CN102091299A

CN102091299A - Method for detecting medicinal composition for treating viral hepatitis

Info

Publication number: CN102091299A
Application number: CN 201110033498
Authority: CN
Inventors: 付立家; 付建家
Original assignee: Beijing Asia East Bio Pharmaceutical Co Ltd
Current assignee: Beijing Asia East Bio Pharmaceutical Co Ltd
Priority date: 2007-08-03
Filing date: 2007-08-03
Publication date: 2011-06-15
Anticipated expiration: 2027-08-03
Also published as: CN102091299B

Abstract

The invention discloses a method for detecting a medicinal composition for treating viral hepatitis. The medicinal composition is prepared from the following raw material medicaments: virgate wormwood herb, indigowoad root, male fern rhizome, Indian buead, root-tuber of aromatic turmeric, barbed skullcap herb, patchouli and the like. The method for detecting the medicinal composition is obtained by screening a great number of specific creative tests. In an identification method, the identification specificity is high by screening sample processing methods and selecting a developing solvent; and the method is economic and practical, is used for quickly acquiring results and can be applied to different thin layer plates. In a content detecting method, the quality of products can be effectively controlled by screening samples and sample supply processing methods and selecting the developing solvent. The products detected by the method have more stable medicinal effects than products detected by other methods.

Description

A kind of detection method for the treatment of the viral hepatitis pharmaceutical composition

The present invention is for dividing an application, and the original bill application number is 200710119920.5, the original bill applying date is 2007 On August 3, in, the original bill name is called A kind of pharmaceutical composition and quality control side for the treatment of viral hepatitis Method

Invention field

The present invention relates to a kind of pharmaceutical composition and preparation method and method of quality control, particularly a kind of pharmaceutical composition and preparation method and method of quality control for the treatment of viral hepatitis.

Background technology

Viral hepatitis is the common transmittable disease that is caused by multiple hepatitis virus, has that infectiousness is strong, the route of transmission is complicated, popular wide general, and sickness rate is than characteristics such as height.Mainly show as weak, loss of appetite clinically, feel sick, vomiting, hepatomegaly and liver function injury, part patient can have jaundice and heating.Urticaria, arthralgia or upper airway symptoms appear in some patient, severe patient even can develop into liver cirrhosis even hepatocarcinoma, the serious harm human health.Though at present the medicine of used clinically treatment viral hepatitis has certain curative effect, exist to some extent body to be developed immunity to drugs and defective such as addiction, side effect be obvious.

Summary of the invention

The object of the invention is to provide a kind of pharmaceutical composition; Another purpose of the present invention is to provide a kind of pharmaceutical composition for the treatment of viral hepatitis; The 3rd purpose of the present invention is to provide this preparation of drug combination method; The 4th purpose of the present invention is to provide the method for quality control of this pharmaceutical composition.

The present invention seeks to 1,2 to realize by following technical solution:

Technical scheme 1:

The crude drug of pharmaceutical composition of the present invention consists of:

Poria 500-1500 weight portion, Herba Artemisiae Scopariae 500-1500 weight portion, Rhizoma Atractylodis Macrocephalae 500-1500 weight portion, Radix Curcumae 150-450 weight portion, Radix Angelicae Sinensis 250-750 weight portion, succinum 50-150 weight portion, the Radix Paeoniae Alba (stir-fry) 500-1500 weight portion, Herba Scutellariae Barbatae 500-1500 weight portion, Fructus Amomi 100-300 weight portion, Rhizoma Polygoni Cuspidati 250-750 weight portion, Radix Salviae Miltiorrhizae 500-1500 weight portion, Herba Lycopi 250-750 weight portion, Radix Bupleuri 150-450 weight portion, Herba Pogostemonis 150-450 weight portion, Herba Eupatorii 250-750 weight portion, Flos Carthami 150-450 weight portion.

The crude drug composition of pharmaceutical composition of the present invention is preferably:

Poria 960 weight portions, Herba Artemisiae Scopariae 960 weight portions, the Rhizoma Atractylodis Macrocephalae 960 weight portions, Radix Curcumae 320 weight portions, Radix Angelicae Sinensis 480 weight portions, succinum 96 weight portions, the Radix Paeoniae Alba (stir-fry) 960 weight portions, Herba Scutellariae Barbatae 960 weight portions, Fructus Amomi 192 weight portions, Rhizoma Polygoni Cuspidati 480 weight portions, Radix Salviae Miltiorrhizae 960 weight portions, Herba Lycopi's 480 weight portions, Radix Bupleuri 320 weight portions, Herba Pogostemonis 320 weight portions, Herba Eupatorii 480 weight portions, Flos Carthami 320 weight portions.

Technical scheme 2:

Poria 500-1500 weight portion, Herba Artemisiae Scopariae 500-1500 weight portion, Radix Curcumae 150-450 weight portion, Radix Angelicae Sinensis 250-750 weight portion, succinum 50-150 weight portion, the Radix Paeoniae Alba (stir-fry) 500-1500 weight portion, Herba Scutellariae Barbatae 500-1500 weight portion, Fructus Amomi 100-300 weight portion, Rhizoma Polygoni Cuspidati 250-750 weight portion, Radix Salviae Miltiorrhizae 500-1500 weight portion, Herba Lycopi 250-750 weight portion, Radix Bupleuri 150-450 weight portion, Herba Pogostemonis 150-450 weight portion, Herba Eupatorii 250-750 weight portion, Flos Carthami 150-450 weight portion, Radix Isatidis 500-1500 weight portion, Rhizoma Dryopteris Crassirhizomatis 250-750 weight portion, Herba Hedyotidis Diffusae 500-1500 weight portion, Rhizoma Paridis 250-750 weight portion.

Poria 960 weight portions, Herba Artemisiae Scopariae 960 weight portions, Radix Curcumae 320 weight portions, Radix Angelicae Sinensis 480 weight portions, succinum 96 weight portions, the Radix Paeoniae Alba (stir-fry) 960 weight portions, Herba Scutellariae Barbatae 960 weight portions, Fructus Amomi 192 weight portions, Rhizoma Polygoni Cuspidati 480 weight portions, Radix Salviae Miltiorrhizae 960 weight portions, Herba Lycopi's 480 weight portions, Radix Bupleuri 320 weight portions, Herba Pogostemonis 320 weight portions, Herba Eupatorii 480 weight portions, Flos Carthami 320 weight portions, Radix Isatidis 960 weight portions, Rhizoma Dryopteris Crassirhizomatis 480 weight portions, Herba Hedyotidis Diffusae 960 weight portions, Rhizoma Paridis 480 weight portions.

Get the pharmaceutical composition crude drug of technical solution of the present invention 1,2, add conventional adjuvant, according to common process, make the dosage form of clinical acceptance, include but not limited to mixture, concentrated pill, capsule, drop pill, granule, tablet, soft capsule, slow releasing agent, oral liquid.

Pharmaceutical composition technical scheme 1 preparation mixture method of the present invention is: Radix Angelicae Sinensis, Radix Curcumae, Herba Pogostemonis, Herba Eupatorii, Fructus Amomi, Herba Lycopi extracted volatile oil 6-10 hour, and the aqueous solution after distillation device is in addition collected; Volatile oil is with the beta-schardinger dextrin-saturated water solution method inclusion of 3-5 times of weight portion, drying, the volatile oil beta-cyclodextrin inclusion complex, standby; Radix Salviae Miltiorrhizae is with 85% alcohol reflux of 4-8 times of weight portion 1-3 time, and each 0.5-1.5 hour, merge extractive liquid, filtered, and filtrate recycling ethanol is not to there being the alcohol flavor, Radix Salviae Miltiorrhizae extract, standby; Medicinal residues and all the other flavour of a drug decoct with water 1-3 time, and the decocting that at every turn adds 6-10 times of weight portion boiled 1-2 hour, and collecting decoction filters; Aqueous solution after filtrate and the above-mentioned distillation merges, be concentrated into 60～65 ℃ of relative densities and be 1.02～1.04 thick paste I, add ethanol and make and contain alcohol amount and reach 60-80%, left standstill 12-36 hour, filter, filtrate recycling ethanol is to an amount of, merge with above-mentioned Radix Salviae Miltiorrhizae extract, and the thick paste II that to be concentrated into 60～65 ℃ of relative densities be 1.10-1.15, add 80% simple syrup 900-1200 parts by volume, the volatile oil beta-cyclodextrin inclusion complex is ground into fine powder, adds in the extractum; Add sodium benzoate 9-10g again, stir evenly, add water to the 3000-4500 parts by volume, filter, fill, sterilization, promptly.

Pharmaceutical composition technical scheme 2 preparation methoies of the present invention are: above 19 flavors, and Radix Angelicae Sinensis, Radix Curcumae, Herba Pogostemonis, Herba Eupatorii, Fructus Amomi, Herba Lycopi extracted volatile oil 6-10 hour, and the aqueous solution after distillation device is in addition collected; Volatile oil is with the beta-schardinger dextrin-saturated water solution method inclusion of 3-5 times of weight portion, drying, the volatile oil beta-cyclodextrin inclusion complex, standby; Radix Salviae Miltiorrhizae is with 85% alcohol reflux of 4-8 times of weight portion 1-3 time, and each 0.5-1.5 hour, merge extractive liquid, filtered, and filtrate recycling ethanol is not to there being the alcohol flavor, Radix Salviae Miltiorrhizae extract, standby; 12 flavors such as medicinal residues and all the other Herba Artemisiae Scopariaes decoct with water 1-3 time, and the decocting that at every turn adds 6-10 times of weight portion boiled 1-2 hour, and collecting decoction filters; Aqueous solution after filtrate and the above-mentioned distillation merges, be concentrated into 60～65 ℃ of relative densities and be 1.02～1.04 thick paste I, adding ethanol makes the alcohol amount of containing reach 60-80%, left standstill 12-36 hour, filter, filtrate recycling ethanol is to an amount of, merge with above-mentioned Radix Salviae Miltiorrhizae extract, being concentrated into relative density is the thick paste II of 1.10-1.15, add the volatile oil beta-cyclodextrin inclusion complex and be ground into fine powder, add conventional adjuvant, according to common process, make the dosage form of clinical acceptance, include but not limited to mixture, concentrated pill, capsule, drop pill, granule, tablet, soft capsule, slow releasing agent, oral liquid or lyophilized injectable powder.

Pharmaceutical composition technical scheme 2 preparation mixture methods of the present invention are preferably: above 19 flavors, and Radix Angelicae Sinensis, Radix Curcumae, Herba Pogostemonis, Herba Eupatorii, Fructus Amomi, Herba Lycopi extracted volatile oil 8 hours, and the aqueous solution after distillation device is in addition collected; Volatile oil is with 40 ℃ of inclusions of beta-schardinger dextrin-saturated water solution method of 4 times of weight portions, 40 ℃ of dryings, the volatile oil beta-cyclodextrin inclusion complex, standby; Radix Salviae Miltiorrhizae is with 85% alcohol reflux of 6 times of weight portions 2 times, and each 1 hour, merge extractive liquid, filtered, and filtrate recycling ethanol is not to there being the alcohol flavor, Radix Salviae Miltiorrhizae extract, standby; 12 flavors such as medicinal residues and all the other Herba Artemisiae Scopariaes decoct with water 2 times, add 8 times of water gagings at every turn and decoct 1.5 hours, and collecting decoction filters; Aqueous solution after filtrate and the above-mentioned distillation merges, be concentrated into 60～65 ℃ of relative densities and be 1.02～1.04 thick paste I, add ethanol and make and contain alcohol amount and reach 70%, left standstill 24 hours, filter, filtrate recycling ethanol is to an amount of, merge with above-mentioned Radix Salviae Miltiorrhizae extract, and the thick paste II that to be concentrated into 60～65 ℃ of relative densities be 1.10-1.15, add 80% simple syrup 960 parts by volume, the volatile oil beta-cyclodextrin inclusion complex is ground into fine powder, adds in the extractum; Add sodium benzoate 9.6g again, add water to 3200 parts by volume, filter, fill, sterilization, promptly.

Pharmaceutical composition method of quality control of the present invention comprises one or more in following discrimination method and/or the assay:

Differentiate: A, get this drug combination preparation 1/100-3/100 day and use dosage, add water 10-30ml and make dissolving, with ethyl acetate extraction 1-3 time, 20-40ml for the first time, 10-30ml for the second time, combined ethyl acetate liquid, water bath method, residue adds ethyl acetate 0.5-1.5ml makes dissolving, as need testing solution; Other gets Herba Artemisiae Scopariae control medicinal material 0.5-1.5g, add water 40-60ml, decocted 0.5-1.5 hour, get supernatant, be concentrated into 10-30ml, with ethyl acetate extraction 1-3 time, 20-40ml, 10-30ml for the second time for the first time, combined ethyl acetate liquid, water bath method, residue add ethyl acetate 0.5-1.5ml makes dissolving, in contrast medical material solution; Draw need testing solution 0.5-1.5 μ l, control medicinal material solution 4-6 μ l, put respectively on same silica gel g thin-layer plate, with 0.5-2: the 60-90 of 0.5-2 ratio ℃ petroleum ether-ethyl acetate is developing solvent, launches, take out, dry, put under the 365nm ultra-violet lamp and observe, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color;

B, get this drug combination preparation 4/1000-8/1000 day and use the dosage porphyrize, add 40-60% ethanol 5-25ml, supersound process 10-30 minute, filter, filtrate is as need testing solution; Other gets the peoniflorin reference substance, adds ethanol and makes the solution that every 1ml contains 0.5-1.5mg, in contrast product solution; Test according to thin layer chromatography, draw each 5-15 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with 30-50: 1-10: 5-15: the chloroform-ethyl acetate of 0.1-0.3 ratio-methanol-formic acid is developing solvent, launch, take out, dry, spray is with 1-10% vanillin sulfuric acid solution, 100-110 ℃ to be heated to speckle colour developing clear, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;

C, get this drug combination preparation 1/100-5/100 day and use dosage, porphyrize, the 20-40ml that adds diethyl ether, reflux, extract, 20-40 minute, filter, filtrate volatilizes, and residue adds ethyl acetate 0.4-0.6ml makes dissolving, as need testing solution; Other gets Radix Angelicae Sinensis control medicinal material 1g, the 5-15ml that adds diethyl ether, and supersound process 5-15 minute, filter, filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, in contrast medical material solution; Test according to thin layer chromatography, draw each 4-6 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with 5-15: the 60-90 of 0.1-1.0 ratio ℃ petroleum ether-ethyl acetate is developing solvent, launches, take out, dry, put under the 365nm ultra-violet lamp and inspect, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color;

D, get the tanshinone IIA reference substance, add ethyl acetate and make the solution that every 1ml contains 1-3mg, in contrast product solution; Get this drug combination preparation 1/100-5/100 day and use dosage, porphyrize, the 20-40ml that adds diethyl ether, reflux, extract, 20-40 minute, filter, filtrate volatilizes, and residue adds ethyl acetate 0.4-0.6ml makes dissolving, as need testing solution, DrawNeed testing solution 5-15 μ l, reference substance solution 4-6 μ l puts respectively on same silica gel g thin-layer plate, and with 4-8: the cyclohexane extraction-ethyl acetate of 1 ratio is developing solvent, launches, and takes out, and dries; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show identical kermesinus speckle;

E, get this drug combination preparation 1/100-3/100 day and use dosage, porphyrize adds methanol 10-30ml, supersound process 10-30 minute, filter the filtrate evaporate to dryness, residue adds 1.5-3.5mol/L sulfuric acid solution 5-15ml makes dissolving, puts in the water-bath and heats 20-40 minute, immediately cooling, extract twice with the chloroform jolting, each 5-15ml merges chloroform liquid, evaporate to dryness, residue adds methanol 0.5-1.5ml makes dissolving, as need testing solution; Other gets the emodin reference substance, adds methanol and makes the solution that every 1ml contains 0.5-1.5mg, in contrast product solution; Test according to thin layer chromatography, draw each 4-8 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with 5-25: 4-6: the upper solution of 30～60 ℃ of petroleum ether-Ethyl formate-formic acid of 0.5-1.5 ratio is developing solvent, launch, take out, dry, put in the ammonia smoked after, inspect under the daylight; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;

Assay: shine high effective liquid chromatography for measuring: chromatographic condition and system suitability test are filler with octadecylsilane chemically bonded silica; 5-25: the acetonitrile-water of 70-95 ratio is a mobile phase, and the detection wavelength is 230nm, and number of theoretical plate calculates by the peoniflorin peak should be not less than 2600;

The preparation of reference substance solution: precision takes by weighing peoniflorin reference substance 4-6mg, puts in the 100ml measuring bottle, adds dissolve with methanol and is diluted to scale, shakes up, and promptly gets the reference substance solution that contains peoniflorin 40-60 μ g among every 1ml;

The need testing solution preparation: get this drug combination preparation under the content uniformity item, porphyrize is got 6/10000-1/1000 day and is used dosage, the accurate title, decide, and puts in the tool plug conical flask, and precision adds 50-70% methanol 15-35ml, close plug claims to decide weight, power 250W, frequency 40KHZ supersound process 20-40 minute is taken out, and puts cold, claim again to decide weight, supply minimizing weight, shake up with 50-70% methanol, get supernatant, filter, promptly;

Algoscopy: accurate respectively reference substance solution and each 5-15 μ l of need testing solution of drawing, inject chromatograph of liquid, measure, promptly;

This drug combination preparation contains the Radix Paeoniae Alba with peoniflorin C with dosage every day ₂₃H ₂₈O ₁₁Meter should be less than 50-70mg.

The pass of weight portion of the present invention and parts by volume is a grams per milliliter.

The daily dosage of the different preparations of pharmaceutical composition of the present invention (every day taking dose or every day using dosage) is different because of preparation, but it is identical to contain suitable crude drug amount in the daily dosage of different preparations.Quality determining method of the present invention is a measurement unit with daily dosage.Convert according to the amount of crude drug, adult every day, the amount with the suitable crude drug of dosage was: 280-360g.

Pharmaceutical composition of the present invention confirms through experimentation: has the remarkable effect that recovers ALT, AST, liver function leading indicators such as GGT, BiL to possess instant effect, has no side effect, and safe in utilization, obtain advantages such as imitating the difficult recurrence in back.

The present composition is compared existing preparation and is possessed good drug effect, and the adjuvant technology of pharmaceutical composition of the present invention is found volatile oil and beta-schardinger dextrin-enclose under certain condition and proportioning through screening, can reach curative effect preferably; 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.

Description of drawings

Figure of description is volatile oil beta-cyclodextrin inclusion experiment flow figure.

The specific embodiment

Following experimental example and embodiment are used to further specify but are not limited to the present invention.

Experimental example 1: the medicine group to the liver protecting and ALT lowering, function of gallbladder promoting, jaundice eliminating, engulf the pharmacodynamics test of the ability of cleaning up

Medicine group I: the pharmaceutical composition mixture of getting the embodiment of the invention 2 preparations

Medicine group II: the pharmaceutical composition mixture of getting the embodiment of the invention 13 preparations

Matched group: commercially available QINGRESHUGAN HEJI.

One. function for protecting liver and reducing enzyme activity

1. to the protective effect of acute liver damage

Get 84 of kunming mices, body weight 18-20g, male and female half and half are divided into 7 groups at random.First group: the normal control group gavages the equivalent drinking water every day; Second group: model group gavages the equivalent drinking water every day; The 3rd group: medicine group I gavages medicine group I medicinal liquid 30ml (being equivalent to crude drug 9.32g)/kg body weight day; The 4th group: medicine group II gavages medicine group II medicinal liquid 30ml (being equivalent to crude drug 9.32g)/kg body weight day; The 5th group: matched group gavages 0.4g/ml QINGRESHUGAN HEJI 0.3ml/10g (being equivalent to contain crude drug 134g/kg) body weight day.The second～five group, at successive administration after 14 days, with D-galactosamine 0.8g/kg body weight mouse peritoneal injection carrying out modeling.After modeling last administration in 24 hours half an hour, get blood from mouse orbit, survey SGPT, SGOT content, the results are shown in Table 1.

Table 1 medicine group of the present invention is to the protective effect of D-galactosamine induced mice acute liver damage (X ± SD)

^*Compare with model group: ^*P＜0.05; ^*P＜0.01;

Experimental result shows, with the normal control group relatively, SGPT, SGOT content obviously raise (P＜0.01) in the serum of model group; Compare with model group, SGPT, the SGOT content of medicine group I of the present invention, II and matched group obviously reduce (P＜0.01); And medicine group of the present invention and matched group and no significant difference.

2. to the protective effect of alcoholic liver injury

Animal grouping and all the same test of administration, in administration, the 2nd～6 group gavages 50% Chinese liquor (strong, colourless liquor distilled from sorghum, 56 °) 0.5ml/20g modeling.Continuously the modeling administration is 30 days, gets blood after last administration half an hour, surveys serum SGPT, SGOT content, the results are shown in Table 2.

Table 2 medicine group of the present invention is to the protective effect of alcoholic liver injury mice (X ± SD)

^*Compare with model group: ^*P＜0.05; ^*P＜0.01;

Experimental result shows, with normal group relatively, SGPT, SGOT content obviously raise (P＜0.01) in the serum of model group.Compare with model group, medicine group of the present invention and matched group all can reduce SGPT in the serum, SGOT content (P＜0.01); And medicine group of the present invention and matched group and no significant difference.

3. to the protective effect of chronic hepatic injury

Get 72 of Wistar rats, body weight 140～150g, male and female half and half are divided into 6 groups at random.First group: the normal control group gavages the equivalent drinking water every day; Second group: model group gavages the equivalent drinking water every day; The 3rd group: medicine group I gavages medicine group I medicinal liquid 30ml (being equivalent to crude drug 9.32g)/kg body weight day; The 4th group: medicine group II gavages medicine group II medicinal liquid 30ml (being equivalent to crude drug 9.32g)/kg body weight day; The 5th group: matched group gavages 0.4g/ml QINGRESHUGAN HEJI 0.3ml/10g (being equivalent to contain crude drug 134g/kg) body weight day.The second～five group, in administration, use 30%CC ₁₄Oil solution 0.3ml/100 subcutaneous injection carries out modeling, twice weekly.The modeling administration was got blood and is surveyed serum SGOT, SGOT, hydroxyproline, total protein, albumin content, and calculate the A/G value after 3 months continuously, the results are shown in Table 3.

Table 3 medicine group of the present invention is to the protective effect of chronic hepatic injury rat (X ± SD)

^*Compare with model group; , ^*P＜0.05; ^*P＜0.01;

Experimental result shows, compares with normal group, and SGPT, SGOT content obviously raise (P＜0.01) in the serum of model group, and the A/G value obviously reduces, and hydroxyproline content obviously raises; Compare with model group, SGPT, the SGOT content of medicine group of the present invention and matched group obviously reduce (P＜0.01); Hydroxyproline content obviously reduces (P＜0.01) in obvious rising of the A/G value of medicine group of the present invention (P＜0.01) and the serum, and the A/G value notable difference (P＜0.05) of matched group; And medicine group of the present invention and matched group and no significant difference.

Two. to of the influence of engulfing clean up ability of mice reticuloendothelial system to inertia carbon granules in the blood flow

Animal, grouping and administration are tested with acute liver damage, more than respectively organized successive administration 20 days, after administration the 7th, 10 day, the 2nd～5 group of mouse peritoneal injection cyclophosphamide 100mg/Kg causes immune function of mice low.After 1 hour, mouse tail vein injects india ink 0.1ml/20g in the last administration, in injecting the back 1,5 minute, gets blood 20 μ l from the vena ophthalmica clump, is dissolved in 2.5ml, in the 0.1%Na2CO3 solution, shakes up, and in 680nm wavelength place colorimetric, measures the OD value.Index K value is cleaned up in calculating.The results are shown in Table 6.

Table 6 medicine group of the present invention is to the influence of macrophage phagocytic function (X ± SD)

^*Compare with model group; ^*P＜0.01

Experimental result shows, compares with normal group, and the K value of model group obviously reduces (P＜0.01).Compare with model group, medicine group of the present invention and matched group K value obviously increase (P＜0.01).

Experimental example 2 adjuvant screening experiment

Volatile oil beta-cyclodextrin inclusion process conditions preferred

Because volatile oil is easy to scatter and disappear, if therefore volatile oil is directly added, to place and with the passing of time or slightly be heated, volatile oil will lose, thereby has reduced content, influences the curative effect of medicine.For addressing this problem, adopted the method for beta-schardinger dextrin-inclusion, and adopted orthogonal experiment that the condition of inclusion has been carried out preferably.

Experiment flow is seen Figure of description.

Adopt the saturated water solution method inclusion: take by weighing predetermined amount of beta-cyclodextrin, add an amount of distilled water, 5% beta-schardinger dextrin-aqueous solution is made in heating in 70 ℃ of water-baths, under the different temperatures (20 ℃, 40 ℃, 60 ℃) stir, rotating speed is 2500 rev/mins, add quantitative volatile oil, fully stir certain hour, get inclusion complex, stop to stir, with the inclusion complex on a small amount of distilled water flushing agitator, airtight, to put in the refrigerator 12 hours, sucking filtration gets the wet product of beta-schardinger dextrin-, through 40 ℃ of dryings of low temperature, get dry product and weigh, use micro-volatile oil extractor, measure by 2005 editions pharmacopeia (appendix X D) first method, record data calculate.

According to Preliminary experiment results, the factor of analyzing influence beta-schardinger dextrin-inclusion is set as follows test, the results are shown in Table 8-10:

Table 8 factor level table

Table 9 result of the test

Table 10 analysis of variance table

F0.1(2，2)＝9，F0.05(2，2)＝19

Result of the test: intuitive analysis is the result know, factor A＞C＞B, optimum condition are A1B2C2; The results of analysis of variance knows, factor A influence is more remarkable, and B, C factor affecting are not remarkable.Should select A1B1C1, but from table 4 result of the test, the inclusion rate of A1B2C2 is greater than A1B1C1, so be asserted A1B2C2, i.e. volatile oil: beta-schardinger dextrin-is 1: 4, and mixing time 30 minutes, inclusion temperature are 40 ℃.

The discrimination test of experimental example 3 Herba Artemisiae Scopariaes

The preparation of blank sample is by medicine group flavour of a drug of the present invention and usage ratio, and autogamy does not contain group's medicine of Herba Artemisiae Scopariae, the Radix Paeoniae Alba, Radix Angelicae Sinensis, Radix Salviae Miltiorrhizae, Rhizoma Polygoni Cuspidati respectively, makes blank preparation by preparation technology.

The preparation of blank solution is got the blank sample of this drug combination preparation Herba Artemisiae Scopariae and was used dosage on 14/1000th, adds water 20ml and makes dissolving, uses ethyl acetate extraction 2 times, 30ml for the first time, 20ml for the second time, combined ethyl acetate liquid, water bath method, residue add ethyl acetate 1ml makes dissolving, as blank solution.

This medicine group and the matter sample of same amount got in the preparation of sample solution, makes sample solution according to the preparation method of blank solution.

Herba Artemisiae Scopariae control medicinal material 1g is got in the preparation of control medicinal material solution, add water 50ml, decocted 1 hour, get supernatant, be concentrated into 20ml, with ethyl acetate extraction 2 times, 30ml, 20ml for the second time for the first time, combined ethyl acetate liquid, water bath method, residue add ethyl acetate 1ml makes dissolving, in contrast medical material solution.

Following developing solvent is prepared in the selection of developing solvent respectively, 60-90 ℃ of petroleum ether-ethyl acetate 1: 4,60-90 ℃ of petroleum ether-ethyl acetate 1: 1,60-90 ℃ of petroleum ether-ethyl acetate 4: 1

Above three kinds of solution are put respectively on same silica gel g thin-layer plate, with 1: 4,1: 1, the 60-90 of 4: 1 ratios ℃ petroleum ether-ethyl acetate was developing solvent respectively, launch, take out, dry, put under the 365nm ultra-violet lamp and observe, when 60-90 ℃ of petroleum ether-ethyl acetate with 1: 1 ratio is developing solvent in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, the fluorescence speckle of tangible same color is arranged, and blank solution no fluorescence speckle demonstration on corresponding position.

The discrimination test of experimental example 4 Radix Paeoniae Albas

The preparation of blank solution is got this pharmaceutical composition blank and preparation 4/1000-8/1000 day is used the dosage porphyrize, adds 40-60% ethanol 5-25ml, and supersound process 10-30 minute, filter, filtrate is as blank solution.

The peoniflorin reference substance is got in the preparation of control medicinal material solution in addition, adds ethanol and makes the solution that every 1ml contains 1mg, in contrast product solution.

The selection preparation of developing solvent 30: 10: 5: 0.3 ratio, 40: 5: 10: 0.2 ratio, 50: 1: 15: the chloroform-ethyl acetate of 0.1 ratio-methanol-formic acid was developing solvent

Test according to thin layer chromatography, draw each 10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, in above-mentioned developing solvent, launch respectively, take out, dry, spray is with 5% vanillin sulfuric acid solution, and 105 ℃ of heating are in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color; Wherein, at 40: 5: 10: the most obvious in the chloroform-ethyl acetate of 0.2 ratio-methanol-formic acid developing solvent.

Experimental example 5 Radix Angelicae Sinensis are differentiated

The preparation of blank solution is got the blank preparation of this pharmaceutical composition and was used the dosage porphyrize on 3/100th, the 30ml that adds diethyl ether, and reflux, extract, 30 minutes filters, and filtrate volatilizes, and residue adds ethyl acetate 0.5ml makes dissolving, as blank solution.

Radix Angelicae Sinensis control medicinal material 1g is got in the preparation of control medicinal material solution, the 10ml that adds diethyl ether, and supersound process 10 minutes filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, in contrast medical material solution.

5: 1 ratios are prepared in the selection of developing solvent respectively, 19: 1 ratios, the 60-90 of 150: 1 ratios ℃ petroleum ether-ethyl acetate is developing solvent.

Test according to thin layer chromatography, draw each 5 μ l of above-mentioned three kinds of solution, put respectively on same silica gel g thin-layer plate, under above-mentioned developing solvent condition, launch respectively, take out, dry, put under the 365nm ultra-violet lamp and inspect, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color; Wherein, speckle is the most obvious when 60-90 ℃ of petroleum ether-ethyl acetate of 19: 1 ratios is developing solvent.

The discrimination test of experimental example 6 Radix Salviae Miltiorrhizaes

The tanshinone IIA reference substance is got in the preparation of control medicinal material solution, adds ethyl acetate and makes the solution that every 1ml contains 2mg, in contrast product solution.

4: 1 ratios are prepared in the selection of developing solvent respectively, 6: 1 ratios, the cyclohexane extraction-ethyl acetate of 8: 1 ratios is developing solvent.

According to the thin layer chromatography test, draw each 10 μ l of blank solution and sample solution, reference substance solution 5 μ l put respectively on same silica gel g thin-layer plate, launch under above-mentioned developing solvent condition respectively, take out, and dry; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show identical kermesinus speckle.Wherein, speckle is the most obvious when the cyclohexane extraction-ethyl acetate of 6: 1 ratios is developing solvent.

Experimental example 7 Rhizoma Polygoni Cuspidati are differentiated

The Rhizoma Polygoni Cuspidati main component is an anthracene derivant, and the emodin of this experiment after with hydrolysis in the Rhizoma Polygoni Cuspidati serves as that index is known in inspection, has set up the TLC inspection knowledge method of Rhizoma Polygoni Cuspidati in the preparation.

The preparation of blank solution is got the blank preparation of this pharmaceutical composition and was used the dosage porphyrize on the 14/1000th, and porphyrize adds methanol 20ml, supersound process 20 minutes filters the filtrate evaporate to dryness, residue adds 2.5mol/L sulfuric acid solution 10ml makes dissolving, puts in the water-bath and heats 30 minutes, immediately cooling, extract twice with the chloroform jolting, each 10ml merges chloroform liquid, evaporate to dryness, residue adds methanol 1ml makes dissolving, as blank solution.

The emodin reference substance is got in the preparation of control medicinal material solution, adds methanol and makes the solution that every 1ml contains 1mg, in contrast product solution.

5: 6: 0.5 ratios are prepared in the selection of developing solvent respectively, 15: 5: 1 ratios, the upper solution of 30～60 ℃ of petroleum ether-Ethyl formate-formic acid of 25: 4: 1.5 ratios.

According to the thin layer chromatography test, draw blank solution and sample solution, each 6 μ l of reference substance solution, put respectively on same silica gel g thin-layer plate, under above-mentioned developing solvent condition, launch respectively, take out, dry; Put in the ammonia smoked after, inspect under the daylight; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color; Wherein, with the upper solution of 30～60 ℃ of petroleum ether-Ethyl formate-formic acid of 15: 5: 1 ratios be developing solvent when launching speckle the most obvious.

Experimental example 8 assays

1. instrument and medicine

The HP1050 of U.S. Hewlett-Packard type high performance liquid chromatograph, quaternary pump, DAD detector, chem workstation; H66025 type ultrasonic cleaner (Ultrasound Electronic Equipment Factory, Wuxi City); Acetonitrile is a chromatographically pure, and mobile phase institute water is a distilled water.

2. the optimization of chromatographic condition

Chromatographic column is YWGC18,10 μ m, and 4.6 * 250mm, the Dalian Chemistry and Physics Institute, post is imitated and is not less than 2600 in its theoretical cam curve of paeoniflorin; Mobile phase is acetonitrile-water (15: 85); Flow velocity is 1ml/min; The detection wavelength is 230nm, and column temperature is a room temperature, and the ratio of acetonitrile in the mobile phase and water is studied, and has optimized chromatographic condition.

Prepare the mobile phase of following ratio respectively: acetonitrile-water (15: 75), acetonitrile-water (15: 80), acetonitrile-water (15: 85), acetonitrile-water (15: 90), acetonitrile-water (15: 95)

By above chromatographic condition sample introduction reference substance, sample, the results are shown in following table 11 respectively:

The optimization of table 11 chromatographic condition

The result as can be known, other chromatograph when mobile phase is acetonitrile-water (15: 75), acetonitrile-water (15: 80) in the sample can not be separated fully with the peoniflorin chromatographic peak, and with acetonitrile-water (15: 90), when acetonitrile-water (15: 95) is mobile phase, holder tail phenomenon appears in the peoniflorin chromatographic peak.Preferred flow is acetonitrile-water (15: 85) mutually

3. sample extraction solvent and optimization for extracting condition

The extraction solvent commonly used of paeoniflorin is water, methanol, ethanol and rare alcohol, and this experiment is adopted methanol aqueous solution can avoid water for the sample extraction solvent and caused the too much phenomenon generation of impurity as extracting solvent.To methanol aqueous solution concentration, solubilizer multiple and ultrasonic treatment time have been done comparative study, have optimized the extraction conditions of sample.

Optimization for extracting condition sample thief powder (lot number 060529), porphyrize (crossing 50 mesh sieves), precision takes by weighing five parts of 0.25g, the accurate 60% methanol 25ml that adds of No.1～No.5 weighs close plug, according to the form below is supersound process (power 250W, frequency 40KHZ) respectively, and extraction finishes, take out, weigh, supply the quantity of solvent that loses, get supernatant with 0.45 μ m filtering with microporous membrane, filtrate is as need testing solution, by above-mentioned chromatographic condition, sample introduction 20 μ l measure, and measurement result is added up, and the results are shown in Table 12.

Table 12 extraction time investigation (content of paeoniflorin mg/g)

The result is as can be known: no significant difference between each group.But the content with ultrasonic 10 minutes and 20 minutes is lower slightly.30 minutes, 40 minutes content basically identicals.

According to above method, be under the condition of 30min in extraction time, investigate the concentration of extracting solution, the results are shown in following table 13

Table 13 extract concentration is investigated (content of paeoniflorin mg/g)

The result is as can be known: no significant difference between each group, to extract with 60% methanol solution, and the paeoniflorin yield is higher.

According to above method, be 30min in extraction time, extracting solution is under the condition of 60% methanol, investigates the solubilizer multiple, the results are shown in Table 14.

The investigation of table 14 solubilizer multiple (content of paeoniflorin mg/g)

The result is as can be known: no significant difference between each group, it is higher to add under 100 times of amount solvent conditions the paeoniflorin yield.

Conclusion: determine that extraction conditions is 60% methanol, 25ml, ultrasonic 30 minutes.

4. stability test

Get need testing solution (060529), respectively at preparation back 0,2,4,6,10,16,24 hour, measure in accordance with the law, the result shows, and is basicly stable in 24 hours, the results are shown in Table 15.

Table 15 stability test

5. precision test

Accurate paeoniflorin reference substance solution (C=0.204mg/ml) the 10 μ l that draw repeat sample introduction 5 times by aforementioned chromatographic condition, record peak area, and trying to achieve RSD is 1.13%, the results are shown in Table 16.

Table 16 Precision test result

6. linear relationship is investigated

Accurate draw the paeoniflorin reference substance solution (2.5,5,7.5,10,12.5,15, the 20 μ l of 2.10mg → 10ml) measure by above-mentioned chromatographic condition sample introduction, measure peak area, the results are shown in Table 9.With peak area the amount of reference substance is carried out linear regression (comprising initial point), trying to achieve regression equation is Y=-39.6975+1574.5933X, and r=0.9999 shows that paeoniflorin is linear in 0.525～4.200 μ g scope, and testing result sees Table 17.

Table 17 paeoniflorin standard curve determination result

7. repeatability test

Get same batch sample (lot number 060529) porphyrize, precision takes by weighing 0.25g, and totally five parts, measure by the text method, try to achieve relative standard deviation.The results are shown in Table 18.

Table 18 reproducible test results

8. blank assay

Do not contain group's medicine of the Radix Paeoniae Alba in the autogamy of prescription taste of Chinese medicine ratio, make blank solution by the text method.Blank solution does not show obvious chromatographic peak at the place of identical retention time with paeoniflorin as a result, thinks noiseless.

9. recovery test

Adopt the application of sample absorption method, precision takes by weighing totally five parts of same lot number sample (lot number 060529) (content the is 6.9mg/g) 0.15g of known content, according to the form below adds paeoniflorin reference substance solution (concentration 0.234mg/ml) 5ml respectively, the accurate again 20ml60% methanol that adds, close plug, extract and measure by sample extraction and condition determination,, the results are shown in Table 19 with the following formula calculate recovery rate.

Table 19 recovery test result

10. sample determination result

Record method by text ten batch samples are measured, the results are shown in Table 20.

Table 20 sample determination result

Lot number content of paeoniflorin (mg/g)

050524 6.20

050529 6.90

050618 5.84

060301 5.98

060306 5.17

060312 5.41

060910 5.22

060914 5.31

070312 5.30

070316 5.33

According to above data, it is that the every gram of this product contains the Radix Paeoniae Alba by paeoniflorin (C that content limit is fixed tentatively ₂₃H ₂₈O ₁₁) meter, should be no less than 5.0mg.

Experimental example 9. content assaying methods of the present invention are the employment and suitability test (E ﹠ ST) in the content of paeoniflorin mensuration in qizhi weitong granules

Qizhi weitong granules mainly is made up of Radix Bupleuri, Rhizoma Corydalis, Fructus Aurantii, Rhizoma Cyperi, the Radix Paeoniae Alba, Radix Glycyrrhizae Preparata Six-element Chinese medicine.Peoniflorin is the index components of control this product quality, and pilot production is measured content of paeoniflorin in the qizhi weitong granules with this method.

(1) repeatability test

Get same batch sample (lot number 060312) porphyrize, precision takes by weighing 0.25g, and totally five parts, measure by the text method, try to achieve relative standard deviation.The results are shown in Table 21.

Table 21 reproducible test results

(2) recovery test

Adopt the application of sample absorption method, precision takes by weighing totally five parts of same lot number sample (lot number 060312) (content the is 1.5mg/g) 0.85g of known content, according to the form below adds paeoniflorin reference substance solution (concentration 0.234mg/ml) 5ml respectively, the accurate again 20ml60% methanol that adds, close plug, extract and measure by sample extraction and condition determination,, the results are shown in Table 22 with the following formula calculate recovery rate.

Table 22 recovery test result

According to above presentation of results, this content assaying method is used for measuring qizhi weitong granules content of paeoniflorin poor reproducibility, the response rate is low.Though illustrate in the multiple Chinese patent medicine and contain peoniflorin, its assay method is often different.

Following embodiment all can realize the described effect of above-mentioned experimental example

Embodiment 1

Poria 960g, Herba Artemisiae Scopariae 960g, Rhizoma Atractylodis Macrocephalae 960g, Radix Curcumae 320g, Radix Angelicae Sinensis 480g, succinum 50-150g, the Radix Paeoniae Alba (stir-fry) 960g, Herba Scutellariae Barbatae 960g, Fructus Amomi 192g, Rhizoma Polygoni Cuspidati 480g, Radix Salviae Miltiorrhizae 960g, Herba Lycopi 480g, Radix Bupleuri 320g, Herba Pogostemonis 320g, Herba Eupatorii 480g, Flos Carthami 320g

This pharmaceutical composition adds conventional adjuvant, makes capsule by common process.

Embodiment 2

Poria 520g, Herba Artemisiae Scopariae 1480g, Rhizoma Atractylodis Macrocephalae 520g, Radix Curcumae 430g, Radix Angelicae Sinensis 270g, succinum 130g, the Radix Paeoniae Alba (stir-fry) 520g, Herba Scutellariae Barbatae 1480g, Fructus Amomi 120g, Rhizoma Polygoni Cuspidati 730g, Radix Salviae Miltiorrhizae 520g, Herba Lycopi 730g, Radix Bupleuri 170g, Herba Pogostemonis 430g, Herba Eupatorii 270g, Flos Carthami 430g

Radix Angelicae Sinensis, Radix Curcumae, Herba Pogostemonis, Herba Eupatorii, Fructus Amomi, Herba Lycopi extracted volatile oil 8 hours, and the aqueous solution after distillation device is in addition collected; Volatile oil is with 40 ℃ of inclusions of beta-schardinger dextrin-saturated water solution method of 4 times of weight portions, 40 ℃ of dryings, the volatile oil beta-cyclodextrin inclusion complex, standby; Radix Salviae Miltiorrhizae is with 85% alcohol reflux of 6 times of weight portions 2 times, and each 1 hour, merge extractive liquid, filtered, and filtrate recycling ethanol is not to there being the alcohol flavor, Radix Salviae Miltiorrhizae extract, standby; Medicinal residues and all the other flavour of a drug decoct with water 2 times, add 8 times of water gagings at every turn and decoct 1.5 hours, and collecting decoction filters; Aqueous solution after filtrate and the above-mentioned distillation merges, be concentrated into 60～65 ℃ of relative densities and be 1.02～1.04 thick paste I, add ethanol and make and contain alcohol amount and reach 70%, left standstill 24 hours, filter, filtrate recycling ethanol is to an amount of, merge with above-mentioned Radix Salviae Miltiorrhizae extract, and the thick paste II that to be concentrated into 60～65 ℃ of relative densities be 1.10-1.15, add 80% simple syrup 960 parts by volume, the volatile oil beta-cyclodextrin inclusion complex is ground into fine powder, adds in the extractum; Add sodium benzoate 9.6g again, stir evenly, add water to 3000 parts by volume, filter, fill, sterilization promptly gets mixture and used dosage on the 30th.

Embodiment 3

Poria 1480g, Herba Artemisiae Scopariae 520g, Rhizoma Atractylodis Macrocephalae 1480g, Radix Curcumae 170g, Radix Angelicae Sinensis 730g, succinum 70g, the Radix Paeoniae Alba (stir-fry) 1480g, Herba Scutellariae Barbatae 520g, Fructus Amomi 280g, Rhizoma Polygoni Cuspidati 270g, Radix Salviae Miltiorrhizae 1480g, Herba Lycopi 270g, Radix Bupleuri 430g, Herba Pogostemonis 170g, Herba Eupatorii 730g, Flos Carthami 170g

Radix Angelicae Sinensis, Radix Curcumae, Herba Pogostemonis, Herba Eupatorii, Fructus Amomi, Herba Lycopi extracted volatile oil 8 hours, and the aqueous solution after distillation device is in addition collected; Volatile oil is with 40 ℃ of inclusions of beta-schardinger dextrin-saturated water solution method of 4 times of weight portions, 40 ℃ of dryings, the volatile oil beta-cyclodextrin inclusion complex, standby; Radix Salviae Miltiorrhizae is with 85% alcohol reflux of 6 times of weight portions 2 times, and each 1 hour, merge extractive liquid, filtered, and filtrate recycling ethanol is not to there being the alcohol flavor, Radix Salviae Miltiorrhizae extract, standby; Medicinal residues and all the other flavour of a drug decoct with water 2 times, add 8 times of water gagings at every turn and decoct 1.5 hours, and collecting decoction filters; Aqueous solution after filtrate and the above-mentioned distillation merges, be concentrated into 60～65 ℃ of relative densities and be 1.02～1.04 thick paste I, add ethanol and make and contain alcohol amount and reach 70%, left standstill 24 hours, filter, filtrate recycling ethanol is to an amount of, merge, and the thick paste II that to be concentrated into 60～65 ℃ of relative densities be 1.25-1.30 drying under reduced pressure under 0.08Mpa, the 65 ℃ of conditions with above-mentioned Radix Salviae Miltiorrhizae extract, add the volatile oil beta-cyclodextrin inclusion complex, be ground into fine powder; 70% ethanol system soft material, extrusion granulator promptly gets and used dosage on the 32nd.

Embodiment 4

Poria 960g, Herba Artemisiae Scopariae 960g, Radix Curcumae 320g, Radix Angelicae Sinensis 480g, succinum 96g, the Radix Paeoniae Alba (stir-fry) 960g, Herba Scutellariae Barbatae 960g, Fructus Amomi 192g, Rhizoma Polygoni Cuspidati 480g, Radix Salviae Miltiorrhizae 960g, Herba Lycopi 480g, Radix Bupleuri 320g, Herba Pogostemonis 320g, Herba Eupatorii 480g, Flos Carthami 320g, Radix Isatidis 960g, Rhizoma Dryopteris Crassirhizomatis 480g, Herba Hedyotidis Diffusae 960g, Rhizoma Paridis 480g

This pharmaceutical composition adds conventional adjuvant, makes mixture by common process.

Embodiment 5

Poria 520g, Herba Artemisiae Scopariae 1480g, Radix Curcumae 170g, Radix Angelicae Sinensis 730g, succinum 70g, the Radix Paeoniae Alba (stir-fry) 1480g, Herba Scutellariae Barbatae 520g, Fructus Amomi 280g, Rhizoma Polygoni Cuspidati 270g, Radix Salviae Miltiorrhizae 1480g, Herba Lycopi 270g, Radix Bupleuri 430g, Herba Pogostemonis 170g, Herba Eupatorii 730g, Flos Carthami 170g, Radix Isatidis 1480g, Rhizoma Dryopteris Crassirhizomatis 270g, Herba Hedyotidis Diffusae 1480g, Rhizoma Paridis 270g

This pharmaceutical composition adds conventional adjuvant, makes tablet by common process.

Embodiment 6

Poria 1480g, Herba Artemisiae Scopariae 520g, Radix Curcumae 430g, Radix Angelicae Sinensis 270g, succinum 130g, the Radix Paeoniae Alba (stir-fry) 520g, Herba Scutellariae Barbatae 1480g, Fructus Amomi 120g, Rhizoma Polygoni Cuspidati 730g, Radix Salviae Miltiorrhizae 520g, Herba Lycopi 730g, Radix Bupleuri 170g, Herba Pogostemonis 430g, Herba Eupatorii 270g, Flos Carthami 430g, Radix Isatidis 520g, Rhizoma Dryopteris Crassirhizomatis 730g, Herba Hedyotidis Diffusae 520g, Rhizoma Paridis 730g

Radix Angelicae Sinensis, Radix Curcumae, Herba Pogostemonis, Herba Eupatorii, Fructus Amomi, Herba Lycopi extracted volatile oil 8 hours, and the aqueous solution after distillation device is in addition collected; Volatile oil is with 40 ℃ of inclusions of beta-schardinger dextrin-saturated water solution method of 4 times of weight portions, 40 ℃ of dryings, the volatile oil beta-cyclodextrin inclusion complex, standby; Radix Salviae Miltiorrhizae is with 85% alcohol reflux of 6 times of weight portions 2 times, and each 1 hour, merge extractive liquid, filtered, and filtrate recycling ethanol is not to there being the alcohol flavor, Radix Salviae Miltiorrhizae extract, standby; 12 flavors such as medicinal residues and all the other Herba Artemisiae Scopariaes decoct with water 2 times, add 8 times of water gagings at every turn and decoct 1.5 hours, and collecting decoction filters; Aqueous solution after filtrate and the above-mentioned distillation merges, be concentrated into 60～65 ℃ of relative densities and be 1.02～1.04 thick paste I, add ethanol and make and contain alcohol amount and reach 70%, left standstill 24 hours, filter, filtrate recycling ethanol is to an amount of, merge with above-mentioned Radix Salviae Miltiorrhizae extract, and the thick paste II that to be concentrated into 60～65 ℃ of relative densities be 1.10-1.15, add 80% simple syrup 1000 parts by volume, the volatile oil beta-cyclodextrin inclusion complex is ground into fine powder, adds in the extractum; Add sodium benzoate 9.6g again, stir evenly, add water to 3600 parts by volume, filter, fill, sterilization promptly gets mixture.

The method of quality control of embodiment 7 preparations of the present invention

Differentiate: A, get the content of 0.014 times of daily dosage of this medicament composition granule agent of embodiment 3 preparations, add water 20ml and make dissolving, with ethyl acetate extraction 2 times, 30ml for the first time, 20ml for the second time, combined ethyl acetate liquid, water bath method, residue adds ethyl acetate 1ml makes dissolving, as need testing solution; Other gets Herba Artemisiae Scopariae control medicinal material 1g, adds water 50ml, decocts 1 hour, gets supernatant, be concentrated into 20ml, use ethyl acetate extraction 2 times, for the first time 30ml, 20ml for the second time, combined ethyl acetate liquid, water bath method, residue add ethyl acetate 1ml makes dissolving, in contrast medical material solution; Draw need testing solution 1 μ l, control medicinal material solution 5 μ l, putting respectively on same silica gel g thin-layer plate, is developing solvent with 60-90 ℃ of petroleum ether-ethyl acetate of 1: 1 ratio, launches, take out, dry, put under the 365nm ultra-violet lamp and observe, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color;

B, get the content of 0.0057 times of daily dosage of this medicament composition granule agent of embodiment 3 preparation, porphyrize adds 50% ethanol 15ml, and supersound process 20 minutes filters, and filtrate is as need testing solution; Other gets the peoniflorin reference substance, adds ethanol and makes the solution that every 1ml contains 1mg, in contrast product solution; Test according to thin layer chromatography, draw each 10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with 40: 5: 10: the chloroform-ethyl acetate of 0.2 ratio-methanol-formic acid is developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, 105 ℃ to be heated to speckle colour developing clear, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;

C, get the content of 0.029 times of daily dosage of this medicament composition granule agent of embodiment 3 preparation, porphyrize, the 30ml that adds diethyl ether, reflux, extract, 30 minutes filters, and filtrate volatilizes, and residue adds ethyl acetate 0.5ml makes dissolving, as need testing solution; Other gets Radix Angelicae Sinensis control medicinal material 1g, the 10ml that adds diethyl ether, and supersound process 10 minutes filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, in contrast medical material solution; Test according to thin layer chromatography, draw each 5 μ l of above-mentioned two kinds of solution, putting respectively on same silica gel g thin-layer plate, is developing solvent with 60-90 ℃ of petroleum ether-ethyl acetate of 9.5: 0.5 ratios, launches, take out, dry, put under the 365nm ultra-violet lamp and inspect, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color;

D, get the tanshinone IIA reference substance, add ethyl acetate and make the solution that every 1ml contains 2mg, in contrast product solution; According to thin layer chromatography test, draw and differentiate C item need testing solution 10 μ l down, reference substance solution 5 μ l put respectively on same silica gel g thin-layer plate, are developing solvent with the cyclohexane extraction-ethyl acetate of 6: 1 ratios, launch, and taking-up is dried; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show identical kermesinus speckle.

The method of quality control of embodiment 8 preparations of the present invention

Assay: shine high effective liquid chromatography for measuring: chromatographic condition and system suitability test are filler with octadecylsilane chemically bonded silica; The acetonitrile-water of 15: 85 ratios is a mobile phase, and the detection wavelength is 230nm, and number of theoretical plate calculates by the peoniflorin peak should be not less than 2600;

The preparation of reference substance solution: precision takes by weighing peoniflorin reference substance 5mg, puts in the 100ml measuring bottle, adds dissolve with methanol and is diluted to scale, shakes up, and promptly gets the reference substance solution that contains peoniflorin 50 μ g among every 1ml;

The need testing solution preparation: get the content of 0.00072 times of daily dosage of this pharmaceutical composition mixture of embodiment 2 preparations under the content uniformity item, the accurate title, decide, and puts in the tool plug conical flask, precision adds 60% methanol 25ml, and close plug claims to decide weight, power 250W, frequency 40KHZ supersound process 30 minutes is taken out, put coldly, claim again to decide weight, supply minimizing weight with 60% methanol, shake up, get supernatant, filter, promptly;

Algoscopy: accurate respectively reference substance solution and each 10 μ l of need testing solution of drawing, inject chromatograph of liquid, measure, promptly;

This pharmaceutical composition mixture contains the Radix Paeoniae Alba with peoniflorin C with dosage every day ₂₃H ₂₈O ₁₁Meter should be less than 60mg.

Embodiment 9

Differentiate: A, get the content of 0.014 times of daily dosage of this pharmaceutical composition mixture of embodiment 2 preparations, add water 20ml and make dissolving, with ethyl acetate extraction 2 times, 30ml for the first time, 20ml for the second time, combined ethyl acetate liquid, water bath method, residue adds ethyl acetate 1ml makes dissolving, as need testing solution; Other gets Herba Artemisiae Scopariae control medicinal material 1g, adds water 50ml, decocts 1 hour, gets supernatant, be concentrated into 20ml, use ethyl acetate extraction 2 times, for the first time 30ml, 20ml for the second time, combined ethyl acetate liquid, water bath method, residue add ethyl acetate 1ml makes dissolving, in contrast medical material solution; Draw need testing solution 1 μ l, control medicinal material solution 5 μ l, putting respectively on same silica gel g thin-layer plate, is developing solvent with 60-90 ℃ of petroleum ether-ethyl acetate of 1: 1 ratio, launches, take out, dry, put under the 365nm ultra-violet lamp and observe, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color;

B, get the content of 0.0057 times of daily dosage of this pharmaceutical composition mixture of embodiment 2 preparation, porphyrize adds 50% ethanol 15ml, and supersound process 20 minutes filters, and filtrate is as need testing solution; Other gets the peoniflorin reference substance, adds ethanol and makes the solution that every 1ml contains 1mg, in contrast product solution; Test according to thin layer chromatography, draw each 10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with 40: 5: 10: the chloroform-ethyl acetate of 0.2 ratio-methanol-formic acid is developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, 105 ℃ to be heated to speckle colour developing clear, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;

C, get the content of 0.029 times of daily dosage of this pharmaceutical composition mixture of embodiment 2 preparation, porphyrize, the 30ml that adds diethyl ether, reflux, extract, 30 minutes filters, and filtrate volatilizes, and residue adds ethyl acetate 0.5ml makes dissolving, as need testing solution; Other gets Radix Angelicae Sinensis control medicinal material 1g, the 10ml that adds diethyl ether, and supersound process 10 minutes filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, in contrast medical material solution; Test according to thin layer chromatography, draw each 5 μ l of above-mentioned two kinds of solution, putting respectively on same silica gel g thin-layer plate, is developing solvent with 60-90 ℃ of petroleum ether-ethyl acetate of 9.5: 0.5 ratios, launches, take out, dry, put under the 365nm ultra-violet lamp and inspect, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color;

The need testing solution preparation: get 0.00072 times of daily dosage content of this pharmaceutical composition mixture of embodiment 12 preparations under the content uniformity item, the accurate title, decide, and puts in the tool plug conical flask, precision adds 60% methanol 25ml, and close plug claims to decide weight, power 250W, frequency 40KHZ supersound process 30 minutes is taken out, put coldly, claim again to decide weight, supply minimizing weight with 60% methanol, shake up, get supernatant, filter, promptly;

The agent of this medicament composition capsule contains the Radix Paeoniae Alba with peoniflorin C with dosage every day ₂₃H ₂₈O ₁₁Meter should be less than 60mg.

Embodiment 10

Herba Artemisiae Scopariae 960g, Radix Isatidis 960g, Rhizoma Dryopteris Crassirhizomatis 480g, Poria 960g, Radix Curcumae 320g, Radix Angelicae Sinensis 480g, Flos Carthami 320g, succinum 96g, the Radix Paeoniae Alba (stir-fry) 960g, Herba Hedyotidis Diffusae 960g, Herba Scutellariae Barbatae 960g, Herba Pogostemonis 320g, Herba Eupatorii 480g, Fructus Amomi 192g, Rhizoma Polygoni Cuspidati 480g, Radix Salviae Miltiorrhizae 960g, Herba Lycopi 480g, Radix Bupleuri 320g, Rhizoma Paridis 480g

More than 19 flavors, Radix Angelicae Sinensis, Radix Curcumae, Herba Pogostemonis, Herba Eupatorii, Fructus Amomi, Herba Lycopi extracted volatile oil 8 hours, the aqueous solution after the distillation in addition device is collected; Volatile oil is with 40 ℃ of inclusions of beta-schardinger dextrin-saturated water solution method of 4 times of weight portions, 40 ℃ of dryings, the volatile oil beta-cyclodextrin inclusion complex, standby; Radix Salviae Miltiorrhizae is with 85% alcohol reflux of 6 times of weight portions 2 times, and each 1 hour, merge extractive liquid, filtered, and filtrate recycling ethanol is not to there being the alcohol flavor, Radix Salviae Miltiorrhizae extract, standby; 12 flavors such as medicinal residues and all the other Herba Artemisiae Scopariaes decoct with water 2 times, add 8 times of water gagings at every turn and decoct 1.5 hours, and collecting decoction filters; Aqueous solution after filtrate and the above-mentioned distillation merges, be concentrated into 60～65 ℃ of relative densities and be 1.02～1.04 thick paste I, adding ethanol makes and contains alcohol amount and reach 70%, left standstill 24 hours, and filtered, filtrate recycling ethanol is to an amount of, merge with above-mentioned Radix Salviae Miltiorrhizae extract, and the thick paste II that to be concentrated into 60～65 ℃ of relative densities be 1.10-1.15, adding 80% simple syrup 960 parts by volume, the volatile oil beta-cyclodextrin inclusion complex is ground into fine powder; Add sodium benzoate 9.6g again, add water to 3200 parts by volume, filter, fill, sterilization promptly gets mixture.

The method of quality control of embodiment 11 preparations of the present invention

The need testing solution preparation: get this medicament composition capsule agent of embodiment 1 preparation under the content uniformity item, porphyrize is got 0.00072 times daily dosage, the accurate title, decide, and puts in the tool plug conical flask, and precision adds 60% methanol 25ml, close plug claims to decide weight, power 250W, frequency 40KHZ supersound process 30 minutes is taken out, and puts cold, claim again to decide weight, supply minimizing weight, shake up with 60% methanol, get supernatant, filter, promptly;

The method of quality control of embodiment 12 preparations of the present invention

Differentiate:

A, get the daily dosage content porphyrize of 0.0057 times in this pharmaceutical composition mixture of embodiment 6 preparation, add 50% ethanol 15ml, supersound process 20 minutes filters, and filtrate is as need testing solution; Other gets the peoniflorin reference substance, adds ethanol and makes the solution that every 1ml contains 1mg, in contrast product solution; Test according to thin layer chromatography, draw each 10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with 40: 5: 10: the chloroform-ethyl acetate of 0.2 ratio-methanol-formic acid is developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, 105 ℃ to be heated to speckle colour developing clear, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;

B, get the tanshinone IIA reference substance, add ethyl acetate and make the solution that every 1ml contains 2mg, in contrast product solution; According to thin layer chromatography test, get the daily dosage content of 0.029 times in this pharmaceutical composition mixture of embodiment 6 preparations, porphyrize, the 30ml that adds diethyl ether, reflux, extract, 30 minutes filters, filtrate volatilizes, and residue adds ethyl acetate 0.5ml makes dissolving, as need testing solution, draw need testing solution 10 μ l, reference substance solution 5 μ l put respectively on same silica gel g thin-layer plate, cyclohexane extraction-ethyl acetate with 6: 1 ratios is developing solvent, launch, take out, dry; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show identical kermesinus speckle;

C, get the daily dosage content of 0.014 times in this pharmaceutical composition mixture of embodiment 6 preparation, porphyrize adds methanol 20ml, ultrasonic place 20 minutes filters the filtrate evaporate to dryness, residue adds 2.5mol/L sulfuric acid solution 10ml makes dissolving, puts in the water-bath and heats 30 minutes, immediately cooling, extract twice with the chloroform jolting, each 10ml merges chloroform liquid, evaporate to dryness, residue adds methanol 1ml makes dissolving, as need testing solution; Other gets the emodin reference substance, adds methanol and makes the solution that every 1ml contains 1mg, in contrast product solution; According to the 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, upper solution with 30～60 ℃ of petroleum ether-Ethyl formate-formic acid of 15: 5: 1 ratios is developing solvent, launches, and takes out, dry, put in the ammonia smoked after, inspect under the daylight; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;

Need testing solution preparation: get the daily dosage of 0.00072 times in this pharmaceutical composition mixture of embodiment 6 preparations the content uniformity item under, the accurate title, decide, and puts in the tool plug conical flask, precision adds 60% methanol 25ml, and close plug claims to decide weight, power 250W, frequency 40KHZ supersound process 30 minutes is taken out, put coldly, claim again to decide weight, supply minimizing weight with 60% methanol, shake up, get supernatant, filter, promptly;

Embodiment 13

Herba Artemisiae Scopariae 960g Radix Isatidis 960g Rhizoma Dryopteris Crassirhizomatis 480g

Poria 960g Radix Curcumae 320g Radix Angelicae Sinensis 480g

The Flos Carthami 320g succinum 96g Radix Paeoniae Alba (stir-fry) 960g

Herba Hedyotidis Diffusae 960g Herba Scutellariae Barbatae 960g Herba Pogostemonis 320g

Herba Eupatorii 480g Fructus Amomi 192g Rhizoma Polygoni Cuspidati 480g

Radix Salviae Miltiorrhizae 960g Herba Lycopi 480g Radix Bupleuri 320g

Rhizoma Paridis 480g

Method for making: above 19 flavors, Radix Angelicae Sinensis, Radix Curcumae, Herba Pogostemonis, Herba Eupatorii, Fructus Amomi, Herba Lycopi extracted volatile oil 8 hours, and the aqueous solution after distillation device is in addition collected, volatile oil 40 ℃ of inclusions of beta-schardinger dextrin-saturated water solution method of 40g, 40 ℃ of dryings get the volatile oil beta-cyclodextrin inclusion complex, and are standby; Radix Salviae Miltiorrhizae is with 85% ethanol 6700ml reflux, extract, secondary, and each 1 hour, merge extractive liquid, filtered, and filtrate recycling ethanol is not to there being the alcohol flavor, Radix Salviae Miltiorrhizae extract, standby; 12 flavors such as medicinal residues and all the other Herba Artemisiae Scopariaes decoct with water secondary, add 8 times of water gagings decocted 1.5 hours at every turn, collecting decoction, filter, the aqueous solution after filtrate and the above-mentioned distillation merges, and being concentrated into relative density is 1.02～1.04 (60～65 ℃), adding ethanol makes and contains alcohol amount and reach 70%, left standstill 24 hours, and filtered, filtrate recycling ethanol is to an amount of, merge with above-mentioned Radix Salviae Miltiorrhizae extract, and be concentrated into the thick paste that relative density is 1.10-1.15 (60～65 ℃), add the volatile oil beta-cyclodextrin inclusion complex and be ground into fine powder, and appropriate amount of auxiliary materials, add water, fill, sterilization, promptly.

[discriminating]

(1) get this product 5g, add water 20ml and make dissolving, with ethyl acetate extraction 2 times, 30ml for the first time, 20ml for the second time, combined ethyl acetate liquid, water bath method, residue add ethyl acetate 1ml makes dissolving, as need testing solution; Other gets Herba Artemisiae Scopariae control medicinal material 1g, adds water 50ml, decocts 1 hour, gets supernatant, is concentrated into 20ml, shines medical material solution with ethyl acetate in pairs with legal system; Draw need testing solution 1 μ l, control medicinal material solution 5 μ l put respectively on same silica gel g thin-layer plate, are developing solvent with petroleum ether (60-90 ℃)-ethyl acetate (1: 1), launch, and take out, and dry, and put under the ultra-violet lamp (365nm) and observe; In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(2) get this product 2g porphyrize, add 50% ethanol 15ml, supersound process 20 minutes filters, and filtrate is as need testing solution; Other gets the peoniflorin reference substance, adds ethanol and makes the solution that every 1ml contains 1mg, in contrast product solution; According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) 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-ethyl acetate-methanol-formic acid (40: 5: 10: 0.2) be developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and 105 ℃ to be heated to the speckle colour developing clear; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.

(3) get this product 10g, porphyrize, the 30ml that adds diethyl ether, reflux, extract, 30 minutes filters, and filtrate volatilizes, and residue adds ethyl acetate 0.5ml makes dissolving, as need testing solution; Other gets Radix Angelicae Sinensis control medicinal material 1g, the 10ml that adds diethyl ether, and supersound process 10 minutes filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, in contrast medical material solution; According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 5 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 (9.5: 0.5) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect; In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(4) get the tanshinone IIA reference substance, add ethyl acetate and make the solution that every 1ml contains 2mg, in contrast product solution; According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw the need testing solution 10 μ l under the item of [discriminatings] (3), reference substance solution 5 μ l, putting respectively on same silica gel g thin-layer plate, is developing solvent with cyclohexane extraction-ethyl acetate (6: 1), launches, take out, dry; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show identical kermesinus speckle.

(5) get this product 5g, porphyrize adds methanol 20ml, supersound process 20 minutes filters the filtrate evaporate to dryness, residue adds 2.5mol/L sulfuric acid solution 10ml makes dissolving, puts in the water-bath and heats 30 minutes, immediately cooling, extract twice with the chloroform jolting, each 10ml merges chloroform liquid, evaporate to dryness, residue adds methanol 1ml makes dissolving, as need testing solution; Other gets the emodin reference substance, adds methanol and makes the solution that every 1ml contains 1mg, in contrast product solution; According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia version in 2005 B) test, draw each 6 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, upper solution with petroleum ether (30～60 ℃)-Ethyl formate-formic acid (15: 5: 1) is developing solvent, launch, take out, dry, put in the ammonia smoked after, inspect under the daylight; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.

Assay: according to high effective liquid chromatography for measuring

Chromatographic condition and system suitability test are filler with octadecylsilane chemically bonded silica; Acetonitrile-water (15: 85) is a mobile phase, and the detection wavelength is 230nm; Number of theoretical plate calculates by the peoniflorin peak should be not less than 2600;

The preparation precision of reference substance solution takes by weighing peoniflorin reference substance 5mg, puts in the 100ml measuring bottle, adds dissolve with methanol and is diluted to scale, shakes up, and promptly gets (containing peoniflorin 50 μ g among every 1ml);

This product under the content uniformity item is got in the need testing solution preparation, and porphyrize is got 0.25g, and accurate the title decides, and puts in the tool plug conical flask, and precision adds 60% methanol 25ml, and close plug claims to decide weight, supersound process (power 250W, frequency 40KH _Z) 30 minutes, take out, put coldly, claim again to decide weight, supply minimizing weight with 60% methanol; Shake up, get supernatant, filter, 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 the Radix Paeoniae Alba with peoniflorin (C ₂₃H ₂₈O ₁₁) meter, must not be less than 0.6mg.

Claims

1. detection method for the treatment of the viral hepatitis pharmaceutical composition is characterized in that the content assaying method in this method is:

Chromatographic condition and system suitability test: with octadecylsilane chemically bonded silica is filler; 5-25: the acetonitrile-water of 70-95 ratio is a mobile phase, and the detection wavelength is 230nm, and number of theoretical plate calculates by the peoniflorin peak should be not less than 2600;

The need testing solution preparation: get this drug combination preparation under the content uniformity item, porphyrize is got 6/10000-1/1000 day and is used dosage, the accurate title, decide, and puts in the tool plug conical flask, and precision adds 50-70% methanol 15-35ml, close plug claims to decide weight, power 250W, frequency 40KHZ supersound process 20-40 minute is taken out, and puts cold, claim again to decide weight, supply minimizing weight, shake up with 50-70% methanol, get supernatant, filter, promptly; Algoscopy: accurate respectively reference substance solution and each 5-15 μ l of need testing solution of drawing, inject chromatograph of liquid, measure, promptly; This drug combination preparation contains the Radix Paeoniae Alba with peoniflorin C with dosage every day ₂₃H ₂₈O ₁₁Meter should be less than 50-70mg;

The crude drug of described pharmaceutical composition consists of:

2. the detection method of pharmaceutical composition as claimed in claim 1 is characterized in that the assay in this method is:

Chromatographic condition and system suitability test: with octadecylsilane chemically bonded silica is filler; The acetonitrile-water of 15: 85 ratios is a mobile phase, and the detection wavelength is 230nm, and number of theoretical plate calculates by the peoniflorin peak should be not less than 2600;

The need testing solution preparation: get this drug combination preparation under the content uniformity item, porphyrize is got and was used dosage on the 7/10000th, the accurate title, decide, and puts in the tool plug conical flask, and precision adds 60% methanol 25ml, close plug claims to decide weight, power 250W, frequency 40KHZ supersound process 30 minutes is taken out, and puts cold, claim again to decide weight, supply minimizing weight, shake up with 60% methanol, get supernatant, filter, promptly;

Algoscopy: accurate respectively reference substance solution and each 10 μ l of need testing solution of drawing, inject chromatograph of liquid, measure, promptly; This drug combination preparation contains the Radix Paeoniae Alba with peoniflorin C with dosage every day ₂₃H ₂₈O ₁₁Meter should be less than 60mg;

The crude drug of described pharmaceutical composition consists of:

3. the detection method of pharmaceutical composition as claimed in claim 1 or 2 is characterized in that this method comprises one or more of following discrimination method:

D, get the tanshinone IIA reference substance, add ethyl acetate and make the solution that every 1ml contains 1-3mg, in contrast product solution; According to thin layer chromatography test, draw and differentiate C item need testing solution 5-15 μ l down, reference substance solution 4-6 μ l puts respectively on same silica gel g thin-layer plate, and with 4-8: the cyclohexane extraction-ethyl acetate of 1 ratio is developing solvent, launches, and taking-up is dried; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show identical kermesinus speckle;

E, get this drug combination preparation 1/100-3/100 day and use dosage, porphyrize adds methanol 10-30ml, supersound process 10-30 minute, filter the filtrate evaporate to dryness, residue adds 1.5-3.5mol/L sulfuric acid solution 5-15ml makes dissolving, puts in the water-bath and heats 20-40 minute, immediately cooling, extract twice with the chloroform jolting, each 5-15ml merges chloroform liquid, evaporate to dryness, residue adds methanol 0.5-1.5ml makes dissolving, as need testing solution; Other gets the emodin reference substance, adds methanol and makes the solution that every 1ml contains 0.5-1.5mg, in contrast product solution; Test according to thin layer chromatography, draw each 4-8 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with 5-25: 4-6: the upper solution of 30～60 ℃ of petroleum ether-Ethyl formate-formic acid of 0.5-1.5 ratio is developing solvent, launch, take out, dry, put in the ammonia smoked after, inspect under the daylight; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.

4. the detection of pharmaceutical composition as claimed in claim 3 is characterized in that this method comprises one or more of following discrimination method:

A, get this drug combination preparation and used dosage on 14/1000th, add water 20ml and make dissolving, with ethyl acetate extraction 2 times, 30ml for the first time, 20ml for the second time, combined ethyl acetate liquid, water bath method, residue add ethyl acetate 1ml makes dissolving, as need testing solution; Other gets Herba Artemisiae Scopariae control medicinal material 1g, adds water 50ml, decocts 1 hour, gets supernatant, be concentrated into 20ml, use ethyl acetate extraction 2 times, for the first time 30ml, 20ml for the second time, combined ethyl acetate liquid, water bath method, residue add ethyl acetate 1ml makes dissolving, in contrast medical material solution; Draw need testing solution 1 μ l, control medicinal material solution 5 μ l, putting respectively on same silica gel g thin-layer plate, is developing solvent with 60-90 ℃ of petroleum ether-ethyl acetate of 1: 1 ratio, launches, take out, dry, put under the 365nm ultra-violet lamp and observe, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color;

B, get this drug combination preparation and used the dosage porphyrize on 6/1000th, add 50% ethanol 15ml, supersound process 20 minutes filters, and filtrate is as need testing solution; Other gets the peoniflorin reference substance, adds ethanol and makes the solution that every 1ml contains 1mg, in contrast product solution; Test according to thin layer chromatography, draw each 10 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with 40: 5: 10: the chloroform-ethyl acetate of 0.2 ratio-methanol-formic acid is developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, 105 ℃ to be heated to speckle colour developing clear, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;

C, get this drug combination preparation and used dosage on 3/100th, porphyrize, the 30ml that adds diethyl ether, reflux, extract, 30 minutes filters, and filtrate volatilizes, and residue adds ethyl acetate 0.5ml makes dissolving, as need testing solution; Other gets Radix Angelicae Sinensis control medicinal material 1g, the 10ml that adds diethyl ether, and supersound process 10 minutes filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, in contrast medical material solution; Test according to thin layer chromatography, draw each 5 μ l of above-mentioned two kinds of solution, putting respectively on same silica gel g thin-layer plate, is developing solvent with 60-90 ℃ of petroleum ether-ethyl acetate of 9.5: 0.5 ratios, launches, take out, dry, put under the 365nm ultra-violet lamp and inspect, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color;

D, get the tanshinone IIA reference substance, add ethyl acetate and make the solution that every 1ml contains 2mg, in contrast product solution; According to thin layer chromatography test, draw and differentiate C item need testing solution 10 μ l down, reference substance solution 5 μ l put respectively on same silica gel g thin-layer plate, are developing solvent with the cyclohexane extraction-ethyl acetate of 6: 1 ratios, launch, and taking-up is dried; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show identical kermesinus speckle;

E, get this drug combination preparation and used dosage on 14/1000th, porphyrize adds methanol 20ml, supersound process 20 minutes filters the filtrate evaporate to dryness, residue adds 2.5mol/L sulfuric acid solution 10ml makes dissolving, puts in the water-bath and heats 30 minutes, immediately cooling, extract twice with the chloroform jolting, each 10ml merges chloroform liquid, evaporate to dryness, residue adds methanol 1ml makes dissolving, as need testing solution; Other gets the emodin reference substance, adds methanol and makes the solution that every 1ml contains 1mg, in contrast product solution; According to the 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, upper solution with 30～60 ℃ of petroleum ether-Ethyl formate-formic acid of 15: 5: 1 ratios is developing solvent, launches, and takes out, dry, put in the ammonia smoked after, inspect under the daylight; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.

5. the detection method of pharmaceutical composition as claimed in claim 1, it is characterized in that described preparation of drug combination method is: above 19 flavors, Radix Angelicae Sinensis, Radix Curcumae, Herba Pogostemonis, Herba Eupatorii, Fructus Amomi, Herba Lycopi extracted volatile oil 6-10 hour, and the aqueous solution after distillation device is in addition collected; Volatile oil is with the beta-schardinger dextrin-saturated water solution method inclusion of 3-5 times of weight portion, drying, the volatile oil beta-cyclodextrin inclusion complex, standby; Radix Salviae Miltiorrhizae is with 85% alcohol reflux of 4-8 times of weight portion 1-3 time, and each 0.5-1.5 hour, merge extractive liquid, filtered, and filtrate recycling ethanol is not to there being the alcohol flavor, Radix Salviae Miltiorrhizae extract, standby; 12 flavors such as medicinal residues and all the other Herba Artemisiae Scopariaes decoct with water 1-3 time, and the decocting that at every turn adds 6-10 times of weight portion boiled 1-2 hour, and collecting decoction filters; Aqueous solution after filtrate and the above-mentioned distillation merges, be concentrated into 60～65 ℃ of relative densities and be 1.02～1.04 thick paste I, adding ethanol makes the alcohol amount of containing reach 60-80%, left standstill 12-36 hour, filter, filtrate recycling ethanol is to an amount of, merge with above-mentioned Radix Salviae Miltiorrhizae extract, being concentrated into relative density is the thick paste II of 1.10-1.15, add the volatile oil beta-cyclodextrin inclusion complex and be ground into fine powder, add conventional adjuvant, according to common process, make the dosage form of clinical acceptance, include but not limited to mixture, concentrated pill, capsule, drop pill, granule, tablet, soft capsule, slow releasing agent, oral liquid or lyophilized injectable powder.