CN102119997A - Method for establishing HPLC (high performance liquid chromatography) finger-print of ophiopogon japonicus and standard finger-print thereof - Google Patents
Method for establishing HPLC (high performance liquid chromatography) finger-print of ophiopogon japonicus and standard finger-print thereof Download PDFInfo
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- CN102119997A CN102119997A CN2010101065857A CN201010106585A CN102119997A CN 102119997 A CN102119997 A CN 102119997A CN 2010101065857 A CN2010101065857 A CN 2010101065857A CN 201010106585 A CN201010106585 A CN 201010106585A CN 102119997 A CN102119997 A CN 102119997A
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Abstract
The invention provides a method for establishing an HPLC (high performance liquid chromatography) finger-print of ophiopogon japonicus and a standard finger-print thereof. The method comprises the following steps of: preparing a test article solution; preparing a reference substance solution; selecting chromatographic conditions; and measuring with a high performance liquid chromatograph to obtain the finger-print of ophiopogon japonicus. 16 batches of HPLC finger-prints of ophiopogon japonicus are compared to determine the common finger-print characteristics of ophiopogon japonicus, thereby obtaining the standard finger-print. The method provided by the invention is simple, convenient, accurate and reliable, and has the characteristics of good reproducibility, multiple characteristic peaks and the like. The established standard finger-print can effectively show the quality of ophiopogon japonicus.
Description
Technical field
The present invention relates to the construction method of Chinese crude drug finger printing, especially the construction method and the standard finger-print thereof of river medical material Radix Ophiopogonis high performance liquid chromatography (HPLC) finger printing.Belong to the pharmaceutical analysis technical field.
Background technology
Chinese medicine fingerprint refers in particular to the collection of illustrative plates or the image of each component population characteristic of Chinese medicine that the Chinese medicine sample obtains through modern analytical techniques such as spectrum or chromatographs.Chinese medicine fingerprint at present, admitted by numerous international medical mechanism as a kind of comprehensive, quantifiable quality control method, become one of effective method of Chinese patent medicine, natural drug quality control.FDA (Food and Drug Adminstration) is in " FDA is about the guide of vegetable products " formulated in 1996, and requiring provides corresponding finger printing to product and plant amedica product in the middle of plant material, the plant amedica.Medicinal and the fragrant plant association of British Herbal Pharmacopoeia, India herbal medicine allusion quotation and Canada, German medicinal plants association also accept chromatographic fingerprinting.China national drug regulatory department has issued " specification requirement (provisional) of the finger printing research of Chinese medicine " in 2000, explicitly call for Chinese medicine of newly declaring and the Chinese medicine that has gone on the market are carried out the finger printing standard.
Chinese medicine fingerprint has passed through the research of many decades, many modern analysis means of testing such as comprising ultraviolet, infrared, gas phase, high performance liquid chromatogram, thin layer, nuclear magnetic resonance, NMR, scanning electron microscope, computer image analysis, electrophoretic techniques, isoenzyme analysis method, Protocols in Molecular Biology, cluster analysis technology have been used, for the material of Chinese medicine is identified and component analysis has accumulated lot of data.But, also reach the purpose that reflects effectively and control the total quality of Chinese medicine far away from the angle of Chinese medicine overall quality control.High performance liquid chromatography (HPLC) finger printing obtains approval in the world easily, comprises that many countries of the U.S., Britain, France, Canada, Germany, Japan and India more are ready to accept the HPLC finger printing.Therefore, setting up corresponding HPLC finger printing for Chinese crude drug, is the effective means of present stage control Chinese medicine inherent quality.
Be the dried root of Liliaceae Ophiopogon herbaceos perennial Ophiopogon Radix Ophiopogonis japonicus (Thunb.) Ker-Gawl. Radix Ophiopogonis, and Sichuan is one of its maximum main producing region.Be usually used in diseases such as dryness of the lung dry cough, cough due to consumptive disease, thirsty, the vexed insomnia of Tianjin wound, interior-heat are quenched one's thirst, dryness of the intestine constipation on the tcm clinical practice.Contain the number of chemical composition Radix Ophiopogonis, since report in 1936 contains vitamin a compound in the tuber Radix Ophiopogonis, successively from Radix Ophiopogonis, isolated chemical compounds such as saccharide, saponin, flavone; At present, " Chinese pharmacopoeia is single to the quality evaluation mode of Radix Ophiopogonis, is not enough to system, intactly shows the total quality of medical material Radix Ophiopogonis, needs more advanced, the more effective quality control method of exploration badly.
Summary of the invention
The method for building up that the purpose of this invention is to provide a kind of river medicinal materials fingerprint Radix Ophiopogonis, and the resulting river of method medical material Radix Ophiopogonis standard finger-print thus.For realizing this purpose, the present invention is by the research to river medical material Radix Ophiopogonis efficient liquid-phase chromatograph finger print atlas, proposed a kind of better river quality of medicinal material Radix Ophiopogonis control method, remedied the deficiency of existing Quality Control Technology, the quality control of river medical material Radix Ophiopogonis is improved and science more.
The method for building up of river of the present invention medical material Radix Ophiopogonis efficient liquid-phase chromatograph finger print atlas comprises the steps:
(1) preparation of need testing solution: precision takes by weighing medicinal powder weighing sample Radix Ophiopogonis 3g to tool plug container, adds 95% ethanol water 50ml, (20 ℃ of ultrasonic concussions, 90min), membrane filtration, the filtrate rotary evaporation is to (50 ℃ of 2-3ml, 80r/min) use the 40ml water dissolution, be transferred in the centrifuge tube extracted with diethyl ether 4 times, each 20ml, with ether layer rotary evaporation to 3～5ml, methanol constant volume is crossed the disposable filter membrane of 0.45 μ m to 10ml, and filtrate is as need testing solution.
(2) efficient liquid phase chromatographic analysis: chromatography column is Symmetry-C
18(250 * 4.6mm, 5 μ m); Mobile phase is methanol-0.2% aqueous acetic acid, gradient elution, and elution program is: 0min → 20min → 38min → 67min → 90min, methanol 15% → 32% → 67% → 74% → 100%; Flow velocity: 1.0mL/min detects wavelength: 280nm, sample size: 15 μ L, column temperature: 30 ℃.Analyze need testing solution with this understanding, obtain the finger printing of river medical material Radix Ophiopogonis.
According to method provided by the invention, 16 batches of river medical materials Radix Ophiopogonis have been set up the HPLC finger printing, by analyzing relatively, determined 25 common characteristic peaks.
The retention time of characteristic peak is respectively: 5.2min, 17.2min, 18.1min, 23.4min, 32.4min, 36.5min, 41.4min, 48.6min, 51.2min, 52.7min, 55.6min, 57.0min, 64.4min, 67.1min, 68.0min, 69.6min, 76.7min, 77.2min, 78.1min, 78.7min, 79.3min, 80.0min, 80.9min, 83.3min, 87.0min, these common characteristic peaks have constituted the fingerprint characteristic of river medical material Radix Ophiopogonis, can be used as the standard finger-print of river medical material Radix Ophiopogonis.
The present invention has following significant advantage and purposes:
(1) river medical material Radix Ophiopogonis is done as a whole treating, can find out nuance between the different medical materials by its total peak relatively;
(2) test sample is simple for production, and chromatographic condition is realized easily;
(3) stability of method and repeatability are all relatively good;
(4) be suitable for discriminating and control to the river medical material Radix Ophiopogonis true and false, the place of production and quality.
Description of drawings
Fig. 1 is the standard finger-print of river provided by the invention medical material Radix Ophiopogonis.From left to right marked No. 1 to 25, its common characteristic peak among the figure respectively.
The specific embodiment
The preferred embodiments of the present invention are as follows:
(1) instrument and medicine: use Agilent 1101 high performance liquid chromatographs; Totally 16 batches of river medical materials Radix Ophiopogonis all are collected in Mianyang City Santai County, Sichuan Province, genuine producing region Radix Ophiopogonis, river; Other reagent is chromatographically pure or analytical pure.
(2) precision takes by weighing medicinal powder weighing sample Radix Ophiopogonis 3g to tool plug container, adds 95% ethanol water 50ml, (20 ℃ of ultrasonic concussions, 90min), membrane filtration, the filtrate rotary evaporation is to (50 ℃ of 2-3ml, 80r/min) use the 40ml water dissolution, be transferred in the centrifuge tube extracted with diethyl ether 4 times, each 20ml, with ether layer rotary evaporation to 3～5ml, methanol constant volume is crossed the disposable filter membrane of 0.45 μ m to 10ml, and filtrate is as need testing solution.
(3) efficient liquid phase chromatographic analysis: the chromatographic isolation column packing is 5 μ m Symmetry-C
18(250 * 4.6mm); Mobile phase is methanol-0.2% aqueous acetic acid, gradient elution, and elution program is: 0min → 20min → 38min → 67min → 90min, methanol 15% → 32% → 67% → 74% → 100%; Flow velocity: 1.0mL/min detects wavelength: 280nm, sample size: 15 μ L, column temperature: 30 ℃.Analyze need testing solution with this understanding, obtain the finger printing of river medical material Radix Ophiopogonis.
(4) according to method provided by the invention, 16 batches of river medical materials Radix Ophiopogonis have been set up the HPLC finger printing, by analyzing relatively, 25 common characteristic peaks have been determined, its retention time is respectively: 5.2min, 17.2min, 18.1min, 23.4min, 32.4min, 36.5min, 41.4min, 48.6min, 51.2min, 52.7min, 55.6min, 57.0min, 64.4min, 67.1min, 68.0min, 69.6min, 76.7min, 77.2min, 78.1min, 78.7min, 79.3min, 80.0min, 80.9min, 83.3min, 87.0min, these common characteristic peaks have constituted the fingerprint characteristic of river medical material Radix Ophiopogonis, can be used as the standard finger-print of river medical material Radix Ophiopogonis.
Claims (4)
1. the method for building up of a river medical material Radix Ophiopogonis efficient liquid-phase chromatograph finger print atlas, its feature comprise preparation, the making of finger printing and the determining of standard finger-print of need testing solution.
2. the method for building up of river according to claim 1 medical material Radix Ophiopogonis efficient liquid-phase chromatograph finger print atlas, the preparation method that it is characterized in that described need testing solution is: precision takes by weighing medicinal powder weighing sample Radix Ophiopogonis 3g to tool plug container, add 95% ethanol water 50ml, (20 ℃ of ultrasonic concussions, 90min), membrane filtration, the filtrate rotary evaporation to 2-3ml (50 ℃, 80r/min) use the 40ml water dissolution, be transferred in the centrifuge tube, extracted with diethyl ether 4 times, each 20ml, with ether layer rotary evaporation to 3～5ml, methanol constant volume is to 10ml, cross the disposable filter membrane of 0.45 μ m, filtrate is as need testing solution.
3. according to the method for building up of claim 1,2 described river medical material Radix Ophiopogonis efficient liquid-phase chromatograph finger print atlas, it is characterized in that the manufacturing process of described finger printing is: chromatography column is Symmetry-C
18(250 * 4.6mm, 5 μ m); Mobile phase is methanol-0.2% aqueous acetic acid, gradient elution, and elution program is: 0min → 20min → 38min → 67min → 90min, methanol 15% → 32% → 67% → 74% → 100%; Flow velocity: 1.0mL/min detects wavelength: 280nm, sample size: 15 μ L, column temperature: 30 ℃.Analyze need testing solution with this understanding, obtain the finger printing of river medical material Radix Ophiopogonis.
4. according to claim 1,2, the method for building up of 3 described river medical material Radix Ophiopogonis efficient liquid-phase chromatograph finger print atlas, the definite method that it is characterized in that described standard finger-print is: according to method provided by the invention, 16 batches of river medical materials Radix Ophiopogonis have been set up the HPLC finger printing, by analyzing relatively, 25 common characteristic peaks have been determined, retention time is respectively: 5.2min, 17.2min, 18.1min, 23.4min, 32.4min, 36.5min, 41.4min, 48.6min, 51.2min, 52.7min, 55.6min, 57.0min, 64.4min, 67.1min, 68.0min, 69.6min, 76.7min, 77.2min, 78.1min, 78.7min, 79.3min, 80.0min, 80.9min, 83.3min, 87.0min, these common characteristic peaks have constituted the fingerprint characteristic of river medical material Radix Ophiopogonis, can be used as the standard finger-print of river medical material Radix Ophiopogonis.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2010101065857A CN102119997B (en) | 2010-01-29 | 2010-01-29 | Method for establishing HPLC (high performance liquid chromatography) finger-print of ophiopogon japonicus and standard finger-print thereof |
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| CN2010101065857A CN102119997B (en) | 2010-01-29 | 2010-01-29 | Method for establishing HPLC (high performance liquid chromatography) finger-print of ophiopogon japonicus and standard finger-print thereof |
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| CN102119997A true CN102119997A (en) | 2011-07-13 |
| CN102119997B CN102119997B (en) | 2012-08-22 |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109541098A (en) * | 2018-11-27 | 2019-03-29 | 华润三九（雅安）药业有限公司 | Radix Ophiopogonis finger-print, its construction method and Radix Ophiopogonis quality determining method |
| CN109633012A (en) * | 2018-12-29 | 2019-04-16 | 正大青春宝药业有限公司 | A kind of discrimination method of Zhejiang Radix Ophiopogonis |
| CN111024855A (en) * | 2019-12-30 | 2020-04-17 | 常熟雷允上制药有限公司 | Fingerprint detection method for traditional Chinese medicine radix ophiopogonis |
| JP6991432B1 (en) * | 2021-04-02 | 2022-01-12 | 中国計量大学 | Dwarf lilyturf origin identification method |
-
2010
- 2010-01-29 CN CN2010101065857A patent/CN102119997B/en not_active Expired - Fee Related
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109541098A (en) * | 2018-11-27 | 2019-03-29 | 华润三九（雅安）药业有限公司 | Radix Ophiopogonis finger-print, its construction method and Radix Ophiopogonis quality determining method |
| CN109541098B (en) * | 2018-11-27 | 2020-11-10 | 华润三九（雅安）药业有限公司 | Radix ophiopogonis fingerprint spectrum, construction method thereof and radix ophiopogonis quality detection method |
| CN109633012A (en) * | 2018-12-29 | 2019-04-16 | 正大青春宝药业有限公司 | A kind of discrimination method of Zhejiang Radix Ophiopogonis |
| CN109633012B (en) * | 2018-12-29 | 2021-11-12 | 正大青春宝药业有限公司 | Identification method of Zhejiang ophiopogon root |
| CN111024855A (en) * | 2019-12-30 | 2020-04-17 | 常熟雷允上制药有限公司 | Fingerprint detection method for traditional Chinese medicine radix ophiopogonis |
| JP6991432B1 (en) * | 2021-04-02 | 2022-01-12 | 中国計量大学 | Dwarf lilyturf origin identification method |
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| CN102119997B (en) | 2012-08-22 |
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