CN108653550A - A kind of fiber crops a kind of reed mentioned in ancient books disappears cough drug and preparation method thereof and detection method - Google Patents

Abstract

The present invention relates to the field of Chinese medicines, and in particular to a kind of fiber crops a kind of reed mentioned in ancient books disappears cough drug and preparation method thereof and detection method.The fiber crops a kind of reed mentioned in ancient books cough drug that disappears is made of the flavour of a drug raw material of following parts by weight：110~125g of Herba ephedrae, 110~125g of the root bark of white mulberry, 225~240g of honeysuckle, 110~125g of radix scutellariae, 205~220g of Thesium chinese, 205~220g of gypsum, 110~125g of fritillaria thunbergii, 110~125g of semen armeniacae amarae, 110~125g of perilla seed, 110~125g of tussilago, 110~125g of lepidium seed, 205~220g of earthworm, 75~90g of pinellia, fry gingko 110~125g of benevolence, 75~90g of honey-fried licorice root.The present invention provides being disappeared the method for cough active constituents of medicine content using hplc simultaneous determination fiber crops a kind of reed mentioned in ancient books, the present invention also provides numb a kind of reed mentioned in ancient books cough drugs that disappears to prepare the new application in treating Chronic Obstructive Pulmonary Disease.

Description

A kind of fiber crops a kind of reed mentioned in ancient books disappears cough drug and preparation method thereof and detection method

Technical field

The present invention relates to the field of Chinese medicines, and in particular to a kind of fiber crops a kind of reed mentioned in ancient books disappears cough drug and preparation method thereof and detection method.

Background technology

Chronic Obstructive Pulmonary Disease (chronic obstructive pulmonary disease, COPD) is a kind of slow Property respiratory disease, using non-specific airway inflammation as the disease of major lesions, feature is in progressive flow limitation, with gas Road height reaction, inflammation cell, inflammatory mediator take part in the pathogenic process of COPD.The incidence of Chronic Obstructive Pulmonary Disease and The death rate rises year by year, and COPD will rise to one of big lethal cause of disease in world three to 2020 for prediction, it has also become one important Public health problem.

In the prior art, the Chinese medicine composition of some treatment Chronic Obstructive Pulmonary Disease is disclosed, such as：

Chinese invention patent disclosed on 01 04th, 2017：A kind of Chinese medicine preparation of Chronic Obstructive Pulmonary Disease, it is open Number：106266501A, a kind of Chronic Obstructive Pulmonary Disease Chinese medicine preparation of the disclosure of the invention, is in including following in parts by weight Medicine ingredient：Cordate houttuynia 30g, radix scutellariae 30g, Fgopyrum dibotrys Hara 30g, campanulaceae 9g, root of purple-flowered peucedanum 9g, fritillaria thunbergii 15g, almond 9g, pinellia 12g。

Chinese invention patent disclosed on 07 27th, 2016：A kind of Chinese medicine preparation for treating chronic obstructive pulmonary disease, it is public The number of opening：105796885A, a kind of Chinese medicine preparation that treating chronic obstructive pulmonary disease of the disclosure of the invention, by following bulk pharmaceutical chemicals system At：35~45 parts of Radix Astragali, 35~45 parts of inular flower, 15~25 parts of Oroxylum indicum, 25~35 parts of cacumen biotae, 25~35 parts of sealwort, female Loyal 35~45 parts of son, 35~45 parts of fritillaria thunbergii, 35~45 parts of Fructus Corni, 25~35 parts of roxburgh rose root, 25~35 parts of sunset abelmoschus root are rung 15~25 parts of bell grass, 25~35 parts of affine cudweed, 15~25 parts of Morinda officinalis.

Chinese invention patent disclosed on 07 06th, 2016：A kind of beach wormwood shellfish trifoliate orange tangerine wood root for treating chronic obstructive pulmonary disease The formula of soup, publication number：105727031A, a kind of beach wormwood shellfish trifoliate orange tangerine wood root soup that treating chronic obstructive pulmonary disease of the disclosure of the invention Formula full Snakegourd Fruit, fritillaria thunbergii, Oroxylum indicum, reed root, the clear water tuber of pinellia, dried orange peel, trifoliate orange have been selected by the reasonable compatibility to Chinese medicine Shell, campanulaceae, Rhizoma Atractylodis Macrocephalae, Poria cocos, are made decoction of medicinal ingredients, have chest qi, reducing phlegm and resolving masses, relieving cough and asthma effect, chronic for treating Obstructive lung disease.

Chinese invention patent disclosed on 09 03rd, 2014：Treat the Chinese traditional medicine composition of type of phlegm-heat stagnating in the lung obstructive emphysema Object and preparation method thereof, publication number：104013888A, the disclosure of the invention is a kind of to treat type of phlegm-heat stagnating in the lung obstructive emphysema Chinese medicine composition and preparation method thereof, wherein Chinese medicine composition includes following raw medicinal material：Fritillaria thunbergii, caulis bambusae in taenian, tabasheer, tangerine Stalk, perilla seed, the tuber of stemona, birthwort, tussilago, the root bark of white mulberry, ardisia japonica, Rhododendron dauricum, Siraitia grosvenorii, rhizoma anemarrhenae, dayflower, radix scutellariae, horse Tail company, Herba Andrographitis, cordate houttuynia, lily, asparagus fern, Radix changii and sealwort.The effect of present invention expels the heat-evil with clearing lung-heat, relieving asthma, Cure mainly chronic obstructive emphysema Phlegm-heat obstructing lung.

Although there is the drug of some treatment Chronic Obstructive Pulmonary Disease in the prior art, the effect is unsatisfactory for it, this hair Bright to provide a kind of Chinese medicine composition, which can effectively treat Chronic Obstructive Pulmonary Disease, curative for effect, slow to treat Property obstructive disease of lung provides a kind of completely new selection.

Applicant inventor has found that the Chinese contained in the peimine and radix scutellariae that contain in fritillaria thunbergii is yellow under study for action A kind of reed mentioned in ancient books glycosides, wogonin, are all that a kind of reed mentioned in ancient books of the invention numb disappears the important activity ingredient of cough drug therapy Chronic Obstructive Pulmonary Disease, so, Applicant inventor has carried out this experimental study repeatedly, proposes and establishes using HPLC-ELSD methods simultaneously to bulb of thunberg fritillary first The method that is detected of content of element, wogonoside, wogonin.

Invention content

Technical problems based on background technology, the present invention propose a kind of numb a kind of reed mentioned in ancient books disappear cough drug and preparation method thereof with Detection method and detection method and pharmaceutical applications.

Disappear cough drug and preparation method thereof the object of the present invention is to provide a kind of numb a kind of reed mentioned in ancient books.

Disappear the detection method of cough drug it is a further object of the present invention to provide numb a kind of reed mentioned in ancient books.

Disappear the new pharmaceutical applications of cough drug the present invention also provides numb a kind of reed mentioned in ancient books.

The purpose of the present invention is what is be accomplished by the following way：

A kind of fiber crops a kind of reed mentioned in ancient books cough drug that disappears is made of the flavour of a drug raw material of following parts by weight：110~125 parts by weight of Herba ephedrae, mulberry 110~125 parts by weight of white skin, 225~240 parts by weight of honeysuckle, 110~125 parts by weight of radix scutellariae, 205~220 weight of Thesium chinese Part, 205~220 parts by weight of gypsum, 110~125 parts by weight of fritillaria thunbergii, 110~125 parts by weight of semen armeniacae amarae, perilla seed 110~ 125 parts by weight, 110~125 parts by weight of tussilago, 110~125 parts by weight of lepidium seed, 205~220 parts by weight of earthworm, pinellia 75~90 parts by weight fry 110~125 parts by weight of gingko benevolence, 75~90 parts by weight of honey-fried licorice root.

Described numb a kind of reed mentioned in ancient books disappear cough drug preferably be made of the flavour of a drug raw material of following parts by weight：117 parts by weight of Herba ephedrae, mulberry 117 parts by weight of white skin, 233 parts by weight of honeysuckle, 117 parts by weight of radix scutellariae, 212 parts by weight of Thesium chinese, 212 parts by weight of gypsum, Zhejiang 117 parts by weight of fritillaria, 117 parts by weight of semen armeniacae amarae, 117 parts by weight of perilla seed, 117 parts by weight of tussilago, 117 weight of lepidium seed Part, 212 parts by weight of earthworm, 83 parts by weight of pinellia fry 117 parts by weight of gingko benevolence, 83 parts by weight of honey-fried licorice root.

A kind of fiber crops a kind of reed mentioned in ancient books disappears the preparation method of cough drug, and the fiber crops a kind of reed mentioned in ancient books cough drug that disappears is flavour of a drug raw material system by following parts by weight At：110~125 parts by weight of Herba ephedrae, 110~125 parts by weight of the root bark of white mulberry, 225~240 parts by weight of honeysuckle, radix scutellariae 110~ 125 parts by weight, 205~220 parts by weight of Thesium chinese, 205~220 parts by weight of gypsum, 110~125 parts by weight of fritillaria thunbergii, semen armeniacae amarae 110~125 parts by weight, 110~125 parts by weight of perilla seed, 110~125 parts by weight of tussilago, 110~125 weight of lepidium seed Part, 205~220 parts by weight of earthworm, 75~90 parts by weight of pinellia fry 110~125 parts by weight of gingko benevolence, honey-fried licorice root 75~90 Parts by weight are prepared with the following method, and steps are as follows：

(1) fritillaria thunbergii crushes, and obtains fritillaria thunbergii fine powder, retains spare；

(2) gypsum adds water to cook secondary, and every time plus 4~6 times of water is measured, 0.5~1.5 hour every time, collecting decoction, filtration, Filtrate retains spare；

(3) radix scutellariae adds water to cook secondary, and for the first time plus 6~10 times of water is measured, and second plus 4~8 times of amounts of water, 1~3 is small every time When, collecting decoction, filtration, filtrate is concentrated into clear cream, with salt acid for adjusting pH value to 1~3, stands, filters to obtain crude extract, with 65~ Washing to pH value is 5~7 to 75% ethyl alcohol by several times, is dried, crushed into fine powder, retains spare；

(4) Herba ephedrae, the root bark of white mulberry, honeysuckle, Thesium chinese, semen armeniacae amarae, perilla seed, tussilago, lepidium seed, earthworm, ginger half Summer fries gingko benevolence, honey-fried licorice root, adds 35~45% ethyl alcohol heating and refluxing extractions three times, for the first time plus 6~10 times of amounts, second, the Three times plus 4~8 times are measured, 0.5~1.5 hour every time, merge extracting solution, filtration, and filtrate merges with the filtrate that step (2) obtains, It is concentrated into clear cream；

(5) by fine powder that fritillaria thunbergii fine powder that step (1) obtains, step (3) obtain, step (4) obtain clear cream mix it is equal It is even, it is dry to get.

Described fiber crops a kind of reed mentioned in ancient books disappears the preparation method of cough drug, and the fiber crops a kind of reed mentioned in ancient books cough drug that disappears is flavour of a drug raw material system by following parts by weight At：117 parts by weight of Herba ephedrae, 117 parts by weight of the root bark of white mulberry, 233 parts by weight of honeysuckle, 117 parts by weight of radix scutellariae, 212 weight of Thesium chinese Measure part, 212 parts by weight of gypsum, 117 parts by weight of fritillaria thunbergii, 117 parts by weight of semen armeniacae amarae, 117 parts by weight of perilla seed, tussilago 117 Parts by weight, 117 parts by weight of lepidium seed, 212 parts by weight of earthworm, 83 parts by weight of pinellia fry 117 parts by weight of gingko benevolence, honey-fried licorice root 83 parts by weight, preferably prepare with the following method, and steps are as follows：

(1) fritillaria thunbergii crushes, and obtains fritillaria thunbergii fine powder, retains spare；

(2) gypsum adds water to cook secondary, and every time plus 5 times of water is measured, and 1 hour every time, collecting decoction, filtration, filtrate retained standby With；

(3) radix scutellariae adds water to cook secondary, and for the first time plus 8 times of amounts of water, second plus 6 times of amounts of water 2 hours every time, merge and decoct Liquid, filtration, filtrate are concentrated into clear cream, with salt acid for adjusting pH value to 2, stand, filter to obtain crude extract, are washed by several times with 70% ethyl alcohol It is 6 to pH value, is dried, crushed into fine powder, retains spare；

(4) Herba ephedrae, the root bark of white mulberry, honeysuckle, Thesium chinese, semen armeniacae amarae, perilla seed, tussilago, lepidium seed, earthworm, ginger half Summer fries gingko benevolence, honey-fried licorice root, adds 40% ethyl alcohol heating and refluxing extraction three times, and for the first time plus 8 times are measured, second, third time plus 6 It measures again, 1 hour every time, merges extracting solution, filtration, filtrate merges with the filtrate that step (2) obtains, and is concentrated into clear cream；

(5) by fine powder that fritillaria thunbergii fine powder that step (1) obtains, step (3) obtain, step (4) obtain clear cream mix it is equal It is even, it is dry to get.

Described numb a kind of reed mentioned in ancient books disappears the preparation method of cough drug, and granule, tablet, hard capsule is made in cough drug that numb a kind of reed mentioned in ancient books can disappear Agent, pill.

A kind of fiber crops a kind of reed mentioned in ancient books disappears the detection method of cough drug, and the fiber crops a kind of reed mentioned in ancient books cough drug that disappears is flavour of a drug raw material system by following parts by weight At：117 parts by weight of Herba ephedrae, 117 parts by weight of the root bark of white mulberry, 233 parts by weight of honeysuckle, 117 parts by weight of radix scutellariae, 212 weight of Thesium chinese Measure part, 212 parts by weight of gypsum, 117 parts by weight of fritillaria thunbergii, 117 parts by weight of semen armeniacae amarae, 117 parts by weight of perilla seed, tussilago 117 Parts by weight, 117 parts by weight of lepidium seed, 212 parts by weight of earthworm, 83 parts by weight of pinellia fry 117 parts by weight of gingko benevolence, honey-fried licorice root 83 parts by weight, the fiber crops a kind of reed mentioned in ancient books cough drug that disappears are prepared with the following method, and steps are as follows：

(1) fritillaria thunbergii crushes, and obtains fritillaria thunbergii fine powder, retains spare；

(2) gypsum adds water to cook secondary, and every time plus 5 times of water is measured, and 1 hour every time, collecting decoction, filtration, filtrate retained standby With；

(3) radix scutellariae adds water to cook secondary, and for the first time plus 8 times of amounts of water, second plus 6 times of amounts of water 2 hours every time, merge and decoct Liquid, filtration, filtrate are concentrated into clear cream, with salt acid for adjusting pH value to 2, stand, filter to obtain crude extract, are washed by several times with 70% ethyl alcohol It is 6 to pH value, is dried, crushed into fine powder, retains spare；

(4) Herba ephedrae, the root bark of white mulberry, honeysuckle, Thesium chinese, semen armeniacae amarae, perilla seed, tussilago, lepidium seed, earthworm, ginger half Summer fries gingko benevolence, honey-fried licorice root, adds 40% ethyl alcohol heating and refluxing extraction three times, and for the first time plus 8 times are measured, second, third time plus 6 It measures again, 1 hour every time, merges extracting solution, filtration, filtrate merges with the filtrate that step (2) obtains, and is concentrated into clear cream；

(5) by fine powder that fritillaria thunbergii fine powder that step (1) obtains, step (3) obtain, step (4) obtain clear cream mix it is equal It is even, it is dry to get；

Numb a kind of reed mentioned in ancient books is measured using HPLC-ELSD methods to disappear the content of peimine in cough drug, wogonoside, wogonin, step It is rapid as follows：

(1) chromatographic condition：C₁₈Chromatographic column, mobile phase are methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution, gradient elution, Eluting order is from 0min to 15min, and the ratio of methanol is from 15% linear rise to 30%, 0.2moLL^-1Trifluoroacetic acid is water-soluble The ratio of liquid is from 85% linear decline to 70%；From 16min to 22min, the ratio of methanol from 30% linear rise to 44%, 0.2moL·L^-1The ratio of trifluoroacetic acid aqueous solution is from 70% linear decline to 56%；From 23min to 32min, the ratio of methanol From 44% linear rise to 54%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution is from 56% linear decline to 46%；From 33min to 50min, the ratio of methanol is from 54% linear rise to 81%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution from 46% linear decline is to 19%；Column temperature is 33~39 DEG C；Flow velocity is 0.5~1.5mLmin^-1；Detector parameters are drift tube temperature Degree is 110~114 DEG C, and carrier gas pressure is 3.0~3.4Lmin^-1；Sample size is 5~15 μ L；

(2) preparation of mixed reference substance solution：Peimine, wogonoside, wogonin reference substance are taken, it is accurately weighed, It is 30 to add volume ratio:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution be made concentration be respectively 0.0500~ 0.0600mg·mL^-1, 0.3100~0.3300mgmL^-1, 0.1800~0.1900mgmL^-1Mixed reference substance solution；

(3) preparation of test solution：Take product to be tested 5.0~7.0g, it is accurately weighed, set in conical flask with cover, precision plus It is 30 to enter volume ratio:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution 50mL, close plug, weighed weight, in 250W, 50kHz Under conditions of, it is ultrasonically treated 25~35min, is let cool, then weighed weight, is 30 with volume ratio:70 methanol -0.2moLL^-1Three Fluoroacetic acid aqueous solution supplies the weight of less loss, shakes up, and filtration, precision measures 25mL, is evaporated, and residue adds water 20mL to make dissolving, uses Water-saturated n-butanol extracts 2~4 times, is every time 25~35mL, merges n-butanol liquid, is washed 2~4 times with ammonia solution, be every time 25~35mL discards ammonia solution, and n-butanol liquid is concentrated to dryness, and residue adds volume ratio to be 30:70 methanol -0.2moLL^-1Trifluoro Acetic acid aqueous solution dissolving and constant volume in 5mL volumetric flasks, shake up to get；

(4) accurate to draw mixed reference substance solution, each 5~15 μ L of test solution, high performance liquid chromatograph is injected, is carried out It measures.

Described numb a kind of reed mentioned in ancient books disappears the detection method of cough drug, and measuring numb a kind of reed mentioned in ancient books using HPLC-ELSD methods disappears bulb of thunberg fritillary first in cough drug The content of element, wogonoside, wogonin, preferred steps are as follows：

(1) chromatographic condition：C18 chromatographic columns, mobile phase are methanol -0.2moLL-1 trifluoroacetic acid aqueous solutions, and gradient is washed De-, eluting order is from 0min to 15min, and the ratio of methanol is from 15% linear rise to 30%, 0.2moLL^-1Trifluoroacetic acid The ratio of aqueous solution is from 85% linear decline to 70%；From 16min to 22min, the ratio of methanol from 30% linear rise to 44%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution is from 70% linear decline to 56%；From 23min to 32min, methanol Ratio is from 44% linear rise to 54%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution is from 56% linear decline to 46%； From 33min to 50min, the ratio of methanol is from 54% linear rise to 81%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution From 46% linear decline to 19%；Column temperature is 36 DEG C；Flow velocity is 1.0mLmin^-1；Detector parameters are that drift tube temperature is 112 DEG C, carrier gas pressure is 3.2Lmin^-1；Sample size is 10 μ L；

(2) preparation of mixed reference substance solution：Peimine, wogonoside, wogonin reference substance are taken, it is accurately weighed, It is 30 to add volume ratio:70 methanol -0.2moLL^-1It is respectively 0.0550mgmL that concentration, which is made, in trifluoroacetic acid aqueous solution^-1、 0.3200mg·mL^-1、0.1850mg·mL^-1Mixed reference substance solution；

(3) preparation of test solution：Product to be tested 6.0g is taken, it is accurately weighed, it sets in conical flask with cover, volume is added in precision Than being 30:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution 50mL, close plug, weighed weight, in 250W, the condition of 50kHz Under, it is ultrasonically treated 30min, is let cool, then weighed weight, is 30 with volume ratio:70 methanol -0.2moLL^-1Trifluoroacetic acid is water-soluble Liquid supplies the weight of less loss, shakes up, and filtration, precision measures 25mL, is evaporated, residue adds water 20mL to make dissolving, with the positive fourth of water saturation Alcohol extracts 3 times, is every time 30mL, merges n-butanol liquid, is washed 3 times with ammonia solution, be every time 30mL, discard ammonia solution, positive fourth Alcohol liquid is concentrated to dryness, and residue adds volume ratio to be 30:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution dissolving and constant volume in In 5mL volumetric flasks, shake up to get；

(4) accurate to draw mixed reference substance solution, each 10 μ L of test solution, high performance liquid chromatograph is injected, is surveyed It is fixed.

A kind of fiber crops a kind of reed mentioned in ancient books cough drug that disappears disappears cough drug preparing the application in treating Chronic Obstructive Pulmonary Disease drug, the fiber crops a kind of reed mentioned in ancient books It is made of the flavour of a drug raw material of following parts by weight：117 parts by weight of Herba ephedrae, 117 parts by weight of the root bark of white mulberry, 233 parts by weight of honeysuckle, 117 parts by weight of radix scutellariae, 212 parts by weight of Thesium chinese, 212 parts by weight of gypsum, 117 parts by weight of fritillaria thunbergii, 117 parts by weight of semen armeniacae amarae, 117 parts by weight of perilla seed, 117 parts by weight of tussilago, 117 parts by weight of lepidium seed, 212 parts by weight of earthworm, 83 parts by weight of pinellia, Fry 117 parts by weight of gingko benevolence, 83 parts by weight of honey-fried licorice root, the fiber crops a kind of reed mentioned in ancient books cough drug that disappears is prepared with the following method, and steps are as follows：

(1) fritillaria thunbergii crushes, and obtains fritillaria thunbergii fine powder, retains spare；

(2) gypsum adds water to cook secondary, and every time plus 5 times of water is measured, and 1 hour every time, collecting decoction, filtration, filtrate retained standby With；

(3) radix scutellariae adds water to cook secondary, and for the first time plus 8 times of amounts of water, second plus 6 times of amounts of water 2 hours every time, merge and decoct Liquid, filtration, filtrate are concentrated into clear cream, with salt acid for adjusting pH value to 2, stand, filter to obtain crude extract, are washed by several times with 70% ethyl alcohol It is 6 to pH value, is dried, crushed into fine powder, retains spare；

(4) Herba ephedrae, the root bark of white mulberry, honeysuckle, Thesium chinese, semen armeniacae amarae, perilla seed, tussilago, lepidium seed, earthworm, ginger half Summer fries gingko benevolence, honey-fried licorice root, adds 40% ethyl alcohol heating and refluxing extraction three times, and for the first time plus 8 times are measured, second, third time plus 6 It measures again, 1 hour every time, merges extracting solution, filtration, filtrate merges with the filtrate that step (2) obtains, and is concentrated into clear cream；

(5) by fine powder that fritillaria thunbergii fine powder that step (1) obtains, step (3) obtain, step (4) obtain clear cream mix it is equal It is even, it is dry to get；

Numb a kind of reed mentioned in ancient books is measured using HPLC-ELSD methods to disappear the content of peimine in cough drug, wogonoside, wogonin, step It is rapid as follows：

(1) chromatographic condition：C₁₈Chromatographic column, mobile phase are methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution, gradient elution, Eluting order is from 0min to 15min, and the ratio of methanol is from 15% linear rise to 30%, 0.2moLL^-1Trifluoroacetic acid is water-soluble The ratio of liquid is from 85% linear decline to 70%；From 16min to 22min, the ratio of methanol from 30% linear rise to 44%, 0.2moL·L^-1The ratio of trifluoroacetic acid aqueous solution is from 70% linear decline to 56%；From 23min to 32min, the ratio of methanol From 44% linear rise to 54%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution is from 56% linear decline to 46%；From 33min to 50min, the ratio of methanol is from 54% linear rise to 81%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution from 46% linear decline is to 19%；Column temperature is 36 DEG C；Flow velocity is 1.0mLmin^-1；Detector parameters are that drift tube temperature is 112 DEG C, carrier gas pressure is 3.2Lmin^-1；Sample size is 10 μ L；

(2) preparation of mixed reference substance solution：Peimine, wogonoside, wogonin reference substance are taken, it is accurately weighed, It is 30 to add volume ratio:70 methanol -0.2moLL^-1It is respectively 0.0550mgmL that concentration, which is made, in trifluoroacetic acid aqueous solution^-1、 0.3200mg·mL^-1、0.1850mg·mL^-1Mixed reference substance solution；

(3) preparation of test solution：Product to be tested 6.0g is taken, it is accurately weighed, it sets in conical flask with cover, volume is added in precision Than being 30:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution 50mL, close plug, weighed weight, in 250W, the condition of 50kHz Under, it is ultrasonically treated 30min, is let cool, then weighed weight, is 30 with volume ratio:70 methanol -0.2moLL^-1Trifluoroacetic acid is water-soluble Liquid supplies the weight of less loss, shakes up, and filtration, precision measures 25mL, is evaporated, residue adds water 20mL to make dissolving, with the positive fourth of water saturation Alcohol extracts 3 times, is every time 30mL, merges n-butanol liquid, is washed 3 times with ammonia solution, be every time 30mL, discard ammonia solution, positive fourth Alcohol liquid is concentrated to dryness, and residue adds volume ratio to be 30:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution dissolving and constant volume in In 5mL volumetric flasks, shake up to get；

(4) accurate to draw mixed reference substance solution, each 10 μ L of test solution, high performance liquid chromatograph is injected, is surveyed It is fixed.

Following experimental study is used to verify the technology contents and technique effect of technical scheme of the present invention, but is not intended to limit this The protection domain of invention.

Experiment one：Numb a kind of reed mentioned in ancient books disappears influence of the cough drug to Chronic Obstructive Pulmonary Disease animal model

This experiment closes the method that lipopolysaccharides air flue instills by sootiness and establishes rat COPD model, and drug application treats one Month, observe its influence to COPD inflammatory reactions and respiratory function.

1 material

1.1 drug

(Shanghai Xinyi Tianping Pharmaceutical Co., Ltd. produces aminophylline tablets, authentication code：Chinese medicines quasi-word H31021799, rule Lattice：0.1g)；IL-1 β, TNF-α, IL-6 radioimmunological kits (manufacture of object Science and Technology Ltd. of upper Haikang nangzan).

Fiber crops a kind of reed mentioned in ancient books of the invention disappears cough drug, is produced by Tian Changyifan pharmaceutical Co. Ltds, prescription and preparation method are as follows：

Prescription：Herba ephedrae 117g, root bark of white mulberry 117g, honeysuckle 233g, radix scutellariae 117g, Thesium chinese 212g, gypsum 212g, Zhejiang Fritillaria 117g, semen armeniacae amarae 117g, perilla seed 117g, tussilago 117g, lepidium seed 117g, earthworm 212g, pinellia 83g are fried white Kernel 117g, honey-fried licorice root 83g.

Preparation method：

(1) fritillaria thunbergii crushes, and obtains fritillaria thunbergii fine powder, retains spare；

(2) gypsum adds water to cook secondary, and every time plus 5 times of water is measured, and 1 hour every time, collecting decoction, filtration, filtrate retained standby With；

(3) radix scutellariae adds water to cook secondary, and for the first time plus 8 times of amounts of water, second plus 6 times of amounts of water 2 hours every time, merge and decoct Liquid, filtration, filtrate are concentrated into clear cream, with salt acid for adjusting pH value to 2, stand, filter to obtain crude extract, are washed by several times with 70% ethyl alcohol It is 6 to pH value, is dried, crushed into fine powder, retains spare；

(4) Herba ephedrae, the root bark of white mulberry, honeysuckle, Thesium chinese, semen armeniacae amarae, perilla seed, tussilago, lepidium seed, earthworm, ginger half Summer fries gingko benevolence, honey-fried licorice root, adds 40% ethyl alcohol heating and refluxing extraction three times, and for the first time plus 8 times are measured, second, third time plus 6 It measures again, 1 hour every time, merges extracting solution, filtration, filtrate merges with the filtrate that step (2) obtains, and is concentrated into clear cream；

(5) by fine powder that fritillaria thunbergii fine powder that step (1) obtains, step (3) obtain, step (4) obtain clear cream mix it is equal It is even, it is dry to get.

1.2 animal

SD rats, male, 180~220g of weight are provided, animal productiong purchased from Anhui University of Chinese Medecine's Experimental Animal Center Credit number：SCXK (Anhui) 2012-2013.SPF grades.Animal feeding is under standard conditions, 12h light and shades alternating, 22 ± 2 DEG C, from By drinking-water of ingesting.Animal starts to test in above-mentioned environment after adaptability is raised 1 week.

1.3 reagents and instrument

Mount Huang board cigarette, tar content 11mg, nicotine content in smoke 1.0mg；16/35 Biological Signal Collecting Systems of PowerLab, Anhui Tobacco Industry Co., Ltd's Bengbu Cigarette Factory production；ALC-V8 animal respirators, Shanghai Alcott bio tech ltd Manufacture；Full automatic biochemical apparatus, the biotech inc Ku Beier of Shenzhen.

2 methods

2.1 groupings and modeling

Male SD rat is randomly divided into 6 groups, every group 10, respectively normal group, model group, aminophylline group (dosage For 0.050gkg^-1), numb a kind of reed mentioned in ancient books disappears cough drug high dose group (dosage 3.00gkg^-1), numb a kind of reed mentioned in ancient books disappears cough drug middle dosage Group (dosage 1.50gkg^-1), numb a kind of reed mentioned in ancient books disappears cough drug low dose group (dosage 0.75gkg^-1)。

LPS is configured to 1mgmL with sterile saline^-1, in testing 1d, 14d is with 10% chloraldurate 0.35g kg^-1Ip anesthetized rats, neck median incision 2cm detach tracheae, and syringe is from tracheae to lung direction instillation LPS (200 μ g/200 μ L), rat is upright, it rotates left and right, lipopolysaccharides is made to be uniformly distributed in intrapulmonary.Rat is put into perfume (or spice) by 2~30d (except 14d) It is smoked 1h/ times in cigarette smog, 2 d^-1, COPD models are replicated, to reduce influence of the produced vapor of cigarette combustion to rat, Proper silica gel drier is placed in bottom.Normal group：In 1d, 14d is the same as method tracheal instillation physiological saline not sootiness.

Numb a kind of reed mentioned in ancient books disappears each dosage group of cough drug and aminophylline group 1d rises by 10mLkg^-1Dosage gastric infusion, normal group and Model group gavage gives equivalent 0.5%CMC-Na solution, 1 time/d, total 30d.After last dose 30min, ip10% chloraldurates 0.35g·kg^-1Anesthetized rat is drawn materials.

2.2 Testing index

2.2.1 pulmonary function detection

By rat with 10% chloraldurate 0.35gkg^-1Anesthesia, longitudinal incision skin of neck, exposure tracheae are simultaneously open, insert Enter to be connected with the trachea cannula of three-way cock, one end connects animal respirator, and one end connection signal acquisition system measures breathing and refers to Mark.Measure the 0.th 3 second forced expiratory volume (FEV0.3), forced vital capacity (FVC), FEV0.3/FVC and End--tidal PCO_2 (PEF) parameter values such as.

2.2.2 serum cytokines measure IL-1 β and IL-6, TNF-a

It has surveyed lung function rat and has opened abdominal cavity, abdominal aorta blood sampling, with 3000rmin at 4 DEG C rapidly^-1Speed, from Heart 15min detaches serum, and -70 DEG C of preservations carry out IL-1 β and IL-6 according to kit illustration method, and TNF-a is measured.

2.2.3 lung morphology inspection

After each group animal is put to death, thoracic cavity is opened rapidly, and the taking-up middle lobe of right lung is placed in 10% formaldehyde fixed, conventional gradients second Dehydration of alcohols, paraffin embedding, slice, HE dyeing, light microscopic observation pathologic change.

2.3 statistical method

Data are indicated with mean ± standard deviationQuantitative response statistics method is examined using one-way analysis of variance Test method, statistic software SPSS 17.0.

3 results

3.1 numb a kind of reed mentioned in ancient books disappear influence of the cough drug to chronic obstructive induced lung injury in rats lung function

Compared with normal group, model group FVC, FEV0.3, FEV0.3/FVC, PEF are decreased obviously, P<0.01, show modeling Success.

Disappear high cough drug, middle dose group and aminophylline group FVC, FEV0.3, FEV0.3/FVC, PEF of numb a kind of reed mentioned in ancient books has compared with model group It is obviously improved (P<0.01, P<0.05).It the results are shown in Table 1.

The numb a kind of reed mentioned in ancient books of table 1 disappears influence of the cough drug to Chronic Obstructive Pulmonary Disease pulmonary function

Note:Compared with normal group^#P ＜ 0.05,^##P ＜ 0.01；Compared with model group^#P ＜ 0.05,^##P ＜ 0.01.

3.2 numb a kind of reed mentioned in ancient books disappear cough drug to -1 β and IL-6 of chronic obstructive induced lung injury in rats serum IL, the influence of TNF-a contents

The results are shown in Table 2, and compared with normal group, model group serum IL -1 β and IL-6, TNF-a content are significantly raised, P< 0.01.Numb a kind of reed mentioned in ancient books disappears cough drug high dose group, middle dose group and aminophylline group and model group comparing difference significantly (P<0.01, P< 0.05), have to improve and reduce cytokine levels effect.It the results are shown in Table 2.

The numb a kind of reed mentioned in ancient books of table 2. disappears influence of the cough drug to the rats with chronic obstructive pulmonary factor

Note:Compared with normal group^#P ＜ 0.05,^##P ＜ 0.01；Compared with model group^#P ＜ 0.05,^##P ＜ 0.01.

3.3 numb a kind of reed mentioned in ancient books disappear influence of the cough drug to chronic obstructive induced lung injury in rats lung morphology

Normal rats lung morphology is without exception；Model group rats alveolar wall hyperemia is apparent, a large amount of cell infiltrations, Goblet cell quantity increases, and mucous secretion is caused to increase.Aminophylline group induced lung steep that wall is slight or mild hyperaemia, with slight Or slight cell infiltration；Numb a kind of reed mentioned in ancient books disappears, and cough drug is high, middle dose group various pulmonary lesions degree is substantially reduced compared with model group, alveolar wall Slight or mild hyperaemia, slight cell infiltration.As a result see Figure of description 1, attached drawing 2, attached drawing 3, attached drawing 4, attached drawing 5, Fig. 6.

4 conclusions

This result of study shows that numb a kind of reed mentioned in ancient books cough drug that disappears can reduce serum levels of inflammatory cytokines IL-1 β and IL-6, and TNF-a contains Amount, shows it with anti-inflammatory effect and improves COPD rat models lung function and histology pathological index, improves COPD moulds Type surviving rats symptom.

Experiment two：HPLC-ELSD methods measure numb a kind of reed mentioned in ancient books and disappear the content of peimine in cough drug, wogonoside, wogonin

Applicant inventor has found under study for action, the Zhejiang contained in fritillaria thunbergii in numb a kind of reed mentioned in ancient books provided by the invention disappears cough drug Wogonoside, the wogonin contained in shellfish A prime and radix scutellariae is all that a kind of reed mentioned in ancient books of the invention numb disappears cough drug therapy chronic obstructive The important activity ingredient of lung disease, so, applicant inventor has carried out this experimental study repeatedly, proposes and establish to adopt With the HPLC-ELSD methods method that is detected to the content of peimine, wogonoside, wogonin simultaneously, disappear cough for numb a kind of reed mentioned in ancient books The detection of drug provides experimental basis.

1 instrument and reagent

1.1 instrument

SHZ-D (III) type circulating water type vacuum pump (is manufactured, from Hefei state wound point by Shanghai Qiang Qiang industry developments Co., Ltd Analysis Instrument Ltd. buys)；HH-6 type digital control constant temperatures water-bath (manufacture of analytical instrument Manufacturing Co., Ltd is led to by Changzhou hundred million, From Hefei, Guo Chuan Analytical Instrument Co., Ltd buys)；U-3000 type Shimadzus high performance liquid chromatograph (6000 type ELSD detectors, U.S.'s match is silent to fly, and from Hefei, Guo Chuan Analytical Instrument Co., Ltd buys)；KH-2200DB type numerical control ultrasonic cleaners are (by Taiwan This special experimental instruments and equipment limited manufacture, from Hefei, Guo Chuan Analytical Instrument Co., Ltd buys)；Electronic analytical balance is (by Germany Sai Duolisi is manufactured, and from Hefei, Guo Chuan Analytical Instrument Co., Ltd buys).

1.2 reagent

Fiber crops a kind of reed mentioned in ancient books of the invention disappears cough drug, is produced by Tian Changyifan pharmaceutical Co. Ltds, prescription and preparation method are as follows：

Prescription：Herba ephedrae 117g, root bark of white mulberry 117g, honeysuckle 233g, radix scutellariae 117g, Thesium chinese 212g, gypsum 212g, Zhejiang Fritillaria 117g, semen armeniacae amarae 117g, perilla seed 117g, tussilago 117g, lepidium seed 117g, earthworm 212g, pinellia 83g are fried white Kernel 117g, honey-fried licorice root 83g.

Preparation method：

(1) fritillaria thunbergii crushes, and obtains fritillaria thunbergii fine powder, retains spare；

(2) gypsum adds water to cook secondary, and every time plus 5 times of water is measured, and 1 hour every time, collecting decoction, filtration, filtrate retained standby With；

(3) radix scutellariae adds water to cook secondary, and for the first time plus 8 times of amounts of water, second plus 6 times of amounts of water 2 hours every time, merge and decoct Liquid, filtration, filtrate are concentrated into clear cream, with salt acid for adjusting pH value to 2, stand, filter to obtain crude extract, are washed by several times with 70% ethyl alcohol It is 6 to pH value, is dried, crushed into fine powder, retains spare；

(4) Herba ephedrae, the root bark of white mulberry, honeysuckle, Thesium chinese, semen armeniacae amarae, perilla seed, tussilago, lepidium seed, earthworm, ginger half Summer fries gingko benevolence, honey-fried licorice root, adds 40% ethyl alcohol heating and refluxing extraction three times, and for the first time plus 8 times are measured, second, third time plus 6 It measures again, 1 hour every time, merges extracting solution, filtration, filtrate merges with the filtrate that step (2) obtains, and is concentrated into clear cream；

(5) by fine powder that fritillaria thunbergii fine powder that step (1) obtains, step (3) obtain, step (4) obtain clear cream mix it is equal It is even, it is dry to get.

Peimine standard items (Heng Hua biological products Co., Ltd buys from Nantong, purity 99.6%), wogonoside mark Quasi- product (Zhi Run Science and Technology Ltd.s buy from Jiangsu, purity 98.7%), wogonin standard items are (from the permanent magnificent biological products in Nantong Co., Ltd buys, purity 99.2%), acetonitrile is chromatographically pure (from Jiangsu, Zhi Run Science and Technology Ltd.s buy)；Methanol is chromatography Pure (from Jiangsu, Zhi Run Science and Technology Ltd.s buy).

2 methods and result

2.1 chromatographic conditions and system suitability

WondaSil C₁₈Chromatographic column (4.6mm × 250mm, 5 μm), mobile phase are methanol -0.2moLL^-1Trifluoroacetic acid Aqueous solution, gradient elution, eluting order be from 0min to 15min, the ratio of methanol from 15% linear rise to 30%, 0.2moL·L^-1The ratio of trifluoroacetic acid aqueous solution is from 85% linear decline to 70%；From 16min to 22min, the ratio of methanol From 30% linear rise to 44%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution is from 70% linear decline to 56%；From 23min to 32min, the ratio of methanol is from 44% linear rise to 54%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution from 56% linear decline is to 46%；From 33min to 50min, the ratio of methanol is from 54% linear rise to 81%, 0.2moLL^-1Three The ratio of fluoroacetic acid aqueous solution is from 46% linear decline to 19%；Column temperature is 36 DEG C；Flow velocity is 1.0mLmin^-1；Detector is joined It is 112 DEG C that number, which is drift tube temperature, and carrier gas pressure is 3.2Lmin^-1；Sample size is 10 μ L.It is surveyed by above-mentioned chromatographic condition Fixed, the separating degree of peimine is 7.23, the separating degree of wogonoside is 7.58, wogonin separating degree is 6.85, separation effect Fruit is preferable.The theoretical number of plates, which is calculated, by peimine, wogonoside, wogonin peak is not less than 5000.

The preparation of 2.2 mixed reference substance solutions

Peimine, wogonoside, wogonin reference substance are taken, accurately weighed, it is 30 to add volume ratio:70 methanol- 0.2moL·L^-1It is respectively 0.0550mgmL that concentration, which is made, in trifluoroacetic acid aqueous solution^-1、0.3200mg·mL^-1、0.1850mg· mL^-1Mixed reference substance solution.

The preparation of 2.3 test solutions

This product 6.0g is taken, it is accurately weighed, it sets in conical flask with cover, it is 30 that volume ratio, which is added, in precision:70 methanol- 0.2moL·L^-1Trifluoroacetic acid aqueous solution 50mL, close plug, weighed weight are ultrasonically treated under conditions of 250W, 50kHz 30min is let cool, then weighed weight, is 30 with volume ratio:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution supplies less loss Weight, shake up, filter, precision measure 25mL, be evaporated, residue adds water 20mL to make dissolving, with water-saturated n-butanol extract 3 times, It is 30mL every time, merges n-butanol liquid, washed 3 times with ammonia solution, be every time 30mL, discard ammonia solution, n-butanol liquid is concentrated into Dry, residue adds volume ratio to be 30:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution dissolves and constant volume is in 5mL volumetric flasks, Shake up to get.

The preparation of 2.4 negative control solutions

Prepare with identical technique by recipe quantity and respectively scarce fritillaria thunbergii, radix scutellariae negative control sample, it is molten according to test sample Liquid and preparation method thereof processing, obtains negative control solution.

2.5 specificities are tested

Precision draws mixing reference substance, test sample and each 10 μ L of negative control solution, and injection liquid chromatograph is measured. As a result in negative control chromatography, to, without corresponding chromatographic peak, show it at reference substance and the corresponding retention time of test sample chromatogram Its component to peimine in the party, wogonoside, the measurement of wogonin is noiseless.See Figure of description 7, attached drawing 8, attached drawing 9。

2.6 linear relationships are investigated

By chromatographic condition provided by the invention, take reference substance solution, respectively use autosampler into 5 μ L, 10 μ L, 15 μ L, 20 μ L, 25 μ L, 30 μ L measure peak area value, record chromatogram.With sample size (g) for abscissa, corresponding peak area is vertical sits Mark, it is as follows to obtain equation of linear regression：

Peimine：Y=23688X+146853 (R²=0.9997)；Sample size is in 0.394~1.215 μ g ranges interior lines Sexual intercourse is good.

Wogonoside：Y=61205X+475863 (R²=0.9998)；Sample size is in 2.368~6.854 μ g ranges interior lines Sexual intercourse is good

Wogonin：Y=51283X+842682 (R²=0.9997)；Sample size is in 1.2665~3.856 μ g ranges interior lines Sexual intercourse is good.

2.7 precision test

Precision claims 20 μ L of reference substance solution, and by chromatographic condition continuous sample introduction provided by the invention 6 times, record fiber crops a kind of reed mentioned in ancient books disappears cough medicine Peimine, wogonoside, wogonin peak area in object, the results showed that, the RSD% of peimine peak area is 1.52%, The RSD% of wogonoside peak area is 1.63%, the RSD% of wogonin peak area is 1.22%.Show that precision is good.

2.8 stability test

Same test solution is taken, respectively at 0h, 1h, 2h, 4h, 8h, 16h after preparation, injects high performance liquid chromatography Instrument is measured the peak area value of peimine, wogonoside, wogonin, bulb of thunberg fritillary first by chromatographic condition provided by the invention The RSD% of plain peak area value is 1.23%, the RSD% of wogonoside peak area value is 1.39%, wogonin peak area value RSD% is 1.57%, shows that peimine, wogonoside, wogonin are good in 16h internal stabilities.

2.9 reappearance test

It takes and disappears cough drug with a batch of numb a kind of reed mentioned in ancient books, press method provided by the invention 6 parts of test solutions of parallel preparations, press The chromatographic condition that invention provides measures, and calculates the average mass fraction of peimine, wogonoside, wogonin, peimine Average content be 0.097mgg^-1, RSD% 1.36%；The average content of wogonoside is 1.582mgg^-1, RSD% It is 1.41%；The average content of wogonin is 0.052mgg^-1, RSD% 1.68%.

2.10 sample recovery rate is tested

Taking numb a kind of reed mentioned in ancient books to be measured of known content to disappear respectively, (wherein peimine content is 0.095mg to cough drug 4.500g g^-1, wogonoside 1.582mgg^-1, wogonin content is 0.052mgg^-1), it is totally 6 parts, accurately weighed, respectively Peimine reference substance, wogonoside reference substance, wogonin reference substance (peimine, wogonoside, the wogonin of addition Concentration is respectively 0.0525mgmL^-1、0.2500mg·mL^-1、0.1520mg·mL^-1), it is molten according to test sample provided by the invention The preparation method and chromatographic condition of liquid are measured, the results show that the sample-adding average recovery rate of peimine is 99.91%, RSD% is 1.32%, and the sample-adding average recovery rate of wogonoside is 99.77%, RSD% 1.53%, the sample-adding of wogonin Average recovery rate is 102.86%, RSD% 1.64%.The RSD% of the rate of recovery of method provided by the invention is respectively less than 3%, Therefore meet quantitative analysis requirement with this method test result.Experimental result is shown in Table 3.

3 sample recovery rate experimental result (n=6) of table

The measurement result of 2.11 samples

It takes 3 batches of numb a kind of reed mentioned in ancient books to disappear cough drug sample, is prepared according to above-mentioned test solution preparation method, in above-mentioned HPLC chromatogram item Chromatographic peak area is measured and recorded under part, is computed, and experimental result is shown in Table 4.

4 sample size measurement result (mgg of table^-1)

3 conclusions

This research establishes numb a kind of reed mentioned in ancient books and disappears the content assaying method of peimine in cough drug, wogonoside, wogonin 3, Method stablizes feasible, high-specificity, and precision is high, reproducible, and other components interference is small, and the rate of recovery is high, can be used for numb a kind of reed mentioned in ancient books and disappears The detection of cough drug.

Description of the drawings：

Fig. 1 is that normal rats lung morphology HE dyes micrograph (200 ×)

Fig. 2 is that model group rats lung morphology HE dyes micrograph (200 ×)

Fig. 3 is that aminophylline group lung tissue of rats morphology HE dyes micrograph (200 ×)

Fig. 4 is that numb a kind of reed mentioned in ancient books cough drug high dose group lung tissue of rats morphology HE that disappears dyes micrograph (200 ×)

Fig. 5 is that numb a kind of reed mentioned in ancient books cough drug middle dose group lung tissue of rats morphology HE that disappears dyes micrograph (200 ×)

Fig. 6 is that numb a kind of reed mentioned in ancient books cough drug low dose group lung tissue of rats morphology HE that disappears dyes micrograph (200 ×)

Fig. 7 is mixed reference substance solution HPLC figures, wherein No. 1 peak is peimine；No. 2 peaks are wogonoside；No. 3 peaks are Wogonin

Fig. 8 is test solution HPLC figures, wherein No. 1 peak is peimine；No. 2 peaks are wogonoside；No. 3 peaks are that the Chinese is yellow A kind of reed mentioned in ancient books element

Fig. 9 schemes for double-negative reference substance solution HPLC.

Specific implementation mode

The present invention is made further to explain with reference to specific embodiment.

Embodiment 1：Numb a kind of reed mentioned in ancient books disappears cough drug

Prescription：Herba ephedrae 117g, root bark of white mulberry 117g, honeysuckle 233g, radix scutellariae 117g, Thesium chinese 212g, gypsum 212g, Zhejiang Fritillaria 117g, semen armeniacae amarae 117g, perilla seed 117g, tussilago 117g, lepidium seed 117g, earthworm 212g, pinellia 83g are fried white Kernel 117g, honey-fried licorice root 83g.

Preparation method：

(1) fritillaria thunbergii crushes, and obtains fritillaria thunbergii fine powder, retains spare；

(2) gypsum adds water to cook secondary, and every time plus 5 times of water is measured, and 1 hour every time, collecting decoction, filtration, filtrate retained standby With；

(3) radix scutellariae adds water to cook secondary, and for the first time plus 8 times of amounts of water, second plus 6 times of amounts of water 2 hours every time, merge and decoct Liquid, filtration, filtrate are concentrated into clear cream, with salt acid for adjusting pH value to 2, stand, filter to obtain crude extract, are washed by several times with 70% ethyl alcohol It is 6 to pH value, is dried, crushed into fine powder, retains spare；

(4) Herba ephedrae, the root bark of white mulberry, honeysuckle, Thesium chinese, semen armeniacae amarae, perilla seed, tussilago, lepidium seed, earthworm, ginger half Summer fries gingko benevolence, honey-fried licorice root, adds 40% ethyl alcohol heating and refluxing extraction three times, and for the first time plus 8 times are measured, second, third time plus 6 It measures again, 1 hour every time, merges extracting solution, filtration, filtrate merges with the filtrate that step (2) obtains, and is concentrated into clear cream；

(5) by fine powder that fritillaria thunbergii fine powder that step (1) obtains, step (3) obtain, step (4) obtain clear cream mix it is equal It is even, it is dry to get.

Numb a kind of reed mentioned in ancient books is measured using HPLC-ELSD methods to disappear the content of peimine in cough drug, wogonoside, wogonin, step It is rapid as follows：

(1) chromatographic condition：C₁₈Chromatographic column, mobile phase are methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution, gradient elution, Eluting order is from 0min to 15min, and the ratio of methanol is from 15% linear rise to 30%, 0.2moLL^-1Trifluoroacetic acid is water-soluble The ratio of liquid is from 85% linear decline to 70%；From 16min to 22min, the ratio of methanol from 30% linear rise to 44%, 0.2moL·L^-1The ratio of trifluoroacetic acid aqueous solution is from 70% linear decline to 56%；From 23min to 32min, the ratio of methanol From 44% linear rise to 54%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution is from 56% linear decline to 46%；From 33min to 50min, the ratio of methanol is from 54% linear rise to 81%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution from 46% linear decline is to 19%；Column temperature is 36 DEG C；Flow velocity is 1.0mLmin^-1；Detector parameters are that drift tube temperature is 112 DEG C, carrier gas pressure is 3.2Lmin^-1；Sample size is 10 μ L；

(2) preparation of mixed reference substance solution：Peimine, wogonoside, wogonin reference substance are taken, it is accurately weighed, It is 30 to add volume ratio:70 methanol -0.2moLL^-1It is respectively 0.0550mgmL that concentration, which is made, in trifluoroacetic acid aqueous solution^-1、 0.3200mg·mL^-1、0.1850mg·mL^-1Mixed reference substance solution；

(3) preparation of test solution：Product to be tested 6.0g is taken, it is accurately weighed, it sets in conical flask with cover, volume is added in precision Than being 30:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution 50mL, close plug, weighed weight, in 250W, the condition of 50kHz Under, it is ultrasonically treated 30min, is let cool, then weighed weight, is 30 with volume ratio:70 methanol -0.2moLL^-1Trifluoroacetic acid is water-soluble Liquid supplies the weight of less loss, shakes up, and filtration, precision measures 25mL, is evaporated, residue adds water 20mL to make dissolving, with the positive fourth of water saturation Alcohol extracts 3 times, is every time 30mL, merges n-butanol liquid, is washed 3 times with ammonia solution, be every time 30mL, discard ammonia solution, positive fourth Alcohol liquid is concentrated to dryness, and residue adds volume ratio to be 30:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution dissolving and constant volume in In 5mL volumetric flasks, shake up to get；

(4) accurate to draw mixed reference substance solution, each 10 μ L of test solution, high performance liquid chromatograph is injected, is surveyed It is fixed；

(5) measurement result：Every gram of numb a kind of reed mentioned in ancient books disappears cough drug 0.096mg containing peimine, wogonoside 1.585mg, Chinese radix scutellariae Plain 0.057mg.

The foregoing is only a preferred embodiment of the present invention, but scope of protection of the present invention is not limited thereto, Any one skilled in the art in the technical scope disclosed by the present invention, according to the technique and scheme of the present invention and its Inventive concept is subject to equivalent substitution or change, should be covered by the protection scope of the present invention.

Claims (8)

1. a kind of fiber crops a kind of reed mentioned in ancient books disappears cough drug, which is characterized in that the fiber crops a kind of reed mentioned in ancient books cough drug that disappears is made of the flavour of a drug raw material of following parts by weight： 110~125 parts by weight of Herba ephedrae, 110~125 parts by weight of the root bark of white mulberry, 225~240 parts by weight of honeysuckle, 110~125 weight of radix scutellariae Amount part, 205~220 parts by weight of Thesium chinese, 205~220 parts by weight of gypsum, 110~125 parts by weight of fritillaria thunbergii, semen armeniacae amarae 110~ 125 parts by weight, 110~125 parts by weight of perilla seed, 110~125 parts by weight of tussilago, 110~125 parts by weight of lepidium seed, earthworm 205~220 parts by weight, 75~90 parts by weight of pinellia fry 110~125 parts by weight of gingko benevolence, 75~90 parts by weight of honey-fried licorice root.

The cough drug 2. fiber crops a kind of reed mentioned in ancient books as described in claim 1 disappears, which is characterized in that the fiber crops a kind of reed mentioned in ancient books cough drug that disappears is by following parts by weight Flavour of a drug raw material is made：117 parts by weight of Herba ephedrae, 117 parts by weight of the root bark of white mulberry, 233 parts by weight of honeysuckle, 117 parts by weight of radix scutellariae, hundred Stamen 212 parts by weight of grass, 212 parts by weight of gypsum, 117 parts by weight of fritillaria thunbergii, 117 parts by weight of semen armeniacae amarae, 117 parts by weight of perilla seed, 117 parts by weight of tussilago, 117 parts by weight of lepidium seed, 212 parts by weight of earthworm, 83 parts by weight of pinellia fry 117 weight of gingko benevolence Part, 83 parts by weight of honey-fried licorice root.

The preparation method of cough drug 3. a kind of fiber crops a kind of reed mentioned in ancient books disappears, the fiber crops a kind of reed mentioned in ancient books cough drug that disappears are made of the flavour of a drug raw material of following parts by weight： 110~125 parts by weight of Herba ephedrae, 110~125 parts by weight of the root bark of white mulberry, 225~240 parts by weight of honeysuckle, 110~125 weight of radix scutellariae Amount part, 205~220 parts by weight of Thesium chinese, 205~220 parts by weight of gypsum, 110~125 parts by weight of fritillaria thunbergii, semen armeniacae amarae 110~ 125 parts by weight, 110~125 parts by weight of perilla seed, 110~125 parts by weight of tussilago, 110~125 parts by weight of lepidium seed, earthworm 205~220 parts by weight, 75~90 parts by weight of pinellia, 110~125 parts by weight of stir-fry gingko benevolence, 75~90 parts by weight of honey-fried licorice root, It is characterized in that, the fiber crops a kind of reed mentioned in ancient books disappears, cough drug is prepared with the following method, and steps are as follows：

(1) fritillaria thunbergii crushes, and obtains fritillaria thunbergii fine powder, retains spare；

(2) gypsum adds water to cook secondary, and every time plus 4~6 times of water is measured, 0.5~1.5 hour every time, collecting decoction, filtration, filtrate Retain spare；

(3) radix scutellariae adds water to cook secondary, and for the first time plus 6~10 times of water is measured, second plus 4~8 times of amounts of water, 1~3 hour every time, Collecting decoction, filtration, filtrate is concentrated into clear cream, with salt acid for adjusting pH value to 1~3, stands, filters to obtain crude extract, with 65~ Washing to pH value is 5~7 to 75% ethyl alcohol by several times, is dried, crushed into fine powder, retains spare；

(4) Herba ephedrae, the root bark of white mulberry, honeysuckle, Thesium chinese, semen armeniacae amarae, perilla seed, tussilago, lepidium seed, earthworm, pinellia, stir-fry Gingko benevolence, honey-fried licorice root add 35~45% ethyl alcohol heating and refluxing extractions three times, and for the first time plus 6~10 times are measured, second, third time Add 4~8 times of amounts, 0.5~1.5 hour every time, merges extracting solution, filtration, filtrate merges with the filtrate that step (2) obtains, and concentrates To clear cream；

(5) fine powder that fritillaria thunbergii fine powder that step (1) obtains, step (3) obtain, step (4) are obtained clear cream and be uniformly mixed, done It is dry to get.

4. fiber crops a kind of reed mentioned in ancient books disappears the preparation method of cough drug as claimed in claim 3, the fiber crops a kind of reed mentioned in ancient books cough drug that disappears is by the medicine of following parts by weight Taste raw material is made：117 parts by weight of Herba ephedrae, 117 parts by weight of the root bark of white mulberry, 233 parts by weight of honeysuckle, 117 parts by weight of radix scutellariae, hundred stamens Careless 212 parts by weight, 212 parts by weight of gypsum, 117 parts by weight of fritillaria thunbergii, 117 parts by weight of semen armeniacae amarae, 117 parts by weight of perilla seed, money 117 parts by weight of coltsfoot, 117 parts by weight of lepidium seed, 212 parts by weight of earthworm, 83 parts by weight of pinellia fry 117 parts by weight of gingko benevolence, 83 parts by weight of honey-fried licorice root, which is characterized in that the fiber crops a kind of reed mentioned in ancient books cough drug that disappears is prepared with the following method, and steps are as follows：

(1) fritillaria thunbergii crushes, and obtains fritillaria thunbergii fine powder, retains spare；

(2) gypsum adds water to cook secondary, and every time plus 5 times of water is measured, and 1 hour every time, collecting decoction, filtration, filtrate retained spare；

(3) radix scutellariae adds water to cook secondary, and for the first time plus 8 times of water is measured, second plus 6 times of amounts of water, 2 hours every time, collecting decoction, filter It crosses, filtrate is concentrated into clear cream, with salt acid for adjusting pH value to 2, stands, filters to obtain crude extract, is washed by several times to pH with 70% ethyl alcohol Value is 6, is dried, crushed into fine powder, is retained spare；

(4) Herba ephedrae, the root bark of white mulberry, honeysuckle, Thesium chinese, semen armeniacae amarae, perilla seed, tussilago, lepidium seed, earthworm, pinellia, stir-fry Gingko benevolence, honey-fried licorice root add 40% ethyl alcohol heating and refluxing extraction three times, and for the first time plus 8 times are measured, second, third time plus 6 times of amounts, 1 hour every time, merge extracting solution, filtration, filtrate merges with the filtrate that step (2) obtains, and is concentrated into clear cream；

(5) fine powder that fritillaria thunbergii fine powder that step (1) obtains, step (3) obtain, step (4) are obtained clear cream and be uniformly mixed, done It is dry to get.

A preparation method of cough drug 5. fiber crops a kind of reed mentioned in ancient books as claimed in claim 4 disappears, which is characterized in that numb a kind of reed mentioned in ancient books cough drug that disappears is made Granula, tablet, hard capsule, pill.

The detection method of cough drug 6. a kind of fiber crops a kind of reed mentioned in ancient books disappears, the fiber crops a kind of reed mentioned in ancient books cough drug that disappears are made of the flavour of a drug raw material of following parts by weight： 117 parts by weight of Herba ephedrae, 117 parts by weight of the root bark of white mulberry, 233 parts by weight of honeysuckle, 117 parts by weight of radix scutellariae, 212 weight of Thesium chinese Part, 212 parts by weight of gypsum, 117 parts by weight of fritillaria thunbergii, 117 parts by weight of semen armeniacae amarae, 117 parts by weight of perilla seed, 117 weight of tussilago Measure part, 117 parts by weight of lepidium seed, 212 parts by weight of earthworm, 83 parts by weight of pinellia, 117 parts by weight of stir-fry gingko benevolence, honey-fried licorice root 83 Parts by weight, the fiber crops a kind of reed mentioned in ancient books cough drug that disappears are prepared with the following method, and steps are as follows：

(1) fritillaria thunbergii crushes, and obtains fritillaria thunbergii fine powder, retains spare；

(2) gypsum adds water to cook secondary, and every time plus 5 times of water is measured, and 1 hour every time, collecting decoction, filtration, filtrate retained spare；

(3) radix scutellariae adds water to cook secondary, and for the first time plus 8 times of water is measured, second plus 6 times of amounts of water, 2 hours every time, collecting decoction, filter It crosses, filtrate is concentrated into clear cream, with salt acid for adjusting pH value to 2, stands, filters to obtain crude extract, is washed by several times to pH with 70% ethyl alcohol Value is 6, is dried, crushed into fine powder, is retained spare；

(4) Herba ephedrae, the root bark of white mulberry, honeysuckle, Thesium chinese, semen armeniacae amarae, perilla seed, tussilago, lepidium seed, earthworm, pinellia, stir-fry Gingko benevolence, honey-fried licorice root add 40% ethyl alcohol heating and refluxing extraction three times, and for the first time plus 8 times are measured, second, third time plus 6 times of amounts, 1 hour every time, merge extracting solution, filtration, filtrate merges with the filtrate that step (2) obtains, and is concentrated into clear cream；

(5) fine powder that fritillaria thunbergii fine powder that step (1) obtains, step (3) obtain, step (4) are obtained clear cream and be uniformly mixed, done It is dry to get；

Disappear peimine in cough drug, wogonoside, wogonin it is characterized in that, measuring numb a kind of reed mentioned in ancient books using HPLC-ELSD methods Content, steps are as follows：

(1) chromatographic condition：C₁₈Chromatographic column, mobile phase are methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution, gradient elution, elution Sequence is from 0min to 15min, and the ratio of methanol is from 15% linear rise to 30%, 0.2moLL^-1Trifluoroacetic acid aqueous solution Ratio is from 85% linear decline to 70%；From 16min to 22min, the ratio of methanol from 30% linear rise to 44%, 0.2moL·L^-1The ratio of trifluoroacetic acid aqueous solution is from 70% linear decline to 56%；From 23min to 32min, the ratio of methanol From 44% linear rise to 54%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution is from 56% linear decline to 46%；From 33min to 50min, the ratio of methanol is from 54% linear rise to 81%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution from 46% linear decline is to 19%；Column temperature is 33~39 DEG C；Flow velocity is 0.5~1.5mLmin^-1；Detector parameters are drift tube temperature Degree is 110~114 DEG C, and carrier gas pressure is 3.0~3.4Lmin^-1；Sample size is 5~15 μ L；

(2) preparation of mixed reference substance solution：Peimine, wogonoside, wogonin reference substance are taken, it is accurately weighed, add body Product is than being 30:70 methanol -0.2moLL^-1It is respectively 0.0500~0.0600mgmL that concentration, which is made, in trifluoroacetic acid aqueous solution^-1, 0.3100~0.3300mgmL^-1, 0.1800~0.1900mgmL^-1Mixed reference substance solution；

(3) preparation of test solution：Product to be tested 5.0~7.0g is taken, it is accurately weighed, it sets in conical flask with cover, body is added in precision Product is than being 30:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution 50mL, close plug, weighed weight, in 250W, the item of 50kHz Under part, it is ultrasonically treated 25~35min, is let cool, then weighed weight, is 30 with volume ratio:70 methanol -0.2moLL^-1Trifluoro second Aqueous acid supplies the weight of less loss, shakes up, and filtration, precision measures 25mL, is evaporated, and residue adds water 20mL to make dissolving, full with water Be every time 25~35mL with extracting n-butyl alcohol 2~4 times, merge n-butanol liquid, 2~4 times are washed with ammonia solution, every time for 25~ 35mL discards ammonia solution, and n-butanol liquid is concentrated to dryness, and residue adds volume ratio to be 30:70 methanol -0.2moLL^-1Trifluoroacetic acid Aqueous dissolution and constant volume in 5mL volumetric flasks, shake up to get；

(4) accurate to draw mixed reference substance solution, each 5~15 μ L of test solution, high performance liquid chromatograph is injected, is surveyed It is fixed.

The detection method of cough drug 7. fiber crops a kind of reed mentioned in ancient books as claimed in claim 6 disappears, which is characterized in that measured using HPLC-ELSD methods Numb a kind of reed mentioned in ancient books disappears the content of peimine in cough drug, wogonoside, wogonin, and steps are as follows：

(1) chromatographic condition：C₁₈Chromatographic column, mobile phase are methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution, gradient elution, elution Sequence is from 0min to 15min, and the ratio of methanol is from 15% linear rise to 30%, 0.2moLL^-1Trifluoroacetic acid aqueous solution Ratio is from 85% linear decline to 70%；From 16min to 22min, the ratio of methanol from 30% linear rise to 44%, 0.2moL·L^-1The ratio of trifluoroacetic acid aqueous solution is from 70% linear decline to 56%；From 23min to 32min, the ratio of methanol From 44% linear rise to 54%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution is from 56% linear decline to 46%；From 33min to 50min, the ratio of methanol is from 54% linear rise to 81%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution from 46% linear decline is to 19%；Column temperature is 36 DEG C；Flow velocity is 1.0mLmin^-1；Detector parameters are that drift tube temperature is 112 DEG C, carrier gas pressure is 3.2Lmin^-1；Sample size is 10 μ L；

(2) preparation of mixed reference substance solution：Peimine, wogonoside, wogonin reference substance are taken, it is accurately weighed, add body Product is than being 30:70 methanol -0.2moLL^-1It is respectively 0.0550mgmL that concentration, which is made, in trifluoroacetic acid aqueous solution^-1、 0.3200mg·mL^-1、0.1850mg·mL^-1Mixed reference substance solution；

(3) preparation of test solution：Product to be tested 6.0g is taken, it is accurately weighed, it sets in conical flask with cover, precision is added volume ratio and is 30:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution 50mL, close plug, weighed weight, under conditions of 250W, 50kHz, It is ultrasonically treated 30min, is let cool, then weighed weight, is 30 with volume ratio:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution The weight for supplying less loss, shakes up, and filtration, precision measures 25mL, is evaporated, residue adds water 20mL to make dissolving, uses water-saturated n-butanol Extraction 3 times, is every time 30mL, merges n-butanol liquid, is washed 3 times with ammonia solution, be every time 30mL, discard ammonia solution, n-butanol Liquid is concentrated to dryness, and residue adds volume ratio to be 30:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution dissolves and constant volume is in 5mL In volumetric flask, shake up to get；

(4) accurate to draw mixed reference substance solution, each 10 μ L of test solution, high performance liquid chromatograph is injected, is measured.

8. a kind of fiber crops a kind of reed mentioned in ancient books disappears, cough drug is preparing the application in treating Chronic Obstructive Pulmonary Disease drug, which is characterized in that the fiber crops A kind of reed mentioned in ancient books cough drug that disappears is made of the flavour of a drug raw material of following parts by weight：117 parts by weight of Herba ephedrae, 117 parts by weight of the root bark of white mulberry, honeysuckle 233 parts by weight, 117 parts by weight of radix scutellariae, 212 parts by weight of Thesium chinese, 212 parts by weight of gypsum, 117 parts by weight of fritillaria thunbergii, semen armeniacae amarae 117 parts by weight, 117 parts by weight of perilla seed, 117 parts by weight of tussilago, 117 parts by weight of lepidium seed, 212 parts by weight of earthworm, ginger half 83 parts by weight of summer, fry 117 parts by weight of gingko benevolence, 83 parts by weight of honey-fried licorice root, and the fiber crops a kind of reed mentioned in ancient books cough drug that disappears is to prepare with the following method , steps are as follows：

(1) fritillaria thunbergii crushes, and obtains fritillaria thunbergii fine powder, retains spare；

(2) gypsum adds water to cook secondary, and every time plus 5 times of water is measured, and 1 hour every time, collecting decoction, filtration, filtrate retained spare；

(3) radix scutellariae adds water to cook secondary, and for the first time plus 8 times of water is measured, second plus 6 times of amounts of water, 2 hours every time, collecting decoction, filter It crosses, filtrate is concentrated into clear cream, with salt acid for adjusting pH value to 2, stands, filters to obtain crude extract, is washed by several times to pH with 70% ethyl alcohol Value is 6, is dried, crushed into fine powder, is retained spare；

(4) Herba ephedrae, the root bark of white mulberry, honeysuckle, Thesium chinese, semen armeniacae amarae, perilla seed, tussilago, lepidium seed, earthworm, pinellia, stir-fry Gingko benevolence, honey-fried licorice root add 40% ethyl alcohol heating and refluxing extraction three times, and for the first time plus 8 times are measured, second, third time plus 6 times of amounts, 1 hour every time, merge extracting solution, filtration, filtrate merges with the filtrate that step (2) obtains, and is concentrated into clear cream；

(5) fine powder that fritillaria thunbergii fine powder that step (1) obtains, step (3) obtain, step (4) are obtained clear cream and be uniformly mixed, done It is dry to get；

Numb a kind of reed mentioned in ancient books is measured using HPLC-ELSD methods to disappear the content of peimine in cough drug, wogonoside, wogonin, step is such as Under：

(1) chromatographic condition：C₁₈Chromatographic column, mobile phase are methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution, gradient elution, elution Sequence is from 0min to 15min, and the ratio of methanol is from 15% linear rise to 30%, 0.2moLL^-1Trifluoroacetic acid aqueous solution Ratio is from 85% linear decline to 70%；From 16min to 22min, the ratio of methanol from 30% linear rise to 44%, 0.2moL·L^-1The ratio of trifluoroacetic acid aqueous solution is from 70% linear decline to 56%；From 23min to 32min, the ratio of methanol From 44% linear rise to 54%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution is from 56% linear decline to 46%；From 33min to 50min, the ratio of methanol is from 54% linear rise to 81%, 0.2moLL^-1The ratio of trifluoroacetic acid aqueous solution from 46% linear decline is to 19%；Column temperature is 36 DEG C；Flow velocity is 1.0mLmin^-1；Detector parameters are that drift tube temperature is 112 DEG C, carrier gas pressure is 3.2Lmin^-1；Sample size is 10 μ L；

(2) preparation of mixed reference substance solution：Peimine, wogonoside, wogonin reference substance are taken, it is accurately weighed, add body Product is than being 30:70 methanol -0.2moLL^-1It is respectively 0.0550mgmL that concentration, which is made, in trifluoroacetic acid aqueous solution^-1、 0.3200mg·mL^-1、0.1850mg·mL^-1Mixed reference substance solution；

(3) preparation of test solution：Product to be tested 6.0g is taken, it is accurately weighed, it sets in conical flask with cover, precision is added volume ratio and is 30:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution 50mL, close plug, weighed weight, under conditions of 250W, 50kHz, It is ultrasonically treated 30min, is let cool, then weighed weight, is 30 with volume ratio:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution The weight for supplying less loss, shakes up, and filtration, precision measures 25mL, is evaporated, residue adds water 20mL to make dissolving, uses water-saturated n-butanol Extraction 3 times, is every time 30mL, merges n-butanol liquid, is washed 3 times with ammonia solution, be every time 30mL, discard ammonia solution, n-butanol Liquid is concentrated to dryness, and residue adds volume ratio to be 30:70 methanol -0.2moLL^-1Trifluoroacetic acid aqueous solution dissolves and constant volume is in 5mL In volumetric flask, shake up to get；

(4) accurate to draw mixed reference substance solution, each 10 μ L of test solution, high performance liquid chromatograph is injected, is measured.