CN100337651C - Chinese medicine composition for preventing bird flu - Google Patents

Abstract

The present invention relates to a Chinese medicine composition for preventing chicken and duck avian influenza viruses. The Chinese medicine composition is composed of effective components and/or a pharmacy acceptable carrier and is characterized in that the effective components are prepared from the following raw material: 90 to 110 portions by the weight of weeping forsythia capsule, 40 to 60 portions by the weight of honeysuckle flower, 25 to 45 portions by the weight of north radix isatidis, 40 to 60 portions by the weight of baical skullcap root, 40 to 60 portions by the weight of male fern rhizome and 70 to 90 portions by the weight of milkvetch root. The prescription can be any oral administration preparation of veterinary medicine. The Chinese medicine composition of the present invention has the advantages of obvious effect for preventing and treating the avian influenza virus of poultry and safe and convenient use.

Description

A kind of traditional Chinese medicine composition for preventing bird flu and its preparation method and application

technical field

The invention relates to an oral Chinese medicinal composition for preventing poultry such as chickens and ducks from avian influenza, a preparation method and application thereof.

Background technique

Avian influenza is an acute and highly fatal infectious disease of poultry caused by type A avian influenza virus. Poultry such as chickens, turkeys, ducks, and quails, as well as wild birds, waterfowl, and seabirds can be infected, but chickens and turkeys are the main ones. Pigs, cats, monkeys and other animals and humans can also be infected.

Avian influenza first occurred in Italy in 1878, and was historically known as fowl plague or true fowl plague. For more than 100 years, bird flu has continuously attacked the poultry industry in the whole world and directly threatened human health and life. In 1997, the outbreak of bird flu in Hong Kong led to large-scale deaths of poultry, and 18 people were infected with bird flu virus, of which 6 died; from December 2003 to March 2004, the outbreak of H5N1 bird flu in Southeast Asia affected almost all In Southeast Asian countries and regions, tens of millions of poultry were infected and died, and more than 20 people were infected with bird flu virus and died.

Although avian influenza has been discovered for more than 100 years, humans have not yet mastered fully effective prevention, treatment and control technologies. Avian influenza vaccine is the main means of preventing outbreaks of avian influenza. If the vaccine used is consistent with the subtype of the invading virus, the protection rate of the vaccine to poultry can reach more than 90%. However, the isolated avian influenza has 15 kinds of H antigens and 9 kinds of N antigens. Different combinations of the two antigens can form hundreds of serotypes, and the popular virus subtypes are unstable, resulting in "drift". The development of vaccines and the diagnosis and control of bird flu have brought great difficulties.

Viruses are microorganisms that parasitize in the living cells of the host. Any drug that can inhibit or kill the virus is also toxic to the host cell, which brings great difficulties to the development of antiviral drugs. Currently, amantadine and rimantadine are the main chemotherapeutic drugs for influenza and avian influenza viruses. Ribavirin is a broad-spectrum antiviral drug, which is also commonly used clinically to treat influenza, but the virus inhibition test shows that ribavirin has no inhibitory effect on influenza virus in vitro. Amantadine and rimantadine have a good effect on preventing avian influenza in chickens and ducks in the early stage of application, but with the widespread and long-term application of this type of drug, the avian influenza virus has developed serious drug resistance to this type of drug, resulting in this type of drug. The drug has no obvious effect in clinical application. The new anti-influenza drugs Zaramivir and Oseltamivir launched in 1999 have definite control effects on human influenza, but there is no report on the prevention of poultry avian influenza. However, these two drugs are expensive, and even if they are effective against bird flu, they are difficult to use in the poultry industry. Therefore, in the prevention and treatment of bird flu, there is no ideal chemotherapeutic drug so far.

In my country, traditional Chinese medicine has been used for the prevention and treatment of human colds and flu for more than a thousand years, and has accumulated rich experience. The traditional Chinese medicines used by human medicine to treat colds and flu are mainly heat-clearing and detoxifying traditional Chinese medicines and exterior traditional Chinese medicines. The former includes forsythia, honeysuckle, scutellaria baicalensis, Houttuynia cordata, Radix Isatidis, Daqingye, wild chrysanthemum, mulberry leaves, burdock, dandelion, Bezoar, etc. The latter includes ginger, cinnamon sticks, ephedra, Bupleurum, nepeta, perilla, magnolia, mint, etc. In addition, traditional Chinese medicines such as Astragalus and Fangfeng can significantly promote and regulate human immunity, and are also commonly used medicines for preventing and treating colds. There are also many kinds of Chinese patent medicines used to prevent and treat colds. The more famous ones include "Yinqiao Tablet", "Shuanghuanglian", "Qingkailing", "Guizhi Mahuang Decoction" and so on.

With the rapid development of my country's livestock and poultry breeding industry and the continuous deepening of veterinary medicine research, the application of traditional Chinese medicine in the prevention and treatment of livestock and poultry diseases has become more and more common, which has played a positive role in improving the survival rate and health level of livestock and poultry. But in general, at present, my country's traditional Chinese veterinary medicine is still at a low technical level with lack of systematic research on drug compatibility, rough product dosage forms, unclear active ingredients, and unstable product quality. There are very few research reports on the application of traditional Chinese medicine to prevent and treat poultry avian influenza, and there are no systematic and in-depth research reports, and there is no Chinese veterinary drug product that has a good control effect on avian influenza in the veterinary drug market.

Contents of the invention

The object of the present invention is to provide a traditional Chinese medicine composition which has a significant preventive effect on chickens, ducks and other poultry avian influenza, as well as its preparation method and application.

In order to achieve the above object, the present invention takes the following technical solutions:

The traditional Chinese medicine composition for preventing bird flu of the present invention is made of the following active ingredients in parts by weight: 90-110 parts by weight of Forsythia, 40-60 parts by weight of Flos Lonicerae, 25-45 parts by weight of Radix Isatidis, and 40-60 parts by weight of Scutellaria baicalensis parts by weight, Mianma Guanzhong 40-60 parts by weight, and Radix Astragali 70-90 parts by weight.

Preferably, 100 parts by weight of forsythia, 50 parts by weight of honeysuckle, 35 parts by weight of banlangen, 50 parts by weight of scutellaria baicalensis, 50 parts by weight of Mianma Guanzhong, and 80 parts by weight of astragalus. More preferably, it is 1000g of forsythia, 500g of honeysuckle, 350g of banlangen, 500g of scutellaria baicalensis, 500g of Mianmaguanzhong, and 800g of astragalus.

The dosage form of the traditional Chinese medicine composition of the present invention is a powder, a drinking water preparation or a granule electuary.

The preparation method of Chinese medicine composition of the present invention, comprises the steps:

(1) Pulverize forsythia, honeysuckle, banlangen, scutellaria baicalensis, Mianma Guanzhong and astragalus into coarse powder respectively;

(2) take step (1) forsythia coarse powder and use ethanol aqueous solution as solvent to carry out percolation extraction, collect percolation liquid;

(3) Take the honeysuckle coarse powder, northern root powder, Scutellaria baicalensis coarse powder, Mianmaguanzhong coarse powder obtained in step (1), use ethanol aqueous solution as solvent, carry out percolation extraction, and collect percolation liquid;

(4) take the astragalus powder obtained through step (1) and use distilled water as a solvent, carry out percolation extraction, and collect percolation liquid;

(5) Merge (2), (3), (4) percolation liquid of step, leave standstill, filter, decompression distillation reclaims ethanol, and remaining liquid medicine is reduced under temperature 65-75 ℃, vacuum tightness 0.02-0.04Mpa Compress and condense into a thick paste, and add auxiliary materials to make the required dosage form.

Yet another preparation method of the traditional Chinese medicine composition of the present invention comprises the steps of: weighing forsythia, honeysuckle, banlangen, scutellaria baicalensis, Mianmaguanzhong, and astragalus in parts by weight according to claim 1, and pulverizing them into coarse powder respectively , decoct according to the water decoction method, collect each decoction liquid, settle and filter, then concentrate under reduced pressure at a temperature of 65-75°C and a vacuum of 0.02-0.04Mpa until the weight of the liquid medicine is 1.0±1.0% of the total weight of the raw materials of traditional Chinese medicine. 0.1 times, add 0.1% sodium benzoate and 0.05% paraben as preservatives to the concentrated medicinal solution to obtain drinking water.

The application of the traditional Chinese medicine composition of the invention in the preparation of medicines for preventing bird flu.

Firstly, the present invention screens more than 40 kinds of traditional Chinese medicines for clearing away heat, detoxifying and expelling symptoms by using the "Chicken Embryo Challenge Test of Avian Influenza Virus" (hereinafter referred to as "Chicken Embryo Test"), and finds Mianmaguanzhong, Forsythia, Honeysuckle, Scutellaria baicalensis, fish More than ten kinds of traditional Chinese medicines such as Xingcao, Radix Isatidis, Folium Isatidis, Licorice, Hypericum perforatum, and Bupleurum have significant inhibitory effects on avian influenza virus in chicken embryos. Then, different formulations are carried out by using the above-mentioned drugs screened out, and the effective components are extracted by two methods of water boiling and alcohol extraction respectively. Finally, the chicken embryo test and artificially infected bird flu chicken treatment test were carried out on the compound formula, and the compound formula with better effect was screened out. According to the test results, it was finally determined that six traditional Chinese medicines such as forsythia, honeysuckle, banlangen, scutellaria baicalensis, Mianmaguanzhong, and astragalus were used as raw materials to make a prescription, which has a significant effect on preventing and controlling avian influenza in chickens and ducks. The first five herbs in the prescription are all essential medicines for clearing away heat and detoxifying, and they all have a strong inhibitory effect on avian influenza virus; the synergistic effect of each herb not only strengthens the inhibitory effect on viruses and bacteria, but also reduces the toxicity of a single herb and the development of viral resistance. Among them, forsythia mainly contains phenylethanol caffeic acid glycosides, Mianma Guanzhong mainly contains mianmasu and trigeminol, honeysuckle mainly contains chlorogenic acid, Radix isatidis mainly contains indigo and indirubin, and Scutellaria baicalensis mainly contains baicalin, Wogonin, these components have different degrees of inhibitory activity on bacteria and viruses. Astragalus contains astragalus polysaccharides, which can significantly promote the immune activity of humans and animals, significantly enhance the phagocytosis of macrophages, increase TNF secretion, and promote the production of virus antibodies.

Detailed ways

Example 1

The preparation method of powder of the present invention, comprises the steps:

(1) Weigh 10000g of forsythia, 5000g of honeysuckle, 3500g of orchid root, 5000g of scutellaria baicalensis, 5000g of Mianmaguanzhong, and 8000g of astragalus, and grind them into coarse powder respectively;

(2) Take the forsythia coarse powder and use 80% ethanol as solvent, carry out percolation according to the liquid extract percolation method, 2.5ml/kg.min, collect percolation liquid;

(3) Get honeysuckle coarse powder, northern banlangen coarse powder, Scutellaria baicalensis coarse powder, Mianmaguanzhong coarse powder through step (1), make solvent with 60% ethanol, carry out percolation according to the liquid extract percolation method, 3ml /kg.min, collect percolate;

(4) Get the astragalus powder obtained through step (1) and use distilled water as a solvent, carry out percolation according to the liquid extract percolation method, 3ml/kg.min, collect percolation liquid;

(5) Combine the percolation liquid in (2), (3), and (4) steps, leave standstill, filter, and distill under reduced pressure

Recover ethanol, concentrate the remaining medicinal solution under reduced pressure at a temperature of 65°C and a vacuum of 0.02Mpa to form a thick paste, spray dry or dry under reduced pressure, and pulverize through a 60-mesh sieve to obtain a dry fine powder;

(6) Add 1.5% white carbon black to the dried fine powder in step (5) to obtain a powder. Among them, the role of silica is to improve fluidity and reduce moisture absorption. The powder of the present invention is powder or powdery granules with good fluidity, brown or tan color, and bitter taste.

Example 2

The preparation method of granule electuary of the present invention, comprises the steps:

(1) Weigh 9000g of forsythia, 5000g of honeysuckle, 3500g of orchid root, 5000g of scutellaria baicalensis, 5000g of Mianma Guanzhong, and 7000g of astragalus, and grind them into coarse powder respectively;

(2) Take the forsythia coarse powder and use 70% ethanol as solvent, carry out percolation according to the liquid extract percolation method, 2.5ml/kg.min, collect percolation liquid;

(3) get through the honeysuckle coarse powder that step (1) makes, northern banlangen coarse powder, Scutellaria baicalensis coarse powder, Mianma Guanzhong coarse powder, make solvent with 50% ethanol, carry out percolation according to the liquid extract percolation method, 2.5 ml/kg.min, collect percolate;

(4) Get the astragalus powder obtained through step (1) and use distilled water as a solvent, carry out percolation according to the liquid extract percolation method, 3ml/kg.min, collect percolation liquid;

(5) Combine (2), (3), (4) percolation liquid of step, leave standstill, filter, underpressure distillation reclaims ethanol, remaining medicinal liquid is concentrated under reduced pressure at temperature 75 ℃, vacuum degree 0.03Mpa to relative thick paste with a density of 1.37;

(6) Add an appropriate amount of sucrose powder to the thick paste in step (5), the sucrose accounts for 28% of the weight of the finished granules, mix well, make granules, and dry in an oven to obtain granules.

Example 3

The preparation method of water-drinking agent of the present invention, comprises the steps:

(1) Weigh 1100g of Forsythia, 500g of Flos Lonicerae, 300g of Banlangen, 600g of Radix Scutellariae, 400g of Mianma Guanzhong, and 800g of Radix Astragali, and grind them into coarse powder respectively;

(2) Take the forsythia coarse powder and use 70% ethanol as solvent, carry out percolation according to the liquid extract percolation method, 2.5ml/kg.min, collect percolation liquid;

(3) get through the honeysuckle coarse powder that step (1) makes, northern banlangen coarse powder, Scutellaria baicalensis coarse powder, Mianma Guanzhong coarse powder, make solvent with 50% ethanol, carry out percolation according to the liquid extract percolation method, 2.5 ml/kg.min, collect percolate;

(4) Get the astragalus powder obtained through step (1) and use distilled water as a solvent, carry out percolation according to the liquid extract percolation method, 3.0ml/kg.min, collect percolation liquid;

(5) Combine the percolation liquid, let it stand, filter, and recover ethanol by distillation under reduced pressure, and the remaining medicinal liquid is concentrated under reduced pressure at a temperature of 65°C and a vacuum of 0.02Mpa to obtain a concentrated medicinal liquid with a weight of 3700g;

(6) Add 0.1% sodium benzoate and 0.05% paraben to the concentrated medicinal solution in step (5) as preservatives to obtain drinking water.

Example 4

Yet another preparation method of the drinking water formulation of the present invention comprises the following steps:

Weigh 1050g forsythia, 400g honeysuckle, 450g beibanlangen, 400g scutellaria baicalensis, 600g mianmaguanzhong, and 700g astragalus membranaceus, and grind them into coarse powder respectively; decoct according to the water decoction method, collect each decoction liquid, settle and filter , concentrated under reduced pressure at a temperature of 72°C and a vacuum of 0.04Mpa until the weight of the medicinal solution is 3600g, and the weight of the concentrated medicinal solution is feasible within the range of 3240g-3960g. Add 0.1% sodium benzoate and 0.05% paraben as preservatives to the concentrated medicinal liquid to obtain drinking water.

Embodiment 5: powder of the present invention

According to the method described in Example 1, the difference is that the raw materials of the following weights are selected: 9500g forsythia, 4500g honeysuckle, 3000g Beibanlangen, 5500g Scutellaria baicalensis, 6000g Mianmaguanzhong, and 8500g Radix Astragali.

Embodiment 6: Granule electuary of the present invention

According to the method described in Example 2, the difference is that the raw materials of the following weights are selected: 10500g forsythia, 5500g honeysuckle, 4000g Beibanlangen, 4500g Scutellaria baicalensis, 5000g Mianmaguanzhong, and 7500g Radix Astragali.

Embodiment 7: drinking water preparation of the present invention

According to the method described in Example 3, the difference is that the following raw materials are selected by weight: 920g forsythia, 420g honeysuckle, 260g northern banlangen, 580g scutellaria baicalensis, 520g Mianmaguanzhong, and 710g astragalus.

Embodiment 8: drinking water preparation of the present invention

According to the method described in Example 4, the difference is that the following raw materials are selected by weight: 1080g forsythia, 580g honeysuckle, 430g Beibanlangen, 430g Scutellaria baicalensis, 590g Mianmaguanzhong, and 870g Radix Astragali.

The medicine of the present invention can be prepared into any conventional oral Chinese medicine preparation by adopting the conventional method of Chinese medicine preparation. For example, ultrafine powder, tablet, etc. But for the convenience of giving chickens and ducks groups medication, the present invention preferably drink water preparations, granule granules, powders and the like. Powders are used as feed additives, and drinking water and granules are used as drinking water additives. But these should not be used to limit the protection scope of the present invention.

The consumption of Chinese medicine composition prevention bird flu of the present invention is respectively:

(1) The powder is used to prevent chicken and duck avian influenza, the amount added to the feed is 1500 grams to 2000 grams per ton of feed; for the treatment of chicken and duck avian influenza, the amount added to the feed is 2500 grams per ton of feed up to 3000 grams;

(2) Drinking water agent is used to prevent chickens and ducks from bird flu, and the amount added in appropriate drinking water is 3-5ml of drinking water agent per 1000 grams of body weight of chickens and ducks per day; The amount added is 6-8ml of drinking water per 1000 grams of body weight for chickens and ducks per day;

(3) Granular granules are used to prevent bird flu in chickens and ducks. Add 0.4-0.6 grams of granules per 1000 grams of body weight of chickens and ducks in an appropriate amount of drinking water; The amount added is 0.7-0.9 grams of granules per 1000 grams of body weight per day for chickens and ducks.

The animal acute toxicity, drug efficacy and pre-clinical control effect on bird flu chickens of the traditional Chinese medicine reconsideration preparation for preventing bird flu described in the present invention are further illustrated below through test examples.

Test 1: Animal acute toxicity test of traditional Chinese medicine composition

Materials and Methods

Animals: Kunming mice, weighing 20.3±1.2g, half male and half male. Purchased from the Military Medical Research Institute of the Logistics Department of the Guangzhou Military Region.

Drugs: Traditional Chinese medicine composition granules, developed and provided by the Guangdong Provincial Key Laboratory of Veterinary Drug Development and Safety Evaluation, School of Veterinary Medicine, South China Agricultural University.

(1) Determination of mouse LD ₅₀

60 mice were randomly divided into 6 groups, 10 in each group, administered by gavage. After taking the medicine, the mice were normal. After continuous observation for 7 days, all the mice survived, and the LD ₅₀ could not be measured.

(2) Maximum tolerance test

Take 30 mice and conduct the test after three days of observation. The day before the test, they were fasted without food and water. Under the condition of room temperature 25±2°C, they were fed once with a drug concentration of 0.5g/ml (maximum suspension concentration). It is 1ml/20g (maximum volume), and the cumulative amount of administration in one day is 25g/kg. After the administration, the animals were kept and observed for two weeks.

During the observation period after administration, the diet, feces, and fur of the mice were normal, and the growth and activity were in good condition. No animal died, and no abnormalities were found in the major organs of the necropsy.

Table 1 Chinese medicine composition drinking water agent _LD50 measurement result group number of animals Drug concentration g/ml Dosing volume ml Dosage g/kg Mouse deaths surviving mice 1 10 0.16 0.5 4.00 0 normal 2 10 0.20 0.5 5.00 0 normal 3 10 0.26 0.5 6.50 0 normal 4 10 0.32 0.5 8.00 0 normal 5 10 0.40 0.5 10.00 0 normal 6 10 0.50 0.5 12.50 0 normal

Discussion and Summary

With the maximum suspension concentration of the drug (0.50g/ml) gavage administration (dose of 12.5g/kg), the LD ₅₀ could not be measured, so the maximum tolerated dose was determined. The acute toxicity test was carried out at the maximum volume (1ml) of the maximum concentration (0.5g/ml) acceptable to mice, and the adverse reactions within two weeks of administration were mainly observed. Results Under the maximum drug dose of 25g/kg, there was no adverse reaction in the mice during the observation period, and no abnormal changes in the main organs were found in the autopsy. The results indicated that the drug was basically non-toxic and safe for clinical use.

Experiment 2: Pharmacodynamics test of oral liquid of traditional Chinese medicine composition

1. Test material

1.1 Test chicken: 50-day-old high-quality yellow chicken. They were purchased from Guangdong Poultry Company of Guangdong Poultry Research Institute at 1 day of age, and were bred to 50 days of age to start the experiment. Hemagglutination inhibition test (HI) was used to detect avian influenza virus (AIV) negative at the age of 40 days.

1.2 Feed: purchased from the experimental chicken farm of the College of Animal Science, South China Agricultural University, without adding any drugs.

1.3 Drugs: All the raw materials of Chinese medicinal materials used were purchased from Guangzhou Medicinal Materials Company, extracted and prepared by the Key Laboratory of Veterinary Drug Development and Safety Evaluation of Guangdong Province, School of Veterinary Medicine, South China Agricultural University, and prepared Chinese medicinal composition drinking water (oral liquid). Each milliliter of oral liquid is equivalent to 1g of crude drug. Amantadine was purchased from Huatian Company of South China Agricultural University, and rimantadine was purchased from Zhejiang Yukang Pharmaceutical Co., Ltd. The melting point and mass spectrometry of the two drugs were consistent with the literature values. Amantadine and rimantadine are directly dissolved in distilled water to prepare a medicinal solution with a mass concentration of 0.2%.

1.4 Challenge virus strain: H5 serotype avian influenza virus (half infection concentration 1.0×10 ^-6 ), preserved in the Laboratory of Poultry Disease, College of Veterinary Medicine, South China Agricultural University.

2. Test method

Test grouping: 90 50-day-old high-quality yellow chickens, half male and half male, were randomly divided into 5 groups, with 10 normal control groups and 20 other groups. See Table 2 for details.

Virus challenge: Dilute chicken embryo allantoic fluid containing avian influenza virus to a concentration of 0.5×10 ^-5 , and inject 0.2ml into abdominal muscle of each chicken for inoculation.

Dosing: On the day of AIV inoculation, each group in the test administration was fed with 5ml, and the healthy control group and the infected control group were fed with normal saline, once a day, for 7 consecutive days .

Drug efficacy evaluation criteria: the test lasted for 21 days, and the death and clinical manifestations of the chickens were observed every day. The dead chickens were autopsied, and each chicken was weighed before and after the test. Drug efficacy was evaluated according to the following criteria: (1) death: during the test period, typical symptoms of bird flu occurred and died, and those with typical pathological changes in autopsy were judged as infected and died. (2) Efficient: After the test ends, all the chickens that are still alive after administration are judged to be effective, and the percentage of the number of surviving chickens in each group to the number of test chickens at the end of the test is the effective rate. (3) Weight gain: Calculate the weight gain of each chicken, the average body weight and standard deviation of each group of test chickens according to the body weight of each chicken at the beginning of the test and the end of the test. The relative weight gain rate is calculated according to the average weight gain ratio of each medication treatment group and the healthy control group, wherein the healthy control group is set as 100%.

3. Animal test results

3.1 Infection control group

Among the 20 infected chickens, 17 died within 15 days of inoculation with the avian influenza virus, and the surviving 3 chickens were still healthy until the 21st day. Among them, 2 died on the 3rd day after inoculation, 3 died on the 4th day, 5 died on the 4th day, 5 died from the 5th to the 10th day, and 2 died from the 11th to the 15th day. Main symptoms: mental depression, loss of appetite or loss of appetite; loose feathers, tears from the eyes; drainage-like feces; reluctance to walk, severe cases lying on the ground, and neurological symptoms such as neck bending, spinning, and jumping. Autopsy features: mucus in the mouth, enlargement of liver, kidney, gallbladder and other organs with bleeding spots, usually purple-black in color.

3.2 Administration group

After the infected chickens were administered and treated, only one died in the traditional Chinese medicine composition group, and the survival rate was 95%. Most of the chickens did not show clinical symptoms, and some chickens showed mild symptoms such as tearing eyes and loose feathers. Symptoms disappeared completely on the 21st day. Eighteen chickens died in the amantadine group, and 15 chickens died in the rimantadine group. See Table 2 for details.

Table 2 The control effect of three kinds of drugs on chickens infected with avian influenza virus group quantity deal with number of deaths mortality rate% Efficient% 50d body weight m/g 71d body weight m/g Weight gain % Relative weight gain% healthy control group 10 No infection, no medication 0 0 - 685.3±58.7 1103.6±63.0 61.04 100 Chinese medicine compound preparation group 20 Infection, feeding with 5.0ml oral liquid 1 5 95 683.9±63.7 1091.2±72.8 59.56 97.58 Amantadine group 20 Infection, 10mg gavage 18 90 10 692.4±67.5 978.2±55.3 41.28 67.63 Rimantadine group 20 Infection, 10mg gavage 15 75 25 679.0±72.5 982.0±81.6 44.62 73.10 Infection control group 20 infection, no medication 17 85 - 677.6±62.0 971.6±35.4 43.39 71.10

4 Conclusion

The test results show that the traditional Chinese medicine composition of the present invention has a good protective effect on chickens artificially infected with avian influenza virus, and the survival rate of infected chickens reaches 95%, while the traditional anti-influenza drugs "amantadine", "amantadine" The effective rates of "ethylamine" are only 10% and 25%, respectively, and the difference is extremely significant. The weight gain of the chickens in the Chinese medicine composition group was comparable to that of the healthy control group, which was significantly better than that of the "amantadine" and "rimantadine" groups, indicating that the Chinese medicine composition not only has a good control effect on avian influenza in chickens, but also has a good effect on chickens. No adverse effect on growth.

Experiment 3: The control effect test of Chinese medicine composition drinking water agent on chickens infected with avian influenza in cohabitation

1. Test material

1.1 Test chicken: 50-day-old high-quality yellow chicken. They were purchased from Guangdong Poultry Company of Guangdong Poultry Research Institute at 1 day of age, and were bred to 50 days of age to start the experiment. Hemagglutination inhibition test (HI) was used to detect avian influenza virus (AIV) negative at the age of 40 days.

1.2 Feed: purchased from the experimental chicken farm of the College of Animal Science, South China Agricultural University, without adding any drugs.

1.3 Drugs: All the raw materials of Chinese medicinal materials used were purchased from Guangzhou Medicinal Materials Company, extracted and prepared by the Key Laboratory of Veterinary Drug Development and Safety Evaluation of Guangdong Province, School of Veterinary Medicine, South China Agricultural University, and formulated drinking water doses of Chinese medicinal compositions. Each milliliter of drinking water is equivalent to 1g of crude drug. Rimantadine was purchased from Zhejiang Yukang Pharmaceutical Co., Ltd. The results of melting point determination and mass spectrometry were consistent with the literature values. Rimantadine is directly dissolved in distilled water to prepare a medicinal solution with a mass concentration of 1%.

1.4 Challenge virus strain: H5 serotype avian influenza virus (half infection concentration 1.0×10 ^-6 ), preserved in the Laboratory of Poultry Disease, College of Veterinary Medicine, South China Agricultural University.

2. Test method

Chickens artificially infected with avian influenza: 50 45-day-old high-quality yellow chickens, half male and half female, dilute the allantoic fluid of chicken embryos containing avian influenza virus to a concentration of 0.5×10 ^-5 , and inject 0.2ml into the abdomen of each chicken for inoculation. On the 3rd day after inoculation, the onset of disease began, and the diseased chickens with obvious clinical symptoms were marked every day, and evenly distributed among the experimental flocks until the 10th day after inoculation.

Test grouping: 110 45-day-old high-quality yellow chickens, half male and half male, were randomly divided into 6 groups, with 10 normal control groups and 20 other groups. See Table 3 for details.

Dosing: On the day when chickens artificially infected with avian influenza were transferred to the test pen, water administration was started. Water was withheld for 3 hours before the administration of drinking water. The Chinese medicine composition drinking water dose is 5ml and 10ml for each chicken respectively, and 1% rimantadine is added to a drinking fountain with 600ml tap water in the amount of 1ml and 2ml for each chicken, once a day, for the chickens to freely drinking water. After drinking the water with the drug added, give tap water for the chickens to drink. The control group was given tap water.

Drug efficacy evaluation criteria: the test lasted for 28 days, and the death and clinical manifestations of the chickens were observed every day. The dead chickens were autopsied, and each chicken was weighed before and after the test. The drug efficacy evaluation standard is the same as that of Experiment 2.

The artificially infected source chickens and cohabitation test chickens were recorded and statistically processed respectively.

3. Animal test results

3.1 Cohabitation infection control group

Thirteen of the 20 cohabiting infected chickens died within 28 days after being transferred to artificially infected chickens. One died on the 3rd day, 2 died on the 4th day, 4 died on the 5th day, 4 died from the 5th to the 10th day, and 2 died from the 11th to the 28th day.

3.2 Administration group

After cohabitation infected chickens were administered and treated, all of the 5ml dosage group of the traditional Chinese medicine composition survived, and one died in the 10ml dosage group, the survival rate was 95%. Nine and eleven chickens died in the rimantadine 10mg and 20mg dose groups, respectively. See Table 3 for details. All the source chickens artificially infected with the virus died.

Table 3 The control effect of Chinese medicine composition and rimantadine on cohabiting chickens infected with avian influenza group quantity deal with number of deaths mortality rate% Survival % 45d body weight m/g 80d body weight m/g Weight gain % Relative weight gain% healthy control group 10 No infection, no medication 0 0 - 617.2±45.3 1356.3±67.2 119.8 100 Chinese medicine composition group I 20 Infection, 5.0ml/drinking water 0 0 100 623.0±60.5 1327.0±76.2 113.0 94.32 Chinese medicine composition group II 20 Infection, 10ml/drinking water 1 5 95 625.5±56.6 1330.5±70.4 112.7 94.07 Riantadine group I 20 Infection, 10mg/drinking water 9 45 55 612.7±50.9 1259.1±63.2 105.5 88.06 Riantadine II group 20 Infection, 20mg/drinking water 11 55 45 622.8±68.7 1179.0±77.8 89.31 75.55 Infection control group 20 infection, no medication 13 65 35 629.5±47.2 1297.6±54.0 106.1 88.56

4 Conclusion

The results of the drug prevention and control test of cohabitation infection show that the Chinese medicine composition of the present invention has a good protective effect on cohabitation infected chickens, and the survival rate of each chicken with 5ml and 10ml drinking water per day reaches 100% and 95% respectively, which is significantly better than that of cohabitation chickens. on the protective effect of rimantadine. In addition, not all chickens in the cohabitation-infected control group became ill, and the survival rate was 35%.

The weight gain of the chickens in the traditional Chinese medicine composition group was slightly lower than that of the healthy control group, but the difference was not significant, and the relative health group was 94.32% and 94.07% respectively. Riantadine has a significant effect on the growth of chickens, and the weight gain of the 10mg and 20mg dose groups is 12% and 24% lower than that of the healthy control group, respectively. The cohabitation infection test shows that the traditional Chinese medicine composition of the present invention has a good control effect on chickens naturally infected with avian influenza.

Claims (7)

1. the Chinese medicine composition of a birds flu-preventing, effective ingredient is wherein made by following raw materials according: Fructus Forsythiae 90-110 weight portion, Flos Lonicerae 40-60 weight portion, northern Radix Isatidis 25-45 weight portion, Radix Scutellariae 40-60 weight portion, Rhizoma Dryopteris Crassirhizomatis 40-60 weight portion, Radix Astragali 70-90 weight portion.

2. Chinese medicine composition according to claim 1 is characterized in that described effective ingredient made by following raw materials according: Fructus Forsythiae 100 weight portions, Flos Lonicerae 50 weight portions, northern Radix Isatidis 35 weight portions, Radix Scutellariae 50 weight portions, Rhizoma Dryopteris Crassirhizomatis 50 weight portions, the Radix Astragali 80 weight portions.

3. Chinese medicine composition according to claim 1 is characterized in that described effective ingredient made by following raw materials according: Fructus Forsythiae 1000g, Flos Lonicerae 500g, northern Radix Isatidis 350g, Radix Scutellariae 500g, Rhizoma Dryopteris Crassirhizomatis 500g, Radix Astragali 800g.

4. Chinese medicine composition according to claim 1 is characterized in that its dosage form is powder, drinking agent or dissolved granule.

5. a method for preparing the described Chinese medicine composition of claim 1 is characterized in that comprising the steps:

(1) Fructus Forsythiae, Flos Lonicerae, northern Radix Isatidis, Radix Scutellariae, Rhizoma Dryopteris Crassirhizomatis, Radix Astragali pulverize separately are become coarse powder;

(2) get step (1) Fructus Forsythiae coarse powder and carry out the percolation extraction as solvent, collect percolate with ethanol water;

(3) get Flos Lonicerae coarse powder, northern Radix Isatidis coarse powder, Radix Scutellariae coarse powder, the Rhizoma Dryopteris Crassirhizomatis coarse powder that step (1) makes, make solvent, carry out percolation and extract, collect percolate with ethanol water;

(4) learn from else's experience radix astragali coarse powder that step (1) makes with the distillation water as solvent, carry out percolation and extract, collect percolate;

(5) merge the percolate of (2), (3), (4) step, leave standstill, filter, ethanol is reclaimed in distilling under reduced pressure, and residue medicinal liquid concentrating under reduced pressure under temperature 65-75 ℃, vacuum 0.02-0.04Mpa becomes thick paste, adds adjuvant and makes required dosage form.

6. method for preparing the described Chinese medicine composition of claim 1, it is characterized in that comprising the steps: taking by weighing Fructus Forsythiae by the described weight portion of claim 1, Flos Lonicerae, the north Radix Isatidis, Radix Scutellariae, Rhizoma Dryopteris Crassirhizomatis, the Radix Astragali, and pulverize separately becomes coarse powder, pressing the decocting cooking method decocts, collect the fried liquid of each time, after sedimentation is filtered, at temperature 65-75 ℃, the weight that is evaporated to medicinal liquid under the vacuum 0.02-0.04Mpa is 1.0 ± 0.1 times of Chinese medicine raw material gross weight, add 0.1% sodium benzoate and 0.05% nipalgin in the concentrated medicament and make antiseptic, promptly get drinking agent.

7. Chinese medicine composition as claimed in claim 1 is used for the application of the medicine of birds flu-preventing in preparation.