CN103207181A - Quality detection method for traditional Chinese medicinal preparation for treating fundus hemorrhage disease - Google Patents

Abstract

The invention relates to a quality detection method for a traditional Chinese medicinal preparation for treating a fundus hemorrhage disease. The traditional Chinese medicinal preparation is pharmaceutically prepared from 8.93 percent of pollen typhae, 8.93 percent of salviae miltiorrhizae, 7.14 percent of radices rehmanniae, 7.14 percent of eclipta, 5.95 percent of chrysanthemum. 5.36 percent of radix scutellariae carbon, 5.36 percent of semen cassiae, 5.36 percent of semen plantaginis, 5.36 percent of fructus leonuri, 5.36 percent of glossy privet fruit, 5.36 percent of selfheal, 5.36 percent of radix gentianae, 3.57 percent of radix curcumae, 5.36 percent of horsetail, 3.57 percent of red peony root, 3.57 percent of moutan bark, 3.57 percent of hawthorn, 3.57 percent of angelica sinensis and 1.19 percent of rhizoma chuanxiong. The fundus hemorrhage disease caused by thermal burn collateral is treated by cooling and stopping blood, nourishing yin, dispersing blood stasis, nourishing the liver and improving eyesight. The quality detection method is high in operability and high in accuracy; whether a product is qualified and whether the quality of the product is high can be effectively detected; and the safety and the effectiveness of drug taking of a patient can be guaranteed.

Description

A kind of quality determining method for the treatment of the Chinese medicine preparation of fundus hemorrhage disease

Technical field

The present invention relates to a kind of quality determining method for the treatment of the Chinese medicine preparation of fundus hemorrhage disease, belong to the pharmaceutical technology field.

Background technology

Fundus hemorrhage is the very high disease of a kind of incidence of disease in our daily life, has a lot of people all to be very easy to suffer from China and sends out this disease of fundus hemorrhage ,Studies show that, cause the reason of fundus hemorrhage a lot, mainly contain CRAO, hypertensive retinopathy, thrombosis of central vein of retina, periphlebitis of retina, retinal periarteritis, renal retinopathy becomes, BDR, Ke thatch disease (coats), retinal blood tuberculation, choroidal hemangioma etc., also has the choroidal rupture due to some eye traumas, commotio retinae, the choroidal hemorrhage of contusion property such as perforatio maculae luteae, the traditional Chinese medical science is commonly considered as the thermal burn channels and collaterals, and blood-head is absurd row then, for example to be that the deficiency of Yin is scorching cause that its fundus hemorrhage is its complication to the diabetes tcm diagnosis.Now retrieve on September 5th, 2007 disclosed publication number and namely provide a kind of quality determining method that fundus hemorrhage Chinese medicine preparation and blood make eye bright for the treatment of for the patent of CN101028487A, this invents the consumption that described preparation for regulating blood and improving eyesight is determined each medicinal material in the prescription composition, so the weight of each dosage unit can't be determined, those skilled in the art adopt the method for this invention record can't realize this quality determining method, i.e. this invention does not have practicality, its creativeness is non-existent, even adopt Chinese medicine preparation of the present invention as detected object, its testing result is also not remarkable, poor reliability.The present invention is according to traditional Chinese medical science cooling blood and hemostasis, the enriching yin stagnation resolvation, the theory of curing the disease that nourishes the liver to improve visual acuity, employing is by cattail pollen 8.93%, the red sage root 8.93%, glutinous rehmannia 7.14%, eclipta 7.14%, chrysanthemum 5.95%, charred Radix Scutellariae 5.36%, cassia seed 5.36%, plantain seed 5.36%, Semen Leonuri 5.36%, the fruit of glossy privet 5.36%, selfheal 5.36%, rough gentian 5.36%, root tuber of aromatic turmeric 3.57%, the scouring rush 5.36%, the radix paeoniae rubrathe 3.57%, moutan bark 3.57%, hawthorn 3.57%, Radix Angelicae Sinensis 3.57%, Ligusticum wallichii 1.19% is made any Chinese medicine preparation on the pharmacy, a kind of quality determining method of this Chinese medicine preparation is provided, this quality determining method is simple and convenient and reliable and stable, effectively guarantees product quality.

Summary of the invention

The purpose of this invention is to provide a kind of quality determining method for the treatment of the Chinese medicine preparation of fundus hemorrhage disease, this quality determining method is simple and convenient and reliable and stable, effectively guarantees product quality.

For achieving the above object, the present invention has adopted following technical scheme:

Chinese medicine preparation of the present invention is by cattail pollen 8.93%, the red sage root 8.93%, glutinous rehmannia 7.14%, eclipta 7.14%, chrysanthemum 5.95%, charred Radix Scutellariae 5.36%, cassia seed 5.36%, plantain seed 5.36%, Semen Leonuri 5.36%, the fruit of glossy privet 5.36%, selfheal 5.36%, rough gentian 5.36%, root tuber of aromatic turmeric 3.57%, the scouring rush 5.36%, the radix paeoniae rubrathe 3.57%, moutan bark 3.57%, hawthorn 3.57%, Radix Angelicae Sinensis 3.57%, Ligusticum wallichii 1.19% is made, this Chinese medicine preparation can be the above any formulation of pharmacy, and its quality determining method can be micro-discrimination method, in thin layer discrimination method or the content assaying method one or more.

1. micro-discrimination method:

Get this product, put microscopically and observe: pollen granule similar round or ellipse, about 17～29 μ m of diameter, there is netted carving line (cattail pollen) on the surface.Bast fiber is faint yellow, fusiformis, wall thickness, hole ditch thin (root of large-flowered skullcap).Plant the intracutaneous epidermal surface and see rectangle like, wall is thin, and wavy is often made zyklopisch and arranged (plantain seed).The pollen granule similar round, diameter 24～34 μ m, outer wall spinosity, long 3～5 μ m of thorn, 3 germinal aperatures of tool (chrysanthemum).

2. the thin layer discrimination method 1:

(1) scutelloside discrimination method:

Get this product that is equivalent to raw medicinal herbs 8.4g, porphyrize, add methyl alcohol 20ml, ultrasonic processing 20 minutes filters the filtrate evaporate to dryness, residue adds methyl alcohol 1ml makes dissolving, get supernatant as need testing solution, other gets the scutelloside reference substance, adds methyl alcohol and makes the solution that every 1ml contains 1mg, product solution in contrast, according to thin-layered chromatography 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 binder with 4% sodium acetate solution, be developping agent with ethyl acetate-butanone-formic acid-water 4～8:1～4:0.5～2:0.5～2, launch, take out, dry, spray is with 1% ferric trichloride ethanolic solution, in the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color.

(2) Chrysophanol discrimination method:

Get this product that is equivalent to raw medicinal herbs 8.4g, porphyrize adds methyl alcohol 20ml, ultrasonic processing 20 minutes filters the filtrate evaporate to dryness, residue adds water 10ml makes dissolving, add hydrochloric acid 1ml again, water-bath added hot reflux 30 minutes, immediately cooling, extract 3 times with the ether jolting, each 15ml merges ether solution, and low temperature volatilizes ether, residue adds methyl alcohol 1ml makes dissolving, as need testing solution, other gets the Chrysophanol reference substance, adds methyl alcohol and makes the solution that every 1ml contains 0.5mg, product solution in contrast, according to the thin-layered chromatography test, draw need testing solution 5 μ l, reference substance solution 1 μ l puts respectively on same silica gel g thin-layer plate, upper solution with 60～90 ℃ of sherwood oil-ethyl formates-formic acid 10～20:3～7:0.5～2 is developping agent, launch, take out, dry, put under the 365nm ultraviolet lamp and inspect, in the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescent spot of same color

3. the thin layer discrimination method 2:

(1) Paeoniflorin discrimination method:

Get this product that is equivalent to raw medicinal herbs 16.8g, porphyrize, add methyl alcohol 50ml, ultrasonic processing 30 minutes, filter, the filtrate evaporate to dryness, residue adds water 20ml makes dissolving, extracts 3 times with water saturated normal butyl alcohol, each 20ml, merge normal butyl alcohol liquid, evaporate to dryness, residue add methyl alcohol 2ml makes dissolving, admixes neutral alumina 3g, drying with water bath, be added in 100～200 orders, internal diameter is 1cm, weight is on the neutral alumina post of 2g, with methyl alcohol 60ml wash-out, collects eluent, evaporate to dryness, residue adds methyl alcohol 1ml makes dissolving, and as need testing solution, other gets the Paeoniflorin reference substance, add methyl alcohol and make the solution that every 1ml contains 1mg, product solution according to the thin-layered chromatography test, is drawn each 5 μ l of above-mentioned two kinds of solution in contrast, put respectively on same silica gel g thin-layer plate, be developping agent with methenyl choloride-ethyl acetate-methyl alcohol-formic acid 30～40:3～5:5～10:0.1～0.2, launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, and it is clear to be heated to the spot colour developing at 105 ℃, in the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color.

(2) typhaneoside discrimination method:

Get the typhaneoside reference substance, add methyl alcohol and make the solution that every 1ml contains 1mg, product solution in contrast, test according to thin-layered chromatography, draw each the 1 μ l of need testing solution under reference substance solution and the Paeoniflorin discrimination method item, putting respectively on same polyamide film, is developping agent with acetone-water 1～3:2～5, launches, take out, dry, spray is placed with the aluminium choride test solution, put under the 365nm ultraviolet lamp and inspect, in the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

(3) rough gentian discrimination method:

Get the need testing solution under the Paeoniflorin discrimination method item, add methyl alcohol 2ml dilution back as need testing solution, other gets rough gentian control medicinal material 0.5g, adds methyl alcohol 20ml, ultrasonic processing 30 minutes, filter, filtrate evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, medicinal material solution in contrast, according to the thin-layered chromatography test, draw each 2 μ l of above-mentioned two kinds of solution, put in same silica G F respectively ₂₅₄On the thin layer plate, be developping agent with methenyl choloride-methanol-water 20～30:5～10:1～3 lower floor's solution, launch, take out, dry, put under the 254nm ultraviolet lamp and inspect, in the test sample chromatogram, with the corresponding position of control medicinal material chromatogram on, show the principal spot of a same color.

(4) chrysanthemum discrimination method:

Get this product that is equivalent to raw medicinal herbs 8.4g, porphyrize adds ethyl acetate 30ml, add hot reflux 1 hour, and filtered the filtrate evaporate to dryness, residue adds ethyl acetate 1ml makes dissolving, as need testing solution, other gets chrysanthemum control medicinal material 1g, shines medicinal material solution in pairs with legal system, test according to thin-layered chromatography, drawing each 2 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, is developping agent with 60～90 ℃ of petroleum ether-ethyl acetates 10～15: 1～3, launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, it is clear to be heated to spot colour developing at 105 ℃, in the test sample chromatogram, with the corresponding position of control medicinal material chromatogram on, show the principal spot of a same color.

4. content assaying method:

Chromatographic condition and system suitability test are filling agent with the octadecylsilane chemically bonded silica; Be the phase that flows with methyl alcohol-0.5% acetic acid 6～8:90～92; The detection wavelength is 281nm, and number of theoretical plate calculates by the Sodium Danshensu peak should be not less than 1500;

It is an amount of that the Sodium Danshensu reference substance is got in the preparation of reference substance solution, adds 50% methyl alcohol and make the solution that every 1ml contains Sodium Danshensu 25 μ g, namely;

This product that is equivalent to raw medicinal herbs 16.8g is got in the preparation of need testing solution, and accurate the title decided porphyrize, get about 0.5g, the accurate title, decide, and puts in the tool plug conical flask, the accurate 50% methyl alcohol 25ml that adds, close plug claims to decide weight, ultrasonic processing 10 minutes, power are that 250W, frequency are 40 kHz, put cold, claim again to decide weight, supply the weight that subtracts mistake with 50% methyl alcohol, shake up, filter, get subsequent filtrate, namely;

Determination method is accurate reference substance solution and each 10 μ l of need testing solution of drawing respectively, injects liquid chromatograph, measures, namely.

5. the detection method that is constituted by micro-discrimination method, thin layer discrimination method and content assaying method:

Micro-discriminating:

Get this product, put microscopically and observe: pollen granule similar round or ellipse, about 17～29 μ m of diameter, there is netted carving line (cattail pollen) on the surface.Bast fiber is faint yellow, fusiformis, wall thickness, hole ditch thin (root of large-flowered skullcap).Plant the intracutaneous epidermal surface and see rectangle like, wall is thin, and wavy is often made zyklopisch and arranged (plantain seed).The pollen granule similar round, diameter 24～34 μ m, outer wall spinosity, long 3～5 μ m of thorn, 3 germinal aperatures of tool (chrysanthemum).

Thin layer is differentiated:

(1) gets this product that is equivalent to raw medicinal herbs 8.4g, porphyrize, add methyl alcohol 20ml, ultrasonic processing 20 minutes filters the filtrate evaporate to dryness, residue adds methyl alcohol 1ml makes dissolving, get supernatant as need testing solution, other gets the scutelloside reference substance, adds methyl alcohol and makes the solution that every 1ml contains 1mg, product solution in contrast, according to thin-layered chromatography 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 binder with 4% sodium acetate solution, be developping agent with ethyl acetate-butanone-formic acid-water 4～8:1～4:0.5～2:0.5～2, launch, take out, dry, spray is with 1% ferric trichloride ethanolic solution, in the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color.

(2) get this product that is equivalent to raw medicinal herbs 8.4g, porphyrize adds methyl alcohol 20ml, ultrasonic processing 20 minutes filters the filtrate evaporate to dryness, residue adds water 10ml makes dissolving, add hydrochloric acid 1ml again, water-bath added hot reflux 30 minutes, immediately cooling, extract 3 times with the ether jolting, each 15ml merges ether solution, and low temperature volatilizes ether, residue adds methyl alcohol 1ml makes dissolving, as need testing solution, other gets the Chrysophanol reference substance, adds methyl alcohol and makes the solution that every 1ml contains 0.5mg, product solution in contrast, according to the thin-layered chromatography test, draw need testing solution 5 μ l, reference substance solution 1 μ l puts respectively on same silica gel g thin-layer plate, upper solution with 60～90 ℃ of sherwood oil-ethyl formates-formic acid 10～20:3～7:0.5～2 is developping agent, launch, take out, dry, put under the 365nm ultraviolet lamp and inspect, in the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

(3) get this product that is equivalent to raw medicinal herbs 16.8g, porphyrize, add methyl alcohol 50ml, ultrasonic processing 30 minutes, filter, the filtrate evaporate to dryness, residue adds water 20ml makes dissolving, extracts 3 times with water saturated normal butyl alcohol, each 20ml, merge normal butyl alcohol liquid, evaporate to dryness, residue add methyl alcohol 2ml makes dissolving, admixes neutral alumina 3g, drying with water bath, be added in 100～200 orders, internal diameter is 1cm, weight is on the neutral alumina post of 2g, with methyl alcohol 60ml wash-out, collects eluent, evaporate to dryness, residue adds methyl alcohol 1ml makes dissolving, and as need testing solution, other gets the Paeoniflorin reference substance, add methyl alcohol and make the solution that every 1ml contains 1mg, product solution according to the thin-layered chromatography test, is drawn each 5 μ l of above-mentioned two kinds of solution in contrast, put respectively on same silica gel g thin-layer plate, be developping agent with methenyl choloride-ethyl acetate-methyl alcohol-formic acid 30～40:3～5:5～10:0.1～0.2, launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, and it is clear to be heated to the spot colour developing at 105 ℃, in the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color.

(4) get the typhaneoside reference substance, add methyl alcohol and make the solution that every 1ml contains 1mg, product solution in contrast, test according to thin-layered chromatography, draw each the 1 μ l of need testing solution under reference substance solution and the Paeoniflorin discrimination method item, putting respectively on same polyamide film, is developping agent with acetone-water 1～3:2～5, launches, take out, dry, spray is placed with the aluminium choride test solution, put under the 365nm ultraviolet lamp and inspect, in the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

(5) get need testing solution under the Paeoniflorin discrimination method item, add methyl alcohol 2ml dilution back as need testing solution, other gets rough gentian control medicinal material 0.5g, adds methyl alcohol 20ml, ultrasonic processing 30 minutes, filter, filtrate evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, medicinal material solution in contrast, according to the thin-layered chromatography test, draw each 2 μ l of above-mentioned two kinds of solution, put in same silica G F respectively ₂₅₄On the thin layer plate, be developping agent with methenyl choloride-methanol-water 20～30:5～10:1～3 lower floor's solution, launch, take out, dry, put under the 254nm ultraviolet lamp and inspect, in the test sample chromatogram, with the corresponding position of control medicinal material chromatogram on, show the principal spot of a same color.

(6) get this product that is equivalent to raw medicinal herbs 8.4g, porphyrize adds ethyl acetate 30ml, add hot reflux 1 hour, and filtered the filtrate evaporate to dryness, residue adds ethyl acetate 1ml makes dissolving, as need testing solution, other gets chrysanthemum control medicinal material 1g, shines medicinal material solution in pairs with legal system, test according to thin-layered chromatography, drawing each 2 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, is developping agent with 60～90 ℃ of petroleum ether-ethyl acetates 10～15: 1～3, launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, it is clear to be heated to spot colour developing at 105 ℃, in the test sample chromatogram, with the corresponding position of control medicinal material chromatogram on, show the principal spot of a same color.

Content assaying method:

Chromatographic condition and system suitability test are filling agent with the octadecylsilane chemically bonded silica; Be the phase that flows with methyl alcohol-0.5% acetic acid 6～8:90～92; The detection wavelength is 281nm, and number of theoretical plate calculates by the Sodium Danshensu peak should be not less than 1500.

It is an amount of that the Sodium Danshensu reference substance is got in the preparation of reference substance solution, adds 50% methyl alcohol and make the solution that every 1ml contains Sodium Danshensu 25 μ g, namely.

This product that is equivalent to raw medicinal herbs 16.8g is got in the preparation of need testing solution, and accurate the title decided porphyrize, get about 0.5g, the accurate title, decide, and puts in the tool plug conical flask, the accurate 50% methyl alcohol 25ml that adds, close plug claims to decide weight, ultrasonic processing 10 minutes, power are that 250W, frequency are 40 kHz, put cold, claim again to decide weight, supply the weight that subtracts mistake with 50% methyl alcohol, shake up, filter, get subsequent filtrate, namely.

Determination method is accurate reference substance solution and each 10 μ l of need testing solution of drawing respectively, injects liquid chromatograph, measures, namely.

The invention provides a kind of quality determining method for the treatment of the Chinese medicine preparation of fundus hemorrhage disease, this Chinese medicine preparation is by cattail pollen 8.93%, the red sage root 8.93%, glutinous rehmannia 7.14%, eclipta 7.14%, chrysanthemum 5.95%, charred Radix Scutellariae 5.36%, cassia seed 5.36%, plantain seed 5.36%, Semen Leonuri 5.36%, the fruit of glossy privet 5.36%, selfheal 5.36%, rough gentian 5.36%, root tuber of aromatic turmeric 3.57%, the scouring rush 5.36%, the radix paeoniae rubrathe 3.57%, moutan bark 3.57%, hawthorn 3.57%, Radix Angelicae Sinensis 3.57%, Ligusticum wallichii 1.19% is made any formulation on the pharmacy, by cooling blood and hemostasis, the enriching yin stagnation resolvation, nourish the liver to improve visual acuity and reach treatment deficiency of Yin liver-yang hyperactivity, the caused fundus hemorrhage of thermal burn channels.The present invention on the basis of existing technology, the relation of measuring between the given each component, by a large amount of experimental studies, contrast, determined a kind of workable, the quality determining method that accuracy is high, deleted the radix scutellariae medicinal materials contrast on the basis of existing technology, the contrast of cassia seed medicinal material, the archen contrast, the contrast of eclipta medicinal material, discrimination methods such as protocatechualdehyde contrast are not remarkable, the thin layer of poor reproducibility is differentiated, increased Paeoniflorin newly, typhaneoside, the rough gentian medicinal material, a series of contrast methods such as chrysanthemum medicinal material are remarkable, reappearance and stability is the thin layer discrimination method reliably, micro-discriminating and content assaying method are provided simultaneously, many-sided, multi-angle guaranteed product quality, can more effective testing product whether qualified, whether quality is good and bad, guaranteed the security of patient's medication, validity.

Now be described below with regard to important research method of the present invention:

One, experimental drug

The experimental drug prescription:

Cattail pollen 75g red sage root 75g glutinous rehmannia 60g eclipta 60g

Chrysanthemum 50g charred Radix Scutellariae 45g cassia seed 45g plantain seed 45g

Semen Leonuri 45g fruit of glossy privet 45g selfheal 45g felwort 45g

Scouring rush 45g radix paeoniae rubrathe 30g moutan bark 30g root tuber of aromatic turmeric 30g

Hawthorn 30g Radix Angelicae Sinensis 30g Ligusticum wallichii 10g

The experimental drug preparation method:

More than 19 the flavor, chrysanthemum, charred Radix Scutellariae, plantain seed 22.5g, cattail pollen 37.5g, mixed powder is broken into fine powder, and is standby; Residue plantain seed, cattail pollen and all the other each flavors were soaked 30 minutes, decocted secondary, and each 2 hours, collecting decoction filtered.Filtrate decompression is concentrated into the clear cream that relative density is 1.10～1.15 (60 ℃), spray drying, and cream powder and above-mentioned fine powder and appropriate amount of auxiliary materials are made particle, add dolomol 1.5g, and mixing is pressed into 1000, sugar coating or film-coating, namely.

Two, research process

The thin layer of scutelloside is differentiated:

Prior art is contrast with scutelloside, root of large-flowered skullcap control medicinal material, now gets 10 of this experiment products, removes dressing, and porphyrize adds methyl alcohol 20ml, and ultrasonic processing 20 minutes filters, and filtrate evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, gets supernatant as need testing solution.Preparation does not simultaneously contain the blank preparation of radix scutellariae medicinal materials ,Make negative control solution with method.According to the thin-layered chromatography test, draw each 5 μ l of above-mentioned four kinds of solution, thin layer plate has compared the method I: the silica gel g thin-layer plate that contains 4% sodium acetate; The method II: silica gel g thin-layer plate is developping agent with ethyl acetate-butanone-formic acid-water (5:3:1:1) all; The method III: polyamide film, acetic acid are developping agent.Spray is with 1% ferric trichloride ethanolic solution.The result: contain silica gel g thin-layer plate and the silica gel g thin-layer plate of 4% sodium acetate, in the test sample chromatogram of three batches of sugar coated tablets and three batches of Film coated tablets, with control medicinal material chromatogram and the corresponding position of reference substance chromatogram on, all show the spot of same color, negative noiseless.Especially be that the sample drawing flocculus point rounding of silica gel g thin-layer plate of binder is clear with 4% sodium acetate solution, the position is moderate, and because the polyamide film volume containing the sample is little, separating effect is bad, so the method for optimizing I.

Result: in the test sample chromatogram, showing the spot of same color with scutelloside reference substance chromatogram relevant position; Showing the spot of basic identical color with root of large-flowered skullcap control medicinal material chromatogram relevant position, but negative control is showing the spot of 3 basically identicals with root of large-flowered skullcap control medicinal material chromatogram and test sample chromatogram relevant position, showing the spot of same color with scutelloside reference substance chromatogram relevant position, scutelloside has specificity for the discriminating of the root of large-flowered skullcap, so select scutelloside to be contrast.

The thin layer of cassia seed is differentiated:

Prior art is with the cassia seed control medicinal material, archen, Chrysophanol is contrast, and the Ji Yuan of a middle cassia seed medicinal material of Chinese Pharmacopoeia version in 2010 is the dry mature seed of Cassia tora and little Cassia tora, get little Cassia tora and each 1g of cassia seed control medicinal material, add methyl alcohol 20ml respectively, ultrasonic processing 20 minutes filters the filtrate evaporate to dryness, residue adds water 10ml makes dissolving, add hydrochloric acid 1ml again, water-bath added hot reflux 30 minutes, immediately cooling, extract 3 times with the ether jolting, each 15ml merges ether solution, and low temperature volatilizes ether, residue adds methyl alcohol 1ml makes dissolving, solution according to the thin-layered chromatography test, is drawn each 5 μ l of comparative solution as a comparison, put respectively on same silica gel g thin-layer plate, upper solution with sherwood oil (60～90 ℃)-ethyl formate-formic acid (15:5:1) is developping agent, launches, and takes out, dry, put under the ultraviolet lamp (365nm) and inspect.The difference of its collection of illustrative plates is that the archen in the little Cassia tora control medicinal material is not too obvious, and all the other collection of illustrative plates spots are consistent with the Cassia tora control medicinal material.Both can not be separated fully in the actual production, cassia seed control medicinal material, archen are removed.

Be reference substance with the Chrysophanol, get 10 of this experimental drugs, remove dressing, porphyrize adds methyl alcohol 20ml, ultrasonic processing 20 minutes, filter, filtrate evaporate to dryness, residue add water 10ml makes dissolving, add hydrochloric acid 1ml again, water-bath added hot reflux 30 minutes, immediately cooling, extract 3 times with the ether jolting, each 15ml merges ether solution, low temperature volatilizes ether, and residue adds methyl alcohol 1ml makes dissolving, as need testing solution.Preparation does not simultaneously contain the blank preparation of cassia seed medicinal material ,Make negative control solution with method.Test according to thin-layered chromatography, draw need testing solution 5 μ l, reference substance solution 1 μ l, put respectively on same silica gel g thin-layer plate, upper solution with sherwood oil (60～90 ℃)-ethyl formate-formic acid (15:5:1) is developping agent, launches, and takes out, dry, put under the ultraviolet lamp (365nm) and inspect.Three batches of sugar coated tablets and three batches of Film coated tablets have all detected Chrysophanol as a result, and are negative noiseless.

The thin layer of Paeoniflorin is differentiated:

Be contrast with the Paeoniflorin, get 20 of this experimental drugs, remove dressing, porphyrize adds methyl alcohol 50ml, ultrasonic processing 30 minutes, filter, filtrate evaporate to dryness, residue add water 20ml makes dissolving, extract 3 times with water saturated normal butyl alcohol, each 20ml merges normal butyl alcohol liquid, evaporate to dryness, residue adds methyl alcohol 2ml makes dissolving, admixes neutral alumina 3g, drying with water bath, be added in neutral alumina post (100～200 orders, 2g, internal diameter are 1cm) on, with methyl alcohol 60ml wash-out, collect eluent, evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution.Other gets the preparation that does not contain the radix paeoniae rubrathe, moutan bark medicinal material, method for making with need testing solution is made negative sample solution with method, according to the thin-layered chromatography test, draw each 5 μ l of above-mentioned three kinds of solution, put respectively on same silica gel g thin-layer plate, be developping agent with methenyl choloride-ethyl acetate-methyl alcohol-formic acid (40:5:10:0.2), launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, and it is clear to be heated to the spot colour developing at 105 ℃.The three batches of sugar coated tablets and the three batches of Film coated tablets have all detected the feature spot of Paeoniflorin as a result, and be negative noiseless, and the collection of illustrative plates clear spot, and separating effect is better.

The thin layer of rough gentian is differentiated:

Be contrast with the rough gentian medicinal material, get the need testing solution of Paeoniflorin thin layer under differentiating, other gets the preparation that does not contain the rough gentian medicinal material, makes negative sample solution with the method for making of need testing solution with method.According to the thin-layered chromatography test, draw each 5 μ l of above-mentioned three kinds of solution, put in same silica G F respectively ₂₅₄On the thin layer plate, developping agent has compared I: ethyl acetate-methanol-water (10:2:1); II: methenyl choloride-methanol-water (30:10:3) lower floor solution is developping agent, launches, and takes out, and dries, and puts under the ultraviolet lamp (254nm) and inspects.The three batches of sugar coated tablets and the three batches of Film coated tablets have all detected the feature spot of rough gentian medicinal material as a result, and be negative noiseless, better with development system II separating effect especially.Again the point sample amount is changed into 2 μ l, launch test sample collection of illustrative plates clear spot with the development system II.

The thin layer of cattail pollen is differentiated:

Be contrast with the typhaneoside, other gets the preparation that does not contain cattail pollen, differentiates that with Paeoniflorin down the method for making of need testing solution is made negative sample solution with method.Test according to thin-layered chromatography, draw each the 1 μ l of need testing solution under above-mentioned two kinds of solution and the Paeoniflorin discriminating item, put respectively on same polyamide film, be developping agent with acetone-water (1:2), launch, take out, dry, spray is placed with the aluminium choride test solution, puts under the ultraviolet lamp (365nm) and inspects.The three batches of sugar coated tablets and the three batches of Film coated tablets have all detected the feature spot of cattail pollen as a result, and be negative noiseless, and the collection of illustrative plates clear spot, and separating effect is better.

The thin layer of chrysanthemum is differentiated:

Be contrast with the chrysanthemum control medicinal material, get 10 of this experimental drugs, remove dressing, porphyrize adds ethyl acetate 30ml, adds hot reflux 1 hour, filters, and filtrate evaporate to dryness, residue add ethyl acetate 1ml makes dissolving, as need testing solution.Other gets the preparation that does not contain the chrysanthemum medicinal material, method for making with need testing solution is made negative sample solution with method, test according to thin-layered chromatography, drawing each 5 μ l of above-mentioned three kinds of solution, put on same silica gel g thin-layer plate respectively, is developping agent with sherwood oil (30～60 ℃)-ethyl acetate (13: 1), launches, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, and it is clear to be heated to spot colour developing at 105 ℃.The three batches of sugar coated tablets and the three batches of Film coated tablets have all detected the feature spot of chrysanthemum control medicinal material as a result, negative noiseless, again the point sample amount is changed into 2 μ l, the developping agent ratio is adjusted into sherwood oil (30～60 ℃)-ethyl acetate (10: 1), test sample collection of illustrative plates clear spot, separating effect is better.

The thin layer of eclipta is differentiated:

Be the method under eclipta is differentiated in the patent of CN101028487A according to publication number, get this experimental drug 0.3g, remove dressing, porphyrize, add ethanol 20ml, flood after 1 hour, ultrasonic processing 30 minutes filters, filtrate evaporate to dryness, residue add absolute ethyl alcohol 1ml makes dissolving, as need testing solution.Other gets eclipta control medicinal material 1g, shines medicinal material solution in pairs with legal system.Drawing each 5 μ l of above-mentioned two kinds of solution, put on same silica gel g thin-layer plate, is developping agent with cyclohexane-ethyl acetate (9:1), launches, and takes out, and dries, and it is smoked clear to the spot colour developing to put in the iodine vapor.Three batch samples are differentiated the result: with test sample chromatogram principal spot relevant position on control medicinal material chromatogram spot do not separate fully, exhibition is not exclusively corresponding with sample apart from increasing back control medicinal material chromatogram principal spot.

The research of content assaying method:

1, instrument, medicine and reagent

1.1, instrument: wear peace: UltiMate 3000 series of high efficiency liquid chromatographs, quaternary gradient pump, diode array detector (DAD), automatic sampler, Chromeleon 6.8 workstations; Agilent1100 type high performance liquid chromatograph, G1311A type quaternary gradient pump, G1314A type UV-detector, G2170AA type chem workstation;

1.2, reagent: methyl alcohol is chromatorgaphy reagent, and water is ultrapure water, other reagent be analyze pure.

1.3, reference substance: Sodium Danshensu, lot number: 110855-200809(for assay with), provided by Nat'l Pharmaceutical ﹠ Biological Products Control Institute.

1.4, test sample and XUEMINGMU PIAN (sugar-coat, lot number: 201202006,201202007,201202008,201203009,201203010; Film-coating, lot number: 201203005,201203006,201203007,201204008,201204009; Produced by Beilin Pharmaceutical Co., Ltd., Xi'an)

2, chromatographic condition

2.1, chromatographic column: Megres C18 (250*4.6mm);

2.2, phase flows: methyl alcohol-0.5% acetic acid (8:92);

Be the phase time that flows with methyl alcohol-0.5% acetic acid (10:90), the danshensu peak can reach baseline separation in the test sample chromatogram, but has with the corresponding chromatographic peak of danshensu in the negative preparation chromatogram of the red sage root; When being adjusted into methyl alcohol-0.5% acetic acid (8:92) mutually when flowing, the danshensu peak can reach baseline separation in the test sample chromatogram, does not have with the corresponding chromatographic peak of Sodium Danshensu in the negative preparation chromatogram of the red sage root.

2.3, detect wavelength: 281nm.

Maximum absorption wavelength is selected to get the Sodium Danshensu reference substance solution, sets 10 μ l, injects high performance liquid chromatograph, by diode array detector, measures maximum absorption wavelength.The result shows that Sodium Danshensu has maximum absorption band at the 282.7nm place.

Under above-mentioned analysis condition, the degree of separation of Sodium Danshensu and adjacent impurity peaks is greater than more than 1.5, and number of theoretical plate calculates by the Sodium Danshensu peak and is not less than 1500.

The selection of 3, pre-treatment condition

3.1, extract choice of Solvent to methyl alcohol, 50% methyl alcohol, water as extracting solvent, investigate.

It is an amount of to get experimental drug, removes dressing, porphyrize, take by weighing three parts, every part of about 0.5g, the accurate title, decide, put in the tool plug conical flask, precision adds 50% methyl alcohol, water, methyl alcohol 25ml respectively, claims to decide weight, ultrasonic processing (power 250W, frequency 40 kHz) 30 minutes is put cold, claim again to decide weight, supply the weight that subtracts mistake with corresponding solvent, shake up, filter, get subsequent filtrate, namely.Measure by chromatographic condition, test findings is asked for an interview table 1 in accordance with the law.

Table 1 extracts choice of Solvent test and result

Test findings shows that water and 50% methyl alcohol are as extracting solvent, and peak area ratio is close, and water is slightly high, but water is as extracting solvent, more sad filter in the specimen preparation process.Select 50% methanol solution as extracting solvent.

3.2, the different solvents amount investigates and to add different volumes multiple 50% methyl alcohol, investigate.

The test sample preparation method: it is an amount of to get experimental drug, removes dressing, porphyrize, the accurate title, decide, and precision adds 50% methyl alcohol 25,50,100 times of amounts respectively, claims to decide weight, ultrasonic processing (power 250W, frequency 40 kHz) 30 minutes, put coldly, claim to decide weight again, supply the weight that subtracts mistake with 50% methyl alcohol, shake up, filter, namely.Experimental result sees Table 2.

Selection test and the result of table 2 different solvents amount

Test findings shows, adds 50 times of amounts of 50% methyl alcohol and can extract fully.Consider from saving solvent, take by weighing test sample 0.5g, add 50% methyl alcohol 25ml, ultrasonic processing gets final product.

3.3, different ultrasonic time investigates

Take by weighing with the about 0.5g of a collection of experimental drug, precision adds each 25ml of 50% methyl alcohol, and ultrasonic processing (power 250W, frequency 40 kHz) is 10,20,30,40 minutes respectively, and test findings is asked for an interview table 3.

The different ultrasonic times of table 3 are investigated test and result

Test findings shows, ultrasonic processing 10,20,30,40 minutes, and the result is close for its assay, illustrates ultrasonic 10 minutes and just can extract fully, so select 10 minutes ultrasonic extraction times as this method of the ultrasonic processing of 50% methanol solvate system.

4, methodological study

4.1, linear relationship investigates

Get danshensu reference substance solution: 0.05861mg/ml, set sampling volume and be respectively 2 μ l, 5 μ l, 10 μ l, 15 μ l, 25 μ l, measure by drafting chromatographic condition, the result asks for an interview table 4.Be horizontal ordinate with the sample size, the peak area integrated value is ordinate, the drawing standard curve, and carry out straight-line regression, regression equation is Y=381.07X+0.7424, r=0.9999.Test findings shows that this method sample size is good linear relationship in 0.11722～1.46525 μ g scope.

Table 4 linear relationship is investigated test and result

4.2, precision test: accurate danshensu reference substance solution and each 10 μ l of need testing solution of drawing, repeat sample introduction continuously 6 times, measure, record its peak area value, the results are shown in Table 5, table 6.

The test of table 5 danshensu precision

The test of table 6 test sample precision

The result shows: danshensu reference substance and need testing solution respectively repeat relative standard deviation RSD that sample introduction records the peak area integrated value for 6 times all less than 2.0%, so this method has good precision.

4.3, specificity test: get the simulation prescription of removing the red sage root, make the negative sample that does not contain the red sage root, make negative control solution by the need testing solution method for making.Other gets need testing solution and reference substance solution, precision is drawn reference substance solution and each 10 μ l of need testing solution respectively, inject liquid chromatograph, measure, test findings shows, in the test sample chromatogram, with the corresponding position of reference substance chromatographic retention on, the single chromatographic peak that one correspondence is arranged, and in the negative control liquid chromatography, do not have corresponding peak in this retention time corresponding position and occur.Illustrate that feminine gender is noiseless to its mensuration, the specificity of its method is better.

4.4, stability test: the accurate need testing solution of drawing, measure at 0,2,4,6,8,24 hour injection liquid chromatograph respectively, test findings sees Table 7.

Table 7 stability test result

Test findings shows that need testing solution is good at 24 hours internal stabilities, and its relative standard deviation is that RSD is 1.48%.

4.5, replica test: get with 6 parts of a collection of experimental drugs, prepare the sample test liquid by the text method, record peak area and calculate content, the results are shown in Table 8.

Table 8 sample replica test and result

Test findings shows that the average content that records this batch sample is the 0.2874mg/ sheet, and its relative standard deviation RSD is 1.91% by statistics.The repeatability of presentation of results this method better.

4.6 recovery test

4.6.1, sugar coated tablet average recovery test: precision take by weighing known content (the 0.2874mg/ sheet, experimental drug 0.958mg/g) is an amount of, accurate claim fixed, precision adds danshensu reference substance solution (0.04312 mg/ml) 5ml respectively, and precision adds 50% methyl alcohol 20ml again, weighs, ultrasonic processing 10 minutes, put coldly, weigh, supply the weight that subtracts mistake with 50% methyl alcohol, shake up, filter, get subsequent filtrate, namely.The results are shown in Table 9.

  

Table 9 sugar coated tablet average recovery test findings

Test findings shows: this method has the higher recovery, and its average recovery rate is 97.5%, and relative standard deviation is 1.30%, and application of sample reclaims good.

4.6.2, Film coated tablets average recovery test: it is an amount of that precision takes by weighing the sample of known content (1.23155mg/g), accurate claim fixed, accurate danshensu reference substance solution (0.05861 mg/ml) 5ml that adds respectively, precision adds 50% methyl alcohol 20ml again, weigh, ultrasonic processing 10 minutes is put cold, weigh, supply the weight that subtracts mistake with 50% methyl alcohol, shake up, filter, get subsequent filtrate, namely.The results are shown in Table 10

Table 10 film-coating average recovery test findings

Test findings shows: this method has the higher recovery, and its average recovery rate is 97.4%, and relative standard deviation is 1.35%, and application of sample reclaims good.

5, sample size is measured

Accurate reference substance solution and the sample solution drawn measured respectively, the results are shown in Table 11.

Content of Danshensu measurement result (n=2) in table 11 sample

In ten batch samples, the measured result of danshensu, high-load is the 0.3823mg/ sheet, minimum is the 0.2620mg/ sheet.Consider factor affecting such as prescription consumption, technology extraction loss and tablet weight variation, tentative is that every of this product contains the red sage root with danshensu (C ₉H ₁₀O ₅) meter, must not be less than 0.25mg.

Embodiment

A kind of quality determining method for the treatment of the Chinese medicine preparation of fundus hemorrhage disease of the present invention is not limited to following examples.

Embodiment 1

Prescription:

Cattail pollen 75g red sage root 75g glutinous rehmannia 60g

Eclipta 60g chrysanthemum 50g charred Radix Scutellariae 45g

Cassia seed 45g plantain seed 45g Semen Leonuri 45g

Fruit of glossy privet 45g selfheal 45g rough gentian 45g

Root tuber of aromatic turmeric 30g scouring rush 45g radix paeoniae rubrathe 30g

Moutan bark 30g hawthorn 30g Radix Angelicae Sinensis 30g

Ligusticum wallichii 10g

The preparation method: above 19 flavors, chrysanthemum, charred Radix Scutellariae, plantain seed 22.5g, cattail pollen 37.5g, mixed powder is broken into fine powder, and is standby; Residue plantain seed, cattail pollen and all the other each flavors were soaked 30 minutes, decocted secondary, and each 2 hours, collecting decoction filtered.Filtrate decompression is concentrated into the clear cream that relative density is 1.10～1.15 (60 ℃), spray drying, and cream powder and above-mentioned fine powder and appropriate amount of auxiliary materials are made particle, add dolomol 1.5g, and mixing is pressed into 1000, sugar coating, namely.

Its quality determining method is as follows:

Get this product, put microscopically and observe: pollen granule similar round or ellipse, about 17～29 μ m of diameter, there is netted carving line (cattail pollen) on the surface.Bast fiber is faint yellow, fusiformis, wall thickness, hole ditch thin (root of large-flowered skullcap).Plant the intracutaneous epidermal surface and see rectangle like, wall is thin, and wavy is often made zyklopisch and arranged (plantain seed).The pollen granule similar round, diameter 24～34 μ m, outer wall spinosity, long 3～5 μ m of thorn, 3 germinal aperatures of tool (chrysanthemum).

Embodiment 2

Prescription:

Cattail pollen 75g red sage root 75g glutinous rehmannia 60g

Eclipta 60g chrysanthemum 50g charred Radix Scutellariae 45g

Cassia seed 45g plantain seed 45g Semen Leonuri 45g

Fruit of glossy privet 45g selfheal 45g rough gentian 45g

Root tuber of aromatic turmeric 30g scouring rush 45g radix paeoniae rubrathe 30g

Moutan bark 30g hawthorn 30g Radix Angelicae Sinensis 30g

Ligusticum wallichii 10g

The preparation method: above 19 flavors, chrysanthemum, charred Radix Scutellariae, plantain seed 22.5g, cattail pollen 37.5g, mixed powder is broken into fine powder, and is standby; Residue plantain seed, cattail pollen and all the other each flavors were soaked 30 minutes, decocted secondary, and each 2 hours, collecting decoction filtered.Filtrate decompression is concentrated into the clear cream that relative density is 1.10～1.15 (60 ℃), spray drying, and cream powder and above-mentioned fine powder and appropriate amount of auxiliary materials are made particle, add dolomol 1.5g, and mixing is pressed into 1000, the bag film-coating, namely.

Its quality determining method is as follows:

(1) get 10 of this product, remove dressing, porphyrize adds methyl alcohol 20ml, and ultrasonic processing 20 minutes filters, and filtrate evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, gets supernatant as need testing solution.Other gets the scutelloside reference substance, adds methyl alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), 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 binder with 4% sodium acetate solution, be developping agent with ethyl acetate-butanone-formic acid-water (5:3:1:1), launch, take out, dry, spray is with 1% ferric trichloride ethanolic solution.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color.

(2) get 10 of this product, remove dressing, porphyrize, add methyl alcohol 20ml, ultrasonic processing 20 minutes filters, filtrate evaporate to dryness, residue add water 10ml makes dissolving, adds hydrochloric acid 1ml again, water-bath added hot reflux 30 minutes, and cooling is immediately extracted 3 times with the ether jolting, each 15ml merges ether solution, and low temperature volatilizes ether, residue adds methyl alcohol 1ml makes dissolving, as need testing solution.Other gets the Chrysophanol reference substance, adds methyl alcohol and makes the solution that every 1ml contains 0.5mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw need testing solution 5 μ l, reference substance solution 1 μ l, put respectively on same silica gel g thin-layer plate, upper solution with sherwood oil (60～90 ℃)-ethyl formate-formic acid (15:5:1) is developping agent, launch, take out, dry, put under the ultraviolet lamp (365nm) and inspect.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

Embodiment 3

Prescription:

Cattail pollen 75g red sage root 75g glutinous rehmannia 60g

Eclipta 60g chrysanthemum 50g charred Radix Scutellariae 45g

Cassia seed 45g plantain seed 45g Semen Leonuri 45g

Fruit of glossy privet 45g selfheal 45g rough gentian 45g

Root tuber of aromatic turmeric 30g scouring rush 45g radix paeoniae rubrathe 30g

Moutan bark 30g hawthorn 30g Radix Angelicae Sinensis 30g

Ligusticum wallichii 10g

The preparation method: above 19 flavors, chrysanthemum, charred Radix Scutellariae, plantain seed 22.5g, cattail pollen 37.5g, mixed powder is broken into fine powder, and is standby; Residue plantain seed, cattail pollen and all the other each flavors were soaked 30 minutes, decocted secondary, and each 2 hours, collecting decoction filtered.Filtrate decompression is concentrated into the clear cream that relative density is 1.10～1.15 (60 ℃), spray drying, and cream powder and above-mentioned fine powder and appropriate amount of auxiliary materials are made particle, add dolomol 1.5g, and mixing is pressed into 1000, the bag film-coating, namely.

Its quality determining method is as follows:

(1) get 10 of this product, remove dressing, porphyrize adds methyl alcohol 20ml, and ultrasonic processing 20 minutes filters, and filtrate evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, gets supernatant as need testing solution.Other gets the scutelloside reference substance, adds methyl alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), 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 binder with 4% sodium acetate solution, be developping agent with ethyl acetate-butanone-formic acid-water (4:1:0.5:0.5), launch, take out, dry, spray is with 1% ferric trichloride ethanolic solution.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color.

(2) get 10 of this product, remove dressing, porphyrize, add methyl alcohol 20ml, ultrasonic processing 20 minutes filters, filtrate evaporate to dryness, residue add water 10ml makes dissolving, adds hydrochloric acid 1ml again, water-bath added hot reflux 30 minutes, and cooling is immediately extracted 3 times with the ether jolting, each 15ml merges ether solution, and low temperature volatilizes ether, residue adds methyl alcohol 1ml makes dissolving, as need testing solution.Other gets the Chrysophanol reference substance, adds methyl alcohol and makes the solution that every 1ml contains 0.5mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw need testing solution 5 μ l, reference substance solution 1 μ l, put respectively on same silica gel g thin-layer plate, upper solution with sherwood oil (60～90 ℃)-ethyl formate-formic acid (10:3:0.5) is developping agent, launch, take out, dry, put under the ultraviolet lamp (365nm) and inspect.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

Embodiment 4

Prescription:

Cattail pollen 75g red sage root 75g glutinous rehmannia 60g

Eclipta 60g chrysanthemum 50g charred Radix Scutellariae 45g

Cassia seed 45g plantain seed 45g Semen Leonuri 45g

Fruit of glossy privet 45g selfheal 45g rough gentian 45g

Root tuber of aromatic turmeric 30g scouring rush 45g radix paeoniae rubrathe 30g

Moutan bark 30g hawthorn 30g Radix Angelicae Sinensis 30g

Ligusticum wallichii 10g

The preparation method: above 19 flavors, chrysanthemum, charred Radix Scutellariae, plantain seed 22.5g, cattail pollen 37.5g, mixed powder is broken into fine powder, and is standby; Residue plantain seed, cattail pollen and all the other each flavors were soaked 30 minutes, decocted secondary, and each 2 hours, collecting decoction filtered.Filtrate decompression is concentrated into the clear cream that relative density is 1.10～1.15 (60 ℃), spray drying, and cream powder and above-mentioned fine powder and appropriate amount of auxiliary materials are made particle, add dolomol 1.5g, and mixing is pressed into 1000, the bag film-coating, namely.

Its quality determining method is as follows:

(1) get 10 of this product, remove dressing, porphyrize adds methyl alcohol 20ml, and ultrasonic processing 20 minutes filters, and filtrate evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, gets supernatant as need testing solution.Other gets the scutelloside reference substance, adds methyl alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), 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 binder with 4% sodium acetate solution, be developping agent with ethyl acetate-butanone-formic acid-water (8:4:2:2), launch, take out, dry, spray is with 1% ferric trichloride ethanolic solution.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color.

(2) get 10 of this product, remove dressing, porphyrize, add methyl alcohol 20ml, ultrasonic processing 20 minutes filters, filtrate evaporate to dryness, residue add water 10ml makes dissolving, adds hydrochloric acid 1ml again, water-bath added hot reflux 30 minutes, and cooling is immediately extracted 3 times with the ether jolting, each 15ml merges ether solution, and low temperature volatilizes ether, residue adds methyl alcohol 1ml makes dissolving, as need testing solution.Other gets the Chrysophanol reference substance, adds methyl alcohol and makes the solution that every 1ml contains 0.5mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw need testing solution 5 μ l, reference substance solution 1 μ l, put respectively on same silica gel g thin-layer plate, upper solution with sherwood oil (60～90 ℃)-ethyl formate-formic acid (20:7:2) is developping agent, launch, take out, dry, put under the ultraviolet lamp (365nm) and inspect.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

Embodiment 5

Prescription:

Cattail pollen 75g red sage root 75g glutinous rehmannia 60g

Eclipta 60g chrysanthemum 50g charred Radix Scutellariae 45g

Cassia seed 45g plantain seed 45g Semen Leonuri 45g

Fruit of glossy privet 45g selfheal 45g rough gentian 45g

Root tuber of aromatic turmeric 30g scouring rush 45g radix paeoniae rubrathe 30g

Moutan bark 30g hawthorn 30g Radix Angelicae Sinensis 30g

Ligusticum wallichii 10g

The preparation method: above 19 flavors, chrysanthemum, charred Radix Scutellariae, plantain seed 22.5g, cattail pollen 37.5g, mixed powder is broken into fine powder, and is standby; Residue plantain seed, cattail pollen and all the other each flavors were soaked 30 minutes, decocted secondary, and each 2 hours, collecting decoction filtered.Filtrate decompression is concentrated into the clear cream that relative density is 1.10～1.15 (60 ℃), spray drying, and cream powder and above-mentioned fine powder and appropriate amount of auxiliary materials are made particle, add dolomol 1.5g, and mixing is pressed into 1000, the bag film-coating, namely.

Its quality determining method is as follows:

(1) get 20 of this product, remove dressing, porphyrize adds methyl alcohol 50ml, ultrasonic processing 30 minutes filters, and filtrate evaporate to dryness, residue add water 20ml makes dissolving, extract 3 times with water saturated normal butyl alcohol, each 20ml merges normal butyl alcohol liquid, evaporate to dryness, residue adds methyl alcohol 2ml makes dissolving, admixes neutral alumina 3g, and drying with water bath is added in neutral alumina post (100～200 orders, 2g, internal diameter are 1cm) on, with methyl alcohol 60ml wash-out, collect eluent, evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution.Other gets the Paeoniflorin reference substance, adds methyl alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, be developping agent with methenyl choloride-ethyl acetate-methyl alcohol-formic acid (40:5:10:0.2), launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, and it is clear to be heated to the spot colour developing at 105 ℃.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color.

(2) getting Paeoniflorin differentiates and a following need testing solution to add methyl alcohol 2ml dilution afterwards as need testing solution.Other gets rough gentian control medicinal material 0.5g, adds methyl alcohol 20ml, and ultrasonic processing 30 minutes filters, and filtrate evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, in contrast medicinal material solution.According to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B) test, draw each 2 μ l of above-mentioned two kinds of solution, put in same silica G F respectively ₂₅₄On the thin layer plate, be developping agent with methenyl choloride-methanol-water (30:10:3) lower floor solution, launch, take out, dry, put under the ultraviolet lamp (254nm) and inspect.In the test sample chromatogram, with the corresponding position of control medicinal material chromatogram on, show the principal spot of a same color.

(3) get the typhaneoside reference substance, add methyl alcohol and make the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw each the 1 μ l of need testing solution under reference substance solution and the Paeoniflorin discriminating item, put respectively on same polyamide film, be developping agent with acetone-water (1:2), launch, take out, dry, spray is placed with the aluminium choride test solution, puts under the ultraviolet lamp (365nm) and inspects.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

(4) get 10 of this product, remove dressing, porphyrize adds ethyl acetate 30ml, adds hot reflux 1 hour, filters, and filtrate evaporate to dryness, residue add ethyl acetate 1ml makes dissolving, as need testing solution.Other gets chrysanthemum control medicinal material 1g, shines medicinal material solution in pairs with legal system.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw each 2 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, be developping agent with sherwood oil (60～90 ℃)-ethyl acetate (10: 1), launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, and it is clear to be heated to the spot colour developing at 105 ℃.In the test sample chromatogram, with the corresponding position of control medicinal material chromatogram on, show the principal spot of a same color.

Embodiment 6

Prescription:

Cattail pollen 75g red sage root 75g glutinous rehmannia 60g

Eclipta 60g chrysanthemum 50g charred Radix Scutellariae 45g

Cassia seed 45g plantain seed 45g Semen Leonuri 45g

Fruit of glossy privet 45g selfheal 45g rough gentian 45g

Root tuber of aromatic turmeric 30g scouring rush 45g radix paeoniae rubrathe 30g

Moutan bark 30g hawthorn 30g Radix Angelicae Sinensis 30g

Ligusticum wallichii 10g

The preparation method: above 19 flavors, chrysanthemum, charred Radix Scutellariae, plantain seed 22.5g, cattail pollen 37.5g, mixed powder is broken into fine powder, and is standby; Residue plantain seed, cattail pollen and all the other each flavors were soaked 30 minutes, decocted secondary, and each 2 hours, collecting decoction filtered.Filtrate decompression is concentrated into the clear cream that relative density is 1.10～1.15 (60 ℃), spray drying, and cream powder and above-mentioned fine powder and appropriate amount of auxiliary materials are made particle, add dolomol 1.5g, and mixing is pressed into 1000, the bag film-coating, namely.

Its quality determining method is as follows:

(1) get 20 of this product, remove dressing, porphyrize adds methyl alcohol 50ml, ultrasonic processing 30 minutes filters, and filtrate evaporate to dryness, residue add water 20ml makes dissolving, extract 3 times with water saturated normal butyl alcohol, each 20ml merges normal butyl alcohol liquid, evaporate to dryness, residue adds methyl alcohol 2ml makes dissolving, admixes neutral alumina 3g, and drying with water bath is added in neutral alumina post (100～200 orders, 2g, internal diameter are 1cm) on, with methyl alcohol 60ml wash-out, collect eluent, evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution.Other gets the Paeoniflorin reference substance, adds methyl alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, be developping agent with methenyl choloride-ethyl acetate-methyl alcohol-formic acid (30:3:5:0.1), launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, and it is clear to be heated to the spot colour developing at 105 ℃.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color.

(2) getting Paeoniflorin differentiates and a following need testing solution to add methyl alcohol 2ml dilution afterwards as need testing solution.Other gets rough gentian control medicinal material 0.5g, adds methyl alcohol 20ml, and ultrasonic processing 30 minutes filters, and filtrate evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, in contrast medicinal material solution.According to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B) test, draw each 2 μ l of above-mentioned two kinds of solution, put in same silica G F respectively ₂₅₄On the thin layer plate, be developping agent with methenyl choloride-methanol-water (20:5:1) lower floor solution, launch, take out, dry, put under the ultraviolet lamp (254nm) and inspect.In the test sample chromatogram, with the corresponding position of control medicinal material chromatogram on, show the principal spot of a same color.

(3) get the typhaneoside reference substance, add methyl alcohol and make the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw each the 1 μ l of need testing solution under reference substance solution and the Paeoniflorin discriminating item, put respectively on same polyamide film, be developping agent with acetone-water (3:5), launch, take out, dry, spray is placed with the aluminium choride test solution, puts under the ultraviolet lamp (365nm) and inspects.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

(4) get 10 of this product, remove dressing, porphyrize adds ethyl acetate 30ml, adds hot reflux 1 hour, filters, and filtrate evaporate to dryness, residue add ethyl acetate 1ml makes dissolving, as need testing solution.Other gets chrysanthemum control medicinal material 1g, shines medicinal material solution in pairs with legal system.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw each 2 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, be developping agent with sherwood oil (60～90 ℃)-ethyl acetate (15: 3), launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, and it is clear to be heated to the spot colour developing at 105 ℃.In the test sample chromatogram, with the corresponding position of control medicinal material chromatogram on, show the principal spot of a same color.

Embodiment 7

Prescription:

Cattail pollen 75g red sage root 75g glutinous rehmannia 60g

Eclipta 60g chrysanthemum 50g charred Radix Scutellariae 45g

Cassia seed 45g plantain seed 45g Semen Leonuri 45g

Fruit of glossy privet 45g selfheal 45g rough gentian 45g

Root tuber of aromatic turmeric 30g scouring rush 45g radix paeoniae rubrathe 30g

Moutan bark 30g hawthorn 30g Radix Angelicae Sinensis 30g

Ligusticum wallichii 10g

The preparation method: above 19 flavors, chrysanthemum, charred Radix Scutellariae, plantain seed 22.5g, cattail pollen 37.5g, mixed powder is broken into fine powder, and is standby; Residue plantain seed, cattail pollen and all the other each flavors were soaked 30 minutes, decocted secondary, and each 2 hours, collecting decoction filtered.Filtrate decompression is concentrated into the clear cream that relative density is 1.10～1.15 (60 ℃), spray drying, and cream powder and above-mentioned fine powder and appropriate amount of auxiliary materials are made particle, add dolomol 1.5g, and mixing is pressed into 1000, sugar coating, namely.

Its quality determining method is as follows:

Measure according to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 D).

Chromatographic condition and system suitability test are filling agent with the octadecylsilane chemically bonded silica; Be the phase that flows with methyl alcohol-0.5% acetic acid (8:92); The detection wavelength is 281nm.Number of theoretical plate calculates by the Sodium Danshensu peak should be not less than 1500.

It is an amount of that the Sodium Danshensu reference substance is got in the preparation of reference substance solution, adds 50% methyl alcohol and make every 1ml and contain Sodium Danshensu 25 μ g(and be equivalent to contain danshensu 22.5 μ g among every 1ml) solution, namely.

20 of this product are got in the preparation of need testing solution, remove dressing, and accurate the title decided porphyrize, get about 0.5g, the accurate title, decide, and puts in the tool plug conical flask, the accurate 50% methyl alcohol 25ml that adds, close plug claims to decide weight, ultrasonic processing (power 250W, frequency 40 kHz) 10 minutes is put cold, claim again to decide weight, supply the weight that subtracts mistake with 50% methyl alcohol, shake up, filter, get subsequent filtrate, namely.

Determination method is accurate reference substance solution and each 10 μ l of need testing solution of drawing respectively, injects liquid chromatograph, measures, namely.

Embodiment 8

Prescription:

Cattail pollen 75g red sage root 75g glutinous rehmannia 60g

Eclipta 60g chrysanthemum 50g charred Radix Scutellariae 45g

Cassia seed 45g plantain seed 45g Semen Leonuri 45g

Fruit of glossy privet 45g selfheal 45g rough gentian 45g

Root tuber of aromatic turmeric 30g scouring rush 45g radix paeoniae rubrathe 30g

Moutan bark 30g hawthorn 30g Radix Angelicae Sinensis 30g

Ligusticum wallichii 10g

The preparation method: above 19 flavors, chrysanthemum, charred Radix Scutellariae, plantain seed 22.5g, cattail pollen 37.5g, mixed powder is broken into fine powder, and is standby; Residue plantain seed, cattail pollen and all the other each flavors were soaked 30 minutes, decocted secondary, and each 2 hours, collecting decoction filtered.Filtrate decompression is concentrated into the clear cream that relative density is 1.10～1.15 (60 ℃), spray drying, and cream powder and above-mentioned fine powder and appropriate amount of auxiliary materials are made particle, add dolomol 1.5g, and mixing is pressed into 1000, sugar coating, namely.

Its quality determining method is as follows:

Measure according to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 D).

Chromatographic condition and system suitability test are filling agent with the octadecylsilane chemically bonded silica; Be the phase that flows with methyl alcohol-0.5% acetic acid (6:90); The detection wavelength is 281nm.Number of theoretical plate calculates by the Sodium Danshensu peak should be not less than 1500.

It is an amount of that the Sodium Danshensu reference substance is got in the preparation of reference substance solution, adds 50% methyl alcohol and make every 1ml and contain Sodium Danshensu 25 μ g(and be equivalent to contain danshensu 22.5 μ g among every 1ml) solution, namely.

20 of this product are got in the preparation of need testing solution, remove dressing, and accurate the title decided porphyrize, get about 0.5g, the accurate title, decide, and puts in the tool plug conical flask, the accurate 50% methyl alcohol 25ml that adds, close plug claims to decide weight, ultrasonic processing (power 250W, frequency 40 kHz) 10 minutes is put cold, claim again to decide weight, supply the weight that subtracts mistake with 50% methyl alcohol, shake up, filter, get subsequent filtrate, namely.

Determination method is accurate reference substance solution and each 10 μ l of need testing solution of drawing respectively, injects liquid chromatograph, measures, namely.

Embodiment 9

Prescription:

Cattail pollen 75g red sage root 75g glutinous rehmannia 60g

Eclipta 60g chrysanthemum 50g charred Radix Scutellariae 45g

Cassia seed 45g plantain seed 45g Semen Leonuri 45g

Fruit of glossy privet 45g selfheal 45g rough gentian 45g

Root tuber of aromatic turmeric 30g scouring rush 45g radix paeoniae rubrathe 30g

Moutan bark 30g hawthorn 30g Radix Angelicae Sinensis 30g

Ligusticum wallichii 10g

The preparation method: above 19 flavors, chrysanthemum, charred Radix Scutellariae, plantain seed 22.5g, cattail pollen 37.5g, mixed powder is broken into fine powder, and is standby; Residue plantain seed, cattail pollen and all the other each flavors were soaked 30 minutes, decocted secondary, and each 2 hours, collecting decoction filtered.Filtrate decompression is concentrated into the clear cream that relative density is 1.10～1.15 (60 ℃), spray drying, and cream powder and above-mentioned fine powder and appropriate amount of auxiliary materials are made particle, add dolomol 1.5g, and mixing is pressed into 1000, sugar coating, namely.

Its quality determining method is as follows:

(1) get this product, put microscopically and observe: pollen granule similar round or ellipse, about 17～29 μ m of diameter, there is netted carving line (cattail pollen) on the surface.Bast fiber is faint yellow, fusiformis, wall thickness, hole ditch thin (root of large-flowered skullcap).Plant the intracutaneous epidermal surface and see rectangle like, wall is thin, and wavy is often made zyklopisch and arranged (plantain seed).The pollen granule similar round, diameter 24～34 μ m, outer wall spinosity, long 3～5 μ m of thorn, 3 germinal aperatures of tool (chrysanthemum).

(2) get 10 of this product, remove dressing, porphyrize adds methyl alcohol 20ml, and ultrasonic processing 20 minutes filters, and filtrate evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, gets supernatant as need testing solution.Other gets the scutelloside reference substance, adds methyl alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), 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 binder with 4% sodium acetate solution, be developping agent with ethyl acetate-butanone-formic acid-water (5:3:1:1), launch, take out, dry, spray is with 1% ferric trichloride ethanolic solution.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color.

(3) get 10 of this product, remove dressing, porphyrize, add methyl alcohol 20ml, ultrasonic processing 20 minutes filters, filtrate evaporate to dryness, residue add water 10ml makes dissolving, adds hydrochloric acid 1ml again, water-bath added hot reflux 30 minutes, and cooling is immediately extracted 3 times with the ether jolting, each 15ml merges ether solution, and low temperature volatilizes ether, residue adds methyl alcohol 1ml makes dissolving, as need testing solution.Other gets the Chrysophanol reference substance, adds methyl alcohol and makes the solution that every 1ml contains 0.5mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw need testing solution 5 μ l, reference substance solution 1 μ l, put respectively on same silica gel g thin-layer plate, upper solution with sherwood oil (60～90 ℃)-ethyl formate-formic acid (15:5:1) is developping agent, launch, take out, dry, put under the ultraviolet lamp (365nm) and inspect.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

(4) get 20 of this product, remove dressing, porphyrize adds methyl alcohol 50ml, ultrasonic processing 30 minutes filters, and filtrate evaporate to dryness, residue add water 20ml makes dissolving, extract 3 times with water saturated normal butyl alcohol, each 20ml merges normal butyl alcohol liquid, evaporate to dryness, residue adds methyl alcohol 2ml makes dissolving, admixes neutral alumina 3g, and drying with water bath is added in neutral alumina post (100～200 orders, 2g, internal diameter are 1cm) on, with methyl alcohol 60ml wash-out, collect eluent, evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution.Other gets the Paeoniflorin reference substance, adds methyl alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, be developping agent with methenyl choloride-ethyl acetate-methyl alcohol-formic acid (40:5:10:0.2), launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, and it is clear to be heated to the spot colour developing at 105 ℃.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color.

(5) getting Paeoniflorin differentiates and a following need testing solution to add methyl alcohol 2ml dilution afterwards as need testing solution.Other gets rough gentian control medicinal material 0.5g, adds methyl alcohol 20ml, and ultrasonic processing 30 minutes filters, and filtrate evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, in contrast medicinal material solution.According to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B) test, draw each 2 μ l of above-mentioned two kinds of solution, put in same silica G F respectively ₂₅₄On the thin layer plate, be developping agent with methenyl choloride-methanol-water (30:10:3) lower floor solution, launch, take out, dry, put under the ultraviolet lamp (254nm) and inspect.In the test sample chromatogram, with the corresponding position of control medicinal material chromatogram on, show the principal spot of a same color.

(6) get the typhaneoside reference substance, add methyl alcohol and make the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw each the 1 μ l of need testing solution under reference substance solution and the Paeoniflorin discriminating item, put respectively on same polyamide film, be developping agent with acetone-water (1:2), launch, take out, dry, spray is placed with the aluminium choride test solution, puts under the ultraviolet lamp (365nm) and inspects.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

(7) get 10 of this product, remove dressing, porphyrize adds ethyl acetate 30ml, adds hot reflux 1 hour, filters, and filtrate evaporate to dryness, residue add ethyl acetate 1ml makes dissolving, as need testing solution.Other gets chrysanthemum control medicinal material 1g, shines medicinal material solution in pairs with legal system.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw each 2 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, be developping agent with sherwood oil (60～90 ℃)-ethyl acetate (10: 1), launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, and it is clear to be heated to the spot colour developing at 105 ℃.In the test sample chromatogram, with the corresponding position of control medicinal material chromatogram on, show the principal spot of a same color.

(8) [assay] measured according to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 D).

Chromatographic condition and system suitability test are filling agent with the octadecylsilane chemically bonded silica; Be the phase that flows with methyl alcohol-0.5% acetic acid (8:92); The detection wavelength is 281nm.Number of theoretical plate calculates by the Sodium Danshensu peak should be not less than 1500.

It is an amount of that the Sodium Danshensu reference substance is got in the preparation of reference substance solution, adds 50% methyl alcohol and make every 1ml and contain Sodium Danshensu 25 μ g(and be equivalent to contain danshensu 22.5 μ g among every 1ml) solution, namely.

20 of this product are got in the preparation of need testing solution, remove dressing, and accurate the title decided porphyrize, get about 0.5g, the accurate title, decide, and puts in the tool plug conical flask, the accurate 50% methyl alcohol 25ml that adds, close plug claims to decide weight, ultrasonic processing (power 250W, frequency 40 kHz) 10 minutes is put cold, claim again to decide weight, supply the weight that subtracts mistake with 50% methyl alcohol, shake up, filter, get subsequent filtrate, namely.

Determination method is accurate reference substance solution and each 10 μ l of need testing solution of drawing respectively, injects liquid chromatograph, measures, namely.

Embodiment 10

Prescription:

Cattail pollen 75g red sage root 75g glutinous rehmannia 60g

Eclipta 60g chrysanthemum 50g charred Radix Scutellariae 45g

Cassia seed 45g plantain seed 45g Semen Leonuri 45g

Fruit of glossy privet 45g selfheal 45g rough gentian 45g

Root tuber of aromatic turmeric 30g scouring rush 45g radix paeoniae rubrathe 30g

Moutan bark 30g hawthorn 30g Radix Angelicae Sinensis 30g

Ligusticum wallichii 10g

The preparation method: above 19 flavors, chrysanthemum, charred Radix Scutellariae, plantain seed 22.5g, cattail pollen 37.5g, mixed powder is broken into fine powder, and is standby; Residue plantain seed, cattail pollen and all the other each flavors were soaked 30 minutes, decocted secondary, and each 2 hours, collecting decoction filtered.Filtrate decompression is concentrated into the clear cream that relative density is 1.10～1.15 (60 ℃), spray drying, and cream powder and above-mentioned fine powder and appropriate amount of auxiliary materials are made particle, add dolomol 1.5g, and mixing is pressed into 1000, sugar coating, namely.

Its quality determining method is as follows:

(1) get this product, put microscopically and observe: pollen granule similar round or ellipse, about 17～29 μ m of diameter, there is netted carving line (cattail pollen) on the surface.Bast fiber is faint yellow, fusiformis, wall thickness, hole ditch thin (root of large-flowered skullcap).Plant the intracutaneous epidermal surface and see rectangle like, wall is thin, and wavy is often made zyklopisch and arranged (plantain seed).The pollen granule similar round, diameter 24～34 μ m, outer wall spinosity, long 3～5 μ m of thorn, 3 germinal aperatures of tool (chrysanthemum).

(2) get 10 of this product, remove dressing, porphyrize adds methyl alcohol 20ml, and ultrasonic processing 20 minutes filters, and filtrate evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, gets supernatant as need testing solution.Other gets the scutelloside reference substance, adds methyl alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), 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 binder with 4% sodium acetate solution, be developping agent with ethyl acetate-butanone-formic acid-water (4:1:0.5:0.5), launch, take out, dry, spray is with 1% ferric trichloride ethanolic solution.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color.

(3) get 10 of this product, remove dressing, porphyrize, add methyl alcohol 20ml, ultrasonic processing 20 minutes filters, filtrate evaporate to dryness, residue add water 10ml makes dissolving, adds hydrochloric acid 1ml again, water-bath added hot reflux 30 minutes, and cooling is immediately extracted 3 times with the ether jolting, each 15ml merges ether solution, and low temperature volatilizes ether, residue adds methyl alcohol 1ml makes dissolving, as need testing solution.Other gets the Chrysophanol reference substance, adds methyl alcohol and makes the solution that every 1ml contains 0.5mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw need testing solution 5 μ l, reference substance solution 1 μ l, put respectively on same silica gel g thin-layer plate, upper solution with sherwood oil (60～90 ℃)-ethyl formate-formic acid (20:7:2) is developping agent, launch, take out, dry, put under the ultraviolet lamp (365nm) and inspect.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

(4) get 20 of this product, remove dressing, porphyrize adds methyl alcohol 50ml, ultrasonic processing 30 minutes filters, and filtrate evaporate to dryness, residue add water 20ml makes dissolving, extract 3 times with water saturated normal butyl alcohol, each 20ml merges normal butyl alcohol liquid, evaporate to dryness, residue adds methyl alcohol 2ml makes dissolving, admixes neutral alumina 3g, and drying with water bath is added in neutral alumina post (100～200 orders, 2g, internal diameter are 1cm) on, with methyl alcohol 60ml wash-out, collect eluent, evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution.Other gets the Paeoniflorin reference substance, adds methyl alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, be developping agent with methenyl choloride-ethyl acetate-methyl alcohol-formic acid (30:3:5:0.1), launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, and it is clear to be heated to the spot colour developing at 105 ℃.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color.

(5) getting Paeoniflorin differentiates and a following need testing solution to add methyl alcohol 2ml dilution afterwards as need testing solution.Other gets rough gentian control medicinal material 0.5g, adds methyl alcohol 20ml, and ultrasonic processing 30 minutes filters, and filtrate evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, in contrast medicinal material solution.According to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B) test, draw each 2 μ l of above-mentioned two kinds of solution, put in same silica G F respectively ₂₅₄On the thin layer plate, be developping agent with methenyl choloride-methanol-water (20:5:1) lower floor solution, launch, take out, dry, put under the ultraviolet lamp (254nm) and inspect.In the test sample chromatogram, with the corresponding position of control medicinal material chromatogram on, show the principal spot of a same color.

(6) get the typhaneoside reference substance, add methyl alcohol and make the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw each the 1 μ l of need testing solution under reference substance solution and the Paeoniflorin discriminating item, put respectively on same polyamide film, be developping agent with acetone-water (3:5), launch, take out, dry, spray is placed with the aluminium choride test solution, puts under the ultraviolet lamp (365nm) and inspects.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

(7) get 10 of this product, remove dressing, porphyrize adds ethyl acetate 30ml, adds hot reflux 1 hour, filters, and filtrate evaporate to dryness, residue add ethyl acetate 1ml makes dissolving, as need testing solution.Other gets chrysanthemum control medicinal material 1g, shines medicinal material solution in pairs with legal system.Test according to thin-layered chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 B), draw each 2 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, be developping agent with sherwood oil (60～90 ℃)-ethyl acetate (15: 3), launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, and it is clear to be heated to the spot colour developing at 105 ℃.In the test sample chromatogram, with the corresponding position of control medicinal material chromatogram on, show the principal spot of a same color.

(8) [assay] measured according to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2010 D).

Chromatographic condition and system suitability test are filling agent with the octadecylsilane chemically bonded silica; Be the phase that flows with methyl alcohol-0.5% acetic acid (6:90); The detection wavelength is 281nm.Number of theoretical plate calculates by the Sodium Danshensu peak should be not less than 1500.

It is an amount of that the Sodium Danshensu reference substance is got in the preparation of reference substance solution, adds 50% methyl alcohol and make every 1ml and contain Sodium Danshensu 25 μ g(and be equivalent to contain danshensu 22.5 μ g among every 1ml) solution, namely.

20 of this product are got in the preparation of need testing solution, remove dressing, and accurate the title decided porphyrize, get about 0.5g, the accurate title, decide, and puts in the tool plug conical flask, the accurate 50% methyl alcohol 25ml that adds, close plug claims to decide weight, ultrasonic processing (power 250W, frequency 40 kHz) 10 minutes is put cold, claim again to decide weight, supply the weight that subtracts mistake with 50% methyl alcohol, shake up, filter, get subsequent filtrate, namely.

Determination method is accurate reference substance solution and each 10 μ l of need testing solution of drawing respectively, injects liquid chromatograph, measures, namely.

  

Claims (10)

1. quality determining method for the treatment of the Chinese medicine preparation of fundus hemorrhage disease, this Chinese medicine preparation is by cattail pollen 8.93%, the red sage root 8.93%, glutinous rehmannia 7.14%, eclipta 7.14%, chrysanthemum 5.95%, charred Radix Scutellariae 5.36%, cassia seed 5.36%, plantain seed 5.36%, Semen Leonuri 5.36%, the fruit of glossy privet 5.36%, selfheal 5.36%, rough gentian 5.36%, root tuber of aromatic turmeric 3.57%, the scouring rush 5.36%, the radix paeoniae rubrathe 3.57%, moutan bark 3.57%, hawthorn 3.57%, Radix Angelicae Sinensis 3.57%, Ligusticum wallichii 1.19% is made, and the quality determining method that it is characterized in that this Chinese medicine preparation is micro-discrimination method, in thin layer discrimination method or the content assaying method one or more.

2. a kind of quality determining method for the treatment of the Chinese medicine preparation of fundus hemorrhage disease according to claim 1, this Chinese medicine preparation is by cattail pollen 8.93%, the red sage root 8.93%, glutinous rehmannia 7.14%, eclipta 7.14%, chrysanthemum 5.95%, charred Radix Scutellariae 5.36%, cassia seed 5.36%, plantain seed 5.36%, Semen Leonuri 5.36%, the fruit of glossy privet 5.36%, selfheal 5.36%, rough gentian 5.36%, root tuber of aromatic turmeric 3.57%, the scouring rush 5.36%, the radix paeoniae rubrathe 3.57%, moutan bark 3.57%, hawthorn 3.57%, Radix Angelicae Sinensis 3.57%, Ligusticum wallichii 1.19% is made, and the quality determining method that it is characterized in that this Chinese medicine preparation is micro-discrimination method.

3. a kind of quality determining method for the treatment of the Chinese medicine preparation of fundus hemorrhage disease according to claim 2, it is characterized in that: get this product, putting microscopically observes: (1) cattail pollen is differentiated: pollen granule similar round or ellipse, and about 17～29 μ m of diameter, there is netted carving line on the surface; (2) root of large-flowered skullcap is differentiated: bast fiber is faint yellow, fusiformis, and wall thickness, the hole ditch is thin; (3) plantain seed is differentiated: plant the intracutaneous epidermal surface and see rectangle like, wall is thin, and wavy is often made zyklopisch and arranged; (4) chrysanthemum is differentiated: pollen granule similar round, diameter 24～34 μ m, outer wall spinosity, long 3～5 μ m of thorn, 3 germinal aperatures of tool.

4. a kind of quality determining method for the treatment of the Chinese medicine preparation of fundus hemorrhage disease according to claim 1, this Chinese medicine preparation is by cattail pollen 8.93%, the red sage root 8.93%, glutinous rehmannia 7.14%, eclipta 7.14%, chrysanthemum 5.95%, charred Radix Scutellariae 5.36%, cassia seed 5.36%, plantain seed 5.36%, Semen Leonuri 5.36%, the fruit of glossy privet 5.36%, selfheal 5.36%, rough gentian 5.36%, root tuber of aromatic turmeric 3.57%, the scouring rush 5.36%, the radix paeoniae rubrathe 3.57%, moutan bark 3.57%, hawthorn 3.57%, Radix Angelicae Sinensis 3.57%, Ligusticum wallichii 1.19% is made, and the quality determining method that it is characterized in that this Chinese medicine preparation is the thin layer discrimination method.

5. a kind of quality determining method for the treatment of the Chinese medicine preparation of fundus hemorrhage disease according to claim 4 is characterized in that differentiating in scutelloside, Chrysophanol, Paeoniflorin, typhaneoside, rough gentian or the chrysanthemum in this Chinese medicine preparation two or more.

6. a kind of quality determining method for the treatment of the Chinese medicine preparation of fundus hemorrhage disease according to claim 5 is characterized in that differentiating scutelloside and Chrysophanol in this Chinese medicine preparation;

The scutelloside discrimination method:

Get this product that is equivalent to raw medicinal herbs 8.4g, porphyrize, add methyl alcohol 20ml, ultrasonic processing 20 minutes filters the filtrate evaporate to dryness, residue adds methyl alcohol 1ml makes dissolving, get supernatant as need testing solution, other gets the scutelloside reference substance, adds methyl alcohol and makes the solution that every 1ml contains 1mg, product solution in contrast, according to thin-layered chromatography 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 binder with 4% sodium acetate solution, be developping agent with ethyl acetate-butanone-formic acid-water 4～8:1～4:0.5～2:0.5～2, launch, take out, dry, spray is with 1% ferric trichloride ethanolic solution, in the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color;

The Chrysophanol discrimination method:

Get this product that is equivalent to raw medicinal herbs 8.4g, porphyrize adds methyl alcohol 20ml, ultrasonic processing 20 minutes filters the filtrate evaporate to dryness, residue adds water 10ml makes dissolving, add hydrochloric acid 1ml again, water-bath added hot reflux 30 minutes, immediately cooling, extract 3 times with the ether jolting, each 15ml merges ether solution, and low temperature volatilizes ether, residue adds methyl alcohol 1ml makes dissolving, as need testing solution, other gets the Chrysophanol reference substance, adds methyl alcohol and makes the solution that every 1ml contains 0.5mg, product solution in contrast, according to the thin-layered chromatography test, draw need testing solution 5 μ l, reference substance solution 1 μ l puts respectively on same silica gel g thin-layer plate, upper solution with 60～90 ℃ of sherwood oil-ethyl formates-formic acid 10～20:3～7:0.5～2 is developping agent, launch, take out, dry, put under the 365nm ultraviolet lamp and inspect, in the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

7. a kind of quality determining method for the treatment of the Chinese medicine preparation of fundus hemorrhage disease according to claim 5 is characterized in that differentiating Paeoniflorin, typhaneoside, rough gentian and chrysanthemum in this Chinese medicine preparation;

The Paeoniflorin discrimination method:

Get this product that is equivalent to raw medicinal herbs 16.8g, porphyrize, add methyl alcohol 50ml, ultrasonic processing 30 minutes, filter, the filtrate evaporate to dryness, residue adds water 20ml makes dissolving, extracts 3 times with water saturated normal butyl alcohol, each 20ml, merge normal butyl alcohol liquid, evaporate to dryness, residue add methyl alcohol 2ml makes dissolving, admixes neutral alumina 3g, drying with water bath, be added in 100～200 orders, internal diameter is 1cm, weight is on the neutral alumina post of 2g, with methyl alcohol 60ml wash-out, collects eluent, evaporate to dryness, residue adds methyl alcohol 1ml makes dissolving, and as need testing solution, other gets the Paeoniflorin reference substance, add methyl alcohol and make the solution that every 1ml contains 1mg, product solution according to the thin-layered chromatography test, is drawn each 5 μ l of above-mentioned two kinds of solution in contrast, put respectively on same silica gel g thin-layer plate, be developping agent with methenyl choloride-ethyl acetate-methyl alcohol-formic acid 30～40:3～5:5～10:0.1～0.2, launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, and it is clear to be heated to the spot colour developing at 105 ℃, in the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color;

The typhaneoside discrimination method:

Get the typhaneoside reference substance, add methyl alcohol and make the solution that every 1ml contains 1mg, product solution in contrast, test according to thin-layered chromatography, draw each the 1 μ l of need testing solution under reference substance solution and the Paeoniflorin discrimination method item, putting respectively on same polyamide film, is developping agent with acetone-water 1～3:2～5, launches, take out, dry, spray is placed with the aluminium choride test solution, put under the 365nm ultraviolet lamp and inspect, in the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color;

The rough gentian discrimination method:

Get the need testing solution under the Paeoniflorin discrimination method item, add methyl alcohol 2ml dilution back as need testing solution, other gets rough gentian control medicinal material 0.5g, adds methyl alcohol 20ml, ultrasonic processing 30 minutes, filter, filtrate evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, medicinal material solution in contrast, according to the thin-layered chromatography test, draw each 2 μ l of above-mentioned two kinds of solution, put in same silica G F respectively ₂₅₄On the thin layer plate, be developping agent with methenyl choloride-methanol-water 20～30:5～10:1～3 lower floor's solution, launch, take out, dry, put under the 254nm ultraviolet lamp and inspect, in the test sample chromatogram, with the corresponding position of control medicinal material chromatogram on, show the principal spot of a same color;

The chrysanthemum discrimination method:

Get this product that is equivalent to raw medicinal herbs 8.4g, porphyrize adds ethyl acetate 30ml, add hot reflux 1 hour, and filtered the filtrate evaporate to dryness, residue adds ethyl acetate 1ml makes dissolving, as need testing solution, other gets chrysanthemum control medicinal material 1g, shines medicinal material solution in pairs with legal system, test according to thin-layered chromatography, drawing each 2 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, is developping agent with 60～90 ℃ of petroleum ether-ethyl acetates 10～15: 1～3, launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, it is clear to be heated to spot colour developing at 105 ℃, in the test sample chromatogram, with the corresponding position of control medicinal material chromatogram on, show the principal spot of a same color.

8. a kind of quality determining method for the treatment of the Chinese medicine preparation of fundus hemorrhage disease according to claim 1, this Chinese medicine preparation is by cattail pollen 8.93%, the red sage root 8.93%, glutinous rehmannia 7.14%, eclipta 7.14%, chrysanthemum 5.95%, charred Radix Scutellariae 5.36%, cassia seed 5.36%, plantain seed 5.36%, Semen Leonuri 5.36%, the fruit of glossy privet 5.36%, selfheal 5.36%, rough gentian 5.36%, root tuber of aromatic turmeric 3.57%, the scouring rush 5.36%, the radix paeoniae rubrathe 3.57%, moutan bark 3.57%, hawthorn 3.57%, Radix Angelicae Sinensis 3.57%, Ligusticum wallichii 1.19% is made, and the quality determining method that it is characterized in that this Chinese medicine preparation is content assaying method.

9. a kind of quality determining method for the treatment of the Chinese medicine preparation of fundus hemorrhage disease according to claim 8 is characterized in that:

Chromatographic condition and system suitability test are filling agent with the octadecylsilane chemically bonded silica; Be the phase that flows with methyl alcohol-0.5% acetic acid 6～8:90～92; The detection wavelength is 281nm, and number of theoretical plate calculates by the Sodium Danshensu peak should be not less than 1500;

It is an amount of that the Sodium Danshensu reference substance is got in the preparation of reference substance solution, adds 50% methyl alcohol and make the solution that every 1ml contains Sodium Danshensu 25 μ g, namely;

This product that is equivalent to raw medicinal herbs 16.8g is got in the preparation of need testing solution, and accurate the title decided porphyrize, get about 0.5g, the accurate title, decide, and puts in the tool plug conical flask, the accurate 50% methyl alcohol 25ml that adds, close plug claims to decide weight, ultrasonic processing 10 minutes, power are that 250W, frequency are 40 kHz, put cold, claim again to decide weight, supply the weight that subtracts mistake with 50% methyl alcohol, shake up, filter, get subsequent filtrate, namely;

Determination method is accurate reference substance solution and each 10 μ l of need testing solution of drawing respectively, injects liquid chromatograph, measures, namely.

10. a kind of quality determining method for the treatment of the Chinese medicine preparation of fundus hemorrhage disease according to claim 1, this Chinese medicine preparation is by cattail pollen 8.93%, the red sage root 8.93%, glutinous rehmannia 7.14%, eclipta 7.14%, chrysanthemum 5.95%, charred Radix Scutellariae 5.36%, cassia seed 5.36%, plantain seed 5.36%, Semen Leonuri 5.36%, the fruit of glossy privet 5.36%, selfheal 5.36%, rough gentian 5.36%, root tuber of aromatic turmeric 3.57%, the scouring rush 5.36%, the radix paeoniae rubrathe 3.57%, moutan bark 3.57%, hawthorn 3.57%, Radix Angelicae Sinensis 3.57%, Ligusticum wallichii 1.19% is made, and it is characterized in that the quality determining method of this Chinese medicine preparation is by micro-discrimination method, thin layer discrimination method and content assaying method are formed;

Micro-discriminating:

Get this product, put microscopically and observe: (1) cattail pollen is differentiated: pollen granule similar round or ellipse, and about 17～29 μ m of diameter, there is netted carving line on the surface; (2) root of large-flowered skullcap is differentiated: bast fiber is faint yellow, fusiformis, and wall thickness, the hole ditch is thin; (3) plantain seed is differentiated: plant the intracutaneous epidermal surface and see rectangle like, wall is thin, and wavy is often made zyklopisch and arranged; (4) chrysanthemum is differentiated: pollen granule similar round, diameter 24～34 μ m, outer wall spinosity, long 3～5 μ m of thorn, 3 germinal aperatures of tool;

Thin layer is differentiated:

(1) gets this product that is equivalent to raw medicinal herbs 8.4g, porphyrize, add methyl alcohol 20ml, ultrasonic processing 20 minutes filters the filtrate evaporate to dryness, residue adds methyl alcohol 1ml makes dissolving, get supernatant as need testing solution, other gets the scutelloside reference substance, adds methyl alcohol and makes the solution that every 1ml contains 1mg, product solution in contrast, according to thin-layered chromatography 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 binder with 4% sodium acetate solution, be developping agent with ethyl acetate-butanone-formic acid-water 4～8:1～4:0.5～2:0.5～2, launch, take out, dry, spray is with 1% ferric trichloride ethanolic solution, in the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color;

(2) get this product that is equivalent to raw medicinal herbs 8.4g, porphyrize adds methyl alcohol 20ml, ultrasonic processing 20 minutes filters the filtrate evaporate to dryness, residue adds water 10ml makes dissolving, add hydrochloric acid 1ml again, water-bath added hot reflux 30 minutes, immediately cooling, extract 3 times with the ether jolting, each 15ml merges ether solution, and low temperature volatilizes ether, residue adds methyl alcohol 1ml makes dissolving, as need testing solution, other gets the Chrysophanol reference substance, adds methyl alcohol and makes the solution that every 1ml contains 0.5mg, product solution in contrast, according to the thin-layered chromatography test, draw need testing solution 5 μ l, reference substance solution 1 μ l puts respectively on same silica gel g thin-layer plate, upper solution with 60～90 ℃ of sherwood oil-ethyl formates-formic acid 10～20:3～7:0.5～2 is developping agent, launch, take out, dry, put under the 365nm ultraviolet lamp and inspect, in the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color;

(3) get this product that is equivalent to raw medicinal herbs 16.8g, porphyrize, add methyl alcohol 50ml, ultrasonic processing 30 minutes, filter, the filtrate evaporate to dryness, residue adds water 20ml makes dissolving, extracts 3 times with water saturated normal butyl alcohol, each 20ml, merge normal butyl alcohol liquid, evaporate to dryness, residue add methyl alcohol 2ml makes dissolving, admixes neutral alumina 3g, drying with water bath, be added in 100～200 orders, internal diameter is 1cm, weight is on the neutral alumina post of 2g, with methyl alcohol 60ml wash-out, collects eluent, evaporate to dryness, residue adds methyl alcohol 1ml makes dissolving, and as need testing solution, other gets the Paeoniflorin reference substance, add methyl alcohol and make the solution that every 1ml contains 1mg, product solution according to the thin-layered chromatography test, is drawn each 5 μ l of above-mentioned two kinds of solution in contrast, put respectively on same silica gel g thin-layer plate, be developping agent with methenyl choloride-ethyl acetate-methyl alcohol-formic acid 30～40:3～5:5～10:0.1～0.2, launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, and it is clear to be heated to the spot colour developing at 105 ℃, in the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the spot of same color;

(4) get the typhaneoside reference substance, add methyl alcohol and make the solution that every 1ml contains 1mg, product solution in contrast, test according to thin-layered chromatography, draw each the 1 μ l of need testing solution under reference substance solution and the Paeoniflorin discrimination method item, putting respectively on same polyamide film, is developping agent with acetone-water 1～3:2～5, launches, take out, dry, spray is placed with the aluminium choride test solution, put under the 365nm ultraviolet lamp and inspect, in the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color;

(5) get need testing solution under the Paeoniflorin discrimination method item, add methyl alcohol 2ml dilution back as need testing solution, other gets rough gentian control medicinal material 0.5g, adds methyl alcohol 20ml, ultrasonic processing 30 minutes, filter, filtrate evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, medicinal material solution in contrast, according to the thin-layered chromatography test, draw each 2 μ l of above-mentioned two kinds of solution, put in same silica G F respectively ₂₅₄On the thin layer plate, be developping agent with methenyl choloride-methanol-water 20～30:5～10:1～3 lower floor's solution, launch, take out, dry, put under the 254nm ultraviolet lamp and inspect, in the test sample chromatogram, with the corresponding position of control medicinal material chromatogram on, show the principal spot of a same color;

(6) get this product that is equivalent to raw medicinal herbs 8.4g, porphyrize adds ethyl acetate 30ml, add hot reflux 1 hour, and filtered the filtrate evaporate to dryness, residue adds ethyl acetate 1ml makes dissolving, as need testing solution, other gets chrysanthemum control medicinal material 1g, shines medicinal material solution in pairs with legal system, test according to thin-layered chromatography, drawing each 2 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, is developping agent with 60～90 ℃ of petroleum ether-ethyl acetates 10～15: 1～3, launch, take out, dry, spray is with 2% vanillic aldehyde sulfuric acid solution, it is clear to be heated to spot colour developing at 105 ℃, in the test sample chromatogram, with the corresponding position of control medicinal material chromatogram on, show the principal spot of a same color;

Content assaying method:

Chromatographic condition and system suitability test are filling agent with the octadecylsilane chemically bonded silica; Be the phase that flows with methyl alcohol-0.5% acetic acid 6～8:90～92; The detection wavelength is 281nm, and number of theoretical plate calculates by the Sodium Danshensu peak should be not less than 1500;

It is an amount of that the Sodium Danshensu reference substance is got in the preparation of reference substance solution, adds 50% methyl alcohol and make the solution that every 1ml contains Sodium Danshensu 25 μ g, namely;

This product that is equivalent to raw medicinal herbs 16.8g is got in the preparation of need testing solution, and accurate the title decided porphyrize, get about 0.5g, the accurate title, decide, and puts in the tool plug conical flask, the accurate 50% methyl alcohol 25ml that adds, close plug claims to decide weight, ultrasonic processing 10 minutes, power are that 250W, frequency are 40 kHz, put cold, claim again to decide weight, supply the weight that subtracts mistake with 50% methyl alcohol, shake up, filter, get subsequent filtrate, namely;

Determination method is accurate reference substance solution and each 10 μ l of need testing solution of drawing respectively, injects liquid chromatograph, measures, namely.