CN109420046A

CN109420046A - A kind of method of herbal prescription preparation prevention and treatment porcine hyperthermia drug

Info

Publication number: CN109420046A
Application number: CN201710740688.0A
Authority: CN
Inventors: 刘莹莹; 肖毅; 肖兵南; 蒋小文
Original assignee: Hunan Minkang Biological Technology Institute
Current assignee: Hunan Minkang Biological Technology Institute
Priority date: 2017-08-25
Filing date: 2017-08-25
Publication date: 2019-03-05

Abstract

The invention discloses a kind of method that herbal prescription prepares prevention and treatment porcine hyperthermia drug, this method is to crush the Chinese herbal medicine for adapting to zymophyte growth in prescription to take into account each disease stage symptom of porcine hyperthermia and carry out herbal prescription, 5-8 times of water is added to extract；The dregs of a decoction add appropriate (10-20%, w/w) nutrition auxiliary material, after high-temperature sterilization, are inoculated with the fermenting microbe of 10-20%, and aerobic fermentation 2-4d is carried out under 26-28 DEG C, relative humidity 70-90%, aseptic condition, and medicinal fungal substance is made.The Chinese herbal medicine and said extracted liquid of its medicinal fungal substance and Fang Zhongwei fermentation (being not suitable with zymophyte growth) carry out preparation by different dosage forms requirement.For the Chinese herbal medicine clearly, to add cold-pungent and diaphoretic drugs disease and evil outlet in clearing heat and detoxicating, purging fire and cooling blood disease and evil, combining the internal organs symptom such as lung is principle, realizes that heat-clearing cooling, eliminating inflammation and expelling toxin, preventing phlegm from forming and stopping coughing, conditioning increase and exempt from, treating both manifestation and root cause of disease；The zymophyte is selected from anti-bacteria and anti-virus, clearing heat and detoxicating, cooling blood and hemostasis, cough-relieving of ventilatining the lung and resolving phlegm, the hot illness for the treatment of and breathes the medicine of reproductive syndrome/fungivorous.

Description

A kind of method of herbal prescription preparation prevention and treatment porcine hyperthermia drug

Technical field

The invention discloses a kind of methods that herbal prescription prepares prevention and treatment porcine hyperthermia drug.

Background technique

Pig " hyperpyrexia disease " be by the virus such as high-pathogenicity blue ear disease and a variety of cause of diseases mixing such as bacterium and helminth or after Acute, hot, highly pathogenic and fatal disease caused by hair infects；To generate heat, not eat, rubefaction and there is respiratory tract disease Shape is main feature.Finishing period pig and part sow are occurred mainly in, the cause of disease detected mainly has: reproductive and respiratory syndrome virus (PRlKSV), swine fever virus (HCV), swine influenza virus (SIV), Pseudorabies virus (ADV), porcine circovirus 2 type (PCV-II) With actinobacillus pleuropneumoniae (APP), streptococcus suis 2-type (S.S- II), Haemophilus parasuis (HP), killing property Pasteur bar more Bacterium (PM), mycoplasma, toxoplasm, eperythrozoon, Salmonella choleraesuls (sc) etc..Body temperature increases after pig illness, and use is a variety of Body temperature is without being decreased obviously after antibiotic treatment, with temperature decline after antibiotic plus analgesic-antipyretic combination therapy, but it is 2nd time most Medication is invalid, and the state of an illness is more serious, systemic die of exhaustion occurs and dies.Pig farm affected pig disease incidence fall ill 50% or more, the death rate Up to 50~90%.

Do not have specific therapeutic agent to the disease at present, mainly by immune epidemic prevention, but because cause of disease is complicated and changeable, effect is immunized Fruit is unstable；Biological agent (interferon, serum etc., expensive) is removed outside to its therapeutic agent, mainly antibiotic and medium-height grass The combination formulations or drug combination of medicine, the former adjusts immune function, amendment is faced for pressing down virus of going out for pressing down germ of going out, the latter Bed symptom etc..A large number of studies show that many Chinese herbal medicines have with good antibiosis and antiviral functions, Chinese medicine in motherland's medicine, though Title without " virus ", but it is early on the books in relation to the treatment to disease of viral infection, and the product in thousands of years of clinical application Experience abundant is tired out.In recent years, domestic and international researcher uses modern technologies, screens and search out one from Chinese herbal medicine Energy antiviral drug a bit, some of them, which has been developed that, is applied to clinic.Qingwen baidu powder, the Jing Fang recorded such as China's veterinary drug allusion quotation Toxin-Vanquishing drink, secondary development preparation " swap buffers " oral solution of Yinqiao San, powder needle etc. are for treating porcine hyperthermia.

Porcine hyperthermia presses Chinese veterinarian's theory, is caused by exogenous disease warm disease and evil, according to the theory of defensive-qi-nutrient-blood, pathological development Process and the state of an illness are shallowly divided into four-stage deeply: disease is being defended point, and the state of an illness is gently shallow；Disease is in gas point, and in heresy has passed, the state of an illness is heavier；Disease In battalion point, the state of an illness is heavier；The disease appearing in blood system, the most seriously.Different phase principle of reatment is different from medication, and individual morbidity treatment can be with In this way, but the exploitation of patent medicine cannot be in this way, each stage of illness need to be taken into account.In addition, the disease generally morbidity is anxious, the course of disease is short, when out Mostly disease and evil has entered gas point, or even battalion point or blood system when existing illness, therefore treatment should promote disease and evil outlet with relieving the exterior syndrome with drugs pungent in flavor and cool in property and battalion, Clearing heat and detoxicating simultaneously, disease and evil is eliminated in cool blood cooling, and conditioning viscera illness is allowed to rehabilitation.But current herbal medicine is to porcine hyperthermia Therapy study is mostly based on traditional herbal medicine theory, the recipe of multiselect heat-clearing class, as qingying decoction, xijiao Dihuang Tang, clear pest lose Poison dissipate etc.；Morbidity early stage selects refrigerant inducing diaphoresis class recipe, as the secondary development preparation " swap buffers " of Yinqiao San and Yinqiao San is oral Liquid, powder needle, Jingfang toxicity-removing drink etc..Lung symptoms significantly with the treatment of clearing heat and freeing lung method, represent prescription such as Ma Xing Shi Gan Tang and add taste Deng.These recipe are only applicable to individual treatment, are difficult to group's administration.

In addition, conventional Chinese veterinary drug preparation, if the release of powder effective component is slow, absorptivity is low, poor to acute disease effect；In Though herbal medicine extraction can heighten the effect of a treatment, the content and bioavilability of effective component are improved, in the method for physics only also to increase Strong curative effect.Modern Chinese herbal medicine biofermentation is on the basis of inheriting tradition herb fermenting method, in conjunction with modern biotechnology to Chinese medicine Development and utilization, the non-active ingredient in Chinese herbal medicine is active constituent by main purpose, so also known as bioconversion；Utilize benefit The strains such as raw bacterium, micromycete and macro fungi ferment under manual control condition, are treated with promoting the clinical of traditional Chinese medicine Effect, the discovery clinical new value of traditional Chinese medicine, the utilization rate for significantly improving Chinese material medicine resource, batch Innovative TCMs, to promote The industrialized development of Chinese medicine.The research of modern Chinese herbal medicine fermentation is related to study of compound prescription from simple and obtains significant achievement. Such as single medicinal material fermentation research: Li Guohong (2006) ferments to 50 kinds of Chinese medicines with bacillus subtilis, investigates to tuberculosis point The activity of branch bacillosis and hazel color mould, antibacterial activity changes after herb fermenting, and part is improved；Gao Kaixian (2007) spirit Sesame solid fermentation ramulus mori, Aqueous extracts reduce the ability of mouse triglycerides and cholesterol, and fermentation kuh-seng improves scavenging hydroxyl Ability；Wang Lin etc. (2004) observes influence and its fermentation liquid of the Chinese medicine to biomass in Fermentation Process of Ganoderma Lucidum Mycelium fermentation process to chronic The curative effect of bronchitis, discovery herb fermenting are better than simple glossy ganoderma fermentation effect；Tao Junyan etc. (2009) is to medicinal white rot in 6 kinds Ganoderma fermentation improves part Chinese medicine and generates new substance, reducing blood lipid ability.For another example in terms of herbal mixture fermentation, Yan Wei etc. (2010) it develops " herb fermenting Moxa patch " treatment chronic superficial gastritis and has received good effect；Take off brilliant (2007) research light Bacterial fermentation Rehmanniae Decoction of Six Ingredients is closed, the spleen weight of hypoimmunity mice is significantly increased after fermentation, and find that photosynthetic bacteria converts small point Sub- substance, HPLC peak number are reduced.Liu Jinzhe (2011) improves memory effect to Chinese medicine and food medicine fungi composite fluid fermentation preparation It is studied, filters out the side's group for improving mouse memory power and fermenting microbe and combinations thereof, and optimization for fermentation technology condition.For another example In terms of veterinary drug, Liu Chunhui (2013) is with bacillus subtilis and produces first Candida ratio mixed solid fermentation prevention and treatment milk cow The herbal mixture of mammitis, discovery fermentation recipe further decrease mastitis for milk cows disease incidence, improve milk-quality, improve milk production Amount.Hou Yongchao^[(2012) report herb fermenting feed addictive (peace times is strong) adjusting body immune system, balance intestinal flora, Intestinal mucosa is repaired, metabolism is promoted, improves production performance and its mechanism.

The new technology of modern Chinese herbal medicine fermentation preparation, including probiotics fermention, mold fermentation and macro fungi (medicine/edible true Bacterium) fermentation etc., especially macro fungi fermentation is the hot spot studied at this stage.Macro fungi polysaccharide polypeptide rich in etc. is living Property substance, have the effect of mostly antiviral and improve immunity, such as split pleat granulose (Schzophyllan) and have and inhibit celestial Platform virus (Sendai virus) intercellular diffusion；For another example Xue opens lotus etc. (2007) report and extracts separation from fungi A kind of substance being JN219 out is shown through anti-human cytomegalovirus study on mechanism: mainly blocking virus invasion host is thin Born of the same parents, action site are mainly human cytomegalovirus outer membrane protein.With macro fungi to herb fermenting, Zhuan Yi proposes " bi-directional fermentation " Concept (Chinese medicine and fungi fermentation two-way interaction influence each other), method is closed in Chinese medicine and fungi fermentation group will carry out greatly The screening (blind choosing) of amount, it is time-consuming and laborious.We have found in years of researches, and cmm of simple prescription or compound are with identical as its nature,taste and action Or similar medicine/fermentation fungivorous, drug effect can be made to increase substantially, method is known as " orientation fermentation ".

Herb fermenting preparation is a kind of effective ways of synergy, is especially had shown that with macro fungi fermentation huge latent Power；But its research is still in the primary stage, and needs deeply to grind in fementative composition screening, transformation mechanism and effect assessment aspects Study carefully.The present invention is based on modern biotechnology fermentation technique, is promoted and innovated, for the purpose of preventing and treating the exploitation of porcine hyperthermia Chinese patent drug, Herbal prescription is outer to rush down in clear, and the principle for the treatment of both manifestation and root cause of disease overcomes treatment to attach undue importance to one thing to the neglect of the other, and is not suitable with the defect of the more symptoms administrations of group； Select have good anti-bacteria and anti-virus it is anti-inflammatory and enhancing immunization Chinese herbal medicine, with the same or similar medicine of nature,taste and action/ Edible fungus carries out fermentation towards synergistic direction；In addition, the effect of its biofermentation mainly passes through microbial enzyme centering Biochemical substances in herbal medicine play biochemical reaction, it is possible to it rises forward reaction (converting functional mass for non-functional substance), Back reaction (converting non-functional substance for functional mass) may also be played, since the diversity of enzyme and reaction are complicated, effect Mechanism is unclear, and institute can only lean on a large amount of test to determine and hold taking human as being difficult to control.For this purpose, we take Chinese herbal medicine The method for first extracting post-fermentation, prevents and treats the inverse change of functional mass, to ensure that drug effect only increases.Meanwhile the medicine after fermentation Property mycoplasma is to digest or self-dissolving method makes macromolecular inert matter biodegrade small molecule active object, the pharmacological property bacterium after fermentation Matter to digest or self-dissolving method makes macromolecular inert matter biodegrade small-molecule active substance (active polysaccharide polypeptide etc.), With extracting concentration or pulverizing technique preparation, the dissolution rate and bioavilability of effective component are improved.Verification experimental verification zymophyte Matter pharmaceutical preparation can imitate prevention and treatment porcine hyperthermia with having, and provide powerful mean for the prevention and treatment of the disease.

Summary of the invention

The present invention is directed to overcome the deficiencies of the prior art and provide a kind of herbal prescription preparation prevention and treatment porcine hyperthermia drug Method.

In order to achieve the above object, technical solution provided by the invention are as follows:

The method is that the Chinese herbal medicine for adapting to zymophyte growth in Chinese herbal medicine prescription is crushed water to extract, and obtains raw medicine extraction Liquid mixes the dregs of a decoction plus nutrition auxiliary material, and the mass percent of the nutrition auxiliary material and Chinese herb residues is 10-20%, high temperature After disinfection, then the inoculum concentration inoculation fermentation bacterium of 10-20% is pressed, is carried out under 26-28 DEG C, relative humidity 70-90%, aseptic condition Medicinal fungal substance is made in aerobic fermentation 2-4d；In the medicinal fungal substance and not fermented (being not suitable with zymophyte growth in prescription) Herbal medicine and raw medicine extracting solution require to carry out processing preparation according to dosage form；The Chinese herbal medicine prescription is with clearing heat and detoxicating, purging fire and cooling blood is sick It is clear in heresy, add cold-pungent and diaphoretic drugs disease and evil outlet, combining the internal organs symptom such as lung is principle, realize heat-clearing cooling, eliminating inflammation and expelling toxin, Preventing phlegm from forming and stopping coughing, conditioning increase exempt from, treating both manifestation and root cause of disease；The zymophyte is selected from anti-bacteria and anti-virus, clearing heat and detoxicating, cooling blood and hemostasis, promoting the dispersing function of the lung Phlegm and relieving cough, treat hot illness with breathe reproductive syndrome medicine/it is fungivorous.

Preferably, the Chinese herbal medicine is selected from gypsum, Radix Rehmanniae, cornu bubali, the coptis, cape jasmine, the root bark of tree peony, the Huang of qingwen baidu powder A kind of reed mentioned in ancient books, radix paeoniae rubra, radix scrophulariae, Fructus Forsythiae, rhizoma anemarrhenae, campanulaceae, lophatherum gracile, 8 kinds or more in Radix Glycyrrhizae add clearing heat and detoxicating (anti-bacteria and anti-virus), cool Blood stops blooding the cold ice stone of class, honeysuckle flower, Radix Isatidis, wild chrysanthemum, folium isatidis, cordate houttuynia, garden burnet, Asian puccoon, rhizome of cyrtomium, the 2-6 in Radix Astragali Kind, add inducing diaphoresis heat-clearing, the radix bupleuri for cough-relieving class of ventilatining the lung and resolving phlegm, the root of purple-flowered peucedanum, reed root, pueraria lobata, blackberry lily, mulberry skin/mulberry leaf, lepidium seed, schizonepeta, Chinese ephedra, sweet wormwood, fritillaria, the 2-6 kind in polygonum cuspidate, amount to 12-29 taste Chinese herbal medicines；The zymophyte is selected from two types and removes from office lid bacterium, white side Ear, Pleurotus nebrodensis, needle mushroom, cordyceps sinensis, ganoderma lucidum, Antrodia camphorata, beef-steak fungus, mushroom, handle mushroom, wild rice bacterium, tabasheer, halimasch, Grifola frondosus, east bolt bacterium, Cyathus striatus, Inonotus obliquus, calvatia gigantea, Lip river bar dried mushroom, golden ear, tremella, pisolithus tinctorius Coker et Coucs, black wood At least one of ear, delicious lactarius.

The parts by weight of each component in the Chinese herbal medicine are as follows: gypsum 100-150, Radix Rehmanniae 20-35, cornu bubali 60-100, the coptis 20-35, cape jasmine 20-35, root bark of tree peony 20-35, radix scutellariae 20-40, radix paeoniae rubra 20-35, radix scrophulariae 20-35, Fructus Forsythiae 20-40, rhizoma anemarrhenae 20-40, Campanulaceae 20-35, lophatherum gracile 25-40, Radix Glycyrrhizae 10-20, tremble with fear ice stone 20-30, honeysuckle flower 20-30, Radix Isatidis 30-50, wild chrysanthemum 20- 30, folium isatidis 30-60, cordate houttuynia 30-60, garden burnet, Asian puccoon 20-30, rhizome of cyrtomium 20-30, Radix Astragali 25-40, radix bupleuri 20-30, the root of purple-flowered peucedanum 20-30, reed root 25-40, pueraria lobata 20-40, blackberry lily 20-30, mulberry skin/mulberry leaf 20-30, lepidium seed 20-30, schizonepeta 20-30, Chinese ephedra 10-20, sweet wormwood 20-40, fritillaria 10-20, polygonum cuspidate 30-50.

The zymophyte is selected from delicious lactarius and Jin Er.

Preferably, complex enzyme zymohydrolysis is added in the medicinal fungal substance, and mass percent of the complex enzyme in medicinal fungal substance is 0.05-0.5%；It mixes with the Chinese herbal medicine of not fermented (being not suitable with zymophyte growth in prescription) or is individually extracted through water again, mentioned Oral solution or granule are made after taking liquid to mix concentration with raw medicine extracting solution.The complex enzyme is non-starch polysaccharide enzyme and protease By (1-3): the complex enzyme of 1 mass ratio composition.

1, after preferably, the medicinal fungal substance and the Chinese herbal medicine of not fermented (being not suitable with zymophyte growth in prescription) crush Mixing, obtains mixture；Raw medicine extracting solution and suitable quantity of water are added into mixture, makes the mass ratio 1:(2- of mixture and water 5)；It is dry after the mycelium self-dissolving of medicinal fungal substance in object to be mixed, pulvis is made after ultramicro grinding.

The invention will be further described below:

1, the selection of herbal prescription and its fermenting microbe:

(1) herbal prescription: with clear disease and evil in clearing heat and detoxicating, purging fire and cooling blood, add relieving the exterior syndrome with drugs pungent in flavor and cool in property, ventilating the lung and resolving phlegm outside pectoral Disease and evil is arranged, realizes that heat-clearing cooling, eliminating inflammation and expelling toxin (virus), conditioning increase and exempt from.Qingwen baidu powder is selected to be added and subtracted, i.e. eliminating pestilence and relieving toxicity Dissipate (gypsum 100-150, Radix Rehmanniae 20-35, cornu bubali 60-100, coptis 20-35, cape jasmine 20-35, root bark of tree peony 20-35, radix scutellariae 20- 40, radix paeoniae rubra 20-35, radix scrophulariae 20-35, Fructus Forsythiae 20-40, rhizoma anemarrhenae 20-40, campanulaceae 20-35, lophatherum gracile 25-40, in Radix Glycyrrhizae 10-20 8 kinds or more), add cold ice stone, honeysuckle flower 20-30, Radix Isatidis 30-50, wild chrysanthemum 20-30, folium isatidis 30-60, cordate houttuynia 30- 60, garden burnet, Asian puccoon 20-30, rhizome of cyrtomium 20-30, the 2-6 kind in Radix Astragali 25-40 are to help removing toxic substances heat-clearing, cooling blood and hemostasis；Add radix bupleuri 20- 30, root of purple-flowered peucedanum 20-30, reed root 25-40, pueraria lobata 20-40, blackberry lily 20-30, mulberry skin (leaf) 20-30, lepidium seed, schizonepeta 20-30, fiber crops Yellow 10-20, sweet wormwood 20-40, fritillaria 10-20, the 2-6 kind in polygonum cuspidate 30-50 are with inducing diaphoresis heat-clearing, cough-relieving of ventilatining the lung and resolving phlegm.Total 12- 29 taste Chinese herbal medicines.

(2) fermenting microbe selects: selecting anti-bacteria and anti-virus, clearing heat and detoxicating, cooling blood and hemostasis, cough-relieving of ventilatining the lung and resolving phlegm, treatment heat Venereal disease disease and medicine/food of breathing reproductive syndrome are true: two types remove from office lid bacterium, foliosous parnassia herb, Pleurotus nebrodensis, needle mushroom, cordyceps sinensis, spirit Sesame, Antrodia camphorata, beef-steak fungus, mushroom, handle mushroom, wild rice bacterium, tabasheer, halimasch, grifola frondosus, east bolt bacterium, Cyathus striatus, Inonotus obliquus, calvatia gigantea, Lip river bar dried mushroom, golden ear, tremella, pisolithus tinctorius Coker et Coucs, black fungus, delicious lactarius etc., single bacterium or more bacterium combined ferments. Project first choice delicious lactarius and Jin Er.Delicious lactarius is commonly called as cold bacterium, has good anti-bacteria and anti-virus, clearing heat and detoxicating, treatment febrile disease Effect；Golden ear, energy resolving sputum, cough-relieving, Dingchuan, regulating the qi flowing in the channels, calming the liver intestines cure mainly lung heat, abundant expectoration, cold cough, asthma, hypertension Deng.

2, preparation process thereof

In order to prevent functional component back reaction and fail, will Chinese herbal medicine prescription carry out water extraction after take the dregs of a decoction to ferment, medicine Medium-height grass liquid medicine to be fermented extracts in side, and the medicinal fungal substance after dregs of a decoction fermentation is non-through cellulase, pectase, zytase etc. Starch-polysaccharides enzyme and protease hydrolytic make non-(or low) active material of some macromoleculars point be converted into small molecule high active substance (active small molecular polysaccharide, polypeptide etc.), is extracted, together with raw medicine extracting solution (containing non-fermentation of Chinese herbal medicine extracting solution in prescription) It is concentrated to improve the effective concentration of drug；Or add unfermentable Chinese herbal medicine and the pharmacological property bacterium fermented in raw medicine extracting solution and prescription Matter carries out mycelium self-dissolving together, and re-dry ultramicro grinding discharges active material sufficiently, to improve the bioavilability of drug And its drug effect.Its technique:

Prescription simple and zymophyte are fermented → are suitble to the simple mixing water extraction of mycelia growth→ the dregs of a decoction and zymophyte Fermentation

Note:Medicinal fungal substance is added as dilution when extracting solution prepares powder and carries out automyophagy, prepare oral solution with It is directly added into when granula in mycoplasma extracting solution and merges preparation

Being not suitable with when the non-fermentation of Chinese herbal medicine that mycelia grows prepares oral solution can not digest, and directly extract, add after concentration Enter.

3, important active substances and pharmacy in vitro measure

Polysaccharide polypeptide is the important active substances of fungi, and content height directly affects drug effect, therefore as ferment effect Main indicator；The power of anti-bacteria and anti-virus power is the key that pharmaceutical preparation success or failure, therefore the important indicator as the evaluation of pesticide effectiveness.

(1) sulfuric acid-phynol method and 3,5- dinitro water hydrochloric acid (DNS) colorimetric method for determining polysaccharide polypeptide assay: are used Total reducing sugar and reduced sugar, acquire polyoses content；Content of peptides is measured using peptide bond measuring method.

(2) Determination of Antibacterial Activity: with Odontothrips loti (every glass plus 50nl), inhibition zone size is measured；Test tube liquid culture method, It measures minimal inhibitory concentration (MIC).

(3) external anti-PRRSV determination of activity: blood medicine-cell culture method measurement antiviral activity is taken.

4, the internal evaluation of pesticide effectiveness

The internal evaluation of pesticide effectiveness is the most reliable method for evaluating effect of drugs, and determines that drug could be applied to clinical elder generation Certainly condition, therefore tested as follows.

(1) therapeutic test of pig artificial challenge PRRSV: epidemic disease is replicated with non-infection piggy injection PRRSV virus liquid, is carried out Drug therapy measures its drug effect.

(2) Clinical Treatment Test of natural infection pig: after making a definite diagnosis morbidity pig farm progress porcine hyperthermia, clinical medicine is carried out Therapeutic test measures drug effect.

5, specific embodiment

The preparation of 1 strain of embodiment is fermented with Chinese herbal medicine

1.1 test method

(1) prepared by culture medium

1) medicine/edible fungus liquid spawn culture medium: corn pulp 1.5%, glucose 2%, (NH4) 2NO30.15%, K2HPO4, MgSO47H2O add murphy juice to 100%；PH6-6.5, sterilize high pressure 30min.

2) Chinese herbal medicine fermentation auxiliaries: wheat bran is pressed: corn flour: dregs of beans: during flour=3:3:2:1.5 (mass ratio) is prepared The auxiliary culture medium of herbal medicine fermentation, is prepared for Chinese herbal medicine and delicious lactarius orientation fermentation culture medium.

3) Chinese herbal medicine fermentation substrate is prepared: the single or compound Chinese herb that the side of taking organizes crush respectively, by Chinese herbal medicine powder: auxiliary The mixing of culture medium=8-9:2-1 ratio, prepares fermentation medium, sets in mushroom culture air-permeable bottle, loading amount is the 1/3 of bottle, high It is spare after pressure sterilizing；

(2) Chinese herbal medicine and delicious lactarius fermentation test of goodness of fit

1) Spawn incubation: taking bacterium culture medium, is inoculated with delicious lactarius and tremella silk respectively, 26-28 DEG C, 150rpm, cultivates 24-48h；

2) Chinese herbal medicine and medicine/edible fungus mixed fermentation are tested: taking the Chinese herbal medicine of single Chinese herbal medicine and the auxiliaries that ferment Fermentation substrate is inoculated with, 26-28 DEG C, relative humidity 70-90%, training by the strain amount of 10% (V/W, delicious lactarius: golden ear=1:1) Support 2-3d.

3) effect measuring: the growing state of mycelia is observed to the fermentation material timing of every taste Chinese herbal medicine and delicious lactarius, and is recorded As a result；

(3) preparation of orientation fermentation medicinal fungal substance

1) culture substrate is prepared: will be suitable in " qingwen baidu powder " addition and subtraction and delicious lactarius/gold ear fermentation single medium-height grass Medicine mixes in proportion, crushes, and 10-20% fermentation auxiliary material is added, prepares fermentation medium (medicinal substrate)；

2) inoculation fermentation: prepared fermentation medium is fitted into tray after autoclave sterilization, is inoculated with 10% delicious lactarius + tremella kind (each 5%) sets relative humidity 70-90%, 2-4d is cultivated under the conditions of 26-28 DEG C of temperature, after mycelia is covered with, 70 DEG C It dry, pulverize, i.e. medicinal fungal substance.

1.2 test results:

Test determines altogether treats related 30 kinds of heat-clearing classes and tonifying Qi cough-relieving class Chinese herbal medicine to loose cream for porcine hyperthermia The influence of mushroom and the unfamiliar to the ear length of gold, wherein 4 kinds of delicious lactarius mycelia do not grow and (account for 13.3%)；9 kinds of growths (30%) reluctantly, that is, grow Slowly, until the 4th day mycelia occurs just now；11 kinds grow (36.7%), have more mycelia to occur within second day, third space-based Originally it covers with；6 kinds grow preferably (20%), and third day has more mycelia to occur.The experimental result of golden ear is similar.Details is shown in Table 1.1

1.1 single Chinese herbal medicine of table and delicious lactarius/gold ear fermentation hypha growth condition

2 orientation fermentation of embodiment and the preparation of high fever seasonal febrile diseases Ling oral liquid/powder

2.1 culture substrates prepare: will in " qingwen baidu powder " addition and subtraction be suitable for delicious lactarius fermentation single Chinese herbal medicine by than Example mixing, crushes, and water extracts to obtain extracting solution (raw medicine extracting solution) and the dregs of a decoction；Its dregs of a decoction is by auxiliary culture medium: the dregs of a decoction (wet)=10: 90 ratio is prepared fermentation medium (medicinal substrate)；

2.2 inoculation fermentations: prepared fermentation medium is fitted into tray after autoclave sterilization, is inoculated with 10% pine cream (each 5%) sets relative humidity 70-80%, 2-3d, after mycelia is covered with, 70 is cultivated under the conditions of 26-28 DEG C of temperature for mushroom and tremella kind It DEG C dry, pulverize, i.e. medicinal fungal substance.

2.3 high fever seasonal febrile diseases spirits preparation: by the medicinal fungal substance fermented processing: (1) prepared by oral solution: adding 0.1-0.2% multiple Synthase (non-starch polysaccharide enzyme: protease=(1-3): 1), PH5, digests 2-3h by 5-6.45-50 DEG C, after 80-90 DEG C of inactivation → plus Chinese herbal medicine → the microwave (120S~180S) or ultrasonic wave (400W, 50-60 for not fermenting and (being not suitable with delicious lactarius growth in prescription) DEG C, 40-60min) processing → filtering, filter residue be extracted with water method extract 2 times, filtrate merges with raw medicine extracting solution is concentrated into every milli Rise 1g containing crude drug, packing, autoclave sterilization, as high fever seasonal febrile diseases Ling oral liquid.

(2) prepared by powder: by the medicinal fungal substance fermented and the medium-height grass that do not ferment and (be not suitable with delicious lactarius growth in prescription) Medicine mixing adds raw medicine extracting solution and suitable quantity of water (material-water ratio 1:2-5), and beater mashing adds 1-2%NaCl, adjusts PH to 7-8, Sealing, handles 12-24h in 40-50 DEG C of water-bath, and mycelium autolytic cleavage is made to discharge polysaccharide polypeptide isoreactivity substance, dry, surpasses Crushing of Ultrafine, i.e. high fever warm disease medicinal powder.

The measurement of the fermentation medicinal fungal substance active constituent of embodiment 3

3.1 test methods:

3.1.1 polysaccharide polypeptide assay:

(1) polysaccharide determination

1. total sugar content measures: using sulfuric acid-phynol method；

2. content of reducing sugar measures: using 3,5- dinitro water hydrochloric acid (DNS) colorimetric method；

3. polyoses content calculates: polyoses content=total sugar content-content of reducing sugar

(2) measuring method of polypeptide: peptide bond measuring method is used.

1. standard curve is drafted: precisely weigh 20mg glutathione, be configured to 0 with distilled water dilution, 40,80,120, 160, each 5mg/ml of standard solution (5ug/ul) of 200,240,280ug/ml；Light absorption value is surveyed at 238nm, calculates standard curve (regression equation)；

2. assay: by diluted sample, drawing 3-4.0ml, light absorption value is surveyed at ultraviolet spectrometer 238nm, by standard Curve calculates total protein content of peptides in sample

3.1.2 Determination of Antibacterial Activity:

The medicinal fungal substance and unfermentable base starting material medicine fermented is taken, by medicine: water=1:10,50 DEG C of ultrasonic wave extractions, Centrifugal filtration takes filtrate, with Odontothrips loti (every glass plus 50nl), measures inhibition zone size.Meanwhile medical fluid is made into doubling dilution, Test tube liquid culture method measures minimal inhibitory concentration (MIC).

3.1.3 anti-PRRSV virus activity external test

(1) Zymocyte active constituent extracts: weighing medicinal fungal substance 50g, adds water 400ml, boil 45min, filter, repeats It extracts, merging filtrate, is concentrated under reduced pressure into 50ml (crude drug 1g/ml) three times, 4 DEG C save backup.

(2) prepared by drug containing mice serum: 20g or so mouse every gavages medicinal fungal substance extracting solution 5ml, and continuous 3 times, every time It is spaced 1h, rear 30min is gavaged for the last time and extracts eyeball method, acquire mouse blood, is centrifugated serum, 56 DEG C of inactivations 30min, -20 DEG C save backup.

(3) it blood medicine-cell culture method measurement: by the method for Xiao Bingnan patent (ZL201010296978.9) report, carries out Marc-145 cell culture and the evaluation of pesticide effectiveness: in 96 holes that cell has grown up to single layer substantially, the old culture solution in each hole is sucked out, presses Three kinds of modes of action: antiviral absorption (after first dosing plus virus), antiviral duplication (first connecing dosing after poison), direct inactivation of viruses Effect (drug and viral interaction in vitro are added afterwards for a period of time), sequentially adds the drug containing blood for being diluted to safe concentration (2.5%) The 100 μ l of virus liquid of cleer and peaceful 1 TCID50, if normal cell controls group, mouse serum control group and virus control group, set 37 DEG C It is cultivated in incubator in 5%CO2 incubator and cultivates 72h, observe cytopathy (CPE) situation with inverted microscope and recorded, used MTT test method(s) measures OD value, calculates drug to the protective rate (CBL) of cell, company is as follows:

3.2 test result

3.2.1 polysaccharide polypeptide assay

Composite Chinese herbal and polysaccharide polypeptide in matrix before and after delicious lactarius orientation fermentation are measured, as the result is shown: fermentation Polysaccharide and content of peptides have increase in matrix afterwards, and especially polysaccharide variation is obvious, increase by 18.42%, the results are shown in Table 3.1.

3.1 composite Chinese herbal of table fermentation front and back polysaccharide polypeptide comparision contents measurement result

3.2.2, Determination of Antibacterial Activity

Antimicrbial power (inhibition zone) size of comparative measurements composite Chinese herbal fermentation front and back；Simultaneously to compound medicinal fungal substance into Row minimal inhibitory concentration (MIC, raw drug content) measures, as the result is shown: the medicinal fungal substance ratio to 4 kinds of pathogens, after fermentation Inhibition zone and minimal inhibitory concentration have improvement before fermenting, and improve 1-2 times of drug effect.It the results are shown in Table 3.2.

The inhibition zone and MIC measurement result of 3.2 composite interstitial substance of table fermentation front and back

3.2.3, Assay of Antiviral Activity

From table 3.3 as it can be seen that qingwen baidu powder plus-minus preparation (do not ferment drug) has stronger restraint to PRRSV virus, Three kinds addition drug method to the protective rate (CBL) of cell 70% or more, especially to suppressing virus replication and directly Killing virus has a stronger effect, and cytoprotection rate is up to 85% or more.Mycoplasma effect after fermentation is stronger, to three kinds of virus The mode of action has nearly 100% effect, especially suppressing virus replication and directly killing virus function very significant, therefore can Therapeutic agent as the disease.

The anti-PRV effect detection (8 hole mean value) of 3.3 mice serum containing Chinese herbal medicine of table

The zoogenetic infection clinical trial of 4 high fever warm disease Ling oral liquid of example

4.1 test method

(1) experimental animal is with grouping: the 18-20kg's that purchase does not vaccinate from non-epidemic disease area hybridizes piglet 30, sets Raising, is randomly divided into 6 groups, every group 5 in closed isolating pig house；First group is not infect Normal group, and second group is sense Control group is not administered in poison of catching an illness, and third group is not ferment qingwen baidu powder plus-minus recipe (with high fever seasonal febrile diseases Ling oral liquid preparation side Method extracts preparation) treatment group, the four, the 5th and the 6th groups of high fever warm disease Ling oral liquid treatment groups for high, medium and low dosage,

(2) artificial challenge and drug therapy: experimental group and infection control group pig inject pig PRRSV virus liquid 2ml respectively, from First pig, which starts to have fallen ill, treats test pig.Medication administration method: drug is admixed in appropriate feed and is fed for that (do not search for food pig It manually feeds)；Dosage, " efficient warm disease Ling oral liquid ": low dose group 0.5ml/kg. weight, middle dose group 1.0ml/kg. body Weight, large dosage group 2ml/kg. weight；It does not ferment qingwen baidu powder addition and subtraction medicament extracting solution 1.0ml/kg. weight；It is early daily, in, 3 dispensings of evening；Medication 3-7 days.

(3) judgement of medicine curative effect: body temperature, spirit, feeding restore normal, and skin spots disappearance is judged as healing, and symptom subtracts Less serious case be it is effective, medication in 7 days does not restore or died is invalid.

4.2 test result

From table 4.1 as it can be seen that the high fever seasonal febrile diseases spirit of the 4th and the 5th group of high middle dosage, which dissipates, can reach fully effective treatment rate, from The temperature decline of beginning in the 2nd day of medication, the returns to normal for 5-6 days；The effect and the agent of 1ml/kg. weight of the treatment group of low dosage The therapeutic equivalence of the unfermentable qingwen baidu powder plus-minus square preparation of amount, up to 80% cure rate.This proves that Chinese herbal medicine passes through and pine Newborn mushroom orientation fermentation is greatly improved curative effect.It is further noted that the pig of non-artificial challenge also occurs falling ill and dead, it is Because with caused by circle infection.

Body temperature and death condition record in 4.1 artificial liver support swinery therapeutic process of table

Note: 1. be Normal group；2. for infection control group；3. for decoction for the treatment group of not fermenting；4-6. is high fever warm disease Spirit treatment height (2ml/kg), in (1ml/kg), low (0.5mlkg) dosage treatment group.

The clinical application of 5 high fever seasonal febrile diseases spirit of example is tested

5.1 test method

(1) experimental animal: selection hyperpyrexia disease morbidity pig farm understands epidemic disease situation, carries out clinical diagnosis, disease takes pathology to send Pass department makees chemical examination detection, is determined as swine high fever syndrome (hyperpyrexia disease).

(2) drug treatment: pressing each 1ml/kg weight of pig, and secondary spice is fed and (do not eat pig manually to gavage), and daily 3 Secondary, administration time is 5-7d；

(3) Clinical evaluation: the clinical change situation after observation medication daily, including body temperature, breathing, feeding, movement And the response situations such as body condition, full recovery in 1 week, it has no repeatedly as healing；Take a favorable turn, but symptom not all disappearance person I have Effect, absolutely not improves or aggravation or died are invalid.

5..2 test result

(1) course of disease and diagnosis: 20-30kg weight piglet 350 is introduced from Hubei in Zhangjiajie in May, 2014 Sangzhi County pig farm Head, the 5th day after showing up starts to fall ill, and shows as that spirit is depressed, and planted agent or does not eat at anorexia, and body temperature is increased to 42.0 DEG C or more, exhales It is purple to inhale rapid or cough and asthma, rubefaction, four limbs, ear and abdomen etc. skin.It is invalid with antibiotic treatment, sick pig after a few days It is dead successively, dead more than 100 head in 10 days.Dissect sick dead pig pulmonary edema is in diffusivity interstitial pneumonia, great Ye or bleeding Property pneumonia, be in piebald shape lung, it is serious then in liver become, petechial hemorrhage；Enlargement of lymph nodes, hyperemia, bleeding, section is wet to turn up, Centre shows marble sample decorative pattern in the exudative infiltration of downright bad sample, part；Tonsillotome hyperemia, bleeding；Gastric mucosa bleeding is burst Ulcer, necrosis, small intestine, caecum, mucous membrane of colon bleeding；There is infarct in spleen enlargement, color blackening, edge or surface；Bladder, kidney Dirty, larynx, sulcus coronarius fat and intracardiac outer membrane, auricle have blutpunkte；Kidney enlargement, brown or khaki, quality are more crisp Or soften, there are extravasated blood phenomenon, the densely covered needle point in surface to the blutpunkte of small rice grain size；Liver swelling, color is thin out, is in loess Color, matter are crisp；Heart enlargement deformation, soft, ventricle, atrial endocardium bleeding.Pig PRRSV virus is diagnosed as through city's Veterinary office With hyperpyrexia disease caused by the main mixed infection of other germs virus (sending pathological material of disease to make a definite diagnosis to Hunan agricultural university afterwards, to be porcine hyperthermia).

(2) treatment and its result: test treats 150 hair sick pig therein using high fever warm disease Ling oral liquid, By each 1ml/kg.bd., spice is fed 3 times a day, is used in conjunction 7 days.50 sick pigs use according to a conventional method antibiotic (Amoxicillin, Ceftiofur etc.)+Chinese herbal medicine compares treatment, and medication 3 times a day is used in conjunction 7 days, is experimental control；Another 30 are not treated, and are Blank control.Therapeutic test the results are shown in Table 5.1: the 3rd day after high fever seasonal febrile diseases spirit medication starts to restore, and returns to normal within the 5th day, Cure rate is up to 92%；And the treatment such as antibiotic+Chinese herbal medicine curative effect is not significant, it is not significant with the mortality difference of not administration group.

5.1 porcine hyperthermia Clinical Treatment Test effect of table

Therapeutic agent	Test pig (head)	It cures (head)	Dead (head)	Cure rate (%)	The death rate (%)
						Efficient high fever warm disease spirit	150	138	18	92	8
Antibiotic+Chinese herbal medicine	50	16	34	32	68
						Control group is not administered	30	9	21	30	70

Example 6；The scattered zooprophylazis test of high fever seasonal febrile diseases spirit

6.1 test methods:

6.1.1 animal selection and grouping

(1) prevent comparative test with field: selecting two adjacent pig houses not yet fallen ill on initial porcine hyperthermia pig farm, one Pig is not administered, and is control group；Another dissipates urgent prevention with high fever seasonal febrile diseases spirit, is test group.

(2) comparative test is prevented in different field: selecting the small-sized pig farm of adjacent two (300 or so), pig is being sent out on a pig farm Hyperpyrexia disease, another pig farm are not yet fallen ill, and are dissipated with high fever seasonal febrile diseases spirit and are promptly prevented.

6.1.2 drug dose usage and effect measuring: taking medicine spice by 2% (W/W), point feed twice sooner or later, when feeding Between 15 days；The health condition, including body temperature, spirit, feeding, fecaluria, breathing etc. of observation pig daily.

6.2 test result

(1) prevent comparative test result: the young pig of 155 25-30kg of test group () weight with field, until the pig farm state of an illness is stopped It ceases (about half a month), it is all normal, do not occur symptom.Pig without 138 identical monthly ages of control group () of prevention are administered, 96 Morbidity, disease incidence 70%；Dead 63, account for the 65.6% of morbid pig.

(2) comparative test result is prevented in different field: during test, fall ill 237, pig farm pig, and dead 152, the death rate reaches 64.1%；Do not fall ill chemoprophylaxis pig farm, 186 none morbidities of pig with it is dead.

Claims

1. a kind of method of herbal prescription preparation prevention and treatment porcine hyperthermia drug, which is characterized in that the method is by Chinese herbal medicine The Chinese herbal medicine that zymophyte growth is adapted in prescription crushes water and extracts, and obtains raw medicine extracting solution, the dregs of a decoction plus nutrition auxiliary material is mixed, institute The mass percent for stating nutrition auxiliary material and Chinese herb residues is 10-20%, after high-temperature sterilization, then is connect by the inoculum concentration of 10-20% Kind zymophyte, carries out aerobic fermentation 2-4d under 26-28 DEG C, relative humidity 70-90%, aseptic condition, and medicinal fungal substance is made；Institute The Chinese herbal medicine for being not suitable with zymophyte growth in medicinal fungal substance and Chinese herbal medicine prescription and raw medicine extracting solution is stated to require to carry out according to dosage form Processing；The Chinese herbal medicine prescription in clearing heat and detoxicating, purging fire and cooling blood disease and evil clearly, to add cold-pungent and diaphoretic drugs disease and evil outlet, combine The internal organs symptom such as lung be principle, realize heat-clearing cooling, eliminating inflammation and expelling toxin, preventing phlegm from forming and stopping coughing, conditioning increase exempt from, treating both manifestation and root cause of disease；The fermentation Bacterium is selected from anti-bacteria and anti-virus, and hot illness and breathing reproductive syndrome are treated in clearing heat and detoxicating, cooling blood and hemostasis, cough-relieving of ventilatining the lung and resolving phlegm Medicine/it is fungivorous.

2. the method as described in claim 1, which is characterized in that the Chinese herbal medicine is selected from gypsum, Radix Rehmanniae, cornu bubali, the coptis, Cape jasmine Son, the root bark of tree peony, radix scutellariae, radix paeoniae rubra, radix scrophulariae, Fructus Forsythiae, rhizoma anemarrhenae, campanulaceae, lophatherum gracile, 8 kinds or more in Radix Glycyrrhizae, add cold ice stone, honeysuckle flower, plate Lan Gen, wild chrysanthemum, folium isatidis, cordate houttuynia, garden burnet, Asian puccoon, rhizome of cyrtomium, the 2-6 kind in Radix Astragali, add radix bupleuri, the root of purple-flowered peucedanum, reed root, pueraria lobata, Blackberry lily, mulberry skin/mulberry leaf, lepidium seed, schizonepeta, Chinese ephedra, sweet wormwood, fritillaria, the 2-6 kind in polygonum cuspidate, amount to 12-29 taste Chinese herbal medicines；Institute State zymophyte be selected from two types leather lid bacterium, foliosous parnassia herb, Pleurotus nebrodensis, needle mushroom, cordyceps sinensis, ganoderma lucidum, Antrodia camphorata, beef-steak fungus, mushroom, Handle mushroom, wild rice bacterium, tabasheer, halimasch, grifola frondosus, east bolt bacterium, Cyathus striatus, Inonotus obliquus, calvatia gigantea, Luo Bakou At least one of mushroom, golden ear, tremella, pisolithus tinctorius Coker et Coucs, black fungus, delicious lactarius.

3. method according to claim 2, which is characterized in that the parts by weight of each component in the Chinese herbal medicine are as follows: gypsum 100- 150, Radix Rehmanniae 20-35, cornu bubali 60-100, coptis 20-35, cape jasmine 20-35, root bark of tree peony 20-35, radix scutellariae 20-40, radix paeoniae rubra 20-35, Radix scrophulariae 20-35, Fructus Forsythiae 20-40, rhizoma anemarrhenae 20-40, campanulaceae 20-35, lophatherum gracile 25-40, Radix Glycyrrhizae 10-20, tremble with fear ice stone 20-30, silver Flower 20-30, Radix Isatidis 30-50, wild chrysanthemum 20-30, folium isatidis 30-60, cordate houttuynia 30-60, garden burnet, Asian puccoon 20-30, rhizome of cyrtomium 20-30, Radix Astragali 25-40, radix bupleuri 20-30, root of purple-flowered peucedanum 20-30, reed root 25-40, pueraria lobata 20-40, blackberry lily 20-30, mulberry skin/mulberry leaf 20-30, lepidium seed 20-30, schizonepeta 20-30, Chinese ephedra 10-20, sweet wormwood 20-40, fritillaria 10-20, polygonum cuspidate 30-50.

4. method as claimed in claim 3, which is characterized in that preferred recipe is gypsum 100-150, Radix Rehmanniae 20-35, cornu bubali 60-100, root bark of tree peony 20-35, radix scutellariae 20-40, radix paeoniae rubra 20-35, radix scrophulariae 20-35, Fructus Forsythiae 20-40, rhizoma anemarrhenae 20-40, campanulaceae 20-35, Lophatherum gracile 25-40, honeysuckle flower 20-30, folium isatidis 30-60, cordate houttuynia 30-60, root of purple-flowered peucedanum 20-30, reed root 25-40, pueraria lobata 20-40, Blackberry lily 20-30, mulberry skin (leaf) 20-30.

5. method according to claim 2, which is characterized in that the zymophyte is selected from delicious lactarius and Jin Er.

6. the method as described in claim 1, which is characterized in that complex enzyme zymohydrolysis is added in the medicinal fungal substance, and complex enzyme is in medicine Property mycoplasma in mass percent be 0.05-0.5%；Again be not suitable in Chinese herbal medicine prescription zymophyte growth medium-height grass of not fermenting Medicine is mixed or is individually extracted through water, and oral solution or granule is made after mixing concentration with raw medicine extracting solution in extracting solution.

7. the method as described in claim 1, which is characterized in that the complex enzyme is that non-starch polysaccharide enzyme and protease press (1- 3): the complex enzyme of 1 mass ratio composition.

8. the method as described in claim 1, which is characterized in that the medicinal fungal substance (is not suitable with hair with not fermented in prescription Yeast-like fungi growth) Chinese herbal medicine crush after mix, obtain mixture；Raw medicine extracting solution and suitable quantity of water are added into mixture, makes to mix The mass ratio of object and water is 1:(2-5)；It is dry after the mycelium self-dissolving of medicinal fungal substance in object to be mixed, it is made after ultramicro grinding Obtain pulvis.