g, cooked rehmannia polysaccharide 16g, tangerine peel 16g, camellia 22g, salvia miltiorrhiza 22g, Poria 24g, Atractylodes macrocephala 26g, Platycodon grandiflorum 26g, Verniciflua 18g, Cyperus rotundus 18g, jujube 18g, licorice 12g, Acanthopanax 14g, Ligustrum lucidum 14g, Bacillus 16g, vitamin C 0.08g, vitamin A 0.06g, Bacillus subtilis powder 1.2g, Lactobacillus powder 0.8g, Bifidobacterium powder 0.6g, Clostridium butyricum powder 0.4g.

[0067] Example 7

[0068] The difference between this embodiment and embodiment 1 is that the weight of each raw material is:

[0069] Wild grass fragrance 40g, astragalus 30g, echinacea 9g, cooked rehmannia polysaccharide 9g, tangerine peel 9g, camellia 15g, salvia miltiorrhiza 15g, Poria 15g, Atractylodes macrocephala 15g, Platycodon grandiflorum 15g, Verniciflua 15g, Herba Cynoglossi 15g, jujube 20g, licorice 9g, Acanthopanax 9g, Ligustrum lucidum 9g, bilberry 9g, vitamin C 0.08g, vitamin A 0.06g, Bacillus subtilis powder 1.2g, Lactobacillus powder 1g, Bifidobacterium powder 0.6g, Clostridium butyricum powder 0.4g.

[0070] Test Example 1

[0071] In vitro virus inhibition assay

[0072] 1 Materials and methods

[0073] 1.1 Experimental materials and drugs

[0074] 1.1.1 Trial Drugs

[0075] Drug 1: 40g of wild grass fragrance, 30g of astragalus, 9g of echinacea, 9g of cooked rehmannia polysaccharide, 9g of tangerine peel, 15g of camellia, 15g of salvia miltiorrhiza, 15g of Poria, 15g of Atractylodes macrocephala, 15g of Platycodon grandiflorum, 15g of Cinnabar Lotus, 15g of Herba Cynoglossi, 20g of jujube, 9g of licorice, 9g of Acanthopanax senticosus, 9g of Ligustrum lucidum, 9g of bilberry, 0.08g of vitamin C, 0.06g of vitamin A, 1.2g of Bacillus subtilis powder, 1g of Lactobacillus powder, 0.6g of Bifidobacterium powder, and 0.4g of Clostridium butyricum powder; the preparation method is prepared according to the method described in Example 1.

[0076] Drug 2: 40g of wild grass fragrance, 30g of astragalus, 9g of echinacea, 9g of cooked rehmannia polysaccharide, 9g of tangerine peel, 15g of camellia, 15g of salvia miltiorrhiza, 15g of poria, 15g of atractylodes, 15g of platycodon, 15g of cinnabar lotus, 15g of big head grass, 20g of jujube, 9g of liquorice, 9g of acanthopanax senticosus, 9g of glossy privet, 9g of bilberry, 0.08g of vitamin C, and 0.06g of vitamin A; the preparation method is prepared according to the method described in Example 1, without the step of probiotic fermentation.

[0077] Drug 3: 40g of wild grass fragrance, 30g of astragalus, 9g of echinacea, 9g of cooked rehmannia polysaccharide, 9g of tangerine peel, 15g of Poria, 15g of Atractylodes macrocephala, 15g of Platycodon grandiflorum, 20g of jujube, 9g of licorice, 9g of Acanthopanax senticosus, 9g of Ligustrum lucidum, 9g of bilberry, 0.08g of vitamin C, 0.06g of vitamin A, 1.2g of Bacillus subtilis powder, 1g of Lactobacillus powder, 0.6g of Bifidobacterium powder, and 0.4g of Clostridium butyricum powder; the preparation method is prepared according to the method described in Example 1.

[0078] Drug 4: 40g of wild grass fragrance, 30g of astragalus, 9g of echinacea, 9g of cooked rehmannia polysaccharide, 15g of camellia, 15g of salvia miltiorrhiza, 15g of Poria, 15g of Atractylodes macrocephala, 15g of cinnabar lotus, 15g of scutellaria baicalensis, 9g of Ligustrum lucidum, 9g of bilberry, 0.08g of vitamin C, 0.06g of vitamin A, 1.2g of Bacillus subtilis powder, 1g of Lactobacillus powder, 0.6g of Bifidobacterium powder, and 0.4g of Clostridium butyricum powder; the preparation method is prepared according to the method described in Example 1.

[0079] Drug 5: 40g of wild grass fragrance, 30g of astragalus, 9g of echinacea, 9g of tangerine peel, 15g of camellia, 15g of salvia miltiorrhiza, 15g of platycodon, 15g of cinnabar lotus, 15g of big head grass, 20g of jujube, 9g of licorice, 9g of acanthopanax, 9g of bilberry, 0.08g of vitamin C, 0.06g of vitamin A, 1.2g of Bacillus subtilis powder, 1g of lactobacillus powder, 0.6g of bifidobacterium powder, and 0.4g of Clostridium butyricum powder; the preparation method is prepared according to the method described in Example 1.

[0080] Drug 6: 9g of Rehmannia glutinosa polysaccharide, 9g of dried tangerine peel, 15g of Camellia japonica, 15g of Salvia miltiorrhiza, 15g of Poria, 15g of Atractylodes macrocephala, 15g of Platycodon grandiflorum, 15g of Scutellaria cinnabarinus, 15g of Herba Cynomorii truncatulae, 20g of Jujube, 9g of Licorice, 9g of Acanthopanax senticosus, 9g of Fructus Ligustri Lucidi, 0.08g of Vitamin C, 0.06g of Vitamin A, 1.2g of Bacillus subtilis powder, 1g of Lactobacillus powder, 0.6g of Bifidobacterium powder, and 0.4g of Clostridium butyricum powder; the preparation method was prepared according to the method described in Example 1.

[0081] 1.1.2 Test materials

[0082] The cells used for the test were primary porcine alveolar macrophages (PAMs). The African swine fever virus strain: ASFV / China / GZ201801 was isolated and identified by the National African Swine Fever Regional Laboratory (Guangzhou). The virus concentration was 10 6 HAD 50 / mL

[0083] 1.2 Test methods

[0084] 1.2.1 WB determination of the effect of fermented Chinese medicine preparations 1-6 on viral activity in PAM cells

[0085] PAM cells were inoculated in a 48-well plate, and after the cells adhered to the wall, the cells were inoculated with a 100-fold diluted ASFV virus solution at 250 μL / well. After culturing in a 37°C, 5% CO2 incubator for 2 hours, the fermented Chinese medicine preparation prepared according to drugs 1-6 was diluted to 1:1000 with 10% FBS RPMI Medium 1640 culture medium, and an equal volume of 1:1 with the virus solution was added to the cell plate. After culturing in a 37°C, 5% CO2 incubator for 48 hours, the cells were collected 24 hours later for Western Blot to determine the content of viral protein ASFV-p72 in the cells.

[0086] 1.2.2 Virus adsorption test

[0087] PAM cells were inoculated in a 24-well plate. After the cells adhered to the wall, the culture medium was discarded. The fermented Chinese medicine preparation prepared according to drugs 1-6 was diluted to 1:1000 with a 100-fold diluted ASFV virus solution. At the same time, a cell negative control group, a virus group and a ribavirin positive drug group were set up, 500 μL / well, and the 24-well plate was placed in a 4°C refrigerator for 2 hours. The plate was washed three times with serum-free RPMI Medium 1640 culture medium, and serum-free culture medium was added again. After culturing in a 37°C, 5% CO2 incubator for 48 hours, the samples were collected for QPCR experiments.

[0088] 1.2.3 Virus penetration test

[0089] PAM cells were inoculated in a 24-well plate, and the culture medium was discarded after the cells adhered to the wall. The cells were inoculated with a 100-fold diluted ASFV virus solution at 500 μL / well. After incubation in a 4°C refrigerator for 1 hour, the cells were washed 3 times with serum-free RPMI Medium 1640 medium. The fermented Chinese medicine preparations prepared according to drugs 1-6 were diluted to 1:1000 with 10% FBS RPMI Medium 1640 medium. At the same time, a cell negative control group, a virus group, and a ribavirin positive drug group were set up. The cells were cultured in a 37°C, 5% CO2 incubator for 1 hour, and then washed 3 times with serum-free RPMI Medium 1640 medium. 10% FBS RPMI Medium 1640 medium was added again. After culture in a 37°C, 5% CO2 incubator for 48 hours, the samples were collected for QPCR experiments.

[0090] 1.2.4 Virus replication assay

[0091] PAM cells were inoculated in a 24-well plate, and the culture medium was discarded after the cells adhered to the wall. The cells were inoculated with a 50-fold diluted ASFV virus solution at 500 μL / well, and cultured in a 37°C, 5% CO2 incubator for 2 h. The fermented Chinese medicine preparation prepared according to drugs 1-6 was diluted to 1:1000 with 10% FBS RPMI Medium 1640 culture medium, and added to the cell plate at an equal volume of 1:1 with the virus solution. After culturing in a 37°C, 5% CO2 incubator for 48 h, the virus supernatant was collected for HAD50 determination and QPCR test.

[0092] 3 Results

[0093] Figure 1 These are the results of the effects of different drugs on the viral activity in PAM cells. The results show that the above six types of fermented Chinese medicine preparations can inhibit the content of African swine fever virus protein ASFV-p72 in cells to varying degrees.

[0094] Figure 2 A is the effect of different drugs on virus adsorption in PAM cells. The results show that the above six fermented Chinese medicine preparations can inhibit the adsorption of viruses by PAM cells to varying degrees. Among them, drug 1, i.e., the fermented Chinese medicine preparation of the present invention, has the most obvious inhibitory effect on virus adsorption, and the difference is significant.

[0095] Figure 2 B is the effect of different drugs on virus penetration in PAM cells. The results show that the above six fermented Chinese medicine preparations can significantly inhibit virus penetration into cells and inhibit virus internalization. Among them, the fermented Chinese medicine preparation of the present invention has the most obvious inhibitory effect on virus penetration, and the difference is significant.

[0096] Figure 2 C and 2D are the results of the effects of different drugs on viral replication in PAM cells. The HAD50 test and QPCR test show that the fermented Chinese medicine preparation of the present invention can inhibit more than 90% of viral replication.

[0097] The above results can illustrate that the fermented Chinese medicine preparation of the present invention has a significant inhibitory effect on African swine fever virus, can effectively inhibit the adsorption, internalization and replication stages of the virus on cells, and can achieve the preventive effect on African swine fever virus from the root. The fermented Chinese medicine preparation of the present invention uses the principle of Chinese medicine dialectical treatment, and the various components complement each other and are indispensable. The fermented Chinese medicine preparation of the present invention has the effects of clearing away heat and detoxifying, and invigorating qi, and can achieve the purpose of strengthening the body without leaving evil, eliminating evil without hurting the body, and keeping the body healthy and the evil cannot interfere. Fundamentally enhance the body's resistance to viruses such as African swine fever, reduce the incidence of the disease, and conform to the idea of ​​"preventive treatment" in Chinese medicine.

[0098] Test Example 2

[0099] Clinical effect test of the fermented Chinese medicine preparation of the present invention on preventing and treating African swine fever:

[0100] Randomly select 187 Duroc pigs with similar growth indicators and age from a certain farm, including 73 boars weighing 360-400kg; 114 sows weighing 320-360kg. Among them, 141 (56 boars and 85 sows) were negative for African swine fever after test strips and fluorescence quantitative PCR tests, that is, they were not sick, and their appearance, fur color, feeding, drinking water, etc. were all normal; 46 (17 boars and 29 sows) were positive for African swine fever after test strips and fluorescence quantitative PCR tests, and were diagnosed as sick pigs with African swine fever by veterinarians. The symptoms of sick pigs: high fever of 40-41℃, high spirits, red ears, sharp drop in feed intake, many red patches on the skin of the buttocks and hind limbs, some sick pigs like to lie on the ground, have difficulty moving, cannot stand on their hind limbs, and discharge loose stools mixed with blood.

[0101] From 46 African swine fever-positive Duclos pigs, 30 pigs with similar body conditions were selected for separate breeding and isolation, including 10 boars and 20 sows, marked as the blank group; 120 pigs were selected from 141 African swine fever-negative Duclos pigs (excluding 21 pigs with poor body conditions) and randomly divided into 3 groups, 30 in each group, each group including 10 boars and 20 sows, namely low-dose group, medium-dose group and high-dose group; wherein, during the blank group test, no drugs were used except basic feed nutrition; the low-dose group, medium-dose group and high-dose group were intragastrically administered with the fermented Chinese medicine preparation of Example 7 of the present invention, with doses of 1.25 g / kg.bt, 1.0 g / kg.bt and 0.75 g / kg.bt, respectively. During the test, the feeding conditions and environment of the pigs in each group remained consistent, and they were free to eat and drink water. The test period was 30 days. At the end of the experiment, the whole group was tested for African swine fever. During the trial, feed consumption and health status were counted, feed intake was calculated, and the behavior and health status of the pigs were observed and recorded. After consultation with the pig farm veterinarian, the clinical effect judgment criteria were:

[0102] 1) Effective: normal spirit, feeding, and body temperature, no skin redness, no respiratory symptoms, no hematuria, no obvious eye mucus and tear stains, moist nose, normal dry and loose stools, and no positive detection of African swine fever;

[0103] 2) Significant effect: The animal is basically in good spirits, has soft and dry feces, eats about normal amounts of food, has normal body temperature, has slight eye mucus and tear stains, has dry nose, and has no positive test results for African swine fever;

[0104] 3) Ineffective: Mental depression, fever, obvious loss of appetite or even loss of appetite, obvious tear stains in the eyes, red patches on the body, and positive test results for African swine fever;

[0105] Among them, "markedly effective" means that there is a difference in effect, "effective" means that the difference is particularly obvious, and "markedly effective" includes "effective".

[0106] The clinical trial results of the present invention are shown in Table 1.

[0107]

[0108] During the test, the blank group gradually developed high fever, loss of appetite, red buttocks, hematuria, and death from the 6th day. At the end of the test, all 30 pigs died. Three and two pigs in the low-dose group and medium-dose group were in poor condition on the 8th and 13th days, respectively. After veterinary diagnosis and testing, five sick pigs were positive for African swine fever, which can be clinically determined to be African swine fever. At the end of the test, one and zero pigs died in the low-dose group and medium-dose group, respectively. The clinical manifestations of the other four sick pigs were significantly relieved and did not die. There were no dead pigs in the high-dose group until the end of the test. The spirit, feeding, and appearance of the pigs in the test group were normal, with no symptoms, and no positive detection of African swine fever.

[0109] From the above test results, it can be seen that the mortality rate of African swine fever positive infected pigs is high without any intervention measures, and can even be as high as 100%. Compared with the blank group, oral administration of the fermented Chinese medicine preparation of the present invention, after 30 days of drug prevention, established an immune barrier for the body, improved the body's antiviral and anti-inflammatory ability, effectively reduced the infection rate, and reduced the mortality rate. With the increase of the oral liquid dosage, its effective rate for the prevention of African swine fever gradually increased, which can reach 93.3%. The above shows that the fermented Chinese medicine preparation of the present invention can be used as a common preparation for the prevention of African swine fever, effectively protecting the health of susceptible animals and reducing economic losses in breeding.

[0110] The above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit the same. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that the technical solutions described in the aforementioned embodiments may still be modified, or some of the technical features may be replaced by equivalents. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A fermented Chinese medicine preparation for preventing and treating African swine fever, characterized in that: The Chinese medicine preparation is prepared from the following raw materials: 30-50g of wild grass incense, 20-50g of astragalus, 5-20g of echinacea, 5-20g of cooked rehmannia polysaccharide, 5-20g of tangerine peel, 10-30g of camellia, 10-30g of salvia miltiorrhiza, 10-30g of poria, 10-30g of atractylodes macrocephala, 10-30g of platycodon, 10-30g of cinnabar lotus, 10-30g of big head grass, 10-30g of jujube, 5-20g of liquorice, 5-20g of acanthopanax senticosus, 5-20g of glossy privet, 5-20g of bilberry, 0.02-0.1g of vitamin C, 0.02-0.1g of vitamin A, 0.5-2g of bacillus subtilis powder, 0.5-2g of lactobacillus powder, 0.2-1g of bifidobacterium powder, and 0.2-1g of clostridium butyricum powder.

2. The fermented Chinese medicine preparation for preventing and treating African swine fever according to claim 1, characterized in that: The Chinese medicine preparation is prepared from the following raw materials: 35-50g of wild grass incense, 25-40g of astragalus, 5-15g of echinacea, 5-15g of cooked rehmannia polysaccharide, 5-15g of tangerine peel, 10-20g of camellia, 10-20g of salvia miltiorrhiza, 10-20g of tuckahoe, 10-20g of atractylodes macrocephala, 10-20g of platycodon, 10-20g of cinnabar lotus, 10-20g of big head grass, 15-25g of jujube, 5-15g of liquorice, 5-15g of acanthopanax senticosus, 5-15g of glossy privet, 5-15g of bilberry, 0.02-0.1g of vitamin C, 0.02-0.1g of vitamin A, 0.5-2g of bacillus subtilis powder, 0.5-2g of lactobacillus powder, 0.2-1g of bifidobacterium powder, and 0.2-1g of clostridium butyricum powder.

3. The fermented Chinese medicine preparation for preventing and treating African swine fever according to claim 1, characterized in that: The Chinese medicine preparation is prepared from the following raw materials: 40g of wild grass fragrance, 30g of astragalus, 9g of echinacea, 9g of cooked rehmannia polysaccharide, 9g of tangerine peel, 15g of camellia, 15g of salvia miltiorrhiza, 15g of poria, 15g of atractylodes, 15g of platycodon, 15g of cinnabar lotus, 15g of scutellaria baicalensis, 20g of jujube, 9g of liquorice, 9g of acanthopanax senticosus, 9g of glossy privet, 9g of bilberry, 0.08g of vitamin C, 0.06g of vitamin A, 1.2g of bacillus subtilis powder, 1g of lactobacillus powder, 0.6g of bifidobacterium powder, and 0.4g of clostridium butyricum powder.

4. The preparation process of the fermented Chinese medicine preparation for preventing and treating African swine fever according to any one of claims 1 to 3, characterized in that: Prepare according to the following steps: (1) Weigh wild grass, astragalus, echinacea, rehmannia polysaccharide, tangerine peel, camellia, salvia miltiorrhiza, tuckahoe, atractylodes, platycodon, cinnabarinum, scutellaria baicalensis, jujube, liquorice, acanthopanax senticosus, glossy privet fruit, and bilberry respectively according to the above weights, wash, dry, and crush the above raw materials, pass through a 60-mesh sieve, and mix them evenly to obtain a mixed Chinese medicine powder for later use; (2) adding 4-6 times the weight of deionized water to the mixed Chinese medicine powder obtained in step (1) and soaking it for more than 40 minutes, boiling it over high heat and then decocting it over low heat for more than 90 minutes, filtering the residue to collect the filtrate, repeatedly extracting the residue 2 to 3 times, and mixing the filtrate collected each time to obtain a Chinese medicine extract; (3) The Chinese medicine extract obtained in step (2) is distilled and concentrated to obtain a final concentration of 1-1.2 g / m Concentrated Chinese herbal medicine extract of L; (4) Weigh vitamin C, vitamin A, Bacillus subtilis powder, Lactobacillus powder, Bifidobacterium powder, and Clostridium butyricum powder respectively according to the above weights, and add them to the concentrated Chinese medicine extract obtained in step (3) in sequence, mix them evenly, and place them in a sealed sterile container for fermentation at 30-37° C. for more than 72 hours; (5) After fermentation is completed, the fermentation liquid is filtered to obtain a clear solution without solid impurities, and spray-dried to obtain a powder or granule to obtain the fermented Chinese medicine preparation.

5. The use of a fermented Chinese medicinal preparation for preventing and treating African swine fever according to any one of claims 1 to 3, characterized in that: The fermented Chinese medicine preparation is used to prepare medicine for preventing and treating African swine fever; the fermented Chinese medicine preparation is used to prepare any drug dosage form acceptable to animals, and the dosage form is powder, granule, decoction, oral liquid.

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