CN101496870A - Chinese medicinal composition for resolving phlegm and suppressing cough as well as preparation method and quality control method thereof - Google Patents
Chinese medicinal composition for resolving phlegm and suppressing cough as well as preparation method and quality control method thereof Download PDFInfo
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Abstract
The invention relates to a Chinese traditional medicine composition for relieving cough and reducing sputum, a method for preparing the same and a method for controlling the quality. The Chinese traditional medicine composition comprises bulk drugs of pummelo peel, dried orange peel, rhizoma pinellinae praeparata, coltsfoot flower, balloon flower, bitter apricot kernel (fried without peel), tuckahoe, trichosanthes bark, liquorice, aster, dwarf lilyturf root, rhizoma anemarrhenae, radix rehmanniae, gypsum, perilla seed (fried), and the like which are added with auxiliary materials and are prepared into clinically acceptable dosage forms including capsules, soft capsules, pills, tablets, powder, injections, granules, mixtures, and the like. The quality control method for the Chinese traditional medicine composition identifies the dwarf lilyturf root, the liquorice, the bitter apricot kernel, the dried orange peel, the balloon flower and the trichosanthes bark, performs content determination on the pummelo peel, and ensures the curative effect of the medicine. The Chinese traditional medicine composition has the efficacy of relieving cough and reducing sputum, and is applied to treating cough with more sputum, fullness sensation in chest with short breath, and dry pharynx and tickling throat caused by phlegm heat lung obstruction.
Description
Technical field
The present invention relates to a kind of Chinese medicine composition and preparation method thereof and method of quality control, particularly Chinese medicine composition of a kind of relieving cough and resolving phlegm and preparation method thereof and method of quality control.
Background technology
From physiological angle, cough is a kind of body protective sexual activity, and it can be discharged the sputum in the respiratory tract, foreign body, thereby the cleaning of maintenance respiratory tract and unobstructed is beneficial to healthy.But more frequent and violent cough will have influence on people's work, living and studying, at this moment just should select the relieving cough and resolving phlegm medicine of suiting the medicine to the illness for use according to symptom.Some more serious diseases usually also are attended by the cough symptom, as pleuritis, spontaneous pneumothorax, pulmonary tuberculosis, pulmonary carcinoma, heart failure etc., cough all can occur.In the medicine of expelling phlegm for arresting cough, Western medicine and Chinese patent medicine have his own strong points: Western medicine is directly suited the medicine to the illness, instant effect now, but addiction is arranged, need associating other drug Comprehensive Treatment more, need use antibiotics simultaneously as respiratory tract infection, flu then need be share anti-cold medicine; Chinese patent medicine commonly used uses as suiting the medicine to the illness, determined curative effect, though take effect not as Western medicine rapid, take stopgap measures and effect a permanent cure.
Summary of the invention
First purpose of the present invention be to provide a kind of relieving cough and resolving phlegm Chinese medicine composition; Second purpose of the present invention is to provide the preparation method of this Chinese medicine composition; The 3rd purpose of the present invention is to provide the method for quality control of this Chinese medicine composition.
Content of the present invention is to be achieved through the following technical solutions:
A kind of relieving cough and resolving phlegm Chinese medicine composition be to make by the crude drug of following weight ratio: Exocarpium Citri Grandis 50-100 weight portion, Pericarpium Citri Reticulatae 50-100 weight portion, Rhizoma Arisaematis 50-100 weight portion, Folium Eriobotryae 50-100 weight portion, Radix Platycodonis 30-70 weight portion, Semen Armeniacae Amarum (peeling is fried) 30-70 weight portion, Poria 30-70 weight portion, Pericarpium Trichosanthis 30-70 weight portion, Radix Glycyrrhizae 20-50 weight portion, Radix Asteris 20-50 weight portion, Radix Ophiopogonis the 20-50 weight portion, Rhizoma Anemarrhenae 20-50 weight portion, Radix Peucedani 10-35 weight portion, Gypsum Fibrosum 10-35 weight portion, Radix Aucklandiae 10-35 weight portion.
Rhizoma Arisaematis in the Chinese medicine composition of relieving cough and resolving phlegm of the present invention can be substituted by Rhizoma Typhonii, Semen Sinapis Albae, Rhizoma Pinelliae Preparatum, Flos Inulae.
Folium Eriobotryae in the Chinese medicine composition of relieving cough and resolving phlegm of the present invention can be substituted by the Radix Stemonae, Flos Farfarae, Semen Lepidii (Semen Descurainiae), Cortex Mori.
The Radix Aucklandiae in the Chinese medicine composition of relieving cough and resolving phlegm of the present invention can be substituted by Fructus Perillae (stir-fry).Radix Peucedani in the Chinese medicine composition of relieving cough and resolving phlegm of the present invention can be substituted by Caulis Bambusae In Taenia, Succus Bambusae, Radix Rehmanniae, Lapis Micae Aureus.
This Chinese medicine composition is to be made by the crude drug of following weight ratio:
Exocarpium Citri Grandis 66 weight portions, Pericarpium Citri Reticulatae 66 weight portions, Rhizoma Arisaematis 66 weight portions, Folium Eriobotryae 66 weight portions, Radix Platycodonis 44 weight portions, Semen Armeniacae Amarum (peeling is fried) 44 weight portions, Poria 44 weight portions, Pericarpium Trichosanthis 44 weight portions, Radix Glycyrrhizae 33 weight portions, Radix Asteris 33 weight portions, Radix Ophiopogonis 33 weight portion, the Rhizoma Anemarrhenae 33 weight portions, Radix Peucedani 22 weight portions, Gypsum Fibrosum 22 weight portions, the Radix Aucklandiae 22 weight portions.
Chinese medicine composition preparation method of the present invention is: Gypsum Fibrosum powder is broken into coarse powder, decocts with water 1-3 time, each 0.5-2 hour, filters filtrate for later use; Exocarpium Citri Grandis, Pericarpium Citri Reticulatae, Folium Eriobotryae, Semen Armeniacae Amarum four flavor vapor distillations are collected distillate 200-300 parts by volume; The distillator inner liquid medicine filters, and filtrate adds ethanol to be made and contain the alcohol amount and reach 50-70%, stirs evenly, and leaves standstill 16-36 hour, filters filtrate for later use; Ten flavors such as all the other Rhizoma Arisaematiss, be ground into coarse powder and above-mentioned medicinal residues mixing, according to the percolation under fluid extract and the extractum item (" appendix IO of Chinese pharmacopoeia version in 2005), make solvent with 50-70% ethanol, flood after 12-36 hour percolation in accordance with the law, the collection liquid 2500-3000 parts by volume of filtering, merge with above-mentioned reserve liquid, decompression recycling ethanol is to there not being the alcohol flavor, merge with the parget water decocting liquid, be concentrated into the clear paste of relative density 1.06 (50 ℃), add conventional adjuvant again, through conventional method, make the preparation of clinical acceptance.
Chinese medicine composition preparation method of the present invention is preferably: Gypsum Fibrosum powder is broken into coarse powder, decocts with water secondary, each 1 hour, filters filtrate for later use; Exocarpium Citri Grandis, Pericarpium Citri Reticulatae, Folium Eriobotryae, Semen Armeniacae Amarum four flavor vapor distillations are collected distillate 250 parts by volume; The distillator inner liquid medicine filters, and filtrate adds ethanol to be made and contain the alcohol amount and reach 60%, stirs evenly, and leaves standstill 24 hours, filters filtrate for later use; Ten flavors such as all the other Rhizoma Arisaematiss, be ground into coarse powder and above-mentioned medicinal residues mixing, according to the percolation under fluid extract and the extractum item (" appendix IO of Chinese pharmacopoeia version in 2005), make solvent with 60% ethanol, flood after 24 hours percolation in accordance with the law, collect liquid 2700 parts by volume of filtering, merge with above-mentioned reserve liquid, decompression recycling ethanol merges with the parget water decocting liquid to there not being the alcohol flavor, is concentrated into the clear paste of relative density 1.06 (50 ℃).Add conventional adjuvant again,, make the oral solid formulation of clinical acceptance through conventional method.
The preparation method of Chinese medicine composition mixture of the present invention is: Gypsum Fibrosum powder is broken into coarse powder, decocts with water secondary, each 1 hour, filters filtrate for later use; Exocarpium Citri Grandis, Pericarpium Citri Reticulatae, Folium Eriobotryae, Semen Armeniacae Amarum four flavor vapor distillations are collected distillate 250 parts by volume; The distillator inner liquid medicine filters, and filtrate adds ethanol to be made and contain the alcohol amount and reach 60%, stirs evenly, and leaves standstill 24 hours, filters filtrate for later use; Ten flavors such as all the other Rhizoma Arisaematiss, be ground into coarse powder and above-mentioned medicinal residues mixing, according to the percolation under fluid extract and the extractum item (" appendix IO of Chinese pharmacopoeia version in 2005), make solvent with 60% ethanol, flood after 24 hours percolation in accordance with the law, collect the liquid 2700ml that filters, merge with above-mentioned reserve liquid, decompression recycling ethanol merges with the parget water decocting liquid to there not being the alcohol flavor, is concentrated into the clear paste of relative density 1.06 (50 ℃).Add sucrose 80 weight portions, boil, left standstill 24 hours, filter, add ethyl hydroxybenzoate 0.3 weight portion, benzoic acid 0.5 weight portion (both use an amount of hot water dissolving earlier) and distillate, add water and adjust total amount to 950 parts by volume, stir evenly cold preservation 48 hours, get supernatant, embedding, sterilization, promptly.
Described parts by volume/weight portion is corresponding with g/ml.
Chinese medicine composition method of quality control of the present invention comprises one or more in following discrimination method and/or the assay:
[discriminating]
(1) get pharmaceutical composition of the present invention and be equivalent to crude drug 20-30g,, add hydrochloric acid 2-4ml, put in the water-bath heating 0.5-2 hour, to put coldly, the 20-40ml jolting that adds diethyl ether is extracted, and divides and gets ether solution, and evaporate to dryness, residue add chloroform 1ml makes dissolving, as need testing solution.Other gets control medicinal material 2g Radix Ophiopogonis, decocts with water 20-40 minute, filters, and filtrate is concentrated into 30-50ml, shines medical material solution in pairs with legal system.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 5～10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with chloroform-acetone (3.5-4.5:0.5-1.5) is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, 100～110 ℃ of heating 4-6 minute.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color.
(2) get pharmaceutical composition of the present invention and be equivalent to crude drug 10-15g, add hydrochloric acid 2-4ml, reflux 0.5-2 hour, put cold, add the chloroform jolting and extract 1-3 time, each 10-20ml merges chloroform extraction liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.Extracting liquorice subacid reference substance adds dehydrated alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution in addition.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 15 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (30～60 ℃)-benzene-ethyl acetate-glacial acetic acid (8-12:15-25:5-10:0-1) is developing solvent, launch, take out, dry, spray is with 10% phosphomolybdic acid ethanol solution, 100～110 ℃ of heating 4-6 minute.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
(3) get pharmaceutical composition of the present invention and be equivalent to crude drug 10-15g, adding the water-saturated n-butanol jolting extracts 1-3 time, each 15-25ml, merge n-butanol extracting liquid, use ammonia scrubbing 1-3 time, each 15-25ml, discard ammonia liquid, n-butanol layer evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution.Other gets the amygdaloside reference substance, adds methanol and makes the solution that 1ml contains 2mg, in contrast product solution.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with chloroform-methanol-water (10-15:5-10:1-3) is developing solvent, launch, take out, dry, (phosphomolybdic acid 2g adds water 20ml and makes dissolving with the phosphomolybdic acid sulfuric acid solution in spray, slowly adding sulphuric acid 30ml, mixing), it is clear to be heated to speckle colour developing at 100～110 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
(4) get pharmaceutical composition of the present invention and be equivalent to crude drug 10-15g, adding petroleum ether (60 ℃～90 ℃) 15-25ml jolting extracts 1 time, discard petroleum ether, adding the ethyl acetate jolting extracts 1-3 time, each 15-25ml merges ethyl acetate extraction liquid, evaporate to dryness, residue adds methanol 2ml makes dissolving, as need testing solution.Other gets Pericarpium Citri Reticulatae glycosides reference substance, adds methanol and makes saturated solution, in contrast product solution.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 6 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with chloroform-methanol-water (10-15:5-10:1-3) is developing solvent, launch about 10-20cm, take out, dry, spray is put under the ultra-violet lamp (365nm) and is inspected with the aluminum chloride test solution.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.
(5) get pharmaceutical composition of the present invention and be equivalent to crude drug 10-15g, add 10% ethanol solution of sulfuric acid 4-6ml, reflux 4-6h, put coldly, extract 1-3 time with the chloroform jolting, at every turn 15-25ml, combined chloroform liquid, add water 20-40ml washing, discard washing liquid, the chloroform solution anhydrous sodium sulfate dehydration, filter, filtrate evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution.Other gets Radix Platycodonis control medicinal material 1g, makes reference substance solution with method.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, drawing each 10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, is developing solvent with chloroform-ether (45-55:45-55), launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, it is clear to be heated to speckle colour developing at 100～110 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
(6) get pharmaceutical composition of the present invention and be equivalent to crude drug 15-20g, add water saturated n-butanol extraction 1-3 time, each 25-40ml at every turn, n-butanol extracting liquid steam in, residue adds methanol 1ml makes dissolving, as need testing solution.Get Pericarpium Trichosanthis control medicinal material 3g, add 60% ethanol 20ml merceration 2h after, supersound process 30min filters, filtrate evaporate to dryness, residue add methanol 1ml makes dissolving, makes control medicinal material solution.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 10 μ L of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (60～90 ℃)-ethyl acetate (35-45:5-15) is developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with Pericarpium Trichosanthis control medicinal material chromatograph relevant position on, show mutually the speckle of color just.
[assay]
Measure according to high performance liquid chromatography (" appendix VID of Chinese pharmacopoeia version in 2005).
Chromatographic condition and system suitability test are filler with the octadecylsilane chemically bonded silica; With methanol-water-acetic acid (30-40:60-70:0-1) is mobile phase; The detection wavelength is 283nm; Column temperature is 40 ℃.Theoretical cam curve is calculated by the naringin peak should be not less than 3000.
The preparation of reference substance solution is taken at 100～120 ℃ of about 15mg of naringin reference substance that are dried to constant weight, and accurate the title decides, and puts in the 100ml measuring bottle, adds methanol and makes dissolving in right amount and be diluted to scale, shakes up; Precision is measured 3ml, puts in the 10ml measuring bottle, adds methanol to scale, shakes up, and promptly gets (every 1ml contains naringin 45 μ g).
The preparation precision of need testing solution is measured this product 1ml, puts in the 25ml measuring bottle, adds methanol and is diluted to scale, shakes up, and filters, and gets subsequent filtrate, promptly.
Accurate respectively reference substance solution and each the 10 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly.
Chinese medicine composition method of quality control of the present invention is preferably as follows one or more in discrimination method and/or the assay:
[discriminating]
(1) get pharmaceutical composition mixture 40ml of the present invention, add hydrochloric acid 3ml, put in the water-bath heating 1 hour, put coldly, the 30ml jolting that adds diethyl ether is extracted, and divides and gets ether solution, and evaporate to dryness, residue add chloroform 1ml makes dissolving, as need testing solution.Other gets control medicinal material 2g Radix Ophiopogonis, decocts with water 30 minutes, filters, and filtrate is concentrated into 40ml, shines medical material solution in pairs with legal system.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 5～10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with chloroform-acetone (4:1) is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, 105 ℃ of heating 5 minutes.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color.
(2) get pharmaceutical composition mixture 20ml of the present invention, add hydrochloric acid 3ml, reflux 1 hour is put coldly, adds the chloroform jolting and extracts 2 times, and each 15ml merges chloroform extraction liquid, and evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.Extracting liquorice subacid reference substance adds dehydrated alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution in addition.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 15 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (30～60 ℃)-benzene-ethyl acetate-glacial acetic acid (10:20:7:0.5) is developing solvent, launch, take out, dry, spray is with 10% phosphomolybdic acid ethanol solution, 105 ℃ of heating 5 minutes.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
(3) get pharmaceutical composition mixture 20ml of the present invention, add the water-saturated n-butanol jolting and extract 2 times, each 20ml merges n-butanol extracting liquid, with ammonia scrubbing 2 times, each 20ml discards ammonia liquid, n-butanol layer evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution.Other gets the amygdaloside reference substance, adds methanol and makes the solution that 1ml contains 2mg, in contrast product solution.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with chloroform-methanol-water (13:7:2) is developing solvent, launch, take out, dry, (phosphomolybdic acid 2g adds water 20ml and makes dissolving with the phosphomolybdic acid sulfuric acid solution in spray, slowly adding sulphuric acid 30ml, mixing), it is clear to be heated to speckle colour developing at 105 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
(4) get pharmaceutical composition mixture 20ml of the present invention, add petroleum ether (60～90 ℃) 20ml jolting and extract 1 time, discard petroleum ether, add the ethyl acetate jolting and extract 2 times, each 20ml merges ethyl acetate extraction liquid, evaporate to dryness, residue add methanol 2ml makes dissolving, as need testing solution.Other gets Pericarpium Citri Reticulatae glycosides reference substance, adds methanol and makes saturated solution, in contrast product solution.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 6 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with chloroform-methanol-water (13:7:2) is developing solvent, launch about 15cm, take out, dry, spray is put under the ultra-violet lamp (365nm) and is inspected with the aluminum chloride test solution.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.
(5) get pharmaceutical composition mixture 20ml of the present invention, add 10% ethanol solution of sulfuric acid 5ml, reflux 3h, put coldly, extract 2 times each 20ml with the chloroform jolting, combined chloroform liquid, add water 30ml washing, discard washing liquid, the chloroform solution anhydrous sodium sulfate dehydration, filter, filtrate evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution.Other gets Radix Platycodonis control medicinal material 1g, makes reference substance solution with method.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with chloroform-ether (1:1) is developing solvent, launches, and takes out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing at 105 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
(6) get pharmaceutical composition mixture 30ml of the present invention, add water saturated n-butanol extraction 2 times, each 30ml at every turn, n-butanol extracting liquid steam in, residue adds methanol 1ml makes dissolving, as need testing solution.Get Pericarpium Trichosanthis control medicinal material 3g, add 60% ethanol 20ml merceration 2h after, supersound process 30min filters, filtrate evaporate to dryness, residue add methanol 1ml makes dissolving, makes control medicinal material solution.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 10 μ L of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (60～90 ℃)-ethyl acetate (4:1) is developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with Pericarpium Trichosanthis control medicinal material chromatograph relevant position on, show the speckle of same color.
[assay]
Measure according to high performance liquid chromatography (" appendix VID of Chinese pharmacopoeia version in 2005).
Chromatographic condition and system suitability test are filler with the octadecylsilane chemically bonded silica; With methanol-water-acetic acid (38:62:0.5) is mobile phase; The detection wavelength is 283nm; Column temperature is 40 ℃.Theoretical cam curve is calculated by the naringin peak should be not less than 3000.
The preparation of reference substance solution is taken at 110 ℃ of about 15mg of naringin reference substance that are dried to constant weight, and accurate the title decides, and puts in the 100ml measuring bottle, adds methanol and makes dissolving in right amount and be diluted to scale, shakes up; Precision is measured 3ml, puts in the 10ml measuring bottle, adds methanol to scale, shakes up, and promptly gets (every 1ml contains naringin 45 μ g).
The preparation precision of need testing solution is measured this product 1ml, puts in the 25ml measuring bottle, adds methanol and is diluted to scale, shakes up, and filters, and gets subsequent filtrate, promptly.
Accurate respectively reference substance solution and each the 10 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly.
Get the every 1ml of pharmaceutical composition mixture of the present invention and contain Exocarpium Citri Grandis with naringin (C
27H
32O
14) meter, must not be less than 0.80mg.
Pharmaceutical composition lung heat clearing of the present invention, cough-relieving is reduced phlegm, and can fundamentally treat because cough with copious phlegm, fullness in the chest and shortness of breath, the dry pharynx itching of the throat that accumulation of phlegm-heat in the lung causes.The present composition is compared existing preparation and is possessed good drug effect, and scope of the present invention through screening, finds in some scope of compositions, to possess more outstanding drug effect unexpectedly when can realizing drug effect of the present invention.Rhizoma Arisaematis in the Chinese medicine composition, Rhizoma Typhonii, Semen Sinapis Albae, Rhizoma Pinelliae Preparatum, Inula medicine for warming and resolving cold-phlegm mainly are applicable to the cough and asthma that cold-phlegm, damp-phlegm cause, excessive dilute and white sputum; It is uncomfortable in chest that Folium Eriobotryae, the Radix Stemonae, Flos Farfarae, Semen Lepidii (Semen Descurainiae), Cortex Mori belong to the cough with asthma that Qinghua heat-phlegm medicine is applicable to that mainly heat-phlegm causes, the yellow thickness of expectorant is difficult for bringing up; The Radix Aucklandiae, Fructus Perillae (stir-fry) belong to the invigorating the spleen for dissipating phlegm medicine and mainly are applicable to fullness in the epigastrium and chest, lack of appetite and vomiting, cough with copious phlegm; It is uncomfortable in chest that Radix Peucedani, Caulis Bambusae In Taenia, Succus Bambusae, Radix Rehmanniae, Lapis Micae Aureus belong to the cough with asthma that Qinghua heat-phlegm medicine is applicable to that mainly heat-phlegm causes, the yellow thickness of expectorant is difficult for bringing up.
The method of quality control of Chinese medicine composition provided by the present invention, be by obtaining behind the creative experiment sieving of big measuring, pass through screening in the discrimination method to sample treatment, the selection of developing solvent, make and differentiate that specificity is fine, and method is economic and practical, the result is quick, and can both use different lamellaes.Pass through screening in the content assaying method to sample, test sample processing method, the selection of developing solvent, make content assaying method effectivelyly to carry out quality control, and compare more stable that product that additive method measures shows on drug effect with the product that this method is measured to product.
The specific embodiment
Following experimental example and embodiment are used to further specify the present invention but are not limited to the present invention
Experimental example 1 pharmaceutical composition pharmacodynamic study of the present invention
1 material
1.1 medicine medicine group of the present invention I makes according to embodiment 6 methods; Medicine group II of the present invention makes according to embodiment 7 methods; Medicine group III of the present invention makes according to embodiment 8 methods; The Chinese medicine positive controls, 'Baikejing ' syrup; Codeine phosphate tablets; Ammonium chloride; Ammonia; Agents useful for same is analytical pure.
1.2 animal NIH mice, Rana nigromaculata.
1.3 statistics is selected for use, and t checks relatively between group.
2 methods
2.1 antitussive effect
2.1.1 ammonia is drawn the influence of coughing
Select 18～22g mice (spraying 20s, making mouse cough is qualified more than 25 times/30s) for use, male and female half and half are divided into 5 groups at random, and the ig administration (first group, the equal-volume distilled water; Second and third group, medicine group I of the present invention, II are equivalent to crude drug 0.3g/kg; The 4th group, Chinese medicine positive controls 0.3g/kg, the 5th group, codeine 10ml), every day 1 time, continuous 3d, 1h behind the last medicine draws the method for coughing by mice ammonia and experimentizes, and compares between the work group, the results are shown in Table 1.
Table 1 pair ammonia draw the influence of coughing number of times (x ± s, n=15)
| Group | The cough number of times (inferior/30s) |
| Distilled water | 29.7±7.8 |
| Medicine group I of the present invention | 15.0±7.8 **△ |
| Medicine group II of the present invention | 22.6±7.2 * |
| The Chinese medicine positive controls | 23.4±9.4 |
| Codeine | 9.9±7.4 **△ |
Compare with the distilled water group
*P＜0.05,
*P＜0.01; Compare △ P＜0.05, △ △ P＜0.01 with the Chinese medicine positive controls;
Medicine group I of the present invention, II can significantly reduce the cough number of times of mice, and relatively there were significant differences with matched group, and relatively there were significant differences with the Chinese medicine positive controls.
2.1.2 sulfur dioxide is drawn the influence of coughing
Select 90 of the NIH mices of cough latent period 15s for use, body weight 18～22g, male and female half and half.Be divided into 5 groups at random, medication is the same, and 1h behind the last medicine experimentizes, and compares between the work group, the results are shown in Table 2.
Table 2 pair sulfur dioxide draw cough preclinical influence (x ± s, n=18)
| Group | Dosage (g/kg) | Cough latent period (s) |
| Distilled water | 20ml | 19.0±4.8 |
| Medicine group I of the present invention | 31.6±4.2 **△ | |
| Medicine group II of the present invention | 30.2±5.3 **△ | |
| The Chinese medicine positive controls | 26.4±6.7 * | |
| Codeine | 10ml | 54.7±6.6 **△ |
Compare with the distilled water group
*P＜0.05,
*P＜0.01; Compare △ P＜0.05, △ △ P＜0.01 with the Chinese medicine positive controls;
Cough incubation period but medicine group I of the present invention, II significant prolongation sulfur dioxide draw, relatively there were significant differences with matched group, and relatively there were significant differences with the Chinese medicine positive controls.
2.2 phlegm-dispelling functions
2.2.1 to the influence of phenol red output of mice trachea
Select 55 of 18～22gNIH mices for use, male and female half and half are divided into 5 groups at random, and medication is the same, and 1h behind the last medicine by the phenol red expelling phlegm method experiment of mice, measures phenol red output, the results are shown in Table 3.
The influence of table 3 pair mice trachea phenol red output (x ± s, n=11)
| Group | Dosage (g/kg) | Phenol red output (OD value) |
| Distilled water | 20ml | 0.152±0.041 |
| Medicine group I of the present invention | 0.235±0.035 **△ | |
| Medicine group II of the present invention | 0.224±0.039 **△ | |
| The Chinese medicine positive controls | 0.194±0.019 ** | |
| Codeine | 10ml | 0.208±0.042 ** |
Compare with the distilled water group
*P＜0.05,
*P＜0.01; Compare △ P＜0.05, △ △ P＜0.01 with the Chinese medicine positive controls;
Medicine group I of the present invention, II and Chinese medicine positive controls can significantly increase mice trachea phenol red output, and relatively there were significant differences with matched group, and there were significant differences with the comparison of Chinese medicine positive controls for medicine group I of the present invention, II.
2.2.2 in body Rana nigromaculata oral mucosa effect on ciliary movement
Select 50～100g Rana nigromaculata for use, destroy spinal cord with pin, be fixed on the frog board, fully expose upper jaw mucosal surface, local medicine group I of the present invention, II, Chinese medicine positive controls, 0.0125% isoproterenol, the equivalent normal saline of dripping surveyed the required time from the start line to the finishing line of 3min wood flour before and after the administration with stopwatch, write down 3 times, average, carry out statistical disposition, the results are shown in Table 4.
The influence of table 4 pair frog oral mucosa ciliary movement speed (x ± s, n=11)
| Group | (mm/min) |
| Normal saline | ↓4.88±7.88 |
| Medicine group I of the present invention | ↑14.16±8.24 *△ |
| Medicine group II of the present invention | ↑12.41±6.27 *△ |
| The Chinese medicine positive controls | ↑5.49±3.26 |
| 0.0125% isoproterenol | ↑12.33±9.58 △ |
Every kg body weight animals administer 0.1ml, ↑ expression is accelerated, and ↓ expression is slowed down
Compare with the normal saline group
*P＜0.05,
*P＜0.01; Compare △ P＜0.05, △ △ P＜0.01 with the Chinese medicine positive controls;
Medicine group I of the present invention, II can significantly increase Rana nigromaculata oral mucosa ciliary movement, and relatively there were significant differences with normal saline, and relatively there were significant differences with the Chinese medicine positive controls, illustrate that said preparation has the effect that promotes expectoration.
3 brief summaries
Medicine group of the present invention can significantly reduce the number of times that ammonia causes mouse cough, energy significant prolongation sulfur dioxide causes the incubation period of mouse cough, can significantly increase mice trachea phenol red output, can significantly increase Rana nigromaculata oral mucosa ciliary movement, point out this medical instrument that the relieving cough and resolving phlegm effect is arranged.
The inhibiting research of experimental example 2 medicine group pair cell cytochrome p 450s of the present invention (CYP3A4 and CYP1A2)
Medicine group of the present invention is a kind of cough medicine that is prepared from by plurality of Chinese composition such as Exocarpium Citri Grandis, Rhizoma Arisaematis, Pericarpium Citri Reticulatae etc., because it has good expelling phlegm for arresting cough effect and is able to extensive use clinically.Medicine group of the present invention contains number of chemical compositions such as flavone compound Hesperidin, citral and monoterpenes chemical compound.CYP3A4 is important Cytochrome P450, the metabolism of the clinical medicine of catalysis 60%.
In experimentation, our surprised discovery medicine group of the present invention has inhibitory action to CYP3A4 and CYP1A2, causes the metabolic medicine of catalysis to produce bad drug interaction.
Materials and methods
1. material
Medicine group of the present invention, Dormicum and 1-hydroxyl Dormicum; 1, the 7-dimethyl xanthine; Caffeine.Chloroform, formic acid and 2-propanol are analytical reagent, and methanol is chromatographically pure reagent.
2. method
Tried: the experimenter is 10 healthy male volunteers, 23 ± 2 years old mean age, body weight standard body weight ± 20% in, all experimenters are through health check-up, and are healthy, tried not take in preceding 2 weeks other any medicines.
Medication adopts cross-over design, 10 experimenters are divided into two groups at random, the medicine group of the present invention (according to prepared among the embodiment 6) that first three day A organizes oral clinical dosage is equivalent to crude drug 15g/ day, B organizes the placebo of oral isodose, and 4d begins two groups of all oral 7.5mg Dormicums of A, B and 100mg caffeine.In the 0th, 0.25,0.5,1,2,3,4,5,6 and 8h gather venous blood 3ml, two groups of intersection behind the eluting 10d repeat previous stage and test.Blood sample is centrifugal, separated plasma, and-40 ℃ of preservations are to be measured.
The mensuration of Dormicum and caffeine and metabolite thereof: Dormicum and caffeine and metabolite thereof are measured with HPLC-MS, chromatographic column is Xterra TM MS C18 (3.9 * 150mm, 5um), mobile phase is that water (contains 0.01% formic acid, pH=4.1)-and methanol, adopt linear gradient elution method, preceding 10min ratio is 52:48, ratio becomes 95:5 then, and flow velocity is 0.2mlmin
-1, column temperature is 40 ℃.Detector is ESI, and voltage is set to capillary voltage: 3.2KV, taper hole voltage: 45v, extraction voltage: 6v.
Sample treatment: get blood plasma 200 μ l and interior mark 10 μ l (alprazolam, 2.03mgL
-1) adding 2ml extraction solvent after the centrifuge tube mesoscale eddies of 15ml mixes (chloroform: the 2-propanol=9:1), and vortex 3min, it is centrifugal that (3000rmin 5min), separates organic facies, and organic facies dries up with nitrogen under 40 ℃ of conditions, and is residual to be measured with dissolve with methanol.
Date processing and statistical analysis: AUC
0-8hCalculate C with trapezoidal method
MaxAnd t
MaxCan directly read from medicine-time curve.Ke and t
1/2Utilization pharmacokinetics special-purpose software 3P87 calculates.The difference of pharmacokinetic parameter is added up with SPSS (10.0version) before and after the administration, and significance test adopts paired t-test, and P＜0.05 thinks that there were significant differences.
The result
Dormicum, 1-hydroxyl Dormicum, caffeine and 1 before and after the administration, the blood drug level and the pharmacokinetic parameter of 7-dimethyl xanthine see Table 5-8.
The influence of table 5 couple Dormicum and 1-hydroxyl Dormicum's blood drug level (μ gL)
Table 6 pair caffeine and 1, the influence of the blood drug level of 7-dimethyl xanthine (μ gL)
Table 7 Dormicum pharmacokinetic parameter
Table 8 caffeine pharmacokinetic parameter
The medicine group of the present invention that gives clinical dosage is after three days, Dormicum's peak concentration (C
Max) and medicine area under curve raising for the moment (116.8 ± 64.32vs127.3 ± 54.96, P〉0.05; 54.20 ± 33.15vs60.00 ± 32.76, P〉0.05), same, peak time (T) slightly improves, but there was no significant difference (0.45 ± 0.10vs0.58 ± 0.35, P〉0.05).Dormicum's elimination half-life (t
1/2) significantly improve (1.86 ± 0.35vs2.07 ± 0.36, P＜0.05).Before and after the oral medicine group of the present invention, the pharmacokinetic parameter of caffeine, except that medicine-time area under curve (10.40 ± 4.23vs12.83 ± 6.00, P=0.163), other parameter all is irregular variation slightly improving.
The result shows: give medicine group of the present invention after three days, Dormicum's AUC, C
Max, t
MaxAll increase t
1/2Significantly improve, point out medicine group of the present invention can suppress Dormicum's metabolism.Because Dormicum's 1 monohydroxy metabolism by CYP3A4 catalysis, can be represented the activity of CYP3A4 with Dormicum's 1 monohydroxy metabolism in the research.Medicine group of the present invention is because it is inhibited to CYP3A4 to Dormicum's inhibitory action.The CYP3A4 that suppresses in the gastrointestinal tract causes C
MaxAnd t
MaxMain cause, the CYP3A4 that suppresses in the liver causes t
1/2The main cause that prolongs, two kinds of factors all can cause the increase of AUC.
The screening of experimental example discrimination test 3 Radix Ophiopogonis
(1) preparation of need testing solution
Method one: get pharmaceutical preparation of the present invention and be equivalent to crude drug 24.08g, porphyrize adds hydrochloric acid 3ml, puts in the water-bath heating 0.5 hour, puts coldly, and the 30ml jolting that adds diethyl ether is extracted, and divides and gets ether solution, and evaporate to dryness, residue add chloroform 1ml makes dissolving, as need testing solution.
Method two: get pharmaceutical preparation of the present invention and be equivalent to crude drug 24.08g, porphyrize adds hydrochloric acid 3ml, puts in the water-bath heating 1 hour, puts coldly, and the 30ml jolting that adds diethyl ether is extracted, and divides and gets ether solution, and evaporate to dryness, residue add chloroform 1ml makes dissolving, as need testing solution.
Method three: get pharmaceutical preparation of the present invention and be equivalent to crude drug 24.08g, porphyrize adds hydrochloric acid 3ml, puts in the water-bath and heats 1.5 hours, put coldly, add petroleum ether 30ml jolting and extract, divide and get petroleum ether liquid, evaporate to dryness, residue add chloroform 1ml makes dissolving, as need testing solution.
Other gets control medicinal material 2g Radix Ophiopogonis, decocts with water 20-40 minute, filters, and filtrate is concentrated into 40ml, shines medical material solution in pairs with legal system.Get to lack and write out a prescription Radix Ophiopogonis, press prepared scarce Radix Ophiopogonis of negative preparation among the embodiment 6, press the preparation of test sample preparation method and lack negative controls Radix Ophiopogonis.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with chloroform-acetone (4:1) is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, 105 ℃ of heating 5 minutes.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color.
The preparation of table 9. need testing solution
| The clear spot degree | The hangover situation | Disturbed condition | |
| Method one: | Unintelligible | Do not have | Do not have |
| Method two: | Clear | Do not have | Do not have |
| Method three: | Clear | Do not have | Have |
(2) developing solvent proportioning preferred:
Developing solvent one: toluene-methanol-glacial acetic acid (80:5:0.1)
Developing solvent two: chloroform-acetone (4:1)
Developing solvent three: chloroform-acetone (4.5:0.5)
Get pharmaceutical preparation of the present invention and be equivalent to crude drug 24.08g, add hydrochloric acid 3ml, put in the water-bath heating 1 hour, put coldly, the 30ml jolting that adds diethyl ether is extracted, and divides and gets ether solution, and evaporate to dryness, residue add chloroform 1ml makes dissolving, as need testing solution.Other gets control medicinal material 2g Radix Ophiopogonis, decocts with water 30 minutes, filters, and filtrate is concentrated into 40ml, shines medical material solution in pairs with legal system.Get to lack and write out a prescription Radix Ophiopogonis, press prepared scarce Radix Ophiopogonis of negative preparation among the embodiment 6, press the preparation of test sample preparation method and lack negative controls Radix Ophiopogonis.According to thin layer chromatography (" Chinese pharmacopoeia version-appendix VI B of portion in 2005) test, draw each 5 μ l of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, developing solvent one, two, three, launch respectively, take out, dry, spray is with 10% ethanol solution of sulfuric acid, 105 ℃ of heating 5 minutes.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color.
Table 10. developing solvent proportioning preferred
| The clear spot degree | The hangover situation | Disturbed condition | |
| Developing solvent one | Unintelligible | Do not have | Do not have |
| Developing solvent two: | Clear | Do not have | Have |
| Developing solvent three: | Clear | Have | Do not have |
When developing solvent is chloroform-acetone (4:1) as can be seen from the above table, launch effectively on lamellae, principal spot is clear, does not have hangover, and is identical with the speckle displacement and the color of reference substance, is fit to test requirements document.
(3) sample solution point sample amount is preferred:
Get pharmaceutical preparation of the present invention and be equivalent to crude drug 24.08g, add hydrochloric acid 3ml, put in the water-bath heating 1 hour, put coldly, the 30ml jolting that adds diethyl ether is extracted, and divides and gets ether solution, and evaporate to dryness, residue add chloroform 1ml makes dissolving, as need testing solution.Other gets control medicinal material 2g Radix Ophiopogonis, decocts with water 30 minutes, filters, and filtrate is concentrated into 40ml, shines medical material solution in pairs with legal system.Get to lack and write out a prescription Radix Ophiopogonis, press prepared scarce Radix Ophiopogonis of negative preparation among the embodiment 6, press the preparation of test sample preparation method and lack negative controls Radix Ophiopogonis.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw above-mentioned two kinds of solution each 1 μ l, 3 μ l, 5 μ l, 10 μ l, put respectively on same silica gel g thin-layer plate, with chloroform-acetone (4:1) is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, 105 ℃ of heating 5 minutes.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color.
Table 11. sample solution point sample amount optimization experiment is table as a result
| The point sample amount | 1 μ l | 3 μ l | 5 μ l | 10 μ l |
| Effect | Test sample is at corresponding reference substance position immaculate | Test sample is very shallow in corresponding reference substance position spot colors | Test sample is identical in corresponding reference substance position spot colors | Test sample is identical in corresponding reference substance position spot colors, but hangover is arranged slightly. |
Test sample point sample amount is when 5 μ l as can be seen from the above table, and color developing effect is good on lamellae, is fit to test requirements document.
The screening of experimental example 4 Radix Glycyrrhizae discrimination tests
(1) preparation of need testing solution
Method one: get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, porphyrize adds hydrochloric acid 3ml, reflux 0.5 hour, put coldly, add the chloroform jolting and extract 2 times, each 10ml merges chloroform extraction liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.
Method two: get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, porphyrize adds hydrochloric acid 3ml, reflux 1 hour, put coldly, add the chloroform jolting and extract 2 times, each 15ml merges chloroform extraction liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.
Method three: get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, porphyrize adds hydrochloric acid 3ml, reflux 2 hours, put coldly, add the chloroform jolting and extract 2 times, each 20ml merges chloroform extraction liquid, evaporate to dryness, residue add methanol 1ml makes dissolving, as need testing solution.
Extracting liquorice subacid reference substance adds dehydrated alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution in addition.Get scarce Radix Glycyrrhizae prescription, press the negative preparation of the scarce Radix Glycyrrhizae of prepared among the embodiment 6, press the scarce Radix Glycyrrhizae negative controls of test sample preparation method preparation.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 15 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (30～60 ℃)-benzene-ethyl acetate-glacial acetic acid (10:20:8:0.5) is developing solvent, launch, take out, dry, spray is with 10% phosphomolybdic acid ethanol solution, 105 ℃ of heating 5 minutes.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
The preparation of table 12. need testing solution
| The clear spot degree | The hangover situation | Disturbed condition | |
| Method one | Unintelligible | Do not have | Do not have |
| Method two: | Clear | Do not have | Do not have |
| Method three: | Clear | Do not have | Have |
(2) developing solvent proportioning preferred:
Developing solvent one: petroleum ether (30～60 ℃)-benzene-ethyl acetate-glacial acetic acid (10:20:8:1.2)
Developing solvent two: petroleum ether (30～60 ℃)-benzene-ethyl acetate-glacial acetic acid (10:20:8:0.5)
Developing solvent three: ethyl acetate-methanol-water (7:2.5:2.5)
Get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, add hydrochloric acid 3ml, reflux 1 hour is put coldly, adds the chloroform jolting and extracts 2 times, and each 15ml merges chloroform extraction liquid, and evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.Extracting liquorice subacid reference substance adds dehydrated alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution in addition.Get scarce Radix Glycyrrhizae prescription, press the negative preparation of the scarce Radix Glycyrrhizae of prepared among the embodiment 6, press the scarce Radix Glycyrrhizae negative controls of test sample preparation method preparation.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 15 μ l of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, developing solvent one, two, three, launch respectively, take out, dry, spray is with 10% phosphomolybdic acid ethanol solution, 105 ℃ of heating 5 minutes.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
Table 13. developing solvent proportioning preferred
| The clear spot degree | The hangover situation | Disturbed condition | |
| Developing solvent one | Unintelligible | Do not have | Do not have |
| Developing solvent two: | Clear | Do not have | Have |
| Developing solvent three: | Clear | Have | Do not have |
When developing solvent is a petroleum ether (30～60 ℃)-benzene-ethyl acetate-glacial acetic acid (10:20:8:0.5) as can be seen from the above table, launch effectively on lamellae, principal spot is clear, does not have hangover, identical with the speckle displacement and the color of reference substance, be fit to test requirements document.
(3) sample solution point sample amount is preferred:
Get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, add hydrochloric acid 3ml, reflux 1 hour is put coldly, adds the chloroform jolting and extracts 2 times, and each 15ml merges chloroform extraction liquid, and evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.Extracting liquorice subacid reference substance adds dehydrated alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution in addition.Get scarce Radix Glycyrrhizae prescription, press the negative preparation of the scarce Radix Glycyrrhizae of prepared among the embodiment 6, press the scarce Radix Glycyrrhizae negative controls of test sample preparation method preparation.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw above-mentioned two kinds of solution each 10 μ l, 15 μ l, 20 μ l, put respectively on same silica gel g thin-layer plate, with petroleum ether (30～60 ℃)-benzene-ethyl acetate-glacial acetic acid (10:20:8:0.5) is developing solvent, launch, take out, dry, spray is with 10% phosphomolybdic acid ethanol solution, 105 ℃ of heating 5 minutes.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
Table 14. sample solution point sample amount optimization experiment is table as a result
| The point sample amount | 10μl | 15μl | 20μl |
| Effect | Test sample is very shallow in corresponding reference substance position spot colors | Test sample is identical in corresponding reference substance position spot colors | Test sample is identical in corresponding reference substance position spot colors, but hangover is arranged slightly |
Test sample point sample amount is when 15 μ l as can be seen from the above table, and color developing effect is good on lamellae, is fit to test requirements document.
The screening of experimental example 5 Semen Armeniacae Amarum discrimination tests
(1) preparation of need testing solution
Method one: get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, porphyrize, add the water-saturated n-butanol jolting and extract 2 times, each 20ml merges n-butanol extracting liquid, with ammonia scrubbing 2 times, each 20ml discards ammonia liquid, the n-butanol layer evaporate to dryness, the residue 5ml that adds diethyl ether makes dissolving, as need testing solution.
Method two: get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, porphyrize, add the water-saturated n-butanol jolting and extract 2 times, each 20ml merges n-butanol extracting liquid, with ammonia scrubbing 2 times, each 20ml discards ammonia liquid, the n-butanol layer evaporate to dryness, residue adds methanol 5ml makes dissolving, as need testing solution.
Method three: get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, porphyrize, the methanol jolting is extracted 2 times, each 20ml, merge methanol extract liquid, with normal hexane washing 2 times, each 20ml discards normal hexane liquid, normal hexane layer evaporate to dryness, the residue 5ml that adds diethyl ether makes dissolving, as need testing solution.
Other gets the amygdaloside reference substance, adds methanol and makes the solution that 1ml contains 2mg, in contrast product solution.Get scarce Semen Armeniacae Amarum prescription, press the negative preparation of the scarce Semen Armeniacae Amarum of prepared among the embodiment 6, press the scarce Semen Armeniacae Amarum negative controls of test sample preparation method preparation.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 10 μ l of above-mentioned 3 kinds of solution, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with chloroform-methanol-water (13:7:2) is developing solvent, launch, take out, dry, (phosphomolybdic acid 2g adds water 20ml and makes dissolving with the phosphomolybdic acid sulfuric acid solution in spray, slowly adding sulphuric acid 30ml, mixing), it is clear to be heated to speckle colour developing at 105 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
The preparation of table 15. need testing solution
| The clear spot degree | The hangover situation | Disturbed condition | |
| Method one | Unintelligible | Do not have | Do not have |
| Method two | Clear | Do not have | Do not have |
| Method three | Clear | Do not have | Have |
(2) developing solvent proportioning preferred:
Developing solvent one: 10 ℃ of lower floor's solution of placing 12 hours of chloroform-ethyl acetate-methanol-water (15:40:22:10)
Developing solvent two: chloroform-methanol-water (10:10:1) is placed stratified lower floor solution below 10 ℃
Developing solvent three: chloroform-methanol-water (13:7:2) is placed stratified lower floor solution below 10 ℃
Get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, add the water-saturated n-butanol jolting and extract 2 times, each 20ml merges n-butanol extracting liquid, with ammonia scrubbing 2 times, each 20ml discards ammonia liquid, n-butanol layer evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution.Other gets the amygdaloside reference substance, adds methanol and makes the solution that 1ml contains 2mg, in contrast product solution.Get scarce Semen Armeniacae Amarum prescription, press the negative preparation of the scarce Semen Armeniacae Amarum of prepared among the embodiment 6, press the scarce Semen Armeniacae Amarum negative controls of test sample preparation method preparation.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 10 μ l of above-mentioned 3 kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, developing solvent one, two, three, launch respectively, take out, dry, (phosphomolybdic acid 2g adds water 20ml and makes dissolving with the phosphomolybdic acid sulfuric acid solution in spray, slowly adding sulphuric acid 30ml, mixing), it is clear to be heated to speckle colour developing at 100～110 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
Table 16 developing solvent proportioning preferred
| The clear spot degree | The hangover situation | Disturbed condition | |
| Developing solvent one | Unintelligible | Do not have | Do not have |
| Developing solvent two: | Clear | Do not have | Have |
| Developing solvent three: | Clear | Do not have | Do not have |
Developing solvent is chloroform-methanol-water (13:7:2) when placing stratified lower floor solution below 10 ℃ as can be seen from the above table, launches effectively on lamellae, and principal spot is clear, does not have hangover, and is identical with the speckle displacement and the color of reference substance, is fit to test requirements document.
(3) sample solution point sample amount is preferred:
Get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, add the water-saturated n-butanol jolting and extract 2 times, each 20ml merges n-butanol extracting liquid, with ammonia scrubbing 2 times, each 20ml discards ammonia liquid, n-butanol layer evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution.Other gets the amygdaloside reference substance, adds methanol and makes the solution that 1ml contains 2mg, in contrast product solution.Get scarce Semen Armeniacae Amarum prescription, press the negative preparation of the scarce Semen Armeniacae Amarum of prepared among the embodiment 6, press the scarce Semen Armeniacae Amarum negative controls of test sample preparation method preparation.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 3 μ l of above-mentioned 3 kinds of solution, 5 μ l, 10 μ l, 15 μ l, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with chloroform-methanol-water (13:7:2) is developing solvent, launch, take out, dry, (phosphomolybdic acid 2g adds water 20ml and makes dissolving with the phosphomolybdic acid sulfuric acid solution in spray, slowly adding sulphuric acid 30ml, mixing), it is clear to be heated to speckle colour developing at 105 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
Table 17 sample solution point sample amount optimization experiment
| The point sample amount | 3 μ l | 5 μ l | 10 μ l | 15 μ l |
| Effect | Test sample is at corresponding reference substance position immaculate | Test sample is very shallow in corresponding reference substance position spot colors | Test sample is identical in corresponding reference substance position spot colors | Test sample is identical in corresponding reference substance position spot colors, but hangover is arranged slightly. |
Test sample point sample amount is when 10 μ l as can be seen from the above table, and color developing effect is good on lamellae, is fit to test requirements document.
The screening of experimental example 6 Pericarpium Trichosanthis discrimination tests
(1) preparation of need testing solution
Method one: get pharmaceutical preparation of the present invention and be equivalent to crude drug 18.06g, porphyrize adds methanol extraction 2 times, each 30ml at every turn, methanol extract liquid steam in, residue adds ethanol 1ml makes dissolving, as need testing solution.
Method two: get pharmaceutical preparation of the present invention and be equivalent to crude drug 18.06g, porphyrize adds water saturated n-butanol extraction 2 times, each 30ml at every turn, n-butanol extracting liquid steam in, residue adds methanol 1ml makes dissolving, as need testing solution.
Method three: get pharmaceutical preparation of the present invention and be equivalent to crude drug 18.06g, porphyrize adds ethanol extraction 2 times, each 30ml at every turn, ethanol extract steam in, residue adds methanol 1ml makes dissolving, as need testing solution.
Get Pericarpium Trichosanthis control medicinal material 3g, add 60% ethanol 20ml merceration 2h after, supersound process 30min filters, filtrate evaporate to dryness, residue add methanol 1ml makes dissolving, makes control medicinal material solution.Get scarce Pericarpium Trichosanthis prescription, press the negative preparation of the scarce Pericarpium Trichosanthis of prepared among the embodiment 6, press the scarce Pericarpium Trichosanthis negative controls of test sample preparation method preparation.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 10 μ L of above-mentioned 3 kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (60～90 ℃)-ethyl acetate (4:1) is developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with Pericarpium Trichosanthis control medicinal material chromatograph relevant position on, show the speckle of same color.
The preparation of table 18 need testing solution
| The clear spot degree | The hangover situation | Disturbed condition | |
| Method one | Unintelligible | Do not have | Do not have |
| Method two: | Clear | Do not have | Do not have |
| Method three: | Clear | Do not have | Have |
(2) developing solvent proportioning preferred:
Developing solvent one: petroleum ether (60～90 ℃)-ethyl acetate (5:1)
Developing solvent two: petroleum ether (60～90 ℃)-ethyl acetate (4:1)
Developing solvent three: petroleum ether (60～90 ℃)-ethyl acetate (3:1)
Get pharmaceutical preparation of the present invention and be equivalent to crude drug 18.06g, add water saturated n-butanol extraction 2 times, each 30ml at every turn, n-butanol extracting liquid steam in, residue adds methanol 1ml makes dissolving, as need testing solution.Get Pericarpium Trichosanthis control medicinal material 3g, add 60% ethanol 20ml merceration 2h after, supersound process 30min filters, filtrate evaporate to dryness, residue add methanol 1ml makes dissolving, makes control medicinal material solution.Get scarce Pericarpium Trichosanthis prescription, press the negative preparation of the scarce Pericarpium Trichosanthis of prepared among the embodiment 6, press the scarce Pericarpium Trichosanthis negative controls of test sample preparation method preparation.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 10 μ L of above-mentioned 3 kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, developing solvent one, two, three, launch respectively, take out, dry, spray is with 5% vanillin sulfuric acid solution, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with Pericarpium Trichosanthis control medicinal material chromatograph relevant position on, show the speckle of same color.
Table 19. developing solvent proportioning preferred
| The clear spot degree | The hangover situation | Disturbed condition | |
| Developing solvent one | Unintelligible | Have | Do not have |
| Developing solvent two: | Clear | Do not have | Do not have |
| Developing solvent three: | Clear | Do not have | Have |
When developing solvent is a petroleum ether (60～90 ℃)-ethyl acetate (4:1) as can be seen from the above table, launch effectively on lamellae, principal spot is clear, does not have hangover, and is identical with the speckle displacement and the color of reference substance, is fit to test requirements document.
(3) sample solution point sample amount is preferred:
Get pharmaceutical preparation of the present invention and be equivalent to crude drug 18.06g, add water saturated n-butanol extraction 2 times, each 30ml at every turn, n-butanol extracting liquid steam in, residue adds methanol 1ml makes dissolving, as need testing solution.Get Pericarpium Trichosanthis control medicinal material 3g, add 60% ethanol 20ml merceration 2h after, supersound process 30min filters, filtrate evaporate to dryness, residue add methanol 1ml makes dissolving, makes control medicinal material solution.Get scarce Pericarpium Trichosanthis prescription, press the negative preparation of the scarce Pericarpium Trichosanthis of prepared among the embodiment 6, press the scarce Pericarpium Trichosanthis negative controls of test sample preparation method preparation.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw above-mentioned 3 kinds of solution each 3 μ l, 5 μ l, 10 μ l, 15 μ l, put respectively on same silica gel g thin-layer plate, with petroleum ether (60～90 ℃)-ethyl acetate (4:1) is developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with Pericarpium Trichosanthis control medicinal material chromatograph relevant position on, show the speckle of same color.
Sample solution point sample amount optimization experiment is table as a result
Table 20 sample solution point sample amount preferred
| The point sample amount | 3 μ l | 5 μ l | 10 μ l | 15 μ l |
| Effect | Test sample is at corresponding reference substance position immaculate | Test sample is very shallow in corresponding reference substance position spot colors | Test sample is identical in corresponding reference substance position spot colors | Test sample is identical in corresponding reference substance position spot colors, but hangover is arranged slightly. |
Test sample point sample amount is when 10 μ l as can be seen from the above table, and color developing effect is good on lamellae, is fit to test requirements document.
The screening of experimental example 7 Pericarpium Citri Reticulatae discrimination tests
(1) preparation of need testing solution
Method one: get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, porphyrize, the 20ml jolting that adds diethyl ether is extracted 1 time, discards ether, add the Ethyl formate jolting and extract 2 times, each 20ml merges the Ethyl formate extracting solution, evaporate to dryness, residue add methanol 2ml makes dissolving, as need testing solution.
Method two: get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, porphyrize, adding petroleum ether (60～90 ℃) 20ml jolting extracts 1 time, discard petroleum ether, add the ethyl acetate jolting and extract 2 times, each 20ml, merge ethyl acetate extraction liquid, evaporate to dryness, residue add methanol 2ml makes dissolving, as need testing solution.
Method three: get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, porphyrize, adding petroleum ether (60～90 ℃) 20ml jolting extracts 1 time, discard petroleum ether, add the Ethyl formate jolting and extract 2 times, each 20ml, merge the Ethyl formate extracting solution, evaporate to dryness, residue add methanol 2ml makes dissolving, as need testing solution.
Other gets Pericarpium Citri Reticulatae glycosides reference substance, adds methanol and makes saturated solution, in contrast product solution.Get scarce Pericarpium Citri Reticulatae prescription, press the negative preparation of the scarce Pericarpium Citri Reticulatae of prepared among the embodiment 6, press the scarce Pericarpium Citri Reticulatae negative controls of test sample preparation method preparation.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 6 μ l of above-mentioned 3 kinds of solution, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with chloroform-methanol-water (13:7:2) is developing solvent, launch about 15cm, take out, dry, spray is put under the ultra-violet lamp (365nm) and is inspected with the aluminum chloride test solution.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.
The preparation of table 21. need testing solution
| The clear spot degree | The hangover situation | Disturbed condition | |
| Method one | Unintelligible | Do not have | Do not have |
| Method two: | Clear | Do not have | Do not have |
| Method three: | Clear | Do not have | Have |
(2) developing solvent proportioning preferred:
Developing solvent one: chloroform-methanol-water (10:10:2) is placed stratified lower floor solution below 10 ℃
Developing solvent two: chloroform-methanol (19:1)
Developing solvent three: chloroform-methanol-water (13:7:2) is placed stratified lower floor solution below 10 ℃
Get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, add petroleum ether (60～90 ℃) 20ml jolting and extract 1 time, discard petroleum ether, add the ethyl acetate jolting and extract 2 times, each 20ml merges ethyl acetate extraction liquid, evaporate to dryness, residue add methanol 2ml makes dissolving, as need testing solution.Other gets Pericarpium Citri Reticulatae glycosides reference substance, adds methanol and makes saturated solution, in contrast product solution.Get scarce Pericarpium Citri Reticulatae prescription, press the negative preparation of the scarce Pericarpium Citri Reticulatae of prepared among the embodiment 6, press the scarce Pericarpium Citri Reticulatae negative controls of test sample preparation method preparation.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 6 μ l of above-mentioned 3 kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, developing solvent one, two, three, launch about 15cm respectively, take out, dry, spray is put under the ultra-violet lamp (365nm) and is inspected with the aluminum chloride test solution.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.
Table 22 developing solvent proportioning preferred
| The clear spot degree | The hangover situation | Disturbed condition | |
| Developing solvent one | Unintelligible | Do not have | Do not have |
| Developing solvent two: | Clear | Do not have | Have |
| Developing solvent three: | Clear | Do not have | Do not have |
Developing solvent is chloroform-methanol-water (13:7:2) when placing stratified lower floor solution below 10 ℃ as can be seen from the above table, launches effectively on lamellae, and principal spot is clear, does not have hangover, and is identical with the speckle displacement and the color of reference substance, is fit to test requirements document.
(3) sample solution point sample amount is preferred:
Get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, add petroleum ether (60～90 ℃) 20ml jolting and extract 1 time, discard petroleum ether, add the ethyl acetate jolting and extract 2 times, each 20ml merges ethyl acetate extraction liquid, evaporate to dryness, residue add methanol 2ml makes dissolving, as need testing solution.Other gets Pericarpium Citri Reticulatae glycosides reference substance, adds methanol and makes saturated solution, in contrast product solution.Get scarce Pericarpium Citri Reticulatae prescription, press the negative preparation of the scarce Pericarpium Citri Reticulatae of prepared among the embodiment 6, press the scarce Pericarpium Citri Reticulatae negative controls of test sample preparation method preparation.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw above-mentioned 3 kinds of solution each 1 μ l, 3 μ l, 6 μ l, 10 μ l, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with chloroform-methanol-water (13:7:2) is developing solvent, launch about 15cm, take out, dry, spray is put under the ultra-violet lamp (365nm) and is inspected with the aluminum chloride test solution.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.
Table 23 sample solution point sample amount optimization experiment result
| The point sample amount | 1 μ l | 3 μ l | 6 μ l | 10 μ l |
| Effect | Test sample is at corresponding reference substance position immaculate | Test sample is very shallow in corresponding reference substance position spot colors | Test sample is identical in corresponding reference substance position spot colors | Test sample is identical in corresponding reference substance position spot colors, but hangover is arranged slightly. |
Test sample point sample amount is when 6 μ l as can be seen from the above table, and color developing effect is good on lamellae, is fit to test requirements document.
The screening of experimental example 8 Radix Platycodonis discrimination tests
(1) preparation of need testing solution
Method one: get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, porphyrize adds 10% ethanol solution of sulfuric acid 5ml, reflux 2h puts coldly, extracts 2 times with the chloroform jolting, each 20ml, combined chloroform liquid adds water 30ml washing, discard washing liquid, the chloroform solution anhydrous sodium sulfate dehydration filters the filtrate evaporate to dryness, residue adds methanol 5ml makes dissolving, as need testing solution.
Method two: get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, porphyrize adds 10% ethanol solution of sulfuric acid 5ml, reflux 3h puts coldly, extracts 2 times with the chloroform jolting, each 20ml, combined chloroform liquid adds water 30ml washing, discard washing liquid, the chloroform solution anhydrous sodium sulfate dehydration filters the filtrate evaporate to dryness, residue adds methanol 5ml makes dissolving, as need testing solution.
Method three: get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, porphyrize adds alcoholic solution 5ml, reflux 4h puts coldly, extracts 2 times with the chloroform jolting, each 20ml, combined chloroform liquid adds water 30ml washing, discard washing liquid, the chloroform solution anhydrous sodium sulfate dehydration filters the filtrate evaporate to dryness, residue adds ethanol 5ml makes dissolving, as need testing solution.
Other gets Radix Platycodonis control medicinal material 1g, makes reference substance solution with method.Get scarce Radix Platycodonis prescription, press the negative preparation of the scarce Radix Platycodonis of prepared among the embodiment 6, press the scarce Radix Platycodonis negative controls of test sample preparation method preparation.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, drawing each 10 μ l of above-mentioned three kinds of solution, put respectively on same silica gel g thin-layer plate, is developing solvent with chloroform-ether (1:1), launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, it is clear to be heated to speckle colour developing at 105 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
The preparation of table 24 need testing solution
| The clear spot degree | The hangover situation | Disturbed condition | |
| Method one | Unintelligible | Do not have | Do not have |
| Method two: | Clear | Do not have | Do not have |
| Method three: | Clear | Do not have | Have |
(2) developing solvent proportioning preferred:
Developing solvent one: chloroform-methanol (20:1)
Developing solvent two: chloroform-methanol-formic acid (16:10:1)
Developing solvent three: chloroform-ether (1:1)
Get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, add 10% ethanol solution of sulfuric acid 5ml, reflux 3h, put coldly, extract 2 times each 20ml with the chloroform jolting, combined chloroform liquid, add water 30ml washing, discard washing liquid, the chloroform solution anhydrous sodium sulfate dehydration, filter, filtrate evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution.Other gets Radix Platycodonis control medicinal material 1g, makes reference substance solution with method.Get scarce Radix Platycodonis prescription, press the negative preparation of the scarce Radix Platycodonis of prepared among the embodiment 6, press the scarce Radix Platycodonis negative controls of test sample preparation method preparation.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 10 μ l of above-mentioned three kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, developing solvent one, two, three, launch respectively, take out, dry, spray is with 10% ethanol solution of sulfuric acid, it is clear to be heated to speckle colour developing at 105 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
Table 25. developing solvent proportioning preferred
| The clear spot degree | The hangover situation | Disturbed condition | |
| Developing solvent one | Unintelligible | Do not have | Do not have |
| Developing solvent two: | Clear | Do not have | Have |
| Developing solvent three: | Clear | Do not have | Do not have |
When developing solvent is chloroform-ether (1:1) as can be seen from the above table, launch effectively on lamellae, principal spot is clear, does not have hangover, and is identical with the speckle displacement and the color of reference substance, is fit to test requirements document.
(3) sample solution point sample amount is preferred:
Get pharmaceutical preparation of the present invention and be equivalent to crude drug 12.04g, add 10% ethanol solution of sulfuric acid 5ml, reflux 3h, put coldly, extract 2 times each 20ml with the chloroform jolting, combined chloroform liquid, add water 30ml washing, discard washing liquid, the chloroform solution anhydrous sodium sulfate dehydration, filter, filtrate evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution.Other gets Radix Platycodonis control medicinal material 1g, makes reference substance solution with method.Get scarce Radix Platycodonis prescription, press the negative preparation of the scarce Radix Platycodonis of prepared among the embodiment 6, press the scarce Radix Platycodonis negative controls of test sample preparation method preparation.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, drawing above-mentioned 3 kinds of solution each 5 μ l, 10 μ l, 15 μ l, put respectively on same silica gel g thin-layer plate, is developing solvent with chloroform-ether (1:1), launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, it is clear to be heated to speckle colour developing at 105 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
Table 26 sample solution point sample amount optimization experiment is table as a result
| The point sample amount | 5μl | 10μl | 15μl |
| Effect | Test sample is very shallow in corresponding reference substance position spot colors | Test sample is identical in corresponding reference substance position spot colors | Test sample is identical in corresponding reference substance position spot colors, but hangover is arranged slightly |
Test sample point sample amount is when 10 μ l as can be seen from the above table, and color developing effect is good on lamellae, is fit to test requirements document.
The content assaying method test of experimental example 9 Exocarpium Citri Grandises
Monarch drug in the assay Exocarpium Citri Grandis side of being, naringin are its effective ingredient, for ensuring drug quality, are the assay object so select naringin, adopt high performance liquid chromatography, and naringin is carried out assay.
1. the preparation method precision of test sample is measured this product 1ml, puts in the 25ml measuring bottle, adds methanol and is diluted to scale, shakes up, and filters, and gets subsequent filtrate, promptly.
2. measure the selection of wavelength and draw naringin reference substance solution 10 μ l, need testing solution 10 μ l, inject high performance liquid chromatograph, through diode array detector the naringin absworption peak is carried out absorption spectromtry in 190nm～370nm, the ultra-violet absorption spectrum basically identical of test sample and reference substance as a result, all strong absorption is arranged, be decided to be 283nm so this product is measured wavelength at the 283nm place.
3. linear relationship is investigated precision and is taken by weighing the naringin reference substance, adds dissolve with methanol, makes the reference substance solution that every 1ml contains 49.8 μ g, and sample introduction 2.5,5,7.5,10,12.5,15 μ l by above-mentioned chromatographic condition, measure naringin peak area integrated value respectively.With the reference substance sample size is abscissa, is vertical coordinate with the naringin peak area.The result shows, in 0.1245 μ g～0.747 μ g scope, the amount of naringin and peak area integrated value are good linear relationship, and be a straight line that was close to initial point, therefore can adopt in test one point external standard method to measure, regression equation Y=1.83556 * 106X-2623.08, γ=0.9995.
4. the precision test is according to prepared need testing solution among the embodiment 6, and replication 5 times writes down naringin absworption peak peak area, and meansigma methods is 870942.8, RSD=1.37% (n=5).
5. to get be example 6 samples for repeatability test, according to 5 parts of test liquids of the parallel preparation of [1] method, carries out assay respectively, and meansigma methods is=1.0252 (mg/ml), RSD=1.49% (n=5).
6. stability test is got embodiment 6 samples according to above-mentioned chromatographic condition, and the preparation need testing solution is respectively at 0,1,2,4,6,8h carries out assay, record naringin absworption peak peak area, RSD=1.24% (n=6).Stable in the need testing solution 8h as a result, can satisfy the mensuration needs.
7. the negative control test is got and is lacked the Exocarpium Citri Grandis prescription, press the negative preparation of the scarce Exocarpium Citri Grandis of prepared among the embodiment 6,, record naringin negative control chromatogram according to the text content assaying method, as a result in the side other composition under this condition determination, noiseless to the assay of naringin.
8. embodiment 6 samples (naringin content is 1.0252mg/ml) 2.5ml is got in the average recovery test, gets 5 parts altogether, adds the naringin reference substance respectively, according to content assaying method among the embodiment 10, measures in accordance with the law, and calculate recovery rate the results are shown in Table 1.
The test of table 27 average recovery
9. sample size is measured according to [1] method and is prepared need testing solution, measures the content of naringin in 3 batch samples, the results are shown in Table 2.
Naringin content in table 28 sample
Following embodiment all can realize the described effect of above-mentioned experimental example
Embodiment 1
Exocarpium Citri Grandis 66g, Pericarpium Citri Reticulatae 66g, Rhizoma Arisaematis 66g, Folium Eriobotryae 66g, Radix Platycodonis 44g, Semen Armeniacae Amarum (peeling is fried) 44g, Poria 44g, Pericarpium Trichosanthis 44g, Radix Glycyrrhizae 33g, Radix Asteris 33g, Radix Ophiopogonis 33g, Rhizoma Anemarrhenae 33g, Radix Peucedani 22g, Gypsum Fibrosum 22g, Radix Aucklandiae 22g
This pharmaceutical composition adds conventional adjuvant, makes capsule, soft capsule, pill, tablet, powder, injection, granule, mixture by common process.
Embodiment 2
Exocarpium Citri Grandis 66g, Pericarpium Citri Reticulatae 66g, Rhizoma Pinelliae Preparatum 55g, Folium Eriobotryae 66g, Radix Platycodonis 44g, Semen Armeniacae Amarum (peeling is fried) 44g, Poria 44g, Pericarpium Trichosanthis 44g, Radix Glycyrrhizae 33g, Radix Asteris 33g, Radix Ophiopogonis 33g, Rhizoma Anemarrhenae 33g, Radix Peucedani 22g, Gypsum Fibrosum 22g, Radix Aucklandiae 22g
This pharmaceutical composition adds conventional adjuvant, makes capsule, soft capsule, pill, tablet, powder, injection, granule, mixture by common process.
Embodiment 3
Exocarpium Citri Grandis 66g, Pericarpium Citri Reticulatae 66g, Rhizoma Arisaematis 66g, Flos Farfarae 66g, Cortex Mori 22g, Radix Platycodonis 44g, Semen Armeniacae Amarum (peeling is fried) 44g, Poria 44g, Pericarpium Trichosanthis 44g, Radix Glycyrrhizae 33g, Radix Asteris 33g, Radix Ophiopogonis 33g, Rhizoma Anemarrhenae 33g, Radix Peucedani 22g, Gypsum Fibrosum 22g, Radix Aucklandiae 22g
Add conventional adjuvant again,, make capsule, soft capsule, pill, tablet, powder, injection, granule, the mixture of clinical acceptance through conventional method.
Embodiment 4
Exocarpium Citri Grandis 66g, Pericarpium Citri Reticulatae 66g, Rhizoma Arisaematis 66g, Folium Eriobotryae 66g, Radix Platycodonis 44g, Semen Armeniacae Amarum (peeling is fried) 44g, Poria 44g, Pericarpium Trichosanthis 44g, Radix Glycyrrhizae 33g, Radix Asteris 33g, Radix Ophiopogonis 33g, Rhizoma Anemarrhenae 33g, Radix Peucedani 22g, Gypsum Fibrosum 22g, Fructus Perillae (stir-fry) 22g
Add conventional adjuvant again,, make capsule, soft capsule, pill, tablet, powder, injection, granule, the mixture of clinical acceptance through conventional method.
Embodiment 5
Exocarpium Citri Grandis 66g, Pericarpium Citri Reticulatae 66g, Rhizoma Arisaematis 66g, Folium Eriobotryae 66g, Radix Platycodonis 44g, Semen Armeniacae Amarum (peeling is fried) 44g, Poria 44g, Pericarpium Trichosanthis 44g, Radix Glycyrrhizae 33g, Radix Asteris 33g, Radix Ophiopogonis 33g, Rhizoma Anemarrhenae 33g, Radix Rehmanniae 22g, Gypsum Fibrosum 22g, Radix Aucklandiae 22g
Add conventional adjuvant again,, make capsule, soft capsule, pill, tablet, powder, injection, granule, the mixture of clinical acceptance through conventional method.
Embodiment 6
Exocarpium Citri Grandis 66g, Pericarpium Citri Reticulatae 66g, Rhizoma Arisaematis 66g, Folium Eriobotryae 66g, Radix Platycodonis 44g, Semen Armeniacae Amarum (peeling is fried) 44g, Poria 44g, Pericarpium Trichosanthis 44g, Radix Glycyrrhizae 33g, Radix Asteris 33g, Radix Ophiopogonis 33g, Rhizoma Anemarrhenae 33g, Radix Peucedani 22g, Gypsum Fibrosum 22g, Radix Aucklandiae 22g
More than ten five tastes, Gypsum Fibrosum powder is broken into coarse powder, decocts with water secondary, each 1 hour, filters filtrate for later use; Exocarpium Citri Grandis, Pericarpium Citri Reticulatae, Folium Eriobotryae, Semen Armeniacae Amarum four flavor vapor distillations are collected distillate 250ml; The distillator inner liquid medicine filters, and filtrate adds ethanol to be made and contain the alcohol amount and reach 60%, stirs evenly, and leaves standstill 24 hours, filters filtrate for later use; Ten flavors such as all the other Rhizoma Arisaematiss, be ground into coarse powder and above-mentioned medicinal residues mixing, according to the percolation under fluid extract and the extractum item (" appendix IO of Chinese pharmacopoeia version in 2005), make solvent with 60% ethanol, flood after 24 hours percolation in accordance with the law, collect the liquid 2700ml that filters, merge with above-mentioned reserve liquid, decompression recycling ethanol merges with the parget water decocting liquid to there not being the alcohol flavor, is concentrated into the clear paste of relative density 1.06 (50 ℃).Add sucrose 80g, boil, left standstill 24 hours, filter, add ethyl hydroxybenzoate 0.3g, benzoic acid 0.5g (both use an amount of hot water dissolving earlier) and distillate, add water and adjust total amount to 950ml, stir evenly, supernatant is got in cold preservation 48 hours, embedding, and sterilization, promptly.
Embodiment 7
Exocarpium Citri Grandis 66g, Pericarpium Citri Reticulatae 66g, Rhizoma Arisaematis 66g, Folium Eriobotryae 66g, Semen Lepidii (Semen Descurainiae) 66g, Cortex Mori 66g, Radix Platycodonis 44g, Semen Armeniacae Amarum (peeling is fried) 44g, Poria 44g, Pericarpium Trichosanthis 44g, Radix Glycyrrhizae 33g, Radix Asteris 33g, Radix Ophiopogonis 33g, Rhizoma Anemarrhenae 33g, Radix Peucedani 22g, Gypsum Fibrosum 22g, Radix Aucklandiae 22g
More than 17 flavors, Gypsum Fibrosum powder is broken into coarse powder, decocts with water secondary, each 1 hour, filters filtrate for later use; Exocarpium Citri Grandis, Pericarpium Citri Reticulatae, Folium Eriobotryae, Semen Lepidii (Semen Descurainiae), Semen Armeniacae Amarum five tastes vapor distillation are collected distillate 250ml; The distillator inner liquid medicine filters, and filtrate adds ethanol to be made and contain the alcohol amount and reach 60%, stirs evenly, and leaves standstill 24 hours, filters filtrate for later use; All the other Rhizoma Arisaematiss etc. ten simply, be ground into coarse powder and above-mentioned medicinal residues mixing, according to the percolation under fluid extract and the extractum item (" appendix IO of Chinese pharmacopoeia version in 2005), make solvent with 60% ethanol, flood after 24 hours percolation in accordance with the law, collect the liquid 2700ml that filters, merge with above-mentioned reserve liquid, decompression recycling ethanol merges with the parget water decocting liquid to there not being the alcohol flavor, is concentrated into the clear paste of relative density 1.06 (50 ℃).Add sucrose 80g, boil, left standstill 24 hours, filter, add ethyl hydroxybenzoate 0.3g, benzoic acid 0.5g (both use an amount of hot water dissolving earlier) and distillate, add water and adjust total amount to 950ml, stir evenly, supernatant is got in cold preservation 48 hours, embedding, and sterilization, promptly.
Embodiment 8
Exocarpium Citri Grandis 66g, Pericarpium Citri Reticulatae 66g, Rhizoma Pinelliae Preparatum 66g, Flos Farfarae 66g, Radix Platycodonis 44g, Semen Armeniacae Amarum (peeling is fried) 44g, Poria 44g, Pericarpium Trichosanthis 44g, Radix Glycyrrhizae 33g, Radix Asteris 33g, Radix Ophiopogonis 33g, Rhizoma Anemarrhenae 33g, Radix Rehmanniae 22g, Gypsum Fibrosum 22g, Fructus Perillae (stir-fry) 22g
More than ten five tastes, Gypsum Fibrosum powder is broken into coarse powder, decocts with water secondary, each 1 hour, filters filtrate for later use; Exocarpium Citri Grandis, Pericarpium Citri Reticulatae, Flos Farfarae, Semen Armeniacae Amarum four flavor vapor distillations are collected distillate 250ml; The distillator inner liquid medicine filters, and filtrate adds ethanol to be made and contain the alcohol amount and reach 60%, stirs evenly, and leaves standstill 24 hours, filters filtrate for later use; Ten flavors such as all the other Rhizoma Pinelliae Preparatum, be ground into coarse powder and above-mentioned medicinal residues mixing, according to the percolation under fluid extract and the extractum item (" an appendix I of Chinese pharmacopoeia version in 2005 O), make solvent with 60% ethanol, flood after 24 hours percolation in accordance with the law, collect the liquid 2700ml that filters, merge with above-mentioned reserve liquid, decompression recycling ethanol merges with the parget water decocting liquid to there not being the alcohol flavor, is concentrated into the clear paste of relative density 1.06 (50 ℃).Add sucrose 80g, boil, left standstill 24 hours, filter, add ethyl hydroxybenzoate 0.3g, benzoic acid 0.5g (both use an amount of hot water dissolving earlier) and distillate, add water and adjust total amount to 950ml, stir evenly, supernatant is got in cold preservation 48 hours, embedding, and sterilization, promptly.
The method of quality control of embodiment 9 preparations of the present invention
Getting embodiment 6 contents differentiates:
[discriminating]
(1) get this product 40ml, add hydrochloric acid 3ml, put in the water-bath heating 1 hour, put coldly, the 30ml jolting that adds diethyl ether is extracted, and divides and gets ether solution, and evaporate to dryness, residue add chloroform 1ml makes dissolving, as need testing solution.Other gets control medicinal material 2g Radix Ophiopogonis, decocts with water 30 minutes, filters, and filtrate is concentrated into 40ml, shines medical material solution in pairs with legal system.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 5～10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with chloroform-acetone (4:1) is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, 105 ℃ of heating 5 minutes.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color.
(2) get this product 20ml, add hydrochloric acid 3ml, reflux 1 hour is put coldly, adds the chloroform jolting and extracts 2 times, and each 15ml merges chloroform extraction liquid, and evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.Extracting liquorice subacid reference substance adds dehydrated alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution in addition.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 15 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (30～60 ℃)-benzene-ethyl acetate-glacial acetic acid (10:20:7:0.5) is developing solvent, launch, take out, dry, spray is with 10% phosphomolybdic acid ethanol solution, 105 ℃ of heating 5 minutes.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
(3) get this product 20ml, add the water-saturated n-butanol jolting and extract 2 times, each 20ml merges n-butanol extracting liquid, uses ammonia scrubbing 2 times, and each 20ml discards ammonia liquid, and n-butanol layer evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution.Other gets the amygdaloside reference substance, adds methanol and makes the solution that 1ml contains 2mg, in contrast product solution.Get scarce Semen Armeniacae Amarum and write out a prescription, lack the negative preparation of Semen Armeniacae Amarum, press the test sample preparation method and prepare scarce Semen Armeniacae Amarum negative controls by prepared.According to thin layer chromatography (" Chinese pharmacopoeia version-appendix VI B of portion in 2005) test, draw each 10 μ l of above-mentioned 3 kinds of solution, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with chloroform-methanol-water (13:7:2) is developing solvent, launch, take out, dry, spray is with phosphomolybdic acid sulfuric acid solution (phosphomolybdic acid 2g, add water 20ml and make dissolving, slowly adding sulphuric acid 30ml, mixing), it is clear to be heated to speckle colour developing at 105 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color, lacking Semen Armeniacae Amarum negative control liquid chromatography does not have corresponding speckle.
(4) get this product 20ml, add petroleum ether (60～90 ℃) 20ml jolting and extract 1 time, discard petroleum ether, add the ethyl acetate jolting and extract 2 times, each 20ml merges ethyl acetate extraction liquid, and evaporate to dryness, residue add methanol 2ml makes dissolving, as need testing solution.Other gets Pericarpium Citri Reticulatae glycosides reference substance, adds methanol and makes saturated solution, in contrast product solution.Get scarce Pericarpium Citri Reticulatae and write out a prescription, lack the negative preparation of Pericarpium Citri Reticulatae, press the test sample preparation method and prepare scarce Pericarpium Citri Reticulatae negative controls by prepared.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 6 μ l of above-mentioned 3 kinds of solution, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with chloroform-methanol-water (13:7:2) is developing solvent, launch about 15cm, take out, dry, spray is put under the ultra-violet lamp (365nm) and is inspected with the aluminum chloride test solution.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.Lack Pericarpium Citri Reticulatae negative control liquid chromatography and do not have corresponding speckle.
(5) get this product 20ml, add 10% ethanol solution of sulfuric acid 5ml, reflux 3h, put coldly, extract 2 times each 20ml with the chloroform jolting, combined chloroform liquid, add water 30ml washing, discard washing liquid, the chloroform solution anhydrous sodium sulfate dehydration, filter, filtrate evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution.Other gets Radix Platycodonis control medicinal material 1g, makes reference substance solution with method.Get scarce Radix Platycodonis and write out a prescription, lack the negative preparation of Radix Platycodonis, press the test sample preparation method and prepare scarce Radix Platycodonis negative controls by prepared.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 10 μ l of above-mentioned three kinds of solution, put respectively on same silica gel g thin-layer plate, with chloroform-ether (1: 1) is developing solvent, launches, and takes out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing at 105 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.Lack Radix Platycodonis negative control liquid chromatography and do not have corresponding speckle.
(6) get this product 30ml and add water saturated n-butanol extraction 2 times, each 30ml at every turn, n-butanol extracting liquid steam in, residue adds methanol 1ml makes dissolving, as need testing solution.Get Pericarpium Trichosanthis control medicinal material 3g, add 60% ethanol 20ml merceration 2h after, supersound process 30min filters, filtrate evaporate to dryness, residue add methanol 1ml makes dissolving, makes control medicinal material solution.Get scarce Pericarpium Trichosanthis and write out a prescription, lack the negative preparation of Pericarpium Trichosanthis, press the test sample preparation method and prepare scarce Pericarpium Trichosanthis negative controls by prepared.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 10 μ L of above-mentioned 3 kinds of solution, put respectively on same silica gel g thin-layer plate, [one is developing solvent with petroleum ether (60～90 ℃) ethyl acetates (4:1), launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with Pericarpium Trichosanthis control medicinal material chromatograph relevant position on, show mutually the speckle of color just; Negative controls does not have this speckle,
The method of quality control of embodiment 10 preparations of the present invention
Get embodiment 6 contents and carry out assay:
[assay] photograph high performance liquid chromatography (" appendix VID of Chinese pharmacopoeia version in 2005) measure.
Chromatographic condition and system suitability test are filler with the octadecylsilane chemically bonded silica; With methanol-water-acetic acid (38: 62:0.5) be mobile phase; The detection wavelength is 283nm; Column temperature is 40 ℃.Theoretical cam curve is calculated by the naringin peak should be not less than 3000.
The preparation of reference substance solution is taken at 110 ℃ of about 15mg of naringin reference substance that are dried to constant weight, and accurate the title decides, and puts in the 100ml measuring bottle, adds methanol and makes dissolving in right amount and be diluted to scale, shakes up; Precision is measured 3ml, puts in the 10ml measuring bottle, adds methanol to scale, shakes up, and promptly gets (every 1ml contains naringin 45 μ g).
The preparation precision of need testing solution is measured this product 1ml, puts in the 25ml measuring bottle, adds methanol and is diluted to scale, shakes up, and filters, and gets subsequent filtrate, promptly.
Accurate respectively reference substance solution and each the 10 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly.
The every 1ml of this product contains Exocarpium Citri Grandis with naringin (C
27H
32O
14) meter, must not be less than 0.80mg.
The method of quality control of embodiment 11 preparations of the present invention
Getting embodiment 6 contents differentiates and assay:
[discriminating]
(1) get this product 40ml, add hydrochloric acid 3ml, put in the water-bath heating 1 hour, put coldly, the 30ml jolting that adds diethyl ether is extracted, and divides and gets ether solution, and evaporate to dryness, residue add chloroform 1ml makes dissolving, as need testing solution.Other gets control medicinal material 2g Radix Ophiopogonis, decocts with water 30 minutes, filters, and filtrate is concentrated into 40ml, shines medical material solution in pairs with legal system.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 5～10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with chloroform-acetone (4:1) is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, 105 ℃ of heating 5 minutes.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color.
(2) get this product 20ml, add hydrochloric acid 3ml, reflux 1 hour is put coldly, adds the chloroform jolting and extracts 2 times, and each 15ml merges chloroform extraction liquid, and evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.Extracting liquorice subacid reference substance adds dehydrated alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution in addition.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 15 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (30～60 ℃)-benzene-ethyl acetate-glacial acetic acid (10:20:7:0.5) is developing solvent, launch, take out, dry, spray is with 10% phosphomolybdic acid ethanol solution, 105 ℃ of heating 5 minutes.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
(3) get this product 20ml, add the water-saturated n-butanol jolting and extract 2 times, each 20ml merges n-butanol extracting liquid, uses ammonia scrubbing 2 times, and each 20ml discards ammonia liquid, and n-butanol layer evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution.Other gets the amygdaloside reference substance, adds methanol and makes the solution that 1ml contains 2mg, in contrast product solution.Get scarce Semen Armeniacae Amarum and write out a prescription, lack the negative preparation of Semen Armeniacae Amarum, press the test sample preparation method and prepare scarce Semen Armeniacae Amarum negative controls by prepared.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 10 μ l of above-mentioned 3 kinds of solution, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with chloroform-methanol-water (13:7:2) is developing solvent, launch, take out, dry, spray is with phosphomolybdic acid sulfuric acid solution (phosphomolybdic acid 2g, add water 20ml and make dissolving, slowly adding sulphuric acid 30ml, mixing), it is clear to be heated to speckle colour developing at 105 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color, lacking Semen Armeniacae Amarum negative control liquid chromatography does not have corresponding speckle.
(4) get this product 20ml, add petroleum ether (60～90 ℃) 20ml jolting and extract 1 time, discard petroleum ether, add the ethyl acetate jolting and extract 2 times, each 20ml merges ethyl acetate extraction liquid, and evaporate to dryness, residue add methanol 2ml makes dissolving, as need testing solution.Other gets Pericarpium Citri Reticulatae glycosides reference substance, adds methanol and makes saturated solution, in contrast product solution.Get scarce Pericarpium Citri Reticulatae and write out a prescription, lack the negative preparation of Pericarpium Citri Reticulatae, press the test sample preparation method and prepare scarce Pericarpium Citri Reticulatae negative controls by prepared.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 6 μ l of above-mentioned 3 kinds of solution, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with chloroform-methanol-water (13:7:2) is developing solvent, launch about 15cm, take out, dry, spray is put under the ultra-violet lamp (365nm) and is inspected with the aluminum chloride test solution.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.Lack Pericarpium Citri Reticulatae negative control liquid chromatography and do not have corresponding speckle.
(5) get this product 20ml, add 10% ethanol solution of sulfuric acid 5ml, reflux 3h, put coldly, extract 2 times each 20ml with the chloroform jolting, combined chloroform liquid, add water 30ml washing, discard washing liquid, the chloroform solution anhydrous sodium sulfate dehydration, filter, filtrate evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution.Other gets Radix Platycodonis control medicinal material 1g, makes reference substance solution with method.Get scarce Radix Platycodonis and write out a prescription, lack the negative preparation of Radix Platycodonis, press the test sample preparation method and prepare scarce Radix Platycodonis negative controls by prepared.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 10 μ l of above-mentioned three kinds of solution, put respectively on same silica gel g thin-layer plate, with chloroform-ether (1: 1) is developing solvent, launches, and takes out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing at 105 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.Lack Radix Platycodonis negative control liquid chromatography and do not have corresponding speckle.
(6) get this product 30ml and add water saturated n-butanol extraction 2 times, each 30ml at every turn, n-butanol extracting liquid steam in, residue adds methanol 1ml makes dissolving, as need testing solution.Get Pericarpium Trichosanthis control medicinal material 3g, add 60% ethanol 20ml merceration 2h after, supersound process 30min filters, filtrate evaporate to dryness, residue add methanol 1ml makes dissolving, makes control medicinal material solution.Get scarce Pericarpium Trichosanthis and write out a prescription, lack the negative preparation of Pericarpium Trichosanthis, press the test sample preparation method and prepare scarce Pericarpium Trichosanthis negative controls by prepared.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 10 μ L of above-mentioned 3 kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (60～90 ℃) ethyl acetates (4:1) is developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with Pericarpium Trichosanthis control medicinal material chromatograph relevant position on, show mutually the speckle of color just; Negative controls does not have this speckle,
[assay] photograph high performance liquid chromatography (" appendix VID of Chinese pharmacopoeia version in 2005) measure.
Chromatographic condition and system suitability test are filler with the octadecylsilane chemically bonded silica; With methanol-water-acetic acid (38: 62:0.5) be mobile phase; The detection wavelength is 283nm; Column temperature is 40 ℃.Theoretical cam curve is calculated by the naringin peak should be not less than 3000.
The preparation of reference substance solution is taken at 110 ℃ of about 15mg of naringin reference substance that are dried to constant weight, and accurate the title decides, and puts in the 100ml measuring bottle, adds methanol and makes dissolving in right amount and be diluted to scale, shakes up; Precision is measured 3ml, puts in the 10ml measuring bottle, adds methanol to scale, shakes up, and promptly gets (every 1ml contains naringin 45 μ g).
The preparation precision of need testing solution is measured this product 1ml, puts in the 25ml measuring bottle, adds methanol and is diluted to scale, shakes up, and filters, and gets subsequent filtrate, promptly.
Accurate respectively reference substance solution and each the 10 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly.
The every 1ml of this product contains Exocarpium Citri Grandis with naringin (C
27H
32O
14) meter, must not be less than 0.80mg.
The preparation method and the method for quality control of the oral liquid of embodiment 12 medicine groups of the present invention
[prescription] Exocarpium Citri Grandis 66g, Pericarpium Citri Reticulatae 66g, Rhizoma Arisaematis 66g, Folium Eriobotryae 66g, Radix Platycodonis 44g, Semen Armeniacae Amarum (peeling is fried) 44g, Poria 44g, Pericarpium Trichosanthis 44g, Radix Glycyrrhizae 33g, Radix Asteris 33g, Radix Ophiopogonis 33g, Rhizoma Anemarrhenae 33g, Radix Peucedani 22g, Gypsum Fibrosum 22g, Radix Aucklandiae 22g
[method for making] above ten five tastes, Gypsum Fibrosum powder is broken into coarse powder, decocts with water secondary, each 1 hour, filters filtrate for later use; Exocarpium Citri Grandis, Pericarpium Citri Reticulatae, Folium Eriobotryae, Semen Armeniacae Amarum four flavor vapor distillations are collected distillate 250ml; The distillator inner liquid medicine filters, and filtrate adds ethanol to be made and contain the alcohol amount and reach 60%, stirs evenly, and leaves standstill 24 hours, filters filtrate for later use; Ten flavors such as all the other Rhizoma Arisaematiss, be ground into coarse powder and above-mentioned medicinal residues mixing, according to the percolation under fluid extract and the extractum item (" appendix IO of Chinese pharmacopoeia version in 2005), make solvent with 60% ethanol, flood after 24 hours percolation in accordance with the law, collect the liquid 2700ml that filters, merge with above-mentioned reserve liquid, decompression recycling ethanol merges with the parget water decocting liquid to there not being the alcohol flavor, is concentrated into the clear paste of relative density 1.06 (50 ℃).Add sucrose 80g, boil, left standstill 24 hours, filter, add ethyl hydroxybenzoate 0.3g, benzoic acid 0.5g (both use an amount of hot water dissolving earlier) and distillate, add water and adjust total amount to 950ml, stir evenly, supernatant is got in cold preservation 48 hours, embedding, and sterilization, promptly.
[discriminating]
(1) get this product 40ml, add hydrochloric acid 3ml, put in the water-bath heating 1 hour, put coldly, the 30ml jolting that adds diethyl ether is extracted, and divides and gets ether solution, and evaporate to dryness, residue add chloroform 1ml makes dissolving, as need testing solution.Other gets control medicinal material 2g Radix Ophiopogonis, decocts with water 30 minutes, filters, and filtrate is concentrated into 40ml, shines medical material solution in pairs with legal system.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 5～10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with chloroform-acetone (4:1) is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, 105 ℃ of heating 5 minutes.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color.
(2) get this product 20ml, add hydrochloric acid 3ml, reflux 1 hour is put coldly, adds the chloroform jolting and extracts 2 times, and each 15ml merges chloroform extraction liquid, and evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.Extracting liquorice subacid reference substance adds dehydrated alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution in addition.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 15 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (30～60 ℃)-benzene-ethyl acetate-glacial acetic acid (10:20:7:0.5) is developing solvent, launch, take out, dry, spray is with 10% phosphomolybdic acid ethanol solution, 105 ℃ of heating 5 minutes.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
(3) get this product 20ml, add the water-saturated n-butanol jolting and extract 2 times, each 20ml merges n-butanol extracting liquid, uses ammonia scrubbing 2 times, and each 20ml discards ammonia liquid, and n-butanol layer evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution.Other gets the amygdaloside reference substance, adds methanol and makes the solution that 1ml contains 2mg, in contrast product solution.Get scarce Semen Armeniacae Amarum and write out a prescription, lack the negative preparation of Semen Armeniacae Amarum, press the test sample preparation method and prepare scarce Semen Armeniacae Amarum negative controls by prepared.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 10 μ l of above-mentioned 3 kinds of solution, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with chloroform-methanol-water (13:7:2) is developing solvent, launch, take out, dry, spray is with phosphomolybdic acid sulfuric acid solution (phosphomolybdic acid 2g, add water 20ml and make dissolving, slowly adding sulphuric acid 30ml, mixing), it is clear to be heated to speckle colour developing at 105 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color, lacking Semen Armeniacae Amarum negative control liquid chromatography does not have corresponding speckle.
(4) get this product 20ml, add petroleum ether (60～90 ℃) 20ml jolting and extract 1 time, discard petroleum ether, add the ethyl acetate jolting and extract 2 times, each 20ml merges ethyl acetate extraction liquid, and evaporate to dryness, residue add methanol 2ml makes dissolving, as need testing solution.Other gets Pericarpium Citri Reticulatae glycosides reference substance, adds methanol and makes saturated solution, in contrast product solution.Get scarce Pericarpium Citri Reticulatae and write out a prescription, lack the negative preparation of Pericarpium Citri Reticulatae, press the test sample preparation method and prepare scarce Pericarpium Citri Reticulatae negative controls by prepared.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 6 μ l of above-mentioned 3 kinds of solution, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with chloroform-methanol-water (13:7:2) is developing solvent, launch about 15cm, take out, dry, spray is put under the ultra-violet lamp (365nm) and is inspected with the aluminum chloride test solution.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.Lack Pericarpium Citri Reticulatae negative control liquid chromatography and do not have corresponding speckle.
(5) get this product 20ml, add 10% ethanol solution of sulfuric acid 5ml, reflux 3h, put coldly, extract 2 times each 20ml with the chloroform jolting, combined chloroform liquid, add water 30ml washing, discard washing liquid, the chloroform solution anhydrous sodium sulfate dehydration, filter, filtrate evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution.Other gets Radix Platycodonis control medicinal material 1g, makes reference substance solution with method.Get scarce Radix Platycodonis and write out a prescription, lack the negative preparation of Radix Platycodonis, press the test sample preparation method and prepare scarce Radix Platycodonis negative controls by prepared.According to thin layer chromatography (" appendix VIB of Chinese pharmacopoeia version in 2005) test, draw each 10 μ l of above-mentioned three kinds of solution, put respectively on same silica gel g thin-layer plate, with chloroform one ether (1: 1) is developing solvent, launches, and takes out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing at 105 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.Lack Radix Platycodonis negative control liquid chromatography and do not have corresponding speckle.
(6) get this product 30ml and add water saturated n-butanol extraction 2 times, each 30ml at every turn, n-butanol extracting liquid steam in, residue adds methanol 1ml makes dissolving, as need testing solution.Get Pericarpium Trichosanthis control medicinal material 3g, add 60% ethanol 20ml merceration 2h after, supersound process 30min filters, filtrate evaporate to dryness, residue add methanol 1ml makes dissolving, makes control medicinal material solution.Get scarce Pericarpium Trichosanthis and write out a prescription, lack the negative preparation of Pericarpium Trichosanthis, press the test sample preparation method and prepare scarce Pericarpium Trichosanthis negative controls by prepared.According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 10 μ L of above-mentioned 3 kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (60～90 ℃) ethyl acetates (4:1) is developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with Pericarpium Trichosanthis control medicinal material chromatograph relevant position on, show mutually the speckle of color just; Negative controls does not have this speckle.
[assay] photograph high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 D) measure.
Chromatographic condition and system suitability test are filler with the octadecylsilane chemically bonded silica; With methanol-water-acetic acid (38: 62:0.5) be mobile phase; The detection wavelength is 283nm; Column temperature is 40 ℃.Theoretical cam curve is calculated by the naringin peak should be not less than 3000.
The preparation of reference substance solution is taken at 110 ℃ of about 15mg of naringin reference substance that are dried to constant weight, and accurate the title decides, and puts in the 100ml measuring bottle, adds methanol and makes dissolving in right amount and be diluted to scale, shakes up; Precision is measured 3ml, puts in the 10ml measuring bottle, adds methanol to scale, shakes up, and promptly gets (every 1ml contains naringin 45 μ g).
The preparation precision of need testing solution is measured this product 1ml, puts in the 25ml measuring bottle, adds methanol and is diluted to scale, shakes up, and filters, and gets subsequent filtrate, promptly.
Accurate respectively reference substance solution and each the 10 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly.
The every 1ml of this product contains Exocarpium Citri Grandis with naringin (C
27H
32O
14) meter, must not be less than 0.80mg.
[function with cure mainly] lung heat clearing, cough-relieving is reduced phlegm.Be used for cough with copious phlegm, fullness in the chest and shortness of breath, dry pharynx itching of the throat that accumulation of phlegm-heat in the lung causes.
[usage and consumption] is oral, a 10ml, 2～3 times on the one; Child's consumption is followed the doctor's advice.
[attention] avoids pungent food greasy.
[specification] every dress 10ml.
Shady and cool place is put in [storage] sealing.
Claims (10)
- A relieving cough and resolving phlegm Chinese medicine composition, it is characterized in that making by the crude drug of following weight ratio:Exocarpium Citri Grandis 50-100 weight portion, Pericarpium Citri Reticulatae 50-100 weight portion, Rhizoma Arisaematis 50-100 weight portion, Folium Eriobotryae 50-100 weight portion, Radix Platycodonis 30-70 weight portion, Semen Armeniacae Amarum (peeling is fried) 30-70 weight portion, Poria 30-70 weight portion, Pericarpium Trichosanthis 30-70 weight portion, Radix Glycyrrhizae 20-50 weight portion, Radix Asteris 20-50 weight portion, Radix Ophiopogonis 20-50 weight portion, Rhizoma Anemarrhenae 20-50 weight portion, Radix Peucedani 10-35 weight portion, Gypsum Fibrosum 10-35 weight portion, Radix Aucklandiae 10-35 weight portion.
- 2. Chinese medicine composition as claimed in claim 1 is characterized in that Rhizoma Arisaematis wherein can be substituted by Rhizoma Typhonii, Semen Sinapis Albae, Rhizoma Pinelliae Preparatum, Flos Inulae.
- 3. Chinese medicine composition as claimed in claim 1 is characterized in that Folium Eriobotryae wherein can be substituted by the Radix Stemonae, Flos Farfarae, Semen Lepidii (Semen Descurainiae), Cortex Mori.
- 4. Chinese medicine composition as claimed in claim 1 is characterized in that the Radix Aucklandiae wherein can be substituted by Fructus Perillae (stir-fry).
- 5. Chinese medicine composition as claimed in claim 1 is characterized in that Radix Peucedani wherein can be substituted by Caulis Bambusae In Taenia, Succus Bambusae, Radix Rehmanniae, Lapis Micae Aureus.
- 6. Chinese medicine composition as claimed in claim 1 is characterized in that being made by the crude drug of following weight ratio:Exocarpium Citri Grandis 66 weight portions, Pericarpium Citri Reticulatae 66 weight portions, Rhizoma Arisaematis 66 weight portions, Folium Eriobotryae 66 weight portions, Radix Platycodonis 44 weight portions, Semen Armeniacae Amarum (peeling is fried) 44 weight portions, Poria 44 weight portions, Pericarpium Trichosanthis 44 weight portions, Radix Glycyrrhizae 33 weight portions, Radix Asteris 33 weight portions, Radix Ophiopogonis 33 weight portion, the Rhizoma Anemarrhenae 33 weight portions, Radix Peucedani 22 weight portions, Gypsum Fibrosum 22 weight portions, the Radix Aucklandiae 22 weight portions.
- 7. as the preparation method of each described Chinese medicine composition of claim 1-6, it is characterized in that this method is:Gypsum Fibrosum powder is broken into coarse powder, decocts with water 1-3 time, each 0.5-2 hour, filters filtrate for later use; Exocarpium Citri Grandis, Pericarpium Citri Reticulatae, Folium Eriobotryae, Semen Armeniacae Amarum four flavor vapor distillations are collected distillate 200-300ml; The distillator inner liquid medicine filters, and filtrate adds ethanol to be made and contain the alcohol amount and reach 50-70%, stirs evenly, and leaves standstill 16-36 hour, filters filtrate for later use; Ten flavors such as all the other Rhizoma Arisaematiss are ground into coarse powder and above-mentioned medicinal residues mixing, according to the percolation under 2005 editions one appendix I O fluid extract of Chinese Pharmacopoeia and the extractum item, make solvent with 50-70% ethanol, flood after 12-36 hour percolation in accordance with the law, collect the liquid 2500-3000ml that filters, merge with above-mentioned reserve liquid, decompression recycling ethanol is to there not being the alcohol flavor, merge with the parget water decocting liquid, the clear paste of relative density 1.06 when being concentrated into 50 ℃ adds conventional adjuvant again, through conventional method, make the preparation of clinical acceptance.
- 8. the preparation method of Chinese medicine composition mixture as claimed in claim 7 is characterized in that this method is:Gypsum Fibrosum powder is broken into coarse powder, decocts with water secondary, each 1 hour, filters filtrate for later use; Exocarpium Citri Grandis, Pericarpium Citri Reticulatae, Folium Eriobotryae, Semen Armeniacae Amarum four flavor vapor distillations are collected distillate 250ml; The distillator inner liquid medicine filters, and filtrate adds ethanol to be made and contain the alcohol amount and reach 60%, stirs evenly, and leaves standstill 24 hours, filters filtrate for later use; Ten flavors such as all the other Rhizoma Arisaematiss, be ground into coarse powder and above-mentioned medicinal residues mixing, according to the percolation under 2005 editions appendix IO fluid extracts of Chinese Pharmacopoeia and the extractum item, make solvent with 60% ethanol, flood after 24 hours percolation in accordance with the law, collect the liquid 2700ml that filters, merge with above-mentioned reserve liquid, decompression recycling ethanol merges the clear paste of relative density 1.06 when being concentrated into 50 ℃ to there not being the alcohol flavor with the parget water decocting liquid; Add sucrose 80g, boil, left standstill 24 hours, filter, add ethyl hydroxybenzoate 0.3g, benzoic acid 0.5g and distillate, add water and adjust total amount to 950ml, stir evenly, supernatant is got in cold preservation 48 hours, embedding, and sterilization, promptly.
- 9. the method for quality control of Chinese medicine composition as claimed in claim 7 is characterized in that this method of quality control comprises one or more in following discrimination method and/or the assay:[discriminating](1) get pharmaceutical composition of the present invention and be equivalent to crude drug 20-30g, add hydrochloric acid 2-4ml, put in the water-bath heating 0.5-2 hour, put coldly, the 20-40ml jolting that adds diethyl ether is extracted, and divides and gets ether solution, and evaporate to dryness, residue add chloroform 1ml makes dissolving, as need testing solution; Other gets control medicinal material 2g Radix Ophiopogonis, decocts with water 20-40 minute, filters, and filtrate is concentrated into 30-50ml, shines medical material solution in pairs with legal system; According to " appendix a VI of Chinese pharmacopoeia version in 2005 B thin layer chromatography test, draw each 5～10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, chloroform-acetone with the 3.5-4.5:0.5-1.5 proportioning is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, 100～110 ℃ of heating 4-6 minute; In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color;(2) get pharmaceutical composition of the present invention and be equivalent to crude drug 10-15g, add hydrochloric acid 2-4ml, reflux 0.5-2 hour, put cold, add the chloroform jolting and extract 1-3 time, each 10-20ml merges chloroform extraction liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution; Extracting liquorice subacid reference substance adds dehydrated alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution in addition; According to " appendix a VI of Chinese pharmacopoeia version in 2005 B thin layer chromatography test, draw each 15 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, petroleum ether (30～60 ℃)-benzene-ethyl acetate-glacial acetic acid with the 8-12:15-25:5-10:0-1 proportioning is developing solvent, launch, take out, dry, spray is with 10% phosphomolybdic acid ethanol solution, 100～110 ℃ of heating 4-6 minute; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;(3) get pharmaceutical composition of the present invention and be equivalent to crude drug 10-15g, adding the water-saturated n-butanol jolting extracts 1-3 time, each 15-25ml, merge n-butanol extracting liquid, use ammonia scrubbing 1-3 time, each 15-25ml, discard ammonia liquid, n-butanol layer evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution; Other gets the amygdaloside reference substance, adds methanol and makes the solution that 1ml contains 2mg, in contrast product solution; According to " appendix a VI of Chinese pharmacopoeia version in 2005 B thin layer chromatography test, draw each 10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with the chloroform-methanol-water of 10-15:5-10:1-3 proportioning is developing solvent, launch, take out, dry, spray is with the phosphomolybdic acid sulfuric acid solution, it is clear to be heated to speckle colour developing at 100～110 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;(4) get pharmaceutical composition of the present invention and be equivalent to crude drug 10-15g, adding petroleum ether (60～90 ℃) 15-25ml jolting extracts 1 time, discard petroleum ether, adding the ethyl acetate jolting extracts 1-3 time, each 15-25ml merges ethyl acetate extraction liquid, evaporate to dryness, residue adds methanol 2ml makes dissolving, as need testing solution; Other gets Pericarpium Citri Reticulatae glycosides reference substance, adds methanol and makes saturated solution, in contrast product solution; According to " appendix a VI of Chinese pharmacopoeia version in 2005 B thin layer chromatography test, draw each 6 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with the chloroform-methanol-water of 10-15:5-10:1-3 proportioning is developing solvent, launch about 10-20cm, take out, dry, spray is put under the ultra-violet lamp (365nm) and is inspected with the aluminum chloride test solution; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color;(5) get pharmaceutical composition of the present invention and be equivalent to crude drug 10-15g, add 10% ethanol solution of sulfuric acid 4-6ml, reflux 4-6h, put coldly, extract 1-3 time with the chloroform jolting, at every turn 15-25ml, combined chloroform liquid, add water 20-40ml washing, discard washing liquid, the chloroform solution anhydrous sodium sulfate dehydration, filter, filtrate evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution; Other gets Radix Platycodonis control medicinal material 1g, makes reference substance solution with method; According to " appendix a VI of Chinese pharmacopoeia version in 2005 B thin layer chromatography test, drawing each 10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, is developing solvent with the chloroform-ether of 45-55:45-55 proportioning, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, it is clear to be heated to speckle colour developing at 100～110 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;(6) get pharmaceutical composition of the present invention and be equivalent to crude drug 15-20g, add water saturated n-butanol extraction 1-3 time, each 25-40ml at every turn, n-butanol extracting liquid steam in, residue adds methanol 1ml makes dissolving, as need testing solution; Get Pericarpium Trichosanthis control medicinal material 3g, add 60% ethanol 20ml merceration 2h after, supersound process 30min filters, filtrate evaporate to dryness, residue add methanol 1ml makes dissolving, makes control medicinal material solution; According to " appendix a VI of Chinese pharmacopoeia version in 2005 B thin layer chromatography test, draw each 10 μ L of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, petroleum ether (60～90 ℃)-ethyl acetate with the 35-45:5-15 proportioning is developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and it is clear to be heated to the speckle colour developing; In the test sample chromatograph, with Pericarpium Trichosanthis control medicinal material chromatograph relevant position on, show the speckle of same color;[assay] is according to " appendix a VI of Chinese pharmacopoeia version in 2005 D high effective liquid chromatography for measuring;Chromatographic condition and system suitability test are filler with the octadecylsilane chemically bonded silica; Methanol-water-acetic acid with the 30-40:60-70:0-1 proportioning is mobile phase; The detection wavelength is 283nm; Column temperature is 40 ℃; Theoretical cam curve is calculated by the naringin peak should be not less than 3000;The preparation of reference substance solution is taken at the 100-120 ℃ of about 15mg of naringin reference substance that is dried to constant weight, and accurate the title decides, and puts in the 100ml measuring bottle, adds methanol and makes dissolving in right amount and be diluted to scale, shakes up; Precision is measured 3ml, puts in the 10ml measuring bottle, adds methanol to scale, shakes up, and promptly gets the reference substance solution that every 1ml contains naringin 45 μ g;The preparation precision of need testing solution is measured pharmaceutical composition of the present invention and is equivalent to crude drug 0.3-1.0g, puts in the 25ml measuring bottle, adds methanol and is diluted to scale, shakes up, and filters, and gets subsequent filtrate, promptly;Accurate respectively reference substance solution and each the 10 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly;The every 1ml of this product contains Exocarpium Citri Grandis with naringin (C 27H 32O 14) meter, must not be less than 0.80mg.
- 10. the method for quality control of Chinese medicine composition mixture as claimed in claim 8 is characterized in that this method of quality control preferably includes one or more in following discrimination method and/or the assay:[discriminating](1) get pharmaceutical composition mixture 40ml of the present invention, add hydrochloric acid 3ml, put in the water-bath heating 1 hour, put coldly, the 30ml jolting that adds diethyl ether is extracted, and divides and gets ether solution, and evaporate to dryness, residue add chloroform 1ml makes dissolving, as need testing solution; Other gets control medicinal material 2g Radix Ophiopogonis, decocts with water 30 minutes, filters, and filtrate is concentrated into 40ml, shines medical material solution in pairs with legal system; According to " appendix a VI of Chinese pharmacopoeia version in 2005 B thin layer chromatography test, draw each 5～10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, chloroform-acetone with the 4:1 proportioning is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, 105 ℃ of heating 5 minutes; In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color;(2) get pharmaceutical composition mixture 20ml of the present invention, add hydrochloric acid 3ml, reflux 1 hour is put coldly, adds the chloroform jolting and extracts 2 times, and each 15ml merges chloroform extraction liquid, and evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution; Extracting liquorice subacid reference substance adds dehydrated alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution in addition; According to " appendix a VI of Chinese pharmacopoeia version in 2005 B thin layer chromatography test, draw each 15 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, petroleum ether (30～60 ℃)-benzene-ethyl acetate-glacial acetic acid with the 10:20:7:0.5 proportioning is developing solvent, launch, take out, dry, spray is with 10% phosphomolybdic acid ethanol solution, 105 ℃ of heating 5 minutes; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;(3) get pharmaceutical composition mixture 20ml of the present invention, add the water-saturated n-butanol jolting and extract 2 times, each 20ml merges n-butanol extracting liquid, with ammonia scrubbing 2 times, each 20ml discards ammonia liquid, n-butanol layer evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution; Other gets the amygdaloside reference substance, adds methanol and makes the solution that 1ml contains 2mg, in contrast product solution; According to " appendix a VI of Chinese pharmacopoeia version in 2005 B thin layer chromatography test, draw each 10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with the chloroform-methanol-water of 13:7:2 proportioning is developing solvent, launch, take out, dry, spray is with the phosphomolybdic acid sulfuric acid solution, it is clear to be heated to speckle colour developing at 105 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;(4) get pharmaceutical composition mixture 20ml of the present invention, add petroleum ether (60～90 ℃) 20ml jolting and extract 1 time, discard petroleum ether, add the ethyl acetate jolting and extract 2 times, each 20ml merges ethyl acetate extraction liquid, evaporate to dryness, residue add methanol 2ml makes dissolving, as need testing solution; Other gets Pericarpium Citri Reticulatae glycosides reference substance, adds methanol and makes saturated solution, in contrast product solution; According to " appendix a VI of Chinese pharmacopoeia version in 2005 B thin layer chromatography test, draw each 6 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, placing stratified lower floor solution below 10 ℃ with the chloroform-methanol-water of 13:7:2 proportioning is developing solvent, launch about 15cm, take out, dry, spray is put under the ultra-violet lamp (365nm) and is inspected with the aluminum chloride test solution; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color;(5) get pharmaceutical composition mixture 20ml of the present invention, add 10% ethanol solution of sulfuric acid 5ml, reflux 3h, put coldly, extract 2 times each 20ml with the chloroform jolting, combined chloroform liquid, add water 30ml washing, discard washing liquid, the chloroform solution anhydrous sodium sulfate dehydration, filter, filtrate evaporate to dryness, residue add methanol 5ml makes dissolving, as need testing solution; Other gets Radix Platycodonis control medicinal material 1g, makes reference substance solution with method; According to " each 10 μ l of above-mentioned two kinds of solution are drawn in the test of an appendix VI of Chinese pharmacopoeia version in 2005 B thin layer chromatography, put respectively on same silica gel g thin-layer plate, chloroform-ether with the 1:1 proportioning is developing solvent, launches, and takes out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing at 105 ℃, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;(6) get pharmaceutical composition mixture 30ml of the present invention, add water saturated n-butanol extraction 2 times, each 30ml at every turn, n-butanol extracting liquid steam in, residue adds methanol 1ml makes dissolving, as need testing solution; Get Pericarpium Trichosanthis control medicinal material 3g, add 60% ethanol 20ml merceration 2h after, supersound process 30min filters, filtrate evaporate to dryness, residue add methanol 1ml makes dissolving, makes control medicinal material solution; According to " appendix a VI of Chinese pharmacopoeia version in 2005 B thin layer chromatography test, draw each 10 μ L of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, petroleum ether (60～90 ℃)-ethyl acetate with the 4:1 proportioning is developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and it is clear to be heated to the speckle colour developing; In the test sample chromatograph, with Pericarpium Trichosanthis control medicinal material chromatograph relevant position on, show the speckle of same color;[assay] is according to " appendix a VI of Chinese pharmacopoeia version in 2005 D high effective liquid chromatography for measuring;Chromatographic condition and system suitability test are filler with the octadecylsilane chemically bonded silica; Methanol-water-acetic acid with the 38:62:0.5 proportioning is mobile phase; The detection wavelength is 283nm; Column temperature is 40 ℃; Theoretical cam curve is calculated by the naringin peak should be not less than 3000;The preparation of reference substance solution is taken at 110 ℃ of about 15mg of naringin reference substance that are dried to constant weight, and accurate the title decides, and puts in the 100ml measuring bottle, adds methanol and makes dissolving in right amount and be diluted to scale, shakes up; Precision is measured 3ml, puts in the 10ml measuring bottle, adds methanol to scale, shakes up, and promptly gets every 1ml and contains naringin 45 μ g;The preparation precision of need testing solution is measured pharmaceutical composition of the present invention and is equivalent to crude drug 0.6g, puts in the 25ml measuring bottle, adds methanol and is diluted to scale, shakes up, and filters, and gets subsequent filtrate, promptly;Accurate respectively reference substance solution and each the 10 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly;The every 1ml of this product contains Exocarpium Citri Grandis with naringin (C 27H 32O 14) meter, must not be less than 0.80mg.
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| CN102370875A (en) * | 2010-08-17 | 2012-03-14 | 北京亚东生物制药有限公司 | Effervescing agent for relieving cough and reducing sputum and preparation method thereof |
| CN102406032A (en) * | 2011-12-01 | 2012-04-11 | 广州潘高寿药业股份有限公司 | Throat-smoothing drop containing traditional Chinese medicine extractives |
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| CN102370875A (en) * | 2010-08-17 | 2012-03-14 | 北京亚东生物制药有限公司 | Effervescing agent for relieving cough and reducing sputum and preparation method thereof |
| CN102370875B (en) * | 2010-08-17 | 2014-03-26 | 北京亚东生物制药有限公司 | Effervescing agent for relieving cough and reducing sputum and preparation method thereof |
| CN102406032A (en) * | 2011-12-01 | 2012-04-11 | 广州潘高寿药业股份有限公司 | Throat-smoothing drop containing traditional Chinese medicine extractives |
| CN102406032B (en) * | 2011-12-01 | 2012-12-12 | 广州白云山潘高寿药业股份有限公司 | Throat-smoothing drop containing traditional Chinese medicine extractives |
| CN102759600A (en) * | 2012-08-06 | 2012-10-31 | 四川逢春制药有限公司 | Detection method of sufi cough syrup |
| CN104382157A (en) * | 2014-10-23 | 2015-03-04 | 广西陆川县泓源食品有限公司 | Aster tataricus health-care beverage |
| CN105232936A (en) * | 2015-10-25 | 2016-01-13 | 四川双鑫生物科技有限公司 | Decoction medicine capable of treating cold, cough and phlegm and preparation method thereof |
| CN110859915A (en) * | 2018-08-28 | 2020-03-06 | 康芝药业股份有限公司 | Cough-relieving exocarpium citri rubrum granules and preparation method thereof |
| CN111632105A (en) * | 2019-02-22 | 2020-09-08 | 康芝药业股份有限公司 | Cough-relieving exocarpium citri rubrum granules and preparation method thereof |
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