CN102068598A - Quality control method of Yangrong Baicao Wan for treating irregular menses caused by hemophthisis - Google Patents
Quality control method of Yangrong Baicao Wan for treating irregular menses caused by hemophthisis Download PDFInfo
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Abstract
The invention belongs to the technical field of Chinese patent medicine, relates to a quality control method of Yangrong Baicao Wan which is prepared from traditional Chinese medicine and used for treating irregular menses caused by hemophthisis, wherein the quality standard comprises the authentication items of dried orange peel and nutgrass galingale rhizome, the content of paeoniflorin (C23H28O11) in the medicine is measured by means of high efficiency liquid chromatography, the medicine of each gram comprises radix paeoniae alba of not less than 3.0 mg by means of quantitative indexes and inspection method based on the paeoniflorin (C23H28O11), so that the quality standard is relatively complete, the quality of the medicine is controlled strictly, and the improvement for the quality of the product is ensured.
Description
Technical field
The invention belongs to the technical field of Chinese patent medicine, relate to the Chinese crude drug method of quality control of the Yangrongbaicao medicinal ball that is the raw material a kind of women's of treatment anemia menoxenia of making.
Background technology
The Yangrongbaicao medicinal ball is a kind of classical Chinese patent medicine, Ministry of Public Health standard WS
3-B-1406-93 record prescription and quality standard:
Prescription: Radix Paeoniae Alba 160g Radix Angelicae Sinensis 80g Herba Taxilli 80g
The Radix Rehmanniae Preparata 80g Cortex Eucommiae (charcoal) 80g Rhizoma Chuanxiong 60g
Rhizoma Cyperi (vinegar system) 60g 40g Radix Ophiopogonis Pericarpium Citri Reticulatae 40g
Poria 40g Colla Corii Asini 20g Radix Glycyrrhizae 20g
Semen sojae atricolor 16g
Method for making: above 13 flavors, be ground into fine powder, sieve, mixing is used water pill, drying, promptly.
At present similar or identical with above-mentioned disclosed prescription, the proportioning difference, patent and document that effect is similar are a lot, as Beijing Fukangren Biopharmaceutical Technology Co., Ltd's application number: the CN200510134437.5 applying date: 2005.12.15 open (bulletin) day: 2006.08.30 " Yangrongbaicao medicinal preparation and new preparation method " the present invention relates to a kind of Chinese medicine composition and preparation technology thereof, be particularly related to a kind of women's of being used for anemia, the deficiency of YIN with the passing of time, menoxenia, expired more than, menalgia, leucorrhea prescription and preparation technology thereof at present.Preferred formulation is drop pill and soft capsule preparation.
Xi'an Ande Pharmacy Co., Ltd. application (patent) number: the CN200410026333.8 applying date: 2004.07.16 open (bulletin) day: flourish capsule is supported by a kind of gynecological of 2006.01.18 " flourish capsule is supported by gynecological ", comprises (consumption is a weight portion): 2～20 parts of Radix Angelicae Sinensis, 4～25 parts of the Rhizoma Atractylodis Macrocephalaes, 3～24 parts of Radix Rehmanniae Preparata, 2～15 parts of Rhizoma Chuanxiongs, 2～20 parts of the Radix Paeoniae Albas, 2～15 parts of Rhizoma Cyperis, 2～16 parts of Herba Leonuris, 1～10 part of the Radix Astragali, 2～14 parts of the Cortexs Eucommiae, 2～16 parts of Folium Artemisiae Argyis, 1～8 part of Radix Ophiopogonis, 1～8 part in Colla Corii Asini, 3～10 parts in Radix Glycyrrhizae, 1～12 part of Pericarpium Citri Reticulatae, 2～11 parts in Poria, Fructus Amomi 1～8.The medicament of formulation method preparation routinely.It is tested through clinical observation, and result of the test shows that the total effective rate of Drug therapy metrorrhagia of the present invention is 95%, and the total effective rate of treatment menoxenia is 93.33%, and the total effective rate of treatment infertility is 47%, and the total effective rate of curative stream back vaginal hemorrhage is 90%.Cure mainly: be used for that insufficiency of vital energy and blood, stagnation of liver-QI are not relaxed, menoxenia, have a dizzy spell, blood leaks metrorrhagia, the anemia body is weak and infertility.
Jiangxi Huiren Pharmaceutical Co., Ltd, application (patent) number: the CN03109431.7 applying date: 2003.04.09, open (bulletin) day: 2003.09.10, " a kind of regulating menstruation and nourishing blood Chinese medicine " the present invention relates to a kind of regulating menstruation and nourishing blood Chinese medicine, the particularly Chinese medicine of being made by following Chinese crude drug.Radix Angelicae Sinensis, the Radix Paeoniae Alba, Rhizoma Chuanxiong, Radix Rehmanniae Preparata, Radix Codonopsis, the Rhizoma Atractylodis Macrocephalae, Poria, Radix Glycyrrhizae, Cortex Cinnamomi, Herba Leonuri, Cortex Moutan, Myrrha (processed), Rhizoma Corydalis (vinegar system), Rhizoma Ligustici, the Radix Angelicae Dahuricae, Radix Scutellariae, Radix Cynanchi Atrati, Rhizoma Cyperi (vinegar system), Fructus Amomi, Pericarpium Citri Reticulatae, Halloysitum Rubrum (forging), Cornu Cervi Degelatinatum, Colla Corii Asini.
It is to set up ginseng osmanthus to support the reversed phase high-performance liquid chromatography analytical method of measuring paeoniflorin content in the flourish ball that the document of putting down in writing relevant quality standard has the refined red Bears English of contemporary Chinese medicinal application the 3rd volume the 6th phase record Song Xiaoxiao king March in 2009 " the RP2HPLC method is measured ginseng osmanthus and supported content of paeoniflorin in the flourish ball " purpose.It is analytical column that method adopts Alltima2C18 (250mm * 416mm, 5 μ m), and mobile phase is acetonitrile 2011% phosphoric acid solution (145: 86), and flow velocity is 018ml/min, and the detection wavelength is 230nm.The result: peoniflorin is good linear relationship (r=1100) with peak area in 01,043 16～11726 4mg/ml concentration range.The average recovery rate of peoniflorin is 9816%, and RSD is 116% (n=6).The average recovery rate of this method is 9816%, and RSD is 116% (n=6).Conclusion: this method is easy and simple to handle, quick, and the result is accurate, can be used for joining the quality control that flourish ball is supported in osmanthus.
Medicine is identified the 6th volume the 9th phase record March in 2009 Chen Bo 1, and Wen Jihui 2, Zhang Guofeng 2 (the 1. four-player people of Daliang City hospital, Dalian 116031; 2. Dongyu Pharmaceutical Co., Ltd., Shenyang City, Shenyang, Liaoning 110036) " the HPLC method is measured to concentrate and supported content of paeoniflorin in the flourish ball " purpose: set up the HPLC method and measure the method for supporting paeoniflorin content in the flourish ball that concentrates.Method: adopt Diamonsil-C18 chromatographic column (200mm * 4.6mm, 5 μ m), mobile phase is methanol-0.2% glacial acetic acid (30: 70), and flow velocity is 1.0ml/min, and the detection wavelength is 230nm.The result: peoniflorin concentration is in 0.041 6～0.2080mg/ml scope internal linear relation good (r=0.9997), and average recovery rate is 98.19%, and RSD is 0.97%.Conclusion: this method good separating effect, easy, quick, accurate, can be used for the quality control of this medicine.
More than disclosed similar or identical with the disclosed prescription of Ministry of Public Health standard WS3-B-1406-93 Yangrongbaicao medicinal ball, the proportioning difference, the similar document of effect is a lot, but also has shortcomings such as discriminating project, discriminating means, content assaying method imperfection in the prior art.
Summary of the invention
The method of quality control that the purpose of this invention is to provide the Yangrongbaicao medicinal ball of a kind of women's of treatment anemia menoxenia makes every effort to develop perfect quality standard, and the quality of strict control medicine is guaranteed curative effect.
In order to reach purpose the technical solution used in the present invention of the present invention be: the method for quality control of the Yangrongbaicao medicinal ball of a kind of women's of treatment anemia menoxenia is provided, and wherein said pharmaceutical formulation is by Chinese crude drug:
Radix Paeoniae Alba 160g Radix Angelicae Sinensis 80g Herba Taxilli 80g
The Radix Rehmanniae Preparata 80g Cortex Eucommiae (charcoal) 80g Rhizoma Chuanxiong 60g
Rhizoma Cyperi (vinegar system) 60g 40g Radix Ophiopogonis Pericarpium Citri Reticulatae 40g
Poria 40g Colla Corii Asini 20g Radix Glycyrrhizae 20g
Semen sojae atricolor 16g forms, and above-mentioned raw materials is ground into fine powder, sieves, and mixing is used water pill, and drying promptly, is characterized in that: the step of its method is:
(1) being reference substance with the Hesperidin, is control medicinal material with the Rhizoma Cyperi, and thin layer chromatography differentiates in the Yangrongbaicao medicinal ball prescription whether contain Pericarpium Citri Reticulatae, Rhizoma Cyperi composition;
(2) with the peoniflorin be reference substance, high performance liquid chromatography detects content of paeoniflorin in the Yangrongbaicao medicinal ball prescription.
The method of quality control of the Yangrongbaicao medicinal ball of described treatment women anemia menoxenia, it is characterized in that: described thin layer chromatography differentiates that the method that whether contains the Pericarpium Citri Reticulatae composition in the Yangrongbaicao medicinal ball prescription is: get this product powder 1g, add water 30mL, supersound process 15 minutes, the 20mL that adds diethyl ether extraction, discard the ether layer, water layer adds n-butyl alcohol 20mL extraction, discards water layer, alcohol layer evaporate to dryness, residue adds methanol 0.5mL makes dissolving, as need testing solution; Other gets the Hesperidin reference substance, adds methanol and makes the solution that every 1mL contains 2mg, in contrast product solution; Test according to thin layer chromatography (first appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 5～10 μ L of above-mentioned five kinds of solution, put respectively on the silica gel g thin-layer plate of same usefulness 0.5% sodium hydroxide solution preparation, with ethyl acetate-formic acid-glacial acetic acid-water is developing solvent, ethyl acetate: formic acid: glacial acetic acid: water=100: 11: 11: 26, launch, take out, dry, spray is with to methoxyl group anisaldehyde solution, and it is clear to be heated to the speckle colour developing at 105 degrees centigrade; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;
Described thin layer chromatography differentiates that the method that whether contains the Rhizoma Cyperi composition in the Yangrongbaicao medicinal ball prescription is: get this product powder 5g, the 5mL that adds diethyl ether places 1h, and jolting constantly filters, and filtrate volatilizes, and residue adds ethyl acetate 0.5mL makes dissolving, as need testing solution; Other gets Rhizoma Cyperi medicinal powder 1g, and the 5mL that adds diethyl ether places 1h, and jolting constantly filters, and filtrate volatilizes, and residue adds ethyl acetate 0.5mL makes dissolving, and filtrate is as Rhizoma Cyperi medical material solution; Test according to thin layer chromatography (first appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 5～10 μ L of above-mentioned freshly prepd three kinds of solution, put respectively on same silica gel g thin-layer plate, with benzene-ethyl acetate-glacial acetic acid is developing solvent, benzene: ethyl acetate: glacial acetic acid=92: 5: 5, launch, take out, dry, put under the ultra-violet lamp 254nm and inspect; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show identical avy blue speckle; Spray is placed a moment with the dinitrophenylhydrazine test solution, and speckle fades to orange red.
The method of quality control of the Yangrongbaicao medicinal ball of described treatment women anemia menoxenia, it is characterized in that: the method that described effect liquid phase chromatogram method is measured paeoniflorin content in the Yangrongbaicao medicinal ball prescription is:
(1) chromatographic condition and system suitability test: chromatographic column: octadecylsilane chemically bonded silica filler; Column temperature: 30 ℃; Acetonitrile: 0.1% phosphoric acid solution=14: 86 is a mobile phase; Flow velocity: 1.0mL/min; The detection wavelength is 230nm; Number of theoretical plate calculates by the peoniflorin peak should be not less than 2500;
(2) preparation of reference substance solution: it is an amount of to take by weighing the peoniflorin reference substance, accurately claims surely, puts in the 100mL measuring bottle, adds 50% methanol and is diluted to scale, shakes up, that is, every 1mL contains peoniflorin 0.1mg;
(3) preparation of need testing solution: take by weighing this product 1g, the accurate title, decide, and puts in the 100mL tool plug conical flask, adds 50% ethanol 50mL, claim decide weight, supersound process 30 minutes, taking-up is put cold, weighs and supplies the ultrasonic vector that subtracts, shake up, filter, get subsequent filtrate, promptly through 0.45 μ m filter membrane;
(4) algoscopy: draw each 5 μ L of above-mentioned two kinds of reference substance solution and need testing solution respectively, inject chromatograph of liquid, measure, promptly; The every gram of this product contains the Radix Paeoniae Alba with peoniflorin (C
23H
28O
11) meter, must not be less than 3.0mg
The invention beneficial effect: Pericarpium Citri Reticulatae that is provided with in the quality standard and Rhizoma Cyperi are differentiated project, with peoniflorin (C in this medicine of high effective liquid chromatography for measuring
23H
28O
11) content; Every gram contains the Radix Paeoniae Alba with peoniflorin (C
23H
28O
11) count, must not be less than the quantitative target and the method for inspection of 3.0mg, make quality standard comparatively perfect, the quality of strict control medicine is guaranteed to improve the quality of products.
Description of drawings
Fig. 1 is the thin-layer chromatogram of Pericarpium Citri Reticulatae in the Yangrongbaicao medicinal ball.
Fig. 1-the 1st, the thin-layer chromatogram of Pericarpium Citri Reticulatae in the Yangrongbaicao medicinal ball.
Fig. 1-2 is the thin-layer chromatogram of Pericarpium Citri Reticulatae in the Yangrongbaicao medicinal ball.
Fig. 2 is the thin-layer chromatogram of Rhizoma Cyperi in the Yangrongbaicao medicinal ball.
Fig. 2-the 1st, the thin-layer chromatogram of Rhizoma Cyperi in the Yangrongbaicao medicinal ball.
Fig. 2-the 2nd, the thin-layer chromatogram of Rhizoma Cyperi in the Yangrongbaicao medicinal ball.
Fig. 3-1 peoniflorin collection of illustrative plates.
Fig. 3-the 2nd, the negative sample collection of illustrative plates.
Fig. 4 peoniflorin standard regression curve chart.
Fig. 5 peoniflorin reference substance (precision test) figure.
Fig. 6-1 peoniflorin reference substance (lot number: 110736-200525) figure.
Fig. 6-2 Yangrongbaicao medicinal ball sample marked graph.
Fig. 7 Yangrongbaicao medicinal ball sample (lot number: 20050401) figure.
The specific embodiment
Below in conjunction with embodiment, the present invention is further described, the Yangrongbaicao medicinal ball quality standard after embodiment 1 improves:
Prescription: Radix Paeoniae Alba 160g Radix Angelicae Sinensis 80g Herba Taxilli 80g
The Radix Rehmanniae Preparata 80g Cortex Eucommiae (charcoal) 80g Rhizoma Chuanxiong 60g
Rhizoma Cyperi (vinegar system) 60g 40g Radix Ophiopogonis Pericarpium Citri Reticulatae 40g
Poria 40g Colla Corii Asini 20g Radix Glycyrrhizae 20g
The above 13 flavor 776g that amount to of Semen sojae atricolor 16g
Method for making: above 13 flavors, be ground into fine powder, sieve, mixing is used water pill, drying, promptly.
Character: this product is the watered pill of yellowish-brown to black; Feeble QI perfume (or spice), sweet in the mouth, hardship.
Differentiate: thin layer chromatography is differentiated
The thin layer chromatography of Pericarpium Citri Reticulatae is differentiated: gets this product powder 1g, adds water 30mL, and supersound process 15 minutes, the 20mL that adds diethyl ether extraction discards the ether layer, and water layer adds n-butyl alcohol 20mL extraction, discards water layer, alcohol layer evaporate to dryness, residue adds methanol 0.5mL makes dissolving, as need testing solution.Other gets the Hesperidin reference substance, adds methanol and makes the solution that every 1mL contains 2mg, in contrast product solution.Test according to thin layer chromatography (first appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 5～10 μ L of above-mentioned five kinds of solution, put respectively on the silica gel g thin-layer plate of same usefulness 0.5% sodium hydroxide solution preparation, with ethyl acetate-formic acid-glacial acetic acid-water=100: 11: 11: 26 was developing solvent, launch, take out, dry, spray is with to methoxyl group anisaldehyde solution, and it is clear to be heated to the speckle colour developing at 105 degrees centigrade.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
The thin layer chromatography of Rhizoma Cyperi is differentiated: get this product powder 5g, the 5mL that adds diethyl ether places 1h, and jolting constantly filters, and filtrate volatilizes, and residue adds ethyl acetate 0.5mL makes dissolving, as need testing solution.Other gets Rhizoma Cyperi medicinal powder 1g, and the 5mL that adds diethyl ether places 1h, and jolting constantly filters, and filtrate volatilizes, and residue adds ethyl acetate 0.5mL makes dissolving, and filtrate is as Rhizoma Cyperi medical material solution.Test according to thin layer chromatography (first appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 5～10 μ L of above-mentioned freshly prepd three kinds of solution, put respectively on same silica gel g thin-layer plate, with benzene-ethyl acetate-glacial acetic acid=92: 5: 5 was developing solvent, launch, take out, dry, put under the ultra-violet lamp (254nm) and inspect.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show identical avy blue speckle; Spray is placed a moment with the dinitrophenylhydrazine test solution, and speckle fades to orange red.
Check: this product should meet relevant every regulation under first appendix of Chinese Pharmacopoeia version in 2005 (appendix I A) pill item.
Assay: measure according to high performance liquid chromatography (first appendix VID of Chinese Pharmacopoeia version in 2005).
Chromatographic condition and system suitability test are filler with octadecylsilane chemically bonded silica; Acetonitrile-0.1% phosphoric acid solution=14: 86; The detection wavelength is 230nm.Number of theoretical plate calculates by the peoniflorin peak should be not less than 2500.
It is an amount of that the preparation of reference substance solution takes by weighing the peoniflorin reference substance, and accurate the title decides, and puts in the 100mL measuring bottle, adds 50% methanol and is diluted to scale, shakes up, promptly.(every 1mL contains peoniflorin 0.1mg)
The preparation of need testing solution takes by weighing this product 1g, and accurate the title decides, and puts in the 100mL tool plug conical flask, adds 50% ethanol 50mL, claim decide weight, supersound process 30 minutes, taking-up is put cold, weighs and supplies the ultrasonic vector that subtracts, shake up, filter, get subsequent filtrate, promptly through 0.45 μ m filter membrane.
Algoscopy is drawn above-mentioned two kinds of reference substance solution and each 5 μ L of need testing solution respectively, injects chromatograph of liquid, measures, promptly.The every gram of this product contains the Radix Paeoniae Alba with peoniflorin (C
23H
28O
11) meter, must not be less than 3.0mg.
Function with cure mainly: regulating menstruation and nourishing blood, nourishing kidney leukorrhagia stopping.Be used for women's anemia, the deficiency of YIN with the passing of time, menstruation water is transferred, and is expired more than, menalgia, leucorrhea is at present.
Usage and consumption: oral, a 5g, 2 times on the one.Specification: the heavy 1g of per 30 balls.Storage: airtight, protection against the tide.
Prescription: Radix Paeoniae Alba 160g Radix Angelicae Sinensis 80g Herba Taxilli 80g
The Radix Rehmanniae Preparata 80g Cortex Eucommiae (charcoal) 80g Rhizoma Chuanxiong 60g
Rhizoma Cyperi (vinegar system) 60g 40g Radix Ophiopogonis Pericarpium Citri Reticulatae 40g
Poria 40g Colla Corii Asini 20g Radix Glycyrrhizae 20g
The above 13 flavor 776g that amount to of Semen sojae atricolor 16g
Method for making: above 13 flavors, be ground into fine powder, sieve, mixing is used water pill, drying, promptly.
Character: this product is the watered pill of yellowish-brown to black; Feeble QI perfume (or spice), sweet in the mouth, hardship.
Thin layer chromatography is differentiated:
1. instrument and reagent
1.1 instrument: Ultrasound Instrument: KUDOS SK8200LH; Electronic balance: Sartorius CP225D/CP224S;
1.2 reagent: silica gel G F
254Lamellae, silica gel g thin-layer plate; Reference substance: (lot number: 110721-200512) above reference substance is all available from Nat'l Pharmaceutical ﹠ Biological Products Control Institute for Hesperidin; (lot number: 000511028) above medical material is all available from Tianjin prepared slices of Chinese crude drugs factory for the Rhizoma Cyperi medical material; Methanol is analytical pure; Ether, n-butyl alcohol, ethyl acetate, sodium bicarbonate, glacial acetic acid, normal hexane, dehydrated alcohol are analytical pure; Hydrochloric acid is analytical pure; Formic acid is analytical pure; Water is deionized water; Yangrongbaicao medicinal ball (lot number: 20050401,20050402,20050403) provide by Lerentang Pharmaceutical Factory, Zhongxin Pharmaceutical Group Co., Ltd., Tianj.
1.3 the thin layer chromatography of Yangrongbaicao medicinal ball is differentiated
1.3.1 the thin layer chromatography of Pericarpium Citri Reticulatae is differentiated: this product powder 1g is got in the preparation of need testing solution, add water 30mL, supersound process 15 minutes, the 20mL that adds diethyl ether extraction discards the ether layer, and water layer adds n-butyl alcohol 20mL extraction, discard water layer, alcohol layer evaporate to dryness, residue adds methanol 0.5mL makes dissolving, as need testing solution.
This product powder 1g is got in the preparation of Pericarpium Citri Reticulatae medical material solution, adds water 30mL, supersound process 15 minutes, and the 20mL that adds diethyl ether extraction discards the ether layer, and water layer adds n-butyl alcohol 20mL extraction, discards water layer, alcohol layer evaporate to dryness, residue adds methanol 0.5mL makes dissolving, as need testing solution.
The Hesperidin reference substance is got in the preparation of reference substance solution, adds methanol and makes the solution that every 1mL contains 2mg, in contrast product solution.
The preparation of negative control solution by all medicines of prescription preparation, according to test sample preparation technology operation, is got 1g and is ground well except that Pericarpium Citri Reticulatae, according to the preparation method item operation down of need testing solution, as the Pericarpium Citri Reticulatae negative control solution.
Test according to thin layer chromatography (first appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 5～10 μ L of above-mentioned five kinds of solution, put respectively on the silica gel g thin-layer plate of same usefulness 0.5% sodium hydroxide solution preparation, with ethyl acetate-formic acid-glacial acetic acid-water=100: 11: 11: 26 was developing solvent, launch, take out, dry, spray is with to methoxyl group anisaldehyde solution, and it is clear to be heated to the speckle colour developing at 105 ℃.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.Illustrate that this method is exclusive, reliable.(the results are shown in Figure 1-1,1-2)
Because by the operation of pharmacopeia method, the exhibition journey of Hesperidin reference substance is lacked (seeing the thin-layer chromatogram of Pericarpium Citri Reticulatae in Fig. 1 Yangrongbaicao medicinal ball), so use above method operation instead.
1. Hesperidin reference substance (111524-200503)
2. negative
3. Yangrongbaicao medicinal ball (20060401) test sample
4. Pericarpium Citri Reticulatae medical material]
The thin-layer chromatogram of Pericarpium Citri Reticulatae in Fig. 1-1 Yangrongbaicao medicinal ball, a Fig. 1-1:[left side → right side: Hesperidin reference substance (110721-200512), Pericarpium Citri Reticulatae medical material, Yangrongbaicao medicinal ball (20060401) test sample, feminine gender]
Fig. 1-2: the thin-layer chromatogram of Pericarpium Citri Reticulatae in the Yangrongbaicao medicinal ball, Fig. 1-2: [left side → right side: Hesperidin reference substance (110721-200512), Yangrongbaicao medicinal ball (20060401), Yangrongbaicao medicinal ball (20060402) Yangrongbaicao medicinal ball (20060403) test sample, feminine gender]
1.3.2 the thin layer chromatography of Rhizoma Cyperi is differentiated:
This product powder 5g is got in the preparation of need testing solution, and the 5mL that adds diethyl ether places 1h, and jolting constantly filters, and filtrate volatilizes, and residue adds ethyl acetate 0.5mL makes dissolving, as need testing solution.
Preparation this product powder 1g of Rhizoma Cyperi medical material solution, the 5mL that adds diethyl ether places 1h, and jolting constantly filters, and filtrate volatilizes, and residue adds ethyl acetate 0.5mL makes dissolving, and filtrate is as need testing solution.
The preparation of negative control solution by all medicines of prescription preparation, according to test sample preparation technology operation, is got 5g and is ground well except that Rhizoma Cyperi, according to the preparation method item operation down of need testing solution, as the Rhizoma Cyperi negative control solution.
Test according to thin layer chromatography (first appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 5～10 μ L of above-mentioned freshly prepd three kinds of solution, put respectively on same silica gel g thin-layer plate, with benzene-ethyl acetate-glacial acetic acid=92: 5: 5 was developing solvent, launch, take out, dry, put under the ultra-violet lamp (254nm) and inspect.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show identical avy blue speckle; Spray is placed a moment with the dinitrophenylhydrazine test solution, and speckle fades to orange red.Illustrate that this method is exclusive, reliable.The results are shown in Figure 2-1,2-2)
Owing to select GF for use with the pharmacopeia method
254Plate is inspected under 254 (nm), and the not obvious (see figure 2) of the corresponding speckle of Rhizoma Cyperi medical material is so select above method for use.
Fig. 2: the thin-layer chromatogram of Rhizoma Cyperi in the Yangrongbaicao medicinal ball, Fig. 2: [left side → right side: 1. Rhizoma Cyperi medical material, 2. Yangrongbaicao medicinal ball (20050401) test sample, 3. feminine gender]
Fig. 2-1: the thin-layer chromatogram of Rhizoma Cyperi in the Yangrongbaicao medicinal ball, a Fig. 2-1:[left side → right side: 1. Rhizoma Cyperi medical material, 2. Yangrongbaicao medicinal ball (20050401) test sample, 3. feminine gender]
Fig. 2-2: the thin-layer chromatogram of Rhizoma Cyperi in the Yangrongbaicao medicinal ball, a Fig. 2-2:[left side → right side: 1. Rhizoma Cyperi medical material, 2. Yangrongbaicao medicinal ball (20060101), 3. Yangrongbaicao medicinal ball (20060202,4. Yangrongbaicao medicinal ball (20060203) test sample, 5. feminine gender).
Check: this product should meet relevant every regulation under first appendix of Chinese Pharmacopoeia version in 2005 (appendix I A) pill item.
Assay: measure according to high performance liquid chromatography (first appendix VID of Chinese Pharmacopoeia version in 2005).
One, instrument and reagent
1. instrument condition: instrument: Tianjin, island LC-2010A
HTThe type high performance liquid chromatograph; Detector: UV/VIS; Detect wavelength: 230nm; Column temperature: 30 ℃; Ultrasound Instrument: KUDOS SK8200LH; Electronic balance: SartoriusCP225D/CP224S.
2. chromatographic condition: chromatographic column:
C18 5 μ m 4.6 * 150mm Column
Mobile phase: acetonitrile-0.1% phosphoric acid solution (14: 86); Flow velocity: 1.0mL/min; Sample size: 5 μ L
3. reagent
Peoniflorin reference substance: available from Nat'l Pharmaceutical ﹠ Biological Products Control Institute (using) for assay
Lot number: 110736-200525
Methanol is chromatographically pure, and other reagent is analytical pure, and water is deionized water.
Yangrongbaicao medicinal ball (lot number: 20050401,20050402,20050403)
Provide by Lerentang Pharmaceutical Factory, Zhongxin Pharmaceutical Group Co., Ltd., Tianj
Two, the investigation of experimental technique
1. chromatographic condition and system suitability test: with octadecylsilane chemically bonded silica is filler; Acetonitrile-0.1% phosphoric acid solution (14: 86) methanol-be mobile phase; The detection wavelength is 230nm.Number of theoretical plate calculates by the peoniflorin peak should be not less than 2500.
2. the investigation of extraction conditions
2.1 the investigation of extracting method
Ultrasonic method: the extraction solvent of water, 30%, 50%, four kinds of opposed polarities of 80% ethanol is adopted in this experiment, utilizes ultrasonic extracting method to investigate; The results are shown in Table 1
The different extraction solvent of table 1 are measured the integral area of peoniflorin
Heat reflow method: the extraction solvent of water, 30%, 50%, four kinds of opposed polarities of 80% ethanol is adopted in this experiment, investigates with the hot reflux method; The results are shown in Table 2
The different extraction solvent of table 2 are measured the integral area of peoniflorin
Though it is higher that heat reflow method extracts paeoniflorin content, both differ and not quite, for to testing simple consideration, select the extracting method of ultrasonic method as peoniflorin for use so follow pharmacopeia.
2.2 the investigation of extraction time
The employing ultrasonic method is handled, and this experiment has been done the gradient of an extraction time and investigated, and is extraction solvent with 50% ethanol, and supersound process was investigated in 10,20,30,40 minutes, the results are shown in Table 3
The investigation result of table 3 extraction time
Experimental result shows: ultrasonic method is handled the 30min peoniflorin and can be extracted fully.
3. the selection of condition determination
3.1 the selection of mobile phase: experiment shows, selects for use under first the 68th page of Radix Paeoniae Alba assay item of Chinese Pharmacopoeia version in 2005 to be mobile phase with acetonitrile-0.1% phosphoric acid solution (14: 86); The detection wavelength is 230nm, and the chromatographic peak of peoniflorin chromatographic peak and other compositions disturbs less.
3.2 the selection of wavelength: select 230nm for detecting wavelength down according to first the 68th page of Radix Paeoniae Alba assay item of Chinese Pharmacopoeia version in 2005.
3.3 the preparation of reference substance solution: it is an amount of to take by weighing the peoniflorin reference substance, accurately claims surely, puts in the 100mL measuring bottle, adds 50% methanol and is diluted to scale, makes the solution that every 1mL contains 0.1mg, in contrast product solution.
3.4 the preparation of need testing solution: take by weighing Yangrongbaicao medicinal ball 1g, the accurate title, decide, and puts in the 100mL tool plug conical flask, adds 50% ethanol 50mL, claim decide weight, supersound process 30 minutes, taking-up is put cold, weighs and supplies the ultrasonic vector that subtracts, shake up, filter through 0.45 μ m filter membrane, as need testing solution.
3.5 the preparation of negative control solution: except that the Radix Paeoniae Alba,,, get 1.0g,, be prepared as negative solution according to the operation of need testing solution preparation method according to the operation of test sample preparation process by all medicines of prescription preparation.
3.6 algoscopy: measure according to high performance liquid chromatography (appendix VID), draw each 5 μ L of reference substance solution, need testing solution and negative control solution respectively, inject chromatograph of liquid, measure, calculate with external standard method, promptly.
3.7 blank assay: by sample prescription take by weighing except that the flavour of a drug of the Radix Paeoniae Alba an amount of, make blank by the test sample preparation method, measure with the chromatographic condition identical with measuring test sample, measure reference substance simultaneously, the result shows that negative sample goes out the place, peak at the peoniflorin reference substance and do not have the peak.(seeing Fig. 3-1, Fig. 3-2)
4. methodological study experiment
4.1 linear relationship is investigated: the preparation of peoniflorin reference substance solution: it is an amount of that precision takes by weighing the peoniflorin reference substance, makes the reference substance solution that every 1mL contains peoniflorin 90 μ g with methanol.Draw above-mentioned solution 2.0,4.0,5.0,6.0,8.0,10 μ L, peak area is measured in sample introduction analysis successively, the results are shown in Table 4
Table 4 linear relationship is investigated the result
Table 5 precision experimental result
With the sample size is abscissa, and peak area is a vertical coordinate, and the drawing standard curve is learned by statistics and calculated regression equation: Y=1.360 * 10
6 X-2.314 * 10
4, r=0.9992.(see figure 4).
4.2 precision test
The accurate peoniflorin reference substance solution 5 μ L (concentration is 0.09mg/mL) that draw repeat sample introduction 5 times, measure the peak area integrated value by above-mentioned chromatographic condition.Calculating the peoniflorin relative standard deviation is RSD=0.64%.The results are shown in Table 5 (see figure 5)s
4.3 recovery test
Take by weighing sample 0.5g (lot number is 20050401, and content is 4.257mg/g), 5 parts, the accurate title, decide, and puts in the 100mL tool plug conical flask, and the adding reference substance is an amount of, presses the test sample preparation and measure operation under the item, and sample introduction is measured, and calculates.Average recovery rate is 100.0% as a result, relative standard deviation RSD=1.74%.The results are shown in Table 6
Table 6 response rate experimental result
Through The effects, the response rate is 100.0%, and RSD is 1.74%.Meet the requirements.(seeing Fig. 6-1, Fig. 6-2)
4.4 repeatability test: the sample (lot number: 20060401), take by weighing 5 parts, extract, analyze, record the peak area integrated value of getting same lot number by the content assaying method of drafting.Calculating relative standard deviation is 0.99%, the results are shown in Table 7
Table 7 repeatability experimental result
4.5 stability test: with the need testing solution sample introduction analysis for preparing, every 1 hour sample introduction, totally 4 times, measure the peak area integrated value by above-mentioned chromatographic condition, calculating relative standard deviation is 0.23%.The results are shown in Table 8
Table 8 stability experiment result
Through The effects, show that test sample is stable in 4 hours, relative standard deviation is 0.23%.
4.6 sample determination
According to drafting method, draw each 5 μ L of reference substance solution and need testing solution, sample introduction is analyzed.Measured content of paeoniflorin in 3 batches of Yangrongbaicao medicinal balls altogether.The results are shown in Table 6 (see figure 7)s
Paeoniflorin content measurement result in table 6 test sample
Limit: the every gram of this product contains the Radix Paeoniae Alba with peoniflorin (C
23H
28O
11) meter, must not be less than 3.0mg.
Claims (3)
1. method of quality control for the treatment of the Yangrongbaicao medicinal ball of women's anemia menoxenia, wherein said pharmaceutical formulation are by Chinese crude drug:
Radix Paeoniae Alba 160g Radix Angelicae Sinensis 80g Herba Taxilli 80g
The Radix Rehmanniae Preparata 80g Cortex Eucommiae (charcoal) 80g Rhizoma Chuanxiong 60g
Rhizoma Cyperi (vinegar system) 60g 40g Radix Ophiopogonis Pericarpium Citri Reticulatae 40g
Poria 40g Colla Corii Asini 20g Radix Glycyrrhizae 20g
Semen sojae atricolor 16g forms, and above-mentioned raw materials is ground into fine powder, sieves, and mixing is used water pill, and drying promptly, is characterized in that: the step of its method is:
(1) being reference substance with the Hesperidin, is control medicinal material with the Rhizoma Cyperi, and thin layer chromatography differentiates in the Yangrongbaicao medicinal ball prescription whether contain Pericarpium Citri Reticulatae, Rhizoma Cyperi composition;
(2) with the peoniflorin be reference substance, high performance liquid chromatography detects content of paeoniflorin in the Yangrongbaicao medicinal ball prescription.
2. the method for quality control of the Yangrongbaicao medicinal ball of treatment women anemia according to claim 1 menoxenia, it is characterized in that: described thin layer chromatography differentiates that the method that whether contains the Pericarpium Citri Reticulatae composition in the Yangrongbaicao medicinal ball prescription is: get this product powder 1g, add water 30mL, supersound process 15 minutes, the 20mL that adds diethyl ether extraction, discard the ether layer, water layer adds n-butyl alcohol 20mL extraction, discards water layer, alcohol layer evaporate to dryness, residue adds methanol 0.5mL makes dissolving, as need testing solution; Other gets the Hesperidin reference substance, adds methanol and makes the solution that every 1mL contains 2mg, in contrast product solution; Test according to thin layer chromatography, draw each 5～10 μ L of above-mentioned five kinds of solution, put respectively on the silica gel g thin-layer plate of same usefulness 0.5% sodium hydroxide solution preparation, with ethyl acetate-formic acid-glacial acetic acid-water is developing solvent, ethyl acetate: formic acid: glacial acetic acid: water=100: 11: 11: 26, launch, take out, dry, spray is with to methoxyl group anisaldehyde solution, and it is clear to be heated to the speckle colour developing at 105 degrees centigrade; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;
Described thin layer chromatography differentiates that the method that whether contains the Rhizoma Cyperi composition in the Yangrongbaicao medicinal ball prescription is: get this product powder 5g, the 5mL that adds diethyl ether places 1h, and jolting constantly filters, and filtrate volatilizes, and residue adds ethyl acetate 0.5mL makes dissolving, as need testing solution; Other gets Rhizoma Cyperi medicinal powder 1g, and the 5mL that adds diethyl ether places 1h, and jolting constantly filters, and filtrate volatilizes, and residue adds ethyl acetate 0.5mL makes dissolving, and filtrate is as Rhizoma Cyperi medical material solution; According to the thin layer chromatography test, draw each 5～10 μ L of above-mentioned freshly prepd three kinds of solution, put respectively on same silica gel g thin-layer plate, with benzene-ethyl acetate-glacial acetic acid is developing solvent, benzene: ethyl acetate: glacial acetic acid=92: 5: 5, launch, take out, dry, put under the ultra-violet lamp 254nm and inspect; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show identical avy blue speckle; Spray is placed a moment with the dinitrophenylhydrazine test solution, and speckle fades to orange red.
3. the method for quality control of the Yangrongbaicao medicinal ball of treatment women anemia according to claim 1 menoxenia is characterized in that: the method for paeoniflorin content is in the described high effective liquid chromatography for measuring Yangrongbaicao medicinal ball prescription:
(1) chromatographic condition and system suitability test: chromatographic column: octadecylsilane chemically bonded silica filler; Column temperature: 30 ℃; Acetonitrile: 0.1% phosphoric acid solution=14: 86 is a mobile phase; Flow velocity: 1.0mL/min; The detection wavelength is 230nm; Number of theoretical plate calculates by the peoniflorin peak should be not less than 2500;
(2) preparation of reference substance solution: it is an amount of to take by weighing the peoniflorin reference substance, accurately claims surely, puts in the 100mL measuring bottle, adds 50% methanol and is diluted to scale, shakes up, that is, every 1mL contains peoniflorin 0.1mg;
(3) preparation of need testing solution: take by weighing this product 1g, the accurate title, decide, and puts in the 100mL tool plug conical flask, adds 50% ethanol 50mL, claim decide weight, supersound process 30 minutes, taking-up is put cold, weighs and supplies the ultrasonic vector that subtracts, shake up, filter, get subsequent filtrate, promptly through 0.45 μ m filter membrane;
(4) algoscopy: draw each 5 μ L of above-mentioned two kinds of reference substance solution and need testing solution respectively, inject chromatograph of liquid, measure, promptly; The every gram of this product contains the Radix Paeoniae Alba with peoniflorin (C
23H
28O
11) meter, must not be less than 3.0mg.
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102507836A (en) * | 2011-10-27 | 2012-06-20 | 天津中新药业集团股份有限公司达仁堂制药厂 | Quality detection method of menstruation regulating pill |
| CN103948766A (en) * | 2014-05-09 | 2014-07-30 | 赵晓宏 | Traditional Chinese medicine for treating irregular menstrual periods |
| CN104758533A (en) * | 2015-04-26 | 2015-07-08 | 王伟伟 | Traditional Chinese medicine composition for treating irregular menstruation |
| CN105203696A (en) * | 2015-10-27 | 2015-12-30 | 河北中医学院 | Multi-developing solvent rapid thin-layer chromatographic method for medicinal material donkey-hide gelatin and water extract thereof |
| CN110579553A (en) * | 2019-09-29 | 2019-12-17 | 江苏省中医院 | Quality detection method of Shenkui Tongmai granules |
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2009
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102507836A (en) * | 2011-10-27 | 2012-06-20 | 天津中新药业集团股份有限公司达仁堂制药厂 | Quality detection method of menstruation regulating pill |
| CN103948766A (en) * | 2014-05-09 | 2014-07-30 | 赵晓宏 | Traditional Chinese medicine for treating irregular menstrual periods |
| CN104758533A (en) * | 2015-04-26 | 2015-07-08 | 王伟伟 | Traditional Chinese medicine composition for treating irregular menstruation |
| CN105203696A (en) * | 2015-10-27 | 2015-12-30 | 河北中医学院 | Multi-developing solvent rapid thin-layer chromatographic method for medicinal material donkey-hide gelatin and water extract thereof |
| CN110579553A (en) * | 2019-09-29 | 2019-12-17 | 江苏省中医院 | Quality detection method of Shenkui Tongmai granules |
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