CN102813713B - Rhizoma dryopteris crassirhizomae and fructus crataegi composition, preparation method and application of composition - Google Patents
Rhizoma dryopteris crassirhizomae and fructus crataegi composition, preparation method and application of composition Download PDFInfo
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Abstract
The invention relates to a rhizoma dryopteris crassirhizomae and fructus crataegi composition, a preparation method and an application of the composition. The effective components of the composition provided by the invention are prepared from the following raw materials by weight: 10-90 parts of rhizoma dryopteris crassirhizomae and 10-90 parts of fructus crataegi. The rhizoma dryopteris crassirhizomae and fructus crataegi composition provided by the invention is good in effect of preventing and treating poulty viral diseases, in particular to the poultry diseases caused by bursal disease viruses, the poultry kidney diseases caused by renal type infectious bronchitis viruses, the influenza virus A diseases of mammals caused by avian influenza viruses and the poultry avian influenza virus diseases; and the rhizoma dryopteris crassirhizomae and fructus crataegi composition provided by the invention has the characteristics of being more liable to obtain the raw materials, good in curative effect, free of residues, green and environment-friendly and the like, so that the composition can replace the chemical antiviral drugs.
Description
Technical field
The invention belongs to field of medicaments, relate to a kind of Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi compositions, its preparation method and application.
Background technology
China's animal epidemic type is many, popular extensively, and viral blight takes place frequently and brings serious impact to social economy and people's lives.Show that according to interrelated data direct economic loss that China causes because of viral disease every year, up to 200～25,000,000,000 yuan, is approximately equivalent to 2.5%～3.1% of the animal husbandry gross output value.From indirect loss, the economic loss of breeding performonce fo animals reduction, the decline of livestock products quality, food consumption increase, artificial waste, expenses for prevention and control increase, environmental nuisance and the related industry causing is thus just huger, and estimation is about damnous 3～5 times of Mortality.Livestock and poultry viral disease occurs to have formed great threat to the mankind's healthy and even existence, has upset the normal orders of life of people, becomes and causes social unsettled key factor.
In addition, livestock and poultry animal viral disease also has a strong impact on China's food safety and animal foods outlet industry.China's livestock products outlet is in recent years had difficulty in walking, and is obstructed repeatedly, and meat outlet only accounts for 1.3% of domestic meat total output, accounts for 3.6% of world's meat total volume of exports.American-European for Chinese animal products, trade barrier are set, during April in August, 1996 to calendar year 2001, in the Fowl meat of European Union with China's export, contain epidemic disease and pesticide, residue of veterinary drug etc. and serve as reasons China's Fowl meat has been descended to ban as long as five years.Although April calendar year 2001, European Union has just announced the import of China Broiler to lift a ban, Korea S is separated to again H5 subtype avian influenza virus from the fresh duck meat of China's somewhere outlet, Korea S and Japan successively announce nonimportation Chinese Poultry meat product, the outlet of European Union is stopped again indefinite duration, cause Chinese exports broiler enterprises ' industry to be absorbed in serious awkward predicament, suffered serious economic wound.Therefore, the new veterinary drug control of developing green environment-friendly type animal viral disease has become the important foundation stone that livestock and poultry breeding industry develops in a healthy way.
Rhizoma Dryopteris Crassirhizomatis is for beginning to be loaded in Shennong's Herbal compared with conventional Chinese medicine, and Rhizoma Osmundae form, the place of production and accompanying drawing feature that successive dynasties book on Chinese herbal medicine is recorded show that Rhizoma Osmundae kind is a lot of.The Rhizoma Dryopteris Crassirhizomatis territory of use that pharmacopeia is recorded is wider, is one of Chinese medicine general types.Rhizoma Dryopteris Crassirhizomatis is dry rhizome and the petiole residual base of Dryopteridaceae plant dryopteris crassirhizoma Dryopteris crassirhizoma Nakai, bitter in the mouth, be slightly cold, slightly poisonous, return liver, stomach warp, there is the function such as heat-clearing and toxic substances removing, anthelmintic.Modern pharmacological research shows to have antiviral, anthelmintic, antitumor, excited uterus, estrogen-like effects etc., is clinically used for the treatment of that pneumonia, hepatitis B, intestinal parasitical diseases, department of obstetrics and gynecology are hemorrhage, herpes zoster etc.In recent years, along with the modern Chinese medicine development that learns a skill, from this plant, get 3 phloroglucinol class monomeric compounds: dryocrassin ABBA, Filixic Acids ABA and albaspidin AA, this plant shoot divides and also contains multiple triterpenoid compound.Calendar year 2001, someone reported that therefrom separation has obtained 4 kinds of ketoside compounds again, in addition also containing compositions (Gao Zengping, 2003, Beijing University of Chinese Medicine's thesis for the doctorate) such as tannin, volatile oil, resins.
The active report of Rhizoma Dryopteris Crassirhizomatis effective ingredient increases gradually: as the water of Rhizoma Dryopteris Crassirhizomatis and alcoholic extract all can suppress the 68-1 of the section strain of influenza virus capital, infected by influenza and herpesvirus are direct killing action (Jiang Yasheng, Yang Ning. pharmacy practice magazine, 2000,18:17).Rhizoma Dryopteris Crassirhizomatis has the effect of anti-HBsAg in the time of 0.3 ~ 5mg/100l concentration, simultaneously to Experimental Hepatic Damage have protective effect (Zheng Minshi, etc. Journal of Chinese Hospital Pharmacy, 1991,11:53; Xie Yunzhang. Hebei medicine, 1995,17:82).Also have experimental results show that Rhizoma Dryopteris Crassirhizomatis have the effect of anti AIDS virus (Zhu Lijiang. Chinese Chinese medicine information magazine, 1996,3:17), Japanese scholars has been isolated 4 flavones ingredient (Byung-Sun Min with anti-HIV activity from Rhizoma Dryopteris Crassirhizomatis, Deng, Pharm.Bull.2001,49:546).In addition experimental results show that Rhizoma Dryopteris Crassirhizomatis to Coxsackie B virus and Echovirus have obvious inhibitory action (Dong Jiede. Shandong College of Traditional Chinese Medicine's journal, 1993,17 (4): 46), test shows that 10 kinds of main flow Rhizoma Osmundae kinds all have pharmacologic action, but the strongest with Rhizoma Dryopteris Crassirhizomatis, Brainea insignis, Blechnum orientale Linn., be secondly Rhizoma Osmundae and Matteuccia strthiopteris.Judging that according to the experimental result that affects on prothrombin time Rhizoma Dryopteris Crassirhizomatis anastalsis is the strongest, is secondly Rhizoma Osmundae and osmund.
In recent years, Rhizoma Dryopteris Crassirhizomatis and compositions antiviral patent of invention thereof emerge in an endless stream, mainly be divided into following a few class: (1) treatment influenza virus aspect: compound rhizome of Cyrtomium aspirin tablet patent is mainly used in treating influenza (application number CN03108749.3, publication number is CN1466981).Vinegar beverage with rhizoma polystichi isatis root has the grippal effect for the treatment of (application number CN200510015819.6, publication number is CN1954731) and the application (application number: CN200910082162.3, publication number be CN10862391A) of Rhizoma Osmundae compositions in preparation treatment human influenza medicine; A kind of prevention of Shanxi Yabao Pharmaceutical Group Corp.'s application and the Chinese medicine composition (application number CN200910210723.3, publication number is CN101695536A) for the treatment of influenza A H1N1.Also there is report Rhizoma Osmundae to have the effectiveness (application number CN200910075211.0, publication number is CN101991694A) of clear and definite anti-influenza A H 1 N 1 virus.(2) atypical pneumonia virus: using Rhizoma Osmundae as Chinese medicine composition as treatment and the prophylactic (application number CN03143211.5, publication number is CN1483463) of atypical pneumonia; (3) viral myocarditis: a kind of Chinese medicine composition of Hebei Yiling Medicine Inst. Co., Ltd's application is in the application of preparation treatment viral myocarditis drug intoxication, this invention clinical experiment confirms that treatment viral myocarditis is had to significant curative effect (application number CN200810117306.X, publication number is CN101637533).(4) hand-foot-mouth disease: the application of a kind of Chinese medicine composition of Hebei Yiling Medicine Inst. Co., Ltd's application in preparation treatment hand-foot-mouth disease medicine, this invention is by effective kill virus, bring down a fever, antiinflammatory, effectively treatment hand-foot-mouth disease (application number is CN200810117302.1, and publication number is CN101637529).(5) herpesvirus: clinical experiment confirms that Rhizoma Osmundae extract has significant curative effect (application number is CN201010271514.2, and publication number is CN102379957A) to treatment herpes zoster.
Patent aspect livestock and poultry concentrates on: the sick control of (1) animal dysentery, this formula of effervescent Chinese medicinal composition of avian dysentery and preparation method thereof report can effectively improve immunity, have no side effect and drug residue (application number CN200910228407.9, publication number is CN101816711A); (2) parasiticide aspect: Rhizoma Dryopteris Crassirhizomatis has heat-clearing and toxic substances removing, anthelmintic hemostatic function (application number CN200510015178.4, publication number is CN1939352); (3) avian influenza prevention: it is remarkable for control poultry bird flu virus prevention effect that Rhizoma Dryopteris Crassirhizomatis is prepared peroral dosage form, uses safety, convenient (application number CN200510011432.3, publication number is CN1682843).A kind of influenza of Agricultural University Of South China's application subsequently and the medicine (application number is CN200610011341.4, and publication number is CN1846717) of bird flu virus; Chinese medicine extract and the preparation method (application number is CN200710123069.3, and publication number is CN101332255) of a kind of birds flu-preventing of Baoding Jizhong Pharmaceutical Co., Ltd.'s application.
Fructus Crataegi is the important medical material in China's Chinese medicine and pharmacy.Fructus Crataegi is the dry mature fruit of the may Fructus Pyri Pashiae Crataegus pinnatifida bge var.majorN.E.Br. of Qiang Wei section or fructus crataegi pinnatifidae C.pinnatifida bge, records in version " Chinese Pharmacopoeia " in 2005.The fruit of Fructus Pyri Pashiae has dividing of " Fructus Crataegi, Fructus Crataegi cuneatae ", and all there is cultivation all parts of the country, are medicinal and edible plant.The acid of Fructus Crataegi sweet in the mouth, slightly warm in nature, returns spleen, stomach, Liver Channel.There is promoting the production of body fluid to quench thirst, removing food stagnancy relieving dyspepsia, blood circulation promoting and blood stasis dispelling, spleen reinforcing digestion promoting function, does not clinically change and postpartum stagnation stomachache for dyspepsia.Fructus Crataegi is rich in Crataegolic acid, anthocyanidin, citric acid, tartaric acid, ascorbic acid, caffeic acid, boots acid, Radix Hyperici Monogyni (Herba Hyperici Monogyni) two, canopy skin element, male dose of element, mushroom alkene, organic acid and flavonoid isoreactivity material.
Fructus Crataegi bioactivity research: the raising of (1) Promote immunity activity.Fructus Crataegi decoct to mouse thymus and spleen weight, T lymhocyte transformation rate, T lymphocyte have remarkable facilitation (Tao little Ping. Liaoning Journal of Traditional Chinese Medicine, 2001, (2): 86; Chang Jiang, etc. packet header medical college journal, 1996,12:10-11).Fructus Crataegi extract can strengthen mice reticuloendothelial system phagocytic activity, improve immune organ index and the rising peripheral blood leucocyte of immunodeficiency models mice, illustrate that Fructus Crataegi extract can significantly improve the immunologic function of immunologic hypofunction mice body, may all there is potentiation (Liu Chunli, 2006 The Fourth Military Medical University's master thesis) to cellular immunization and humoral immunization.(2) antiviral activity: a kind of patent of invention of preparing high purity maslinic acid taking Fructus oleae europaeae as raw material shows that high purity maslinic acid can be used as the crude drug (application number is CN200810125935.2, and publication number is CN101418031) of preparing antitumor, antiviral, anti-HIV and anti-cardiovascular disease; Preventing and treating aspect hepatitis virus, the Chinese medicine composition that utilizes Fructus Crataegi and Lentinus Edodes to prepare, can be used for the treatment (application number is CN200710137338.4, and publication number is CN101057901) of fatty liver; In addition, utilize pure natural haw berry nuclear extract, can be for the preparation for the treatment of hemorrhoid (application number: CN03143103.8, publication number CN1457875).
Aspect livestock and poultry viral disease, having declared patent comprises following: (1) prevents and treats Avian pneumo-encephalitis virus and bird flu virus: a kind of Chinese medicine composition for the treatment of Newcastle disease of patent and preparation method thereof (application number CN201110119524.9, publication number CN102205036A), mainly with 14 kinds of ingredients such as the Radix Astragali, Radix Codonopsis, Radix et Caulis Opuntiae Dillenii, Rhizoma Coptidis, Fructus Crataegi, the Radix Aucklandiae, report the medicine effective percentage 99% of its treatment newcastle disease epidemic disease, cure rate 96%.Chinese medicine (the application number: CN200910062932.8 of patent treatment fowl flu, publication number: CN101933998A), this patent is characterized in that it is made up of Rhizoma Coptidis, Cortex Phellodendri, the Radix Astragali, Radix Scutellariae, Herba Ephedrae, Radix Glycyrrhizae, Fructus Forsythiae, Semen Armeniacae Amarum, Folium Isatidis, Radix Bupleuri, herba houttuyniae, Radix Platycodonis, Ramulus Cinnamomi, Radix Saposhnikoviae, the Rhizoma Atractylodis Macrocephalae, Fructus Crataegi, Flos Lonicerae, 18 kinds of Chinese medicines of Bufo siccus.Advantage of the present invention is to feed ill fowl or be ground into powder mix fodder with this Chinese medicine decocting to feed ill fowl, can effectively treat viral fowl flu.Laying ducks Chinese herbal medicine additive and application (application number CN200910103647.6, publication number is CN101543260), main component is by 23 kinds of compositions such as Cortex Phellodendri, the Cortex Eucommiae, Flos Lonicerae, Radix Codonopsis, Fructus Crataegis, then adds calcium bicarbonate, phytase, vitamin E and methionine, can preventing bird flu effectively.Semen Crataegi extract is to viral effect and the application of antiviral drugs; this invention is to find that first the extract of Semen Crataegi is to bird flu H9N2; H7N2; H6N2; inhibition and the counteracting toxic substances protective effect (application number: CN200510131943.9, publication number is CN1985907) of H5N1 and H5N2 virus and newcastle (F4) virus.(2) prevent and treat infectious bronchitis: patent is for preventing and treating a kind of Chinese herbal and crude drugs preparations (application number CN95100946.X of livestock and poultry, publication number is CN1116536), can prevent and treat chicken, the fowl such as Columba livia are raiseeed because of respiratory tract, and the disease that escherichia coli etc. cause has significant curative effect.Medicine (the application number CN200610069485.5 for the treatment of Nephropathogenic infectious bronchitis, publication number is CN1966009), the invention discloses a kind of medicine for the treatment of Nephropathogenic infectious bronchitis, be by the Radix Astragali, Flos Lonicerae, Herba Hedyotidis Diffusae, Rhizoma Imperatae, Fructus Crataegi, Rhizoma Dioscoreae, Radix Glycyrrhizae according to a certain weight ratio prescription form, taking convenience, without any side effects, after curing, be difficult for recurrence, take rear instant effect, apply Drug therapy Nephropathogenic infectious bronchitis effective percentage 100% of the present invention, cure rate 95%.(3) control unknown high fever of pigs: a kind of Chinese herbal medicine additive (application number: CN200910018731.8 of preventing and treating porcine hyperthermia, notification number: CN101653494), the invention discloses a kind of Chinese herbal medicine additive of preventing and treating porcine hyperthermia, it is made up of following herbal raw material: the Radix Astragali, Radix Codonopsis, Fructus Crataegi, the Radix Aucklandiae, Flos Lonicerae, Radix Isatidis, Echinacea, Periostracum Cicadae, Venenum Bufonis, Radix Glycyrrhizae, garlicin and 12 kinds of Chinese medicines of Radix et Caulis Opuntiae Dillenii form, preventing swine high fever effective percentage 94%, treated effect 96%, cure rate 82%.
In sum, utilize Rhizoma Dryopteris Crassirhizomatis, Fructus Crataegi to prepare separately medicine and had report and the patent of many anti-virus aspects.But, there is no both at home and abroad about Rhizoma Dryopteris Crassirhizomatis and Fructus Crataegi and carry out prescription, and two kinds of prescription extracts report of compatibility together, have no the report in curative effect aspect control animal viral disease at compatibility.
Summary of the invention
The object of this invention is to provide a kind of Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi compositions that can prevent and treat poultry, mammalian disease toxicity disease.
Another object of the present invention is to provide the preparation method of above-mentioned Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi compositions.
A further object of the present invention is to provide above-mentioned Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi compositions and applies in preparation prevention with in treating poultry, mammiferous viral disease medicine.
Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi compositions provided by the invention, its effective ingredient is made up of the raw material of following weight portion: 10 ~ 90 parts of 10 ~ 90 parts of Rhizoma Dryopteris Crassirhizomatiss and Fructus Crataegis.
Preferably, described effective ingredient is made up of the raw material of following weight portion: 20 ~ 80 parts of 20 ~ 80 parts of Rhizoma Dryopteris Crassirhizomatiss and Fructus Crataegis.
More preferably, described effective ingredient is made up of the raw material of following weight portion: 60 parts of 40 parts of Rhizoma Dryopteris Crassirhizomatiss and Fructus Crataegis.
Described Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi compositions is Rhizoma Dryopteris Crassirhizomatis, Fructus Crataegi and conventional adjuvant or carrier, is prepared into soluble powder, granule, oral liquid or tablet according to conventional formulation method.Wherein be preferably granule, soluble powder.
Described Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi compositions, comprises that conventional pharmaceutical excipient is as one or more in starch, dextrin, lactose, stevioside, sucrose, glycyrrhizin, high fructose syrup, sucralose, xylitol, Sorbitol, glucose, pregelatinized Starch, carboxymethyl starch sodium.
The preparation method of described Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi compositions, comprises the following steps:
(1) take Rhizoma Dryopteris Crassirhizomatis, Fructus Crataegi by above-mentioned weight portion, and be ground into respectively coarse powder, use ethyl acetate lixiviate, extracting solution concentrating under reduced pressure, obtains ethyl acetate extract;
(2) 75% ~ 95% soak with ethanol for medicinal residues after lixiviate, boils, and by lixiviating solution sucking filtration, concentrating under reduced pressure evaporate to dryness, obtains ethanol extract;
(3) ethyl acetate extract and ethanol extract are mixed.
Particularly, take 10 ~ 90 parts of Rhizoma Dryopteris Crassirhizomatiss, 10 ~ 90 parts of Fructus Crataegis, pulverize, mix, add the ethyl acetate of 4 ~ 8 times of amounts, lixiviate 24 ~ 48h, collects lixiviating solution, reclaims ethyl acetate, obtains extractum; Medicinal residues add 4 ~ 8 times of volumes, 75 ~ 95% soak with ethanol 2 ~ 8h, boil 1h ~ 2h, and extracting solution sucking filtration is obtained to supernatant, merge lixiviating solution and supernatant and revolve steaming, and concentrating under reduced pressure, obtains extractum.The extractum that two kinds of organic solvents are obtained is prepared into soluble powder, granule, tablet or oral liquid with conventional formulation after mixing.
The another kind of preparation method of described Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi compositions, comprises the following steps:
(1) take Rhizoma Dryopteris Crassirhizomatis, use ethyl acetate lixiviate, concentrating under reduced pressure obtains extractum; 75 ~ 95% soak with ethanol for medicinal residues, boil and extract 1 ~ 2 time, merging filtrate, and concentrating under reduced pressure evaporate to dryness obtains extractum; Merge extractum;
(2) take Fructus Crataegi, use ethyl acetate lixiviate, concentrating under reduced pressure obtains extractum; 75 ~ 95% soak with ethanol for medicinal residues, boil, and by lixiviating solution sucking filtration, concentrating under reduced pressure evaporate to dryness obtains extractum, merge extractum;
(3) getting Rhizoma Dryopteris Crassirhizomatis extract and Fructus Crataegi extract extractum mixes.
Particularly, Rhizoma Dryopteris Crassirhizomatis extract preparation: take 10 ~ 90 weight portion Rhizoma Dryopteris Crassirhizomatiss, pulverize, with 4 ~ 8 times of volumes of acetic acid ethyl esters immersion 24 ~ 48h, collect lixiviating solution, reclaim ethyl acetate, obtain extractum; Medicinal residues add 4 ~ 8 times of 75 ~ 95% soak with ethanol 2 ~ 8h, boil and extract 1 ~ 2h, extract 1 ~ 2 time, and merging filtrate, by above-mentioned filtrate rotary evaporation, concentrating under reduced pressure obtains extractum.By the extractum mixing for standby use of 2 collections.
Fructus Crataegi extract preparation: take 10 ~ 90 weight portion Fructus Crataegis, pulverize, with after 4 ~ 8 times of volumes of acetic acid ethyl esters immersion 24 ~ 48h, reclaim ethyl acetate, obtain extractum; Medicinal residues are with after 4 ~ 8 times of volumes, 75 ~ 95% soak with ethanol 2 ~ 8h, boil and extract 1 ~ 2h, lixiviating solution is poured out and sucking filtration to obtain supernatant for subsequent use, merge supernatant, concentrating under reduced pressure obtains extractum; By 2 extract extractum mixing for standby use.Ethyl acetate extract and ethanol extract are mixed, be prepared into soluble powder, granule, tablet or oral liquid with conventional formulation.
The preparation of described Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi compositions can be applicable to preparation prevention and treatment animal viral disease or bacterial disease medicine.
Wherein, livestock and poultry virus comprises bird flu virus, bursal disease virus and renal type infectious bronchitis.Bird flu virus is H5, H9 subtype avian influenza virus; Bursal disease virus is BCLX strain; Renal type infectious bronchitis is strain.
Above-mentioned preparation according to every day 100mg/kg ~ 400mg/kg body weight to poultry, mammal administration, continuous 4 ~ 5 days.
Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi compositions of the present invention can be for the control of the viral disease of livestock and poultry breeding industry; making up veterinary vaccines immunoprotection deficiency and immuning failure, strain virulence strengthens the virosis outburst causing and distributes; for livestock and poultry cultivation provides technical support, improve the competitiveness of China's livestock and poultry breeding industry products export.Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi compositions of the present invention, its raw material be easy to get and cost lower, good effect, the features such as noresidue and environmental protection, can instead of chemical class antiviral drugs.
Detailed description of the invention
Following examples are used for illustrating the present invention, but are not used for limiting the scope of the invention.The preparation of embodiment 1 granule of the present invention
(1) take Rhizoma Dryopteris Crassirhizomatis 4g, Fructus Crataegi 6g, and be ground into respectively coarse powder.
(2) get the Rhizoma Dryopteris Crassirhizomatis coarse powder that step (1) makes, with 4 times of volumes of acetic acid ethyl esters immersion 24h, collect lixiviating solution, reclaim ethyl acetate, obtain extractum; Medicinal residues add 4 times of 75% soak with ethanol 2h, boil and extract 1h, extract 2 times, and merging filtrate, by above-mentioned filtrate rotary evaporation, concentrating under reduced pressure obtains extractum.By the extractum mixing for standby use of 2 collections.
(3) get the Fructus Crataegi coarse powder that step (1) makes, with after 4 times of volumes of acetic acid ethyl esters immersion 24h, reclaim ethyl acetate, obtain extractum; Medicinal residues are with after 4 times of volumes, 75% soak with ethanol 2h, boil and extract 2h, lixiviating solution is poured out and sucking filtration to obtain supernatant for subsequent use, merge supernatant, concentrating under reduced pressure obtains extractum; By 2 extract extractum mixing for standby use.
(4) getting Rhizoma Dryopteris Crassirhizomatis extract and Fructus Crataegi extract extractum mixes.
(5) get extractum: sucrose: beta cyclodextrin=1:1:2 fully mixes, prepare granule by wet granulation mode, dry and granulate, obtain Rhizoma Dryopteris Crassirhizomatis granule.
The preparation of embodiment 2 soluble powders of the present invention
(1) take Rhizoma Dryopteris Crassirhizomatis 4g, Fructus Crataegi 6g, and be ground into respectively coarse powder.
(2) get the Rhizoma Dryopteris Crassirhizomatis coarse powder that step (1) makes, with 6 times of volumes of acetic acid ethyl esters immersion 48h, collect lixiviating solution, reclaim ethyl acetate, obtain extractum; Medicinal residues add 4 times of 85% soak with ethanol 4h, boil and extract 2h, extract 2 times, and merging filtrate, by above-mentioned filtrate rotary evaporation, concentrating under reduced pressure obtains extractum.By the extractum mixing for standby use of 2 collections.
(3) get the Fructus Crataegi coarse powder that step (1) makes, with after 6 times of volumes of acetic acid ethyl esters immersion 48h, reclaim ethyl acetate, obtain extractum; Medicinal residues are with after 4 times of volumes, 95% soak with ethanol 4h, boil and extract 2h, lixiviating solution is poured out and sucking filtration to obtain supernatant for subsequent use, merge supernatant, concentrating under reduced pressure obtains extractum; By 2 extract extractum mixing for standby use.
(4) get Rhizoma Dryopteris Crassirhizomatis extract and Fructus Crataegi extract extractum is placed in 100ml container, add normal hexane 5ml, tween 20 solution 5ml, be prepared into Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi solution 20ml; Meanwhile, take 4g beta cyclodextrin and be dissolved in water; Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi solution, beta cyclodextrin solution are mixed, stir 3 ~ 5min at 3000 revs/min; After drying, pulverize, obtain Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi powder.
The preparation of embodiment 3 tablets of the present invention
(1) take Rhizoma Dryopteris Crassirhizomatis 5g, Fructus Crataegi 5g, and be ground into respectively coarse powder.
(2) get the Rhizoma Dryopteris Crassirhizomatis coarse powder that step (1) makes, with 8 times of volumes of acetic acid ethyl esters immersion 24h, collect lixiviating solution, reclaim ethyl acetate, obtain extractum; Medicinal residues add 8 times of 95% soak with ethanol 2h, boil and extract 1h, extract 2 times, and merging filtrate, by above-mentioned filtrate rotary evaporation, concentrating under reduced pressure obtains extractum.By the extractum mixing for standby use of 2 collections.
(3) get the Fructus Crataegi coarse powder that step (1) makes, with after 8 times of volumes of acetic acid ethyl esters immersion 24h, reclaim ethyl acetate, obtain extractum; Medicinal residues are with after 8 times of volumes, 95% soak with ethanol 2h, boil and extract 2h, lixiviating solution is poured out and sucking filtration to obtain supernatant for subsequent use, merge supernatant, concentrating under reduced pressure obtains extractum; 2 extract extractum are mixed.
(4) getting Rhizoma Dryopteris Crassirhizomatis extract and Fructus Crataegi extract extractum mixes.
(5) get extractum: beta cyclodextrin=1:4 fully mixes, prepare granule by wet granulation mode, dry, compacting in flakes.
The preparation of embodiment 4 oral liquids of the present invention
(1) take Rhizoma Dryopteris Crassirhizomatis 2g, Fructus Crataegi 8g, and be ground into respectively coarse powder.
(2) get the Rhizoma Dryopteris Crassirhizomatis coarse powder that step (1) makes, with 4 times of volumes of acetic acid ethyl esters immersion 24h, collect lixiviating solution, reclaim ethyl acetate, obtain extractum; Medicinal residues add 4 times of 95% soak with ethanol 2h, boil and extract 1h, extract 2 times, and merging filtrate, by above-mentioned filtrate rotary evaporation, concentrating under reduced pressure obtains extractum.By the extractum mixing for standby use of 2 collections.
(3) get the Fructus Crataegi coarse powder that step (1) makes, with after 4 times of volumes of acetic acid ethyl esters immersion 24h, reclaim ethyl acetate, obtain extractum; Medicinal residues are with after 4 times of volumes, 75% soak with ethanol 2h, boil and extract 2h, lixiviating solution is poured out and sucking filtration to obtain supernatant for subsequent use, merge supernatant, concentrating under reduced pressure obtains extractum; By 2 extract extractum mixing for standby use.
(4) get Rhizoma Dryopteris Crassirhizomatis extract and Fructus Crataegi extract extractum is placed in 100ml container, add normal hexane 5ml, tween 20 solution 5ml, be prepared into Rhizoma Dryopteris Crassirhizomatis haw oral liquid 20ml.
The preparation of embodiment 5 granules of the present invention
The another kind of preparation method of granule: take Rhizoma Dryopteris Crassirhizomatis 1g, Fructus Crataegi 9g, pulverize, mix, add the ethyl acetate of 8 times of amounts, lixiviate 48h, collects lixiviating solution, reclaims ethyl acetate, obtains extractum; Medicinal residues add 4 times of volumes, 95% soak with ethanol 8h, boil 2h, and extracting solution sucking filtration is obtained to supernatant, merge lixiviating solution and supernatant and revolve steaming, and concentrating under reduced pressure, obtains extractum.The extractum that two kinds of organic solvents are obtained mixes.Get extractum: sucrose: beta cyclodextrin=1:1:2 fully mixes, prepare granule by wet granulation mode, mixed dry, obtain compound recipe Rhizoma Dryopteris Crassirhizomatis granule.
The preparation of embodiment 6 soluble powders of the present invention
The another kind of preparation method of soluble powder: take Rhizoma Dryopteris Crassirhizomatis 5g, Fructus Crataegi 5g, pulverize, mix, add the ethyl acetate of 8 times of amounts, lixiviate 48h, collects lixiviating solution, reclaims ethyl acetate, obtains extractum; Medicinal residues add 8 times of volumes, 95% soak with ethanol 8h, boil 2h, and extracting solution sucking filtration is obtained to supernatant, merge lixiviating solution and supernatant and revolve steaming, and concentrating under reduced pressure, obtains extractum 1.7 ~ 2.4g.The extractum mixing that two kinds of organic solvents are obtained is placed in 100ml container, adds 95% ethanol 20ml, tween 20 solution 20ml, is prepared into Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi solution 40ml; Meanwhile, take 4g beta cyclodextrin and be dissolved in water; Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi solution, beta cyclodextrin solution are mixed, stir 3 ~ 5min at 3000 revs/min; After drying, pulverize, obtain compound recipe Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi soluble powder.
The preparation of embodiment 7 granules of the present invention
According to the method described in embodiment 1, difference is to select the raw material of following weight: Rhizoma Dryopteris Crassirhizomatis 9g, Fructus Crataegi 1g.
The preparation of embodiment 8 granules of the present invention
According to the method described in embodiment 1, difference is to select the raw material of following weight: Rhizoma Dryopteris Crassirhizomatis 8g, Fructus Crataegi 2g.
The preparation of embodiment 9 granules of the present invention
According to the method described in embodiment 1, difference is to select the raw material of following weight: Rhizoma Dryopteris Crassirhizomatis 7g, Fructus Crataegi 3g.
The preparation of embodiment 10 granules of the present invention
According to the method described in embodiment 1, difference is to select the raw material of following weight: Rhizoma Dryopteris Crassirhizomatis 6g, Fructus Crataegi 4g.
The preparation of embodiment 11 granules of the present invention
According to the method described in embodiment 1, difference is to select the raw material of following weight: Rhizoma Dryopteris Crassirhizomatis 5g, Fructus Crataegi 5g.
The preparation of embodiment 12 granules of the present invention
According to the method described in embodiment 1, difference is to select the raw material of following weight: Rhizoma Dryopteris Crassirhizomatis 3g, Fructus Crataegi 7g.
The preparation of embodiment 13 granules of the present invention
According to the method described in embodiment 1, difference is to select the raw material of following weight: Rhizoma Dryopteris Crassirhizomatis 2g, Fructus Crataegi 8g.
Further illustrate below the drug action of compositions of the present invention by test example.
The different proportioning compositions treatment of test example 1 Nephropathogenic infectious bronchitis effect test
1 materials and methods
1.1 laboratory animals: 21 age in days SPF chickling are divided into 11 groups at random,, amount to 110 by 10 every group.
1.2 medicine groupings: Rhizoma Osmundae Fructus Crataegi 9:1 group (embodiment 7), Rhizoma Osmundae Fructus Crataegi 8:2 group (embodiment 8), Rhizoma Osmundae Fructus Crataegi 7:3 group (embodiment 9), Rhizoma Osmundae Fructus Crataegi 6:4 group (embodiment 10), Rhizoma Osmundae Fructus Crataegi 5:5 group (embodiment 11), Rhizoma Osmundae Fructus Crataegi 4:6(embodiment 1), Rhizoma Osmundae Fructus Crataegi 3:7(embodiment 12), Rhizoma Osmundae Fructus Crataegi 2:8(embodiment 13), Rhizoma Osmundae Fructus Crataegi 1:9(embodiment 5), astragalus polysaccharides matched group and infect matched group.Above medicine 1-10 group all gives medicine 200mg/kg body weight.Astragalus polysaccharides powder, purchased from Beijing Centre Biology Co., Ltd., lot number 20100605.
1.3 strains and counteracting toxic substances: infectious bronchitis virus BJ1 strain, associate professor Wang Jingyu of northwest agricultural university give (Wang Jingyu, etc. the Isolation and Identification of. Nephropathogenic infectious bronchitis virus 18 strains. Chinese veterinary's magazine, 2006,42:19-20).The ELD of 10 age in days SPF Embryo Gallus domesticus
50for 107.0/0.2ml.10 age in days SPF chicken LD
50=103ELD
50/ 0.2ml.Every chickling eye dripping, collunarium are total to 0.2mL.After counteracting toxic substances, observe chicken mental status, wait to fall ill three/for the moment, start medication, use continuously 5 days.
1.4 evaluation indexes:
Clinical onset feature and body weight: spirit, appetite, activity and the feces situation of duration of test viewing test every day chicken, and make a record, in the time of on-test and end, test chicken, by only weighing, is calculated to the relative weight gain rate.
Nephropathy is of science to be changed: administration, after 5 days, continues to observe after 3 days, and survival chicken is implemented euthanasia, measures kidney swelling number/observation kidney.
Table 1 compositions and drug dose situation
| Group | Medicine preparation method | Chicken | Dosage (mg/Kg) | Administering mode |
| Pass through mountain 9:1 | Embodiment 7 | 10 | 200 | Oral administration 5 days |
| Pass through mountain 8:2 | Embodiment 8 | 10 | 200 | Oral administration 5 days |
| Pass through mountain 7:3 | Embodiment 9 | 10 | 200 | Oral administration 5 days |
| Pass through mountain 6:4 | Embodiment 10 | 10 | 200 | Oral administration 5 days |
| Pass through mountain 5:5 | Embodiment 11 | 10 | 200 | Oral administration 5 days |
| Pass through mountain 4:6 | Embodiment 1 | 10 | 200 | Oral administration 5 days |
| Pass through mountain 3:7 | Embodiment 12 | 10 | 200 | Oral administration 5 days |
| Pass through mountain 2:8 | Embodiment 13 | 10 | 200 | Oral administration 5 days |
| Pass through mountain 1:9 | Embodiment 5 | 10 | 200 | Oral administration 5 days |
| Astragalus polysaccharides contrast | 10 | 200 | Oral administration 5 days | |
| Infect contrast | 10 | Normal saline | Oral administration 5 days |
2 experimental results:
The impact of 2.1 different components on body weight and survival rate
Different components the relative weight gain rate is respectively 45.8%, 63.9%, 69.1%, 81.9%, 48.6%, 95.8%, 83.3%, 44.4% and 47.2%.All there is significant difference (P<0.01) with astragalus polysaccharides matched group (compared with 62.5%, can find Rhizoma Osmundae Fructus Crataegi 6:4 group (embodiment 10), Rhizoma Osmundae Fructus Crataegi 4:6 group (embodiment 1), Rhizoma Osmundae Fructus Crataegi 3:7(embodiment 12).The results are shown in Table 2.
The impact of table 2 compositions on chicken body weight
Note: body weight before body weight-test after average weight gain=test; The relative weight gain rate=test group average weight gain/infection matched group average weight gain × 100%.
Aspect survival rate, the results are shown in Table 3.With astragalus polysaccharides be reference object, exceed being combined as of astragalus polysaccharides survival rate: Rhizoma Osmundae Fructus Crataegi 4:6 group (embodiment 1) 90.0%, Rhizoma Osmundae Fructus Crataegi 3:7 group (embodiment 12) 80.0%; The compositions that approaches astragalus polysaccharides group is Rhizoma Osmundae Fructus Crataegi 6:4 group (embodiment 10), the results are shown in Table 3.
Investigate from kidney swelling rate aspect, swelling rate lower than or approach 40% mainly contain Rhizoma Osmundae Fructus Crataegi 6:4 group (embodiment 10), Rhizoma Osmundae Fructus Crataegi 5:5 group (embodiment 11) and Rhizoma Osmundae Fructus Crataegi 4:6 group (embodiment 1), the results are shown in Table 3.
Table 3 pharmaceutical composition on artificial challenge after the impact of survival rate and pathological changes
| Group | Chicken | Survival number | Survival rate % | Kidney swelling rate % |
| Embodiment 7 | 10 | 2 | 20.0 | 50.0 |
| Embodiment 8 | 10 | 1 | 10.0 | 100.0 |
| Embodiment 9 | 10 | 6 | 60 | 66.6 |
| Embodiment 10 | 10 | 7 | 70 | 40.0 |
| Embodiment 11 | 10 | 6 | 60.0 | 33.3 |
| Embodiment 1 | 10 | 9 | 90.0 | 22.2 |
| Embodiment 12 | 10 | 8 | 80.0 | 37.5 |
| Embodiment 13 | 10 | 6 | 60.0 | 50.0 |
| Embodiment 5 | 10 | 5 | 50.0 | 60.0 |
| Astragalus polysaccharides contrast | 10 | 7 | 70.0 | 42.8 |
| Infect contrast | 10 | 4 | 40.0 | 75.0 |
| Normal healthy controls | 10 | 10 | 100.0 | 0 |
Note: survival kidney swelling number/observation kidney quantity x100
3 conclusions:
After artificial challenge SPF Nephropathogenic infectious bronchitis virus, successfully copy typical renal type infectious bronchitis model.The index of comprehensive proportion of weight to height, survival rate and three aspects of kidney swelling, Rhizoma Osmundae Fructus Crataegi 4:6 group (embodiment 1) curative effect is best, below in test, all furthers investigate with this compositions.
Test example 2 compositions treatment Nephropathogenic infectious bronchitis effects
Object: the renal type infectious bronchitis of evaluating composition granule treatment artificial challenge
1 materials and methods
1.1 laboratory animals: 21 Japanese instar chicklings are divided into 6 groups at random, 15 every group.
1.2 medicine groupings: composition grain high dose group (400mg/kg), middle dosage group (200mg/kg), low dose group (100mg/kg), infection matched group, astragalus polysaccharides matched group and infection contrast.Astragalus polysaccharides powder, purchased from Beijing Centre Biology Co., Ltd., lot number 20100605; It is trial drug that embodiment 1 prepares granule.
1.3 strains and counteracting toxic substances: infectious bronchitis virus BJ1 strain, associate professor Wang Jingyu of northwest agricultural university give (Wang Jingyu, etc. the Isolation and Identification of. Nephropathogenic infectious bronchitis virus 18 strains. Chinese veterinary's magazine, 2006,42:19-20).The ELD of 10 age in days SPF Embryo Gallus domesticus
50for 107.0/0.2ml.10 age in days SPF chicken LD
50=103ELD
50/ 0.2ml.Every chickling eye dripping, collunarium are total to 0.2mL.After counteracting toxic substances, observe chicken mental status, wait to fall ill three/for the moment, start medication, use continuously 5 days.
1.4 evaluation indexes:
1.4.1 clinical onset feature and body weight: spirit, appetite, activity and the feces situation of duration of test viewing test every day chicken, and make a record, in the time of on-test and end, test chicken, by only weighing, is calculated to the relative weight gain rate.
1.4.2 kidney is observed the index of pathological changes: medication finishes latter the 7th day, utilizes euthanasia mode to put to death survival chicken.Cut open inspection, and observe one by one kidney and bronchial pathological changes situation.Primary part observation nephropathy, whether enlargement is hemorrhage, is piebaldism kidney, whether occurs white urate deposition.Nephropathy index is: 1 point-one-sided single-row kidney swelling; 2 points-one-sided 2 row kidney swelling; 3 points-one-sided 3 row kidney swelling; The 2 row kidney swelling of 4 points-bilateral; The 3 row kidney swelling of 6 points-bilateral.
Table 4 test grouping and drug dose situation
| Grouping | Chicken | Dosage (mg/kg) | Administering mode |
| Embodiment 1 high dose group | 15 | 400mg/kg | After counteracting toxic substances, continuous oral administration 5 days |
| Dosage group in embodiment 1 | 15 | 200mg/kg | After counteracting toxic substances, continuous oral administration 5 days |
| Embodiment 1 low dose group | 15 | 100mg/kg | After counteracting toxic substances, continuous oral administration 5 days |
| Astragalus polysaccharides group | 15 | 0.1g/ only | After counteracting toxic substances, continuous oral administration 5 days |
| Infect matched group | 15 | - | Infect not administration |
| Blank group | 15 | - | Do not infect not administration |
2 experimental results
2.1 impacts of Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi various dose on SPF chicken rate of body weight gain
Aspect rate of body weight gain, Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi high dose group, middle dosage group, low dose group are respectively 45.8%, 126.9% and 69.0%, during statistical analysis shows, dosage group significance is higher than high dose group and low dose group (P<0.05), and utmost point significance is higher than astragalus polysaccharides group (P<0.01).The results are shown in Table 5.
Table 5 test grouping and drug dose situation
Note: body weight before body weight-test after average weight gain=test; The relative weight gain rate=test group average weight gain/infection matched group average weight gain × 100%.
2.2 impacts of different prescriptions on survival rate and nephropathy
Aspect survival rate, as can be seen from Table 6, Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi high dose group, middle dosage group, low dose group are respectively 80.0%, 93.3% and 86.6%, and astragalus polysaccharides matched group and infection matched group are respectively 66.6% and 53.3%.Statistical analysis discovery, high dose group and low dose group difference are not remarkable, but middle dosage group significance is higher than high and low dose group and astragalus polysaccharides matched group.In addition, cut open inspection after Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi high dose group, middle dosage group, three dosage group nephropathy indexes of low dose group be respectively 6.5,4.1 and 5.9, significance is lower than astragalus polysaccharides group nephropathy index 8.5.
After table 6 drug oral to survival rate and nephropathy variable effect of science
3 conclusions:
Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi can effectively be treated the infection that Nephropathogenic infectious bronchitis virus causes in oral 5 days with 200mg/kg, and effect is better than the astragalus polysaccharides of 250mg/kg.
The treatment verification the verifying results of test example 3 compositionss to SPF artificial challenge bursal disease virus
Object: the therapeutic effect of inspection Rhizoma Dryopteris Crassirhizomatis haw thorn granule to artificial challenge's bursal disease virus
1 materials and methods
1.1 animals: 21 Japanese instar chicklings are divided into 6 groups at random, 15 every group.
1.2 infect strain and dosage: artificial challenge's strain IBDV LX strain (Academy of Agricultural Sciences of Beijing animal and veterinary institute), every chickling eye dripping of tissue homogenate, collunarium 0.2mL altogether.After 24h, start medication, use continuously 5 days.
1.3 medicine grouping and administering modes: be divided into 6 groups: infection group and viral infection group, blank group, astragalus polysaccharides matched group (being positive drug matched group), Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi high dose group, middle dosage group and low dose group, every group of only SPF chicken of 24 ages in days of 10-15.Administering mode: water way administration; In table 4.
Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi high dose group, gives the granule of embodiment 1,400mg/kg;
Dosage group in Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi, gives the granule of embodiment 1,200mg/kg;
Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi low dose group, gives the granule of embodiment 1,100mg/kg;
Astragalus polysaccharides group, gives astragalus polysaccharides, 250mg/kg;
Infection group and viral infection group, gives diluent (0.9% normal saline solution) group, a counteracting toxic substances, not administration;
Blank group: not counteracting toxic substances, also not administration.
1.4 evaluation indexes:
1.4.1 survival rate and weight gain index: duration of test, every day viewing test chicken spirit, appetite, activity and feces situation, and make a record, in on-test with while finishing, test chicken, by only weighing, is calculated to the relative weight gain rate.The relative weight gain rate=(the front body weight of body weight-test after the test group)/blank group test forebody-afterbody method of double differences × 100%.
1.4.2 pathological changes index assessment: medication finishes to cut open for latter 5 days inspection, observes chest muscle lower limb myopathy and becomes situation, measures fabricius bursa weight, measures its pathological changes index.
Table 7 test grouping and drug dose situation
| Grouping | Chicken | Dosage (mg/kg) | Administering mode |
| Embodiment 1 high dose group | 15 | 400mg/kg | After counteracting toxic substances, continuous oral administration 5 days |
| Dosage group in embodiment 1 | 15 | 200mg/kg | After counteracting toxic substances, continuous oral administration 5 days |
| Embodiment 1 low dose group | 15 | 100mg/kg | After counteracting toxic substances, continuous oral administration 5 days |
| Astragalus polysaccharides powder | 15 | 0.5mL/ only | Intramuscular injection, continuous 2 days |
| Infect matched group | 15 | 0.2mL/ only | Infect not administration |
| Blank group | 10 | - | Do not infect not administration |
2 results
The impact of Drug therapy on chicken body weight after 2.1 artificial challenge's fabricius bursa
The results are shown in Table 8, Rhizoma Dryopteris Crassirhizomatis haw thorn granule high dose, middle dosage group and low dose group rate of body weight gain are respectively 85.2%, 88.8% and 79.6%, astragalus polysaccharides matched group is 76.3%, through statistical analysis, in Rhizoma Dryopteris Crassirhizomatis haw thorn granule, dosage group significance is higher than low dose group and astragalus polysaccharides group, but compared with high dose group, difference is not remarkable.
The impact of medicine on chicken organ pathology index after 2.2 artificial challenge's fabricius bursa
After artificial challenge's fabricius bursa virulent strain LX, after infection chicken 36h, there is symptom, comprise necking down, close one's eyes, stand transfixed to the ground and draw white loose stool.After counteracting toxic substances there is death in 48h, and 96h reaches peak mortality.Rhizoma Dryopteris Crassirhizomatis haw thorn granule high dose, middle dosage group and low dose group survival rate are respectively 80.0%, 86.6% and 80.0%, and astragalus polysaccharides group is 66.6%.Statistical analysis shows that between three dosage groups of Rhizoma Dryopteris Crassirhizomatis haw thorn granule, difference is not remarkable, but significance is higher than astragalus polysaccharides group.The results are shown in Table 9.
Aspect the variation of science of target organ-bursal disease, the high, medium and low dosage group of Rhizoma Dryopteris Crassirhizomatis haw thorn granule pathological changes suppression ratio is respectively 66.6%, 73.3% and 60.0%, and significance is higher than astragalus polysaccharides group (50.0%).
In sum, the Rhizoma Dryopteris Crassirhizomatis haw thorn granule of embodiments of the invention 1 can reduce bursal disease varying index, improves survival rate and improves weightening finish, demonstrates certain anti-fabricius bursa effect.
Table 8 compositions treatment fabricius bursa infects body weight situation of change
Survival rate and pathological changes index result after the treatment of table 9 compositions
Note: bursal disease varying index: 4 points of severe haemorrhages; 3 points of moderates; Slight 1 point; Pathological changes slip relatively=(infecting contrast-experimental group pathological changes index)/infection matched group pathological changes index × 100%
3, conclusion:
Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi, with 200mg/kg body weight continuous oral 5 days, can effectively be improved the survival rate of artificial challenge's fabricius bursa chicken, improves the relative weight gain rate, reduces pathological changes index.Can substitute the infection of astragalus polysaccharides control bursal disease virus.
Test example 4 compositions treatment artificial challenge SPF avian influenza virus H5N1 therapeutic tests
Object: determine whether Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi compositions has treatment H 5 N 1 avian influenza effect
1 materials and methods
1.1 laboratory animals: 5 week age 90 of SPF chickens, purchased from Jinan Si Pafasi poultry company limited.Raise in negative pressure isolator.
1.2 medicine groupings: dosage group, compositions low dose group, astragalus polysaccharides group, infection matched group and blank group in compositions high dose group, compositions.Amantadine matched group, content 99%, purchased from Johnson Matthey Company, USA, Lot:10118375.Each dosage group drug dose is in table 7.
Table 10 test grouping and processing
| Grouping | Quantity (only) | Administration concentration | Route of administration | Administration number of times |
| Embodiment 2 high dose group | 15 | 400mg/kg | Oral | 2/ day |
| Dosage group in embodiment 2 | 15 | 200mg/kg | Oral | 2/ day |
| Embodiment 2 low dose group | 15 | 100mg/kg | Oral | 2/ day |
| Amantadine matched group | 15 | 250mg/kg | Oral | 2/ day |
| Virus control group | 15 | 0 | - | - |
| Blank group | 15 | 0 | - | - |
1.3 strains and counteracting toxic substances avian influenza A/Chicken/Hebei/126/2005 (He Guimei, Deng .Amantadine-resistance among H5N1 avian influenza viruses isolated in Northern China.Antiviral Research, 2008,77:7276).By viral dilution, to 10-5, nasal cavity gives 0.1ml, has 30% appearance to play listless rear beginning administration, and continuous use uses 5 days.
1.4 evaluation indexes:
1.4.1 survival rate and weight gain index: duration of test, every day viewing test chicken spirit, appetite, active situation, makes a record.In the time of on-test and end, test chicken is weighed.Calculate the relative weight gain rate, survival rate.
1.4.2 virus isolated rate: after counteracting toxic substances the 9th day, the chicken of survival was implemented euthanasia, aseptic chicken liver, the lungs taked, inoculation 9-10 age in days SPF Embryo Gallus domesticus, measures its viral level.
2 results
The body weight impact of 2.1 compositionss on SPF chicken artificial challenge bird flu
After infecting by collunarium, after artificial challenge's H 5 N 1 avian influenza hypotype, compositions high dose group, middle dosage group and low dose group the relative weight gain rate are respectively 44.0%, 57.38% and 16.8%, and astragalus polysaccharides matched group is 30.1%.In statistical analysis compositions, dosage group significance is higher than high dose group and amantadine matched group (P<0.05), and utmost point significance is higher than low dose group (P<0.01).
The impact of SPF chicken rate of body weight gain in the therapeutic test of table 11 compositions
2.2 compositionss are on SPF chicken artificial challenge bird flu survival rate and the impact of time-to-live
After the strong poison of artificial challenge, compositions high dose group, middle dosage group, low dose group survival rate are respectively 73.3%, 80.0% and 73.3%, and astragalus polysaccharides matched group is 53.3%.On the time-to-live, three dosage group mean survival times of compositions are 7.9 days, 8.5 days and 8.4 days, and amantadine matched group is 6.3 days.
The impact of table 12 compositions on artificial challenge SPF adult livability and time-to-live
2.3 compositionss separate with liver organization virus SPF chicken artificial challenge bird flu lungs
Compositions high dose group, middle dosage group, low dose group lungs viral level 102.2ELD respectively after the strong poison of artificial challenge
50/ 0.1ml, 101.2ELD
50/ 0.1ml and 101.9ELD
50/ 0.1ml, liver viral level is 102.1ELD
50/ 0.1ml, 100.9ELD
50/ 0.1ml and 102.4ELD
50/ 0.1ml.In obvious lungs and liver organization, viral residual quantity significance is lower than amantadine matched group.
Table 13 compositions is organized the impact of virus isolated rate on lungs
| Grouping | Quantity | Lungs viral level | Liver viral level |
| Embodiment 2 high dose group | 5 | 102.2ELD 50/0.1ml | 102.1ELD 50/0.1ml |
| Dosage group in embodiment 2 | 5 | 101.2ELD 50/0.1ml | 100.9ELD 50/0.1ml |
| Embodiment 2 low dose group | 5 | 101.9ELD 50/0.1ml | 102.4ELD 50/0.1ml |
| Amantadine matched group | 5 | 104.3ELD 50/0.1ml | 104.1ELD 50/0.1ml |
| Virus control group | 5 | 105.7ELD 50/0.1ml | 106.3ELD 50/0.1ml |
| Blank group | 5 | 0 | 0 |
3 conclusions:
This result of the test shows after artificial challenge high pathogenic avian influenza, and compositions is with the oral SPF chicken of 200mg/kg, continuous oral 5 days, and the survival rate of chicken reaches 80%, and significance is higher than amantadine matched group.
The therapeutic test of test example 5 compositionss to BALB/C mice H5N1 influenza virus
Object: determine whether compositions has treatment mice H 5 N 1 avian influenza effect
1 materials and methods
Each 60 of male, the female BALB/C mice of 1.1 animals: 18g ~ 22g, is divided into 6 groups at random, and 20 every group, wherein each 10 of male and female.
1.2 infect strain and dosage: artificial challenge H5N1A/Chicken/Hebei/126/2005 (He Guimei, Deng .Amantadine-resistance among H5N1 avian influenza viruses isolated in Northern China.Antiviral Research, 2008,77:72-76).Embryo Gallus domesticus ELD50 allantoic fluid eye dripping, collunarium 0.2mL altogether.After having 1/3rd morbidities, start administration, use continuously 5 days.
1.3 medicine groupings: compositions high dose group, middle dosage group and low dose group, amantadine matched group, infect matched group and blank group.Random every group of 20 mices, the situation of specifically dividing into groups and process as shown in table 14.
Table 14 test grouping and processing
| Grouping | Quantity (only) | Administration concentration | Route of administration | Administration number of times |
| Embodiment 2 high dose group | 20 | 400mg/kg | Oral | 2/ day |
| Dosage group in embodiment 2 | 20 | 200mg/kg | Oral | 2/ day |
| Embodiment 2 low dose group | 20 | 100mg/kg | Oral | 2/ day |
| Amantadine matched group | 20 | 50mg/kg | Oral | 2/ day |
| Virus control group | 20 | 0 | - | - |
| Blank group | 20 | 0 | - | - |
1.4 evaluation indexes:
1.4.1 clinical indices: the initial body weight and the opisthosoma weight that take respectively every mice; Observe M & M; Medication finishes to cut open inspection after latter 5 days, gets lung, and observes lung one by one, liver pathological changes situation.
The relative weight gain rate=the relative weight gain/normal healthy controls weight gain × 100%; Mean survival time=total survival natural law/number of elements
1.4.2 survival mice lungs, liver bird flu virus content: each group of aseptic liver, the lungs of getting 5 mices respectively, grind respectively, grind centrifugal liquid and add gentamycin, 4000 units/ml, 37 DEG C of effects 30 minutes; Do titre dilution (10
-1, 10
-2, 10
-3, 10
-4, 10
-5) inoculation, 3 pieces of Embryo Gallus domesticus of each dilution factor inoculation, 120h observes Embryo Gallus domesticus survival continuously, records dead Embryo Gallus domesticus quantity; After 120h, collect the allantoic fluid of survival Embryo Gallus domesticus, measure its coagulation erythrocyte feature.
2 experimental results
Impact on Mouse Weight after 2.1 artificial challenge's bird flus
From rate of body weight gain, high, the middle dosage group of Rhizoma Dryopteris Crassirhizomatis and low dose group the relative weight gain rate are respectively 20.7%, 68.1% and 85.6%, amantadine matched group is 41.4%, through statistical analysis, in Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi, dosage and low dose group are significantly higher than amantadine matched group (P<0.05), and utmost point significance is higher than high dose group (P<0.01), still, in Rhizoma Dryopteris Crassirhizomatis, dosage group, low dose group are compared, and difference is not remarkable.
The impact of table 15 compositions on mice weightening finish
| Grouping | Just body weight (g) | Opisthosoma heavy (g) | The relative weight gain (g) | The relative weight gain rate (%) |
| Embodiment 2 high doses | 14.23±0.90 | 14.75±3.12 | 0.52 | 20.7 a |
| Dosage in embodiment 2 | 14.20±1.07 | 15.91±4.13 | 1.71 | 68.1 b |
| Embodiment 2 low dosages | 13.10±0.75 | 15.25±2.19 | 2.15 | 85.6 b |
| Amantadine group | 14.10±1.08 | 15.14±1.61 | 1.04 | 41.4 c |
| Infect contrast | 13.38±0.83 | 13.05±1.63 | -0.33 | -13.1 |
| Blank | 14.16±0.22 | 16.67±0.7 | 2.51 | 100.0 |
Impact on mice survival rate and time-to-live after 2.2 artificial challenge's bird flus
As can be seen from Table 16, combination object height, middle dosage group and low dose group, amantadine contrast, infection contrast, blank survival rate are respectively 70.0%, 85.0% and 80.0%, and amantadine is 55.0%.From mean survival time, combination object height, middle dosage group and low dose group are respectively 8.2,9.2 and 8.5 days, and significance is better than amantadine matched group (6.5 days).
The impact of table 16 compositions on mice survival rate and mean survival time
The impact of the lungs index on mice after 2.3 artificial challenge's bird flus
Table 17 shows that compositions weighs BALB/c mouse artificial challenge H5N1 virus model mouse lung and the impact of pneumonopathy varying index: compositions high dose group, middle dosage group and low dose group lungs index are respectively 0.88%, 0.69% and 0.92%, and significance is lower than amantadine matched group.But between compositions high dose group, low dose group, difference is not remarkable.
The impact of table 17 compositions on mice lungs index
| Grouping | Measure quantity (only) | Lungs weight (g) | Lungs index (%) |
| Embodiment 2 high doses | 8 | 0.13±0.013 | 0.88±0.05 |
| Dosage in embodiment 2 | 8 | 0.11±0.007 | 0.69±0.12 |
| Embodiment 2 low dosages | 8 | 0.14±0.007 | 0.92±0.08 |
| Amantadine matched group | 8 | 0.18±0.014 | 1.18±0.11 |
| Infect contrast | 8 | 0.26±0.024 | 1.99±0.13 |
| Blank | 8 | 0.11±0.007 | 0.66±0.03 |
Note: lung index=lung weight/body weight; Know and be not all significant difference (P<0.05) with the medium and small female mark of writing of string, be not all extremely significantly (P<0.01) of difference with capitalization mark in string, be all mutually not remarkable (P>0.05) of difference with letter mark in string.
After 2.4 administrations, the lungs of mice and liver bird flu virus are measured
After artificial challenge mice high pathogenic avian influenza, compositions high dose group, middle dosage group, low dose group lungs viral level 100.2ELD respectively
50/ 0.1ml, 100.3ELD
50/ 0.1ml and 100.9ELD
50/ 0.1ml, liver viral level is 100.5ELD
50/ 0.1ml, 100.2ELD
50/ 0.1ml and 100.6ELD
50/ 0.1ml.In obvious lungs and liver organization, viral residual quantity significance is lower than amantadine matched group (103.1ELD
50/ 0.1ml).The results are shown in Table 18.
Table 18 compositions is organized the impact of virus isolated rate on mice lungs
| Grouping | Quantity | Lungs viral level | Liver viral level |
| Embodiment 2 high dose group | 5 | 100.2ELD 50/0.1ml | 100.5ELD 50/0.1ml |
| Dosage group in embodiment 2 | 5 | 100.3ELD 50/0.1ml | 100.2ELD 50/0.1ml |
| Embodiment 2 low dose group | 5 | 100.9ELD 50/0.1ml | 100.6ELD 50/0.1ml |
| Amantadine matched group | 5 | 102.5ELD 50/0.1ml | 103.1ELD 50/0.1ml |
| Virus control group | 5 | 103.3ELD 50/0.1ml | 104.3ELD 50/0.1ml |
| Blank group | 5 | 0 | 0 |
3 conclusions:
Compositions is oral with 200mg/kg body weight, and continuous 5 days, can effectively improve the survival rate of the rear mice of bird flu infection, can reduce the infection of virus at lungs and liver organization simultaneously.
Claims (5)
1. treat a pharmaceutical composition for Nephropathogenic infectious bronchitis, infectious bursal disease, its effective ingredient is to be made up of the raw material of following weight portion: 20～80 parts of 20～80 parts of Rhizoma Dryopteris Crassirhizomatiss and Fructus Crataegis;
The preparation method of described compositions comprises the following steps:
1) take by weight Rhizoma Dryopteris Crassirhizomatis, Fructus Crataegi, and pulverize respectively, use ethyl acetate lixiviate, extracting solution concentrating under reduced pressure, obtains ethyl acetate extract;
2) 75%～95% soak with ethanol for medicinal residues after lixiviate, boils, and by lixiviating solution sucking filtration, concentrating under reduced pressure evaporate to dryness, obtains ethanol extract;
3) ethyl acetate extract and ethanol extract are mixed, add adjuvant to make dosage form.
2. pharmaceutical composition according to claim 1, is characterized in that, described effective ingredient is made up of the raw material of following weight portion: 60 parts of 40 parts of Rhizoma Dryopteris Crassirhizomatiss and Fructus Crataegis.
3. pharmaceutical composition according to claim 1 and 2, is characterized in that, the dosage form of described Rhizoma Dryopteris Crassirhizomatis Fructus Crataegi compositions is granule, soluble powder, tablet or oral liquid.
4. pharmaceutical composition according to claim 3, it is characterized in that, also comprise one or more in starch, beta cyclodextrin, lactose, stevioside, sucrose, glycyrrhizin, high fructose syrup, sucralose, xylitol, Sorbitol, glucose, pregelatinized Starch, carboxymethyl starch sodium.
5. pharmaceutical composition according to claim 1, is characterized in that, the preparation method of described compositions comprises the following steps:
1) take Rhizoma Dryopteris Crassirhizomatis, use ethyl acetate lixiviate, concentrating under reduced pressure obtains extractum; 75～95% soak with ethanol for medicinal residues, boil and extract 1～2 time, merging filtrate, and concentrating under reduced pressure evaporate to dryness obtains extractum, merges extractum;
2) take Fructus Crataegi, use ethyl acetate lixiviate, concentrating under reduced pressure obtains extractum; 75～95% soak with ethanol for medicinal residues, boil, and by lixiviating solution sucking filtration, concentrating under reduced pressure evaporate to dryness obtains extractum, merge extractum;
3) get Rhizoma Dryopteris Crassirhizomatis extract and Fructus Crataegi extract extractum and mix, add adjuvant to make dosage form.
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Non-Patent Citations (4)
| Title |
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| 山楂粉治疗70例病毒性肝炎;杨健萍;《人民军医》;19770831;全文 * |
| 杨健萍.山楂粉治疗70例病毒性肝炎.《人民军医》.1977, |
| 绵马贯众抗禽流感病毒活性部位的高效液相色谱-质谱法分析;陈建新等;《分析测试学报》;20080630;摘要 * |
| 陈建新等.绵马贯众抗禽流感病毒活性部位的高效液相色谱-质谱法分析.《分析测试学报》.2008, |
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