CN101574477A - Method for controlling quality of menstruation regulation and macula removal tablet - Google Patents

Abstract

The invention discloses a method for controlling the quality of a traditional Chinese medicine preparation-menstruation regulation and macula removal tablet. The quality control method includes the steps of adopting the thin-layer chromatography to identify fruit of glossy privet, medlar, angelica, radix paeoniae alba and radix bupleuri and adopting the high performance liquid chromatography to determine the content of paeoniflorin in the preparation. The quality control method is reliable and feasible, and can guarantee the product quality and the clinical efficacy.

Description

The method of quality control of menstruation regulation and macula removal tablet

Invention field

The present invention relates to a kind of method of quality control of Chinese medicine preparation, be specifically related to the method for quality control of menstruation regulation and macula removal tablet.

Background technology

Menstruation regulation and macula removal tablet is that menstruation-regulating and speckle-dispelling capsule dosage changing form comes, prescription records in " Chinese patent medicine provincial standard rising national standard " (surgery gynecological fascicle) 582 pages, standard code is WS-10710 (ZD-0710)-2002, has nourishing blood for regulating menstruation, the effect of dissolving stasis and dispelling spots, be used for nutrient QI and blood being insufficient, the menorrhagia of caused by energy stagnation and blood stasis, chloasma.Its prescription is: Radix Astragali 80g, Semen Cuscutae 40g, Herba Ecliptae 50g, Fructus Ligustri Lucidi 40g, Fructus Lycii 40g, Radix Angelicae Sinensis 40g, Radix Paeoniae Alba 50g, Radix Polygoni Multiflori 40g, Radix Rehmanniae 50g, Radix Rehmanniae Preparata 60g, Semen Persicae 30g, Radix Bupleuri 30g, Colla Corii Asini 12g, Rhizoma Gymnadeniae 40g, Flos Carthami 10g; The tablet method for making is: above ten five tastes medical materials, and to get Fructus Ligustri Lucidi, Fructus Lycii, Radix Angelicae Sinensis, the Radix Paeoniae Alba, Radix Bupleuri, Colla Corii Asini, be broken into fine powder in ginseng, Flos Carthami powder, seven flavor medicine materials such as all the other Semen Cuscutae decoct with water secondary, each 2 hours, collecting decoction filtered, filtrate is concentrated into the thick paste that relative density is 1.30～1.35 (50 ℃), adds above-mentioned fine powder, oven dry, add right amount of auxiliary materials such as calcium hydrogen phosphate, mixing is granulated, dry, tabletting, the bag film-coat, promptly.Pulverize, make granule, incapsulate, promptly.

Exist in the former menstruation-regulating and speckle-dispelling capsule standard Fructus Ligustri Lucidi thin layer differentiate in test sample chromatographic isolation degree poor; The Fructus Lycii thin layer has used chloroform in differentiating, toxicity is bigger; The flavour of a drug of differentiating in the side are less; Be difficult to problems such as filtration when test liquid prepares in the content assaying method, need optimize all sidedly and replenish quality control method.The menstruation regulation and macula removal tablet extraction process is consistent with capsule, but preparation process is different with adjuvant, needs to formulate truly feasible method of quality control to guarantee product quality and clinical efficacy.

Summary of the invention

The object of the invention is to provide a kind of method of quality control of menstruation regulation and macula removal tablet, and then guarantees product quality and clinical efficacy.

The invention solves the deficiency that exists in the existing menstruation-regulating and speckle-dispelling quality of the pharmaceutical preparations control method, the Fructus Ligustri Lucidi thin layer has been changed developing solvent in differentiating, has improved the Thin-layer separation effect; During differentiating, used the Fructus Lycii thin layer hypotoxic solvent to substitute the big chloroform of toxicity; The thin layer chromatography that has increased the Radix Bupleuri and the Radix Paeoniae Alba is differentiated; Solved problems such as being difficult to filtration when test liquid prepares in the content assaying method, the quality of the pharmaceutical preparations is well controlled.

The menstruation regulation and macula removal tablet method of quality control comprises following all or part of content: thin layer is differentiated Fructus Ligustri Lucidi; Thin layer is differentiated Fructus Lycii; Thin layer is differentiated Radix Angelicae Sinensis; Thin layer is differentiated the Radix Paeoniae Alba; Thin layer is differentiated Radix Bupleuri; Measure content of paeoniflorin in this product.

During the menstruation regulation and macula removal tablet method of quality control may further comprise the steps one or more:

(1) thin layer is differentiated Fructus Ligustri Lucidi: it is an amount of to get this product, removes film-coat, and porphyrize adds alcohol heating reflux, puts coldly, filters, and filtrate is concentrated into 2ml, as need testing solution.It is an amount of that other gets the Fructus Ligustri Lucidi control medicinal material, shines medical material solution in pairs with legal system.Even up pier fruit acid reference substance again, add ethanol and make the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw above-mentioned three kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene-ethyl acetate-formic acid proper proportion is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with control medicinal material chromatograph and the corresponding position of reference substance chromatograph on, show the speckle of same color.

(2) thin layer is differentiated Fructus Lycii: it is an amount of to get this product, removes film-coat, and porphyrize adds the water heated and boiled, puts coldly, filters, and filtrate is extracted with the ethyl acetate jolting, and acetic acid ethyl fluid is concentrated into 1ml, as need testing solution.It is an amount of that other gets the Fructus Lycii control medicinal material, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene-ethyl acetate-formic acid proper proportion is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(3) thin layer is differentiated Radix Angelicae Sinensis: it is an amount of to get this product, removes film-coat, porphyrize, and the supersound process that adds diethyl ether filters, and filtrate volatilizes, and residue adds methanol 2ml makes dissolving, as need testing solution.It is an amount of that other gets the Radix Angelicae Sinensis control medicinal material, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with petroleum ether (60～90 ℃)-ethyl acetate (17: 2) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(4) thin layer is differentiated the Radix Paeoniae Alba: it is an amount of to get this product, removes film-coat, porphyrize, add the methanol supersound process, filter, filtrate evaporate to dryness, residue add the saturated water dissolution of n-butyl alcohol, put in the separatory funnel, extract with water saturated n-butyl alcohol jolting, divide and get n-butyl alcohol liquid, evaporate to dryness, residue adds ethanol 1ml makes dissolving, as need testing solution.Other gets the peoniflorin reference substance, adds ethanol and makes the solution that every 1ml contains 2mg, in contrast product solution.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw need testing solution and reference substance solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with chloroform-ethyl acetate-methanol-formic acid proper proportion is developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.

(5) thin layer is differentiated Radix Bupleuri: it is an amount of to get this product, removes film-coat, porphyrize, and it is an amount of to add methanol, supersound process filters the filtrate evaporate to dryness, residue adds the saturated water dissolution of n-butyl alcohol, puts in the separatory funnel, extracts with water saturated n-butyl alcohol jolting, divide and get n-butyl alcohol liquid, wash in right amount, discard the ammonia washing liquid with ammonia solution, the water that the reuse n-butyl alcohol is saturated washs in right amount, discards water lotion, divides and gets n-butyl alcohol liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.It is an amount of that other gets the Radix Bupleuri control medicinal material, adds the methanol supersound process, filters, and filtrate evaporate to dryness, residue add water saturated n-butyl alcohol makes dissolving, with the ammonia solution washing, discards the ammonia washing liquid, divides and get n-butyl alcohol liquid, and evaporate to dryness, residue add ethanol 1ml makes dissolving, in contrast medical material solution.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with chloroform-methanol-water proper proportion is developing solvent, launch, take out, dry, spray is with 40% sulfuric acid solution of 2% paradime thylaminobenzaldehyde, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color; Put under the ultra-violet lamp (365nm) and inspect, show the fluorescence speckle of same color.

(6) measure content of paeoniflorin in this product:

Measure according to high performance liquid chromatography (the corresponding appendix of Chinese Pharmacopoeia).

Chromatographic condition and system suitability test are filler with octadecylsilane chemically bonded silica; Methanol-water (25: 75) is a mobile phase; The detection wavelength is 230nm.Number of theoretical plate calculates by peoniflorin should be not less than 2000.

It is an amount of that the preparation precision of reference substance solution takes by weighing the peoniflorin reference substance, makes the solution that every 1ml contains 0.1mg with Diluted Alcohol, promptly.

This product is got in the preparation of need testing solution, removes film-coat, and accurate the title decided porphyrize, get in right amount, the accurate title, decide, and puts in the tool plug conical flask, accurate adding Diluted Alcohol 50ml, close plug claims to decide weight, supersound process, put coldly, claim again to decide weight, supply the weight that subtracts mistake with Diluted Alcohol, shake up, centrifugal, get supernatant, filter, get subsequent filtrate, promptly.

Accurate respectively reference substance solution and the need testing solution drawn of algoscopy injects chromatograph of liquid, measures, promptly.

During the method for quality control of menstruation regulation and macula removal tablet may further comprise the steps one or more:

(1) thin layer is differentiated Fructus Ligustri Lucidi: get 5～30 of this product, remove film-coat, porphyrize adds ethanol 10～50ml, and reflux 10～60 minutes is put coldly, filters, and filtrate is concentrated into 2ml, as need testing solution.Other gets Fructus Ligustri Lucidi control medicinal material 0.2～1g, shines medical material solution in pairs with legal system.Even up pier fruit acid reference substance again, add ethanol and make the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw each 2～10 μ l of above-mentioned three kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene-ethyl acetate-formic acid (15～25: 5～10: 0.2～0.8) be developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and 100 ℃～120 ℃ to be heated to the speckle colour developing clear.In the test sample chromatograph, with control medicinal material chromatograph and the corresponding position of reference substance chromatograph on, show the speckle of same color.

(2) thin layer is differentiated Fructus Lycii: get 5～30 of this product, remove film-coat, porphyrize adds water 50～200ml, heated and boiled 10～60 minutes is put coldly, filters, and filtrate is extracted 1～3 time with the ethyl acetate jolting, each 10～50ml, combined ethyl acetate liquid is concentrated into 1ml, as need testing solution.Other gets Fructus Lycii control medicinal material 0.5～2g, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw each 2～10 μ l of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene-ethyl acetate-formic acid (15～25: 5～10: 0.2～0.8) be developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(3) thin layer is differentiated Radix Angelicae Sinensis: get 5～30 of this product, remove film-coat, and porphyrize, the 20～100ml that adds diethyl ether, supersound process 10～60 minutes filters, and filtrate volatilizes, and residue adds methanol 2ml makes dissolving, as need testing solution.Other gets Radix Angelicae Sinensis control medicinal material 0.5～2g, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw each 2～10 μ l of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with petroleum ether (60～90 ℃)-ethyl acetate (17: 2) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(4) thin layer is differentiated the Radix Paeoniae Alba: get 5～30 of this product, remove film-coat, porphyrize, add methanol 20～100ml, supersound process 10～60 minutes filters, filtrate evaporate to dryness, residue add the saturated water 10～50ml gradation dissolving of n-butyl alcohol, put in the separatory funnel, extract 1～3 time with water saturated n-butyl alcohol jolting, each 10～50ml divides and gets n-butyl alcohol liquid, evaporate to dryness, residue adds ethanol 1ml makes dissolving, as need testing solution.Other gets the peoniflorin reference substance, adds ethanol and makes the solution that every 1ml contains 2mg, in contrast product solution.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw need testing solution and reference substance solution 2～10 μ l, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with chloroform-ethyl acetate-methanol-formic acid (35～45: 3～7: 8～12: 0.1～0.5) be developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and 100 ℃～120 ℃ to be heated to the speckle colour developing clear.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.

(5) thin layer is differentiated Radix Bupleuri: get 10～50 of this product, remove film-coat, porphyrize adds methanol 20～100ml, supersound process 10～60 minutes, filter, filtrate evaporate to dryness, residue add the saturated water 10～50ml gradation dissolving of n-butyl alcohol, put in the separatory funnel, extract 1～3 time with water saturated n-butyl alcohol jolting, each 10～50ml divides and gets n-butyl alcohol liquid, wash with ammonia solution 10～100ml, discard the ammonia washing liquid, water 10～100ml washing that the reuse n-butyl alcohol is saturated discards water lotion, divide and get n-butyl alcohol liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.Other gets Radix Bupleuri control medicinal material 0.2～2g, add methanol 10～50ml, supersound process 10～60 minutes filters, the filtrate evaporate to dryness, residue adds water saturated n-butyl alcohol 10～50ml makes dissolving, with ammonia solution 10～50ml washing, discards the ammonia washing liquid, divide and get n-butyl alcohol liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, in contrast medical material solution.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw above-mentioned two kinds of solution, 2～10 μ l, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with chloroform-methanol-water (25～35: 8～12: 0.5～1.5) be developing solvent, launch, take out, dry, spray is with 40% sulfuric acid solution of 2% paradime thylaminobenzaldehyde, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color; Put under the ultra-violet lamp (365nm) and inspect, show the fluorescence speckle of same color.

(6) measure content of paeoniflorin in this product:

Measure according to high performance liquid chromatography (the corresponding appendix of Chinese Pharmacopoeia).

Chromatographic condition and system suitability test are filler with octadecylsilane chemically bonded silica; Methanol-water (25: 75) is a mobile phase; The detection wavelength is 230nm.Number of theoretical plate calculates by peoniflorin should be not less than 2000.

It is an amount of that the preparation precision of reference substance solution takes by weighing the peoniflorin reference substance, makes the solution that every 1ml contains 0.1mg with Diluted Alcohol, promptly.

This product is got in the preparation of need testing solution, removes film-coat, and accurate the title decided porphyrize, get in right amount, the accurate title, decide, and puts in the tool plug conical flask, accurate adding Diluted Alcohol 50ml, close plug claims to decide weight, supersound process, put coldly, claim again to decide weight, supply the weight that subtracts mistake with Diluted Alcohol, shake up, centrifugal 5～30 minutes, get supernatant, filter, get subsequent filtrate, promptly.

Accurate respectively reference substance solution and each 5～20 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly.

The specific embodiment

Method of quality control of the present invention is the preferred plan that obtains through a large amount of screenings, and following experimental example is used to further specify technical scheme of the present invention and technique effect.

Experimental example 1: for the thin layer chromatography of Fructus Ligustri Lucidi and characteristic component oleanolic acid thereof is differentiated

Developing solvent that the present invention is preferred.Also investigated following development system: 1. 2. cyclohexane extraction-acetone-ethyl acetate-methanol (5: 2: 1: 0.5) 3. toluene-ethyl acetate-formic acid (20: 4: 0.5) of cyclohexane extraction-acetone-ethyl acetate (5: 2: 1), result of the test shows that development system 1. 2. can't be with test sample and each component separating of medical material, and 3. development system also can reach separating effect.

Test with menstruation regulation and macula removal tablet by embodiment 1 Fructus Ligustri Lucidi thin layer discrimination method, as a result in the test sample chromatograph, with control medicinal material chromatograph and the corresponding position of reference substance chromatograph on, show identical aubergine speckle.This method thin layer chromatography good separating effect, clear spot, rounding, Rf value is moderate, and repeatability and specificity are good, and negative control is noiseless.

Experimental example 2: for the thin layer chromatography of Fructus Lycii is differentiated

Once following method for extraction and purification had been carried out contrast test: 1. get 16 of this product, remove film-coat, porphyrize adds water 50ml, and heated and boiled 15 minutes is put coldly, filters, and filtrate is extracted with ethyl acetate 15ml jolting, and extracting solution is concentrated into 1ml, as need testing solution.(extracting method of proper mass standard) 2. gets 16 of this product, removes film-coat, and porphyrize adds water 50ml, and heated and boiled 15 minutes is put coldly, filters, and filtrate is extracted 2 times with the ethyl acetate jolting, each 15ml, and extracting solution is concentrated into 1ml, as need testing solution.3. get 16 of this product, remove film-coat, porphyrize adds water 100ml, and heated and boiled 15 minutes is put coldly, filters, and filtrate is extracted with ethyl acetate 15ml jolting, and extracting solution is concentrated into 1ml, as need testing solution.4. get 16 of this product, remove film-coat, porphyrize adds ethyl acetate 20ml, and reflux 30 minutes filters, and filtrate is concentrated into 1ml, as need testing solution.5. get 16 of this product, remove film-coat, porphyrize adds ethanol 20ml, and supersound process 20 minutes filters, the filtrate evaporate to dryness, and residue is dissolved in water, and extracts 2 times with the ethyl acetate jolting, each 15ml, extracting solution is concentrated into 1ml, as need testing solution.Result of the test show method for extraction and purification 1. and 2. sample be difficult to filter, emulsifying especially easily during with ethyl acetate extraction, speckle is very weak.Method for extraction and purification 3. speckle a little less than.4. and 5. the impurity speckle is more for method for extraction and purification.

Developing solvent that the present invention is preferred.Development system is contrasted: 1. 2. 3. petroleum ether (60～90 ℃)-ethyl acetate (17: 3) of ethyl acetate-chloroform-formic acid (3: 2: 0.3) of ethyl acetate-chloroform-formic acid (3: 2: 1).Result of the test shows development system 1., 2. Rf value is bigger, and development system 3. Rf value is less.

Test with menstruation regulation and macula removal tablet by embodiment 1 Fructus Lycii thin layer discrimination method, as a result in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical blue-fluorescence speckle.This method thin layer chromatography good separating effect, clear spot, rounding, Rf value is moderate, and repeatability and specificity are good, and negative control is noiseless.

Experimental example 3: for the thin layer chromatography of Radix Angelicae Sinensis is differentiated

Investigated development system petroleum ether (60～90 ℃)-ethyl acetate (17: 3), Rf value is too big as a result.

Test with menstruation regulation and macula removal tablet by embodiment 1 Radix Angelicae Sinensis thin layer discrimination method, as a result in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical blue-fluorescence principal spot.This method thin layer chromatography good separating effect, clear spot, rounding, Rf value is moderate, and repeatability and specificity are good, and negative control is noiseless.

Experimental example 4: for the thin layer chromatography feature of Radix Paeoniae Alba characteristic component peoniflorin is differentiated

Following method for extraction and purification has been carried out contrast test: 1. get 5 of this product, remove film-coat, porphyrize adds ethanol 30ml, and reflux 30 minutes filters, and filtrate evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.2. get 20 of this product, remove film-coat, porphyrize, add methanol 40ml, supersound process 30 minutes filters, filtrate evaporate to dryness, residue add water 2ml makes dissolving, is added on D101 type macroporous adsorptive resins (internal diameter 1.0cm, the high 15cm of post), it is colourless that water is eluted to eluent, discards water lotion, reuse 40% ethanol 50ml eluting is collected 40% ethanol elution, evaporate to dryness, residue adds ethanol 1ml makes dissolving, as need testing solution.3. get 10 of this product, remove film-coat, porphyrize, add methanol 30ml, supersound process 20 minutes filters, filtrate evaporate to dryness, residue add water 20ml makes dissolving, puts in the separatory funnel, extract 2 times with the ether jolting, each 20ml discards ether solution, aqueous solution extracts 2 times with water saturated n-butyl alcohol jolting, and each 20ml merges n-butyl alcohol liquid, with the saturated water washing of n-butyl alcohol 2 times, each 20ml discards water liquid, n-butyl alcohol liquid evaporate to dryness, residue add methanol 1ml makes dissolving, adds neutral alumina 1g (100～200 order), mix thoroughly, evaporate to dryness is added on neutral alumina post (100～200 orders, 3g, internal diameter 1.0cm dry column-packing) on, with 80% methanol 30ml eluting, collect eluent, evaporate to dryness, residue adds ethanol 1ml makes dissolving, as need testing solution.1. 2. in the test sample chromatograph, impurity is more for method as a result, and speckle is clear inadequately; Method is 3. in the test sample chromatograph, the clear spot rounding, but extraction step is many, is awkward, so do not adopt.

Test with menstruation regulation and macula removal tablet by embodiment 1 Radix Paeoniae Alba thin layer discrimination method, as a result in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show identical bluish violet speckle.This method thin layer chromatography good separating effect, the clear spot rounding, repeatability and specificity are good, and be negative noiseless.

Experimental example 5: for the thin layer chromatography feature of Radix Bupleuri is differentiated

Following method for extraction and purification has been carried out contrast test: 1. get 30 of this product, remove film-coat, porphyrize, add methanol 50ml, supersound process 30 minutes filters, filtrate evaporate to dryness, residue add the saturated water 30ml gradation dissolving of n-butyl alcohol, put in the separatory funnel, extract 2 times with water saturated n-butyl alcohol jolting, each 20ml merges n-butyl alcohol liquid, divide and get 10ml n-butyl alcohol liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.2. get 20 of this product, remove film-coat, porphyrize, add methanol 40ml, supersound process 20 minutes filters, filtrate evaporate to dryness, residue add water 5ml makes dissolving, filters, filtrate is added on the D101 type macroporous adsorptive resins (internal diameter 1.0cm, the high 18cm of post), water 30ml eluting, discard water lotion, reuse ethanol 30ml eluting is collected ethanol elution, evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution; 3. get 10 of this product, remove film-coat, porphyrize, add methanol 30ml, supersound process 20 minutes filters, filtrate evaporate to dryness, residue add water 20ml makes dissolving, puts in the separatory funnel, extract 2 times with the ether jolting, each 20ml discards ether solution, aqueous solution extracts 2 times with water saturated n-butyl alcohol jolting, and each 20ml merges n-butyl alcohol liquid, with the saturated water washing of n-butyl alcohol 2 times, each 20ml discards water liquid, n-butyl alcohol liquid evaporate to dryness, residue add methanol 1ml makes dissolving, adds neutral alumina 1g (100～200 order), mix thoroughly, evaporate to dryness is added on neutral alumina post (100～200 orders, 3g, internal diameter 1.0cm dry column-packing) on, with 80% methanol 30ml eluting, collect eluent, evaporate to dryness, residue adds ethanol 1ml makes dissolving, as need testing solution; 4. get 15 of this product, remove film-coat, porphyrize, add methanol 20ml, supersound process 30 minutes filters, the filtrate evaporate to dryness, residue adds 2% sodium hydroxide solution 20ml makes dissolving, puts coldly, extracts with n-butyl alcohol 20ml jolting, divide and get n-butyl alcohol liquid, evaporate to dryness, residue add methanol 2ml makes dissolving, as need testing solution; 5. get 20 of this product, remove film-coat, porphyrize, add methanol 40ml, supersound process 30 minutes filters, filtrate evaporate to dryness, residue add water 2ml makes dissolving, is added on D101 type macroporous adsorptive resins (internal diameter 1.0cm, the high 15cm of post), it is colourless that water is eluted to eluent, discards water lotion, reuse 40% ethanol 50ml eluting, discard 40% ethanol elution, continue, collect eluent with 70% ethanol 70ml eluting, evaporate to dryness, residue adds water saturated n-butyl alcohol 20ml makes dissolving, with ammonia solution 10ml washing, discards ammonia solution, divide and get n-butyl alcohol liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.1. 2. 3. in the test sample chromatograph, background is darker for method as a result, and feminine gender has the impurity speckle to disturb, multiple development system on probation, and separating effect is all bad; Method 4. feature speckle is clear inadequately; Method is 5. in the test sample chromatograph, the clear spot rounding, and effect is better, but extraction step is loaded down with trivial details, so do not adopt.

Test with menstruation regulation and macula removal tablet by embodiment 1 Radix Bupleuri thin layer discrimination method, the result inspects under the daylight, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical punctation; Put under the ultra-violet lamp (365nm) and inspect, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical yellow fluorescence speckle.This method thin layer chromatography good separating effect, clear spot, rounding, repeatability and specificity are good, and negative control is noiseless.

Experimental example 6: paeoniflorin content is measured research

The preparation of 1 reference substance and sample solution

The preparation precision of reference substance solution takes by weighing peoniflorin reference substance 10.05mg, puts in the 10ml measuring bottle, adds the Diluted Alcohol dissolving and is diluted to scale, shake up, precision is measured 1ml and is put in the 10ml measuring bottle, adds Diluted Alcohol to scale, shake up, promptly get (every 1ml contains peoniflorin 100.5 μ g).

This product is got in the preparation of need testing solution, removes film-coat, and accurate the title decided porphyrize, get about 1.5g, the accurate title, decide, and puts in the tool plug conical flask, the accurate Diluted Alcohol 50ml that adds, close plug claims to decide weight, supersound process (power 260W, frequency 50Hz) 30 minutes, put coldly, claim again to decide weight, supply the weight that subtracts mistake with Diluted Alcohol, shake up, centrifugal 10 minutes (3000 rev/mins) get supernatant, filter, get subsequent filtrate, promptly.

The preparation negative sample solution of negative sample solution is to take by weighing flavour of a drug except that the Radix Paeoniae Alba in the prescription ratio, makes the negative preparation of the Radix Paeoniae Alba by method for making, gets this negative preparation does not contain the Radix Paeoniae Alba by the preparation of above-mentioned " preparation of need testing solution " method negative sample solution again.

Investigated the time (10 minutes, 20 minutes, 30 minutes, 40 minutes) of supersound process in the test, the result shows that supersound process can extract fully in 30 minutes.

Solution after the supersound extraction is difficult to filter, and easily filters after centrifugal, has reduced operation easier, has shortened the test period.

2 chromatographic conditions and system suitability test

Instrument: day island proper Tianjin LC-10Avp high performance liquid chromatograph; Detector: day island proper Tianjin SPD-10Atvp UV-detector; Chromatographic column: octadecylsilane chemically bonded silica chromatographic column (Tianjin, island VP-ODS post, 4.6 * 150mm), guard column: YWG C ₁₈10 * 4.6mm; Weil-McLain dragon chromatographic work station; Mobile phase: methanol-water (25: 75) is a mobile phase; Flow velocity: 1.0ml/min, column temperature: room temperature.The detection wavelength is 230nm.

Draw peoniflorin reference substance solution, need testing solution respectively and do not contain each 10 μ l of negative sample solution of the Radix Paeoniae Alba, inject chromatograph of liquid, measure.Peoniflorin and other component reach baseline separation under this condition, separating degree R＞1.5.Number of theoretical plate is pressed the peoniflorin peak and is calculated greater than 2000, and the peoniflorin retention time is about 16 minutes.Negative sample solution chromatograph does not have absworption peak in the relevant position, and is visible negative noiseless.

3 wavelength are selected

Precision takes by weighing peoniflorin reference substance 10.05mg, put in the 10ml measuring bottle, add the Diluted Alcohol dissolving and be diluted to scale, shake up, precision is measured 0.2ml and is put in the 10ml measuring bottle, adds methanol to scale, shake up, with the scanning of uv-spectrophotometric instrument, peoniflorin has absorption maximum at the 229nm place as a result, is the mensuration wavelength with reference to pharmacopeia and former dosage form Standard Selection 230nm.

4 reference substance purity tests

Precision takes by weighing peoniflorin reference substance 10.05mg, puts in the 10ml measuring bottle, adds the Diluted Alcohol dissolving and is diluted to scale, shakes up, by above-mentioned chromatographic condition, sample introduction 10 μ l measure, and calculate with the peak area normalization method, content is 99.27%, meets the assay requirement, calculates by 100% during calculating.

The checking of 5 methodologies

The precision test: calculating peoniflorin peak area meansigma methods is 1181505, and relative standard deviation is 0.24%, shows that precision is better; Linear relationship is investigated: with peak area integrated value A peoniflorin sample size C (μ g) is carried out regression analysis, get regression equation: A=1.16 * 10 ⁶C+1.14 * 10 ⁴, r=0.9999.The range of linearity is 0.201～4.02 μ g, and the straight-line pass initial point, and available single-point external standard method is measured and calculated; Stability test: calculate in a few days that the peak area meansigma methods is 956424, relative standard deviation is that the peak area meansigma methods is 953734 between 0.89%, three day, and relative standard deviation is 1.13%, shows that need testing solution is stable in 72h; Replica test: calculating content of paeoniflorin is the 1.084mg/ sheet, and relative standard deviation is 1.44%, shows method repeatability better; The average recovery test: calculating average recovery rate is 99.81%, and relative standard deviation is 1.74%, shows that the method response rate is better.

Above methodology checking shows that this method is accurate, and specificity, good reproducibility meet the assay requirement.

Specific embodiment is as follows:

Embodiment 1 menstruation regulation and macula removal tablet method of quality control

(1) get 16 of this product, remove film-coat, porphyrize adds ethanol 20ml, and reflux 30 minutes is put coldly, filters, and filtrate is concentrated into 2ml, as need testing solution.Other gets Fructus Ligustri Lucidi control medicinal material 0.5g, shines medical material solution in pairs with legal system.Even up pier fruit acid reference substance again, add ethanol and make the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw above-mentioned three kinds of each 5ul of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene-ethyl acetate-formic acid (20: 7: 0.5) is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing at 110 ℃.In the test sample chromatograph, with control medicinal material chromatograph and the corresponding position of reference substance chromatograph on, show the speckle of same color.

(2) get 16 of this product, remove film-coat, porphyrize adds water 100ml, and heated and boiled 15 minutes is put coldly, filters, and filtrate is extracted 2 times with the ethyl acetate jolting, each 15ml, and combined ethyl acetate liquid is concentrated into 1ml, as need testing solution.Other gets Fructus Lycii control medicinal material 1g, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw above-mentioned two kinds of each 5ul of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene-ethyl acetate-formic acid (20: 7: 0.5) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(3) get 16 of this product, remove film-coat, porphyrize, the 40ml that adds diethyl ether, supersound process 15 minutes filters, and filtrate volatilizes, and residue adds methanol 2ml makes dissolving, as need testing solution.Other gets Radix Angelicae Sinensis control medicinal material 1g, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw above-mentioned two kinds of each 5ul of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with petroleum ether (60～90 ℃)-ethyl acetate (17: 2) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(4) get 30 of this product, remove film-coat, porphyrize, add methanol 50ml, supersound process 30 minutes filters, filtrate evaporate to dryness, residue add the saturated water 30ml gradation dissolving of n-butyl alcohol, put in the separatory funnel, extract 2 times with water saturated n-butyl alcohol jolting, each 20ml merges n-butyl alcohol liquid, divide and get 10ml n-butyl alcohol liquid, surplus liquid is continued to employ, evaporate to dryness, residue adds ethanol 1ml makes dissolving, as need testing solution.Other gets the peoniflorin reference substance, adds ethanol and makes the solution that every 1ml contains 2mg, in contrast product solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw need testing solution 3ul, reference substance solution 5ul, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with chloroform-ethyl acetate-methanol-formic acid (40: 5: 10: 0.2) be developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and 105 ℃ to be heated to the speckle colour developing clear.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.

(5) get discriminating (a 5) n-butyl alcohol liquid of continuing to employ down, with ammonia solution 30ml washing, discard the ammonia washing liquid, the water 30ml washing that the reuse n-butyl alcohol is saturated discards water lotion, divides and gets n-butyl alcohol liquid, and evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.Other gets Radix Bupleuri control medicinal material 0.5g, adds methanol 30ml, and supersound process 30 minutes filters, filtrate evaporate to dryness, residue add water saturated n-butyl alcohol 20ml makes dissolving, with ammonia solution 10ml washing, discards the ammonia washing liquid, divide and get n-butyl alcohol liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, in contrast medical material solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 5～8ul of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with chloroform-methanol-water (30: 10: 1) is developing solvent, launch, take out, dry, spray is with 40% sulfuric acid solution of 2% paradime thylaminobenzaldehyde, and 105 ℃ to be heated to the speckle colour developing clear.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color; Put under the ultra-violet lamp (365nm) and inspect, show the fluorescence speckle of same color.

(6) content of paeoniflorin in mensuration this product

Measure according to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 D).

Chromatographic condition and system suitability test are filler with octadecylsilane chemically bonded silica; Methanol-water (25: 75) is a mobile phase; The detection wavelength is 230nm.Number of theoretical plate calculates by peoniflorin should be not less than 2000.

It is an amount of that the preparation precision of reference substance solution takes by weighing the peoniflorin reference substance, makes the solution that every 1ml contains 0.1mg with Diluted Alcohol, promptly.

This product is got in the preparation of need testing solution, removes film-coat, and accurate the title decided porphyrize, get about 1.5g, the accurate title, decide, and puts in the tool plug conical flask, the accurate Diluted Alcohol 50ml that adds, close plug claims to decide weight, supersound process (power 260W, frequency 50Hz) 30 minutes, put coldly, claim again to decide weight, supply the weight that subtracts mistake with Diluted Alcohol, shake up, centrifugal 10 minutes (3000 rev/mins) get supernatant, filter, get subsequent filtrate, promptly.

Accurate respectively reference substance solution and each the 10 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly.

Every of this product contains the Radix Paeoniae Alba with peoniflorin (C ₂₃H ₂₈O ₁₁) meter, must not be less than 0.60mg.

Embodiment 2 menstruation regulation and macula removal tablet method of quality control

(1) thin layer is differentiated Fructus Ligustri Lucidi: get 30 of this product, remove film-coat, porphyrize adds ethanol 50ml, and reflux 60 minutes is put coldly, filters, and filtrate is concentrated into 2ml, as need testing solution.Other gets Fructus Ligustri Lucidi control medicinal material 1g, shines medical material solution in pairs with legal system.Even up pier fruit acid reference substance again, add ethanol and make the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 2 μ l of above-mentioned three kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene-ethyl acetate-formic acid (25: 5: 0.8) is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and 120 ℃ to be heated to the speckle colour developing clear.In the test sample chromatograph, with control medicinal material chromatograph and the corresponding position of reference substance chromatograph on, show the speckle of same color.

(2) thin layer is differentiated Fructus Lycii: get 30 of this product, remove film-coat, porphyrize adds water 200ml, and heated and boiled 60 minutes is put coldly, filters, and filtrate is extracted 3 times with the ethyl acetate jolting, each 50ml, and combined ethyl acetate liquid is concentrated into 1ml, as need testing solution.Other gets Fructus Lycii control medicinal material 2g, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 2 μ l of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene-ethyl acetate-formic acid (25: 5: 0.8) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(3) thin layer is differentiated Radix Angelicae Sinensis: get 30 of this product, remove film-coat, and porphyrize, the 100ml that adds diethyl ether, supersound process 60 minutes filters, and filtrate volatilizes, and residue adds methanol 2ml makes dissolving, as need testing solution.Other gets Radix Angelicae Sinensis control medicinal material 2g, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 2 μ l of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with petroleum ether (60～90 ℃)-ethyl acetate (17: 2) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(4) thin layer is differentiated the Radix Paeoniae Alba: get 30 of this product, remove film-coat, porphyrize, add methanol 100ml, supersound process 60 minutes filters, filtrate evaporate to dryness, residue add the saturated water 50ml gradation dissolving of n-butyl alcohol, put in the separatory funnel, extract 3 times with water saturated n-butyl alcohol jolting, each 50ml divides and gets n-butyl alcohol liquid, evaporate to dryness, residue adds ethanol 1ml makes dissolving, as need testing solution.Other gets the peoniflorin reference substance, adds ethanol and makes the solution that every 1ml contains 2mg, in contrast product solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw need testing solution and reference substance solution 2 μ l, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with chloroform-ethyl acetate-methanol-formic acid (45: 3: 12: 0.1) be developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and 120 ℃ to be heated to the speckle colour developing clear.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.

(5) thin layer is differentiated Radix Bupleuri: get 50 of this product, remove film-coat, porphyrize adds methanol 100ml, supersound process 60 minutes, filter, filtrate evaporate to dryness, residue add the saturated water 50ml gradation dissolving of n-butyl alcohol, put in the separatory funnel, extract 3 times with water saturated n-butyl alcohol jolting, each 50ml divides and gets n-butyl alcohol liquid, wash with ammonia solution 100ml, discard the ammonia washing liquid, the water 100ml washing that the reuse n-butyl alcohol is saturated discards water lotion, divide and get n-butyl alcohol liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.Other gets Radix Bupleuri control medicinal material 2g, adds methanol 50ml, and supersound process 60 minutes filters, filtrate evaporate to dryness, residue add water saturated n-butyl alcohol 50ml makes dissolving, with ammonia solution 50ml washing, discards the ammonia washing liquid, divide and get n-butyl alcohol liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, in contrast medical material solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw above-mentioned two kinds of solution, 2 μ l, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with chloroform-methanol-water (35: 8: 1.5) is developing solvent, launch, take out, dry, spray is with 40% sulfuric acid solution of 2% paradime thylaminobenzaldehyde, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color; Put under the ultra-violet lamp (365nm) and inspect, show the fluorescence speckle of same color.

(6) measure content of paeoniflorin in this product:

Measure according to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 D).

Chromatographic condition and system suitability test are filler with octadecylsilane chemically bonded silica; Methanol-water (25: 75) is a mobile phase; The detection wavelength is 230nm.Number of theoretical plate calculates by peoniflorin should be not less than 2000.

It is an amount of that the preparation precision of reference substance solution takes by weighing the peoniflorin reference substance, makes the solution that every 1ml contains 0.1mg with Diluted Alcohol, promptly.

This product is got in the preparation of need testing solution, removes film-coat, and accurate the title decided porphyrize, get in right amount, the accurate title, decide, and puts in the tool plug conical flask, accurate adding Diluted Alcohol 50ml, close plug claims to decide weight, supersound process, put coldly, claim again to decide weight, supply the weight that subtracts mistake with Diluted Alcohol, shake up, centrifugal 30 minutes, get supernatant, filter, get subsequent filtrate, promptly.

Accurate respectively reference substance solution and each the 20 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure promptly.

Embodiment 3 menstruation regulation and macula removal tablet method of quality control

(1) thin layer is differentiated Fructus Ligustri Lucidi: get 5 of this product, remove film-coat, porphyrize adds ethanol 10ml, and reflux 10 minutes is put coldly, filters, and filtrate is concentrated into 2ml, as need testing solution.Other gets Fructus Ligustri Lucidi control medicinal material 0.2g, shines medical material solution in pairs with legal system.Even up pier fruit acid reference substance again, add ethanol and make the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 10 μ l of above-mentioned three kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene-ethyl acetate-formic acid (15: 10: 0.2) is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and 100 ℃ to be heated to the speckle colour developing clear.In the test sample chromatograph, with control medicinal material chromatograph and the corresponding position of reference substance chromatograph on, show the speckle of same color.

(2) thin layer is differentiated Fructus Lycii: get 5 of this product, remove film-coat, porphyrize adds water 50ml, and heated and boiled 10 minutes is put coldly, filters, and filtrate is extracted with ethyl acetate 10ml jolting, and acetic acid ethyl fluid is concentrated into 1ml, as need testing solution.Other gets Fructus Lycii control medicinal material 0.5, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 10 μ l of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene-ethyl acetate-formic acid (15: 10: 0.2) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(3) thin layer is differentiated Radix Angelicae Sinensis: get 5 of this product, remove film-coat, and porphyrize, the 20ml that adds diethyl ether, supersound process 10 minutes filters, and filtrate volatilizes, and residue adds methanol 2ml makes dissolving, as need testing solution.Other gets Radix Angelicae Sinensis control medicinal material 0.5g, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 10 μ l of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with petroleum ether (60～90 ℃)-ethyl acetate (17: 2) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(4) thin layer is differentiated the Radix Paeoniae Alba: get 5 of this product, remove film-coat, porphyrize, add methanol 20ml, supersound process 10 minutes filters, the filtrate evaporate to dryness, residue adds the saturated water 10ml gradation dissolving of n-butyl alcohol, puts in the separatory funnel, extracts with water saturated n-butyl alcohol 10ml jolting, divide and get n-butyl alcohol liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.Other gets the peoniflorin reference substance, adds ethanol and makes the solution that every 1ml contains 2mg, in contrast product solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw need testing solution and reference substance solution 10 μ l, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with chloroform-ethyl acetate-methanol-formic acid (35: 7: 8: 0.5) be developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and 100 ℃ to be heated to the speckle colour developing clear.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.

(5) thin layer is differentiated Radix Bupleuri: get 10 of this product, remove film-coat, porphyrize adds methanol 20ml, supersound process 10 minutes filters the filtrate evaporate to dryness, residue adds the saturated water 10ml gradation dissolving of n-butyl alcohol, puts in the separatory funnel, extracts with water saturated n-butyl alcohol 10ml jolting, divide and get n-butyl alcohol liquid,, discard the ammonia washing liquid with ammonia solution 10ml washing, the water 10ml washing that the reuse n-butyl alcohol is saturated discards water lotion, divides and gets n-butyl alcohol liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.Other gets Radix Bupleuri control medicinal material 0.2g, adds methanol 10ml, and supersound process 10 minutes filters, filtrate evaporate to dryness, residue add water saturated n-butyl alcohol 10ml makes dissolving, with ammonia solution 10ml washing, discards the ammonia washing liquid, divide and get n-butyl alcohol liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, in contrast medical material solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw above-mentioned two kinds of solution, 10 μ l, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with chloroform-methanol-water (25: 12: 0.5) is developing solvent, launch, take out, dry, spray is with 40% sulfuric acid solution of 2% paradime thylaminobenzaldehyde, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color; Put under the ultra-violet lamp (365nm) and inspect, show the fluorescence speckle of same color.

(6) measure content of paeoniflorin in this product:

Measure according to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 D).

Chromatographic condition and system suitability test are filler with octadecylsilane chemically bonded silica; Methanol-water (25: 75) is a mobile phase; The detection wavelength is 230nm.Number of theoretical plate calculates by peoniflorin should be not less than 2000.

It is an amount of that the preparation precision of reference substance solution takes by weighing the peoniflorin reference substance, makes the solution that every 1ml contains 0.1mg with Diluted Alcohol, promptly.

This product is got in the preparation of need testing solution, removes film-coat, and accurate the title decided porphyrize, get in right amount, the accurate title, decide, and puts in the tool plug conical flask, accurate adding Diluted Alcohol 50ml, close plug claims to decide weight, supersound process, put coldly, claim again to decide weight, supply the weight that subtracts mistake with Diluted Alcohol, shake up, centrifugal 5 minutes, get supernatant, filter, get subsequent filtrate, promptly.

Accurate respectively reference substance solution and each the 5 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly.

Claims (3)

1. the method for quality control of a menstruation regulation and macula removal tablet is characterized in that, this method comprises following all or part of content: thin layer is differentiated Fructus Ligustri Lucidi; Thin layer is differentiated Fructus Lycii; Thin layer is differentiated Radix Angelicae Sinensis; Thin layer is differentiated the Radix Paeoniae Alba; Thin layer is differentiated Radix Bupleuri; Measure content of paeoniflorin in this product.

2. method of quality control as claimed in claim 1 is characterized in that, one or more in may further comprise the steps:

(1) thin layer is differentiated Fructus Ligustri Lucidi: it is an amount of to get this product, removes film-coat, and porphyrize adds alcohol heating reflux, puts coldly, filters, and filtrate is concentrated into 2ml, as need testing solution.It is an amount of that other gets the Fructus Ligustri Lucidi control medicinal material, shines medical material solution in pairs with legal system.Even up pier fruit acid reference substance again, add ethanol and make the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw above-mentioned three kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene-ethyl acetate-formic acid proper proportion is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with control medicinal material chromatograph and the corresponding position of reference substance chromatograph on, show the speckle of same color.

(2) thin layer is differentiated Fructus Lycii: it is an amount of to get this product, removes film-coat, and porphyrize adds the water heated and boiled, puts coldly, filters, and filtrate is extracted with the ethyl acetate jolting, and acetic acid ethyl fluid is concentrated into 1ml, as need testing solution.It is an amount of that other gets the Fructus Lycii control medicinal material, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene-ethyl acetate-formic acid proper proportion is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(3) thin layer is differentiated Radix Angelicae Sinensis: it is an amount of to get this product, removes film-coat, porphyrize, and the supersound process that adds diethyl ether filters, and filtrate volatilizes, and residue adds methanol 2ml makes dissolving, as need testing solution.It is an amount of that other gets the Radix Angelicae Sinensis control medicinal material, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with petroleum ether (60～90 ℃)-ethyl acetate (17: 2) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(4) thin layer is differentiated the Radix Paeoniae Alba: it is an amount of to get this product, removes film-coat, porphyrize, add the methanol supersound process, filter, filtrate evaporate to dryness, residue add the saturated water dissolution of n-butyl alcohol, put in the separatory funnel, extract with water saturated n-butyl alcohol jolting, divide and get n-butyl alcohol liquid, evaporate to dryness, residue adds ethanol 1ml makes dissolving, as need testing solution.Other gets the peoniflorin reference substance, adds ethanol and makes the solution that every 1ml contains 2mg, in contrast product solution.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw need testing solution and reference substance solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with chloroform-ethyl acetate-methanol-formic acid proper proportion is developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.

(5) thin layer is differentiated Radix Bupleuri: it is an amount of to get this product, removes film-coat, porphyrize, and it is an amount of to add methanol, supersound process filters the filtrate evaporate to dryness, residue adds the saturated water dissolution of n-butyl alcohol, puts in the separatory funnel, extracts with water saturated n-butyl alcohol jolting, divide and get n-butyl alcohol liquid, wash in right amount, discard the ammonia washing liquid with ammonia solution, the water that the reuse n-butyl alcohol is saturated washs in right amount, discards water lotion, divides and gets n-butyl alcohol liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.It is an amount of that other gets the Radix Bupleuri control medicinal material, adds the methanol supersound process, filters, and filtrate evaporate to dryness, residue add water saturated n-butyl alcohol makes dissolving, with the ammonia solution washing, discards the ammonia washing liquid, divides and get n-butyl alcohol liquid, and evaporate to dryness, residue add ethanol 1ml makes dissolving, in contrast medical material solution.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with chloroform-methanol-water proper proportion is developing solvent, launch, take out, dry, spray is with 40% sulfuric acid solution of 2% paradime thylaminobenzaldehyde, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color; Put under the ultra-violet lamp (365nm) and inspect, show the fluorescence speckle of same color.

(6) measure content of paeoniflorin in this product:

Measure according to high performance liquid chromatography (the corresponding appendix of Chinese Pharmacopoeia).

Chromatographic condition and system suitability test are filler with octadecylsilane chemically bonded silica; Methanol-water (25: 75) is a mobile phase; The detection wavelength is 230nm.Number of theoretical plate calculates by peoniflorin should be not less than 2000.

It is an amount of that the preparation precision of reference substance solution takes by weighing the peoniflorin reference substance, makes the solution that every 1ml contains 0.1mg with Diluted Alcohol, promptly.

This product is got in the preparation of need testing solution, removes film-coat, and accurate the title decided porphyrize, get in right amount, the accurate title, decide, and puts in the tool plug conical flask, accurate adding Diluted Alcohol 50ml, close plug claims to decide weight, supersound process, put coldly, claim again to decide weight, supply the weight that subtracts mistake with Diluted Alcohol, shake up, centrifugal, get supernatant, filter, get subsequent filtrate, promptly.

Accurate respectively reference substance solution and the need testing solution drawn of algoscopy injects chromatograph of liquid, measures, promptly.

3. method of quality control as claimed in claim 1 is characterized in that, one or more in may further comprise the steps:

(1) thin layer is differentiated Fructus Ligustri Lucidi: get 5～30 of this product, remove film-coat, porphyrize adds ethanol 10～50ml, and reflux 10～60 minutes is put coldly, filters, and filtrate is concentrated into 2ml, as need testing solution.Other gets Fructus Ligustri Lucidi control medicinal material 0.2～1g, shines medical material solution in pairs with legal system.Even up pier fruit acid reference substance again, add ethanol and make the solution that every 1ml contains 1mg, in contrast product solution.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw each 2～10 μ l of above-mentioned three kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene-ethyl acetate-formic acid (15～25: 5～10: 0.2～0.8) be developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and 100 ℃～120 ℃ to be heated to the speckle colour developing clear.In the test sample chromatograph, with control medicinal material chromatograph and the corresponding position of reference substance chromatograph on, show the speckle of same color.

(2) thin layer is differentiated Fructus Lycii: get 5～30 of this product, remove film-coat, porphyrize adds water 50～200ml, heated and boiled 10～60 minutes is put coldly, filters, and filtrate is extracted 1～3 time with the ethyl acetate jolting, each 10～50ml, combined ethyl acetate liquid is concentrated into 1ml, as need testing solution.Other gets Fructus Lycii control medicinal material 0.5～2g, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw each 2～10 μ l of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene-ethyl acetate-formic acid (15～25: 5～10: 0.2～0.8) be developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(3) thin layer is differentiated Radix Angelicae Sinensis: get 5～30 of this product, remove film-coat, and porphyrize, the 20～100ml that adds diethyl ether, supersound process 10～60 minutes filters, and filtrate volatilizes, and residue adds methanol 2ml makes dissolving, as need testing solution.Other gets Radix Angelicae Sinensis control medicinal material 0.5～2g, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw each 2～10 μ l of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with petroleum ether (60～90 ℃)-ethyl acetate (17: 2) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(4) thin layer is differentiated the Radix Paeoniae Alba: get 5～30 of this product, remove film-coat, porphyrize, add methanol 20～100ml, supersound process 10～60 minutes filters, filtrate evaporate to dryness, residue add the saturated water 10～50ml gradation dissolving of n-butyl alcohol, put in the separatory funnel, extract 1～3 time with water saturated n-butyl alcohol jolting, each 10～50ml divides and gets n-butyl alcohol liquid, evaporate to dryness, residue adds ethanol 1ml makes dissolving, as need testing solution.Other gets the peoniflorin reference substance, adds ethanol and makes the solution that every 1ml contains 2mg, in contrast product solution.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw need testing solution and reference substance solution 2～10 μ l, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with chloroform-ethyl acetate-methanol-formic acid (35～45: 3～7: 8～12: 0.1～0.5) be developing solvent, launch, take out, dry, spray is with 5% vanillin sulfuric acid solution, and 100 ℃～120 ℃ to be heated to the speckle colour developing clear.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.

(5) thin layer is differentiated Radix Bupleuri: get 10～50 of this product, remove film-coat, porphyrize adds methanol 20～100ml, supersound process 10～60 minutes, filter, filtrate evaporate to dryness, residue add the saturated water 10～50ml gradation dissolving of n-butyl alcohol, put in the separatory funnel, extract 1～3 time with water saturated n-butyl alcohol jolting, each 10～50ml divides and gets n-butyl alcohol liquid, wash with ammonia solution 10～100ml, discard the ammonia washing liquid, water 10～100ml washing that the reuse n-butyl alcohol is saturated discards water lotion, divide and get n-butyl alcohol liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.Other gets Radix Bupleuri control medicinal material 0.2～2g, add methanol 10～50ml, supersound process 10～60 minutes filters, the filtrate evaporate to dryness, residue adds water saturated n-butyl alcohol 10～50ml makes dissolving, with ammonia solution 10～50ml washing, discards the ammonia washing liquid, divide and get n-butyl alcohol liquid, evaporate to dryness, residue add ethanol 1ml makes dissolving, in contrast medical material solution.Test according to thin layer chromatography (the corresponding appendix of Chinese Pharmacopoeia), draw above-mentioned two kinds of solution, 2～10 μ l, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with chloroform-methanol-water (25～35: 8～12: 0.5～1.5) be developing solvent, launch, take out, dry, spray is with 40% sulfuric acid solution of 2% paradime thylaminobenzaldehyde, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color; Put under the ultra-violet lamp (365nm) and inspect, show the fluorescence speckle of same color.

(6) measure content of paeoniflorin in this product:

Measure according to high performance liquid chromatography (the corresponding appendix of Chinese Pharmacopoeia).

Chromatographic condition and system suitability test are filler with octadecylsilane chemically bonded silica; Methanol-water (25: 75) is a mobile phase; The detection wavelength is 230nm.Number of theoretical plate calculates by peoniflorin should be not less than 2000.

It is an amount of that the preparation precision of reference substance solution takes by weighing the peoniflorin reference substance, makes the solution that every 1ml contains 0.1mg with Diluted Alcohol, promptly.

This product is got in the preparation of need testing solution, removes film-coat, and accurate the title decided porphyrize, get in right amount, the accurate title, decide, and puts in the tool plug conical flask, accurate adding Diluted Alcohol 50ml, close plug claims to decide weight, supersound process, put coldly, claim again to decide weight, supply the weight that subtracts mistake with Diluted Alcohol, shake up, centrifugal 5～30 minutes, get supernatant, filter, get subsequent filtrate, promptly.

Accurate respectively reference substance solution and each 5～20 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly.