CN101991661B - Method for detecting Chinese patent drug containing at least two of white paeony root, ginseng, salvia miltiorrhiza, sweet wormwood, liquorice and angelica sinensis - Google Patents
Method for detecting Chinese patent drug containing at least two of white paeony root, ginseng, salvia miltiorrhiza, sweet wormwood, liquorice and angelica sinensis Download PDFInfo
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- CN101991661B CN101991661B CN2009101664780A CN200910166478A CN101991661B CN 101991661 B CN101991661 B CN 101991661B CN 2009101664780 A CN2009101664780 A CN 2009101664780A CN 200910166478 A CN200910166478 A CN 200910166478A CN 101991661 B CN101991661 B CN 101991661B
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Abstract
The invention provides a method for detecting a Chinese patent drug containing at least two of white paeony root, ginseng, salvia miltiorrhiza, sweet wormwood, liquorice and angelica sinensis, comprising the steps as follows: preparing a sample solution and a comparison product drug solution, adopting acetonitrile and water as mobile phases, and performing high performance liquid chromatography (HPLC) measurement, wherein the detection wavelength is 201-290nm; and detecting whether a corresponding chromatographic peak exists at a position, which has the same reservation time with the chromatograph of a comparison drug, in the chromatography of a sample. Compared with the traditional thin-layer chromatography identification method, the detection method in the invention has the advantages that the mode of processing a sample containing various drugs in a liquid phase chromatograph is changed, the mixed peak in the liquid phase chromatography except the components to be detected in the Chinese patent drug is successfully eliminated, and results are more accurate and stable.
Description
Technical field
The invention provides the detection method of effective constituent in a kind of medicine, it is a kind of detection method of middle pharmaceutically active ingredient, refers in particular to the detection method of effective constituent in the Chinese medicine preparation that contains two or more medicinal ingredient in the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii, the Radix Angelicae Sinensis.
Background technology
Chinese medicine compound prescription is principal mode and the means of tcm clinical practice medication, the medicine of respectively distinguishing the flavor of in the side contains the number of chemical composition again, these intrinsic chemical constitutions are material bases of compound performance drug action, but the whole curative effect that medical drugs embodied during arbitrary active component can not reflect again comprehensively in the side, this is Chinese medicine compound prescription and the fundamental difference of chemosynthesis medicine in formulating the quality standard process, so the analysis-by-synthesis of macroscopic view becomes the inexorable trend of Chinese medicine compound prescription development.Chinese medicine compound prescription has been used more than one thousand years as the marrow of traditional Chinese medicine, and from the angle of modern study, a Chinese medicine compound prescription has characteristics such as multicomponent, multiaction, many target spots.Put down in writing in very will discussing greatly " element is asked ": " the meaning monarch of main disease, the drug in a prescription helping monarch-drug is called miniSter answers the meaning of minister to make ".Along with the modernization of Chinese medicine, international deeply development, press for set up a kind of to Chinese medicine and Chinese medicine compound prescription product from the former method of expecting that finished product carries out quality control and its complicated ingredient is detected, finger-print arises at the historic moment.
" fingerprint " (finger printing) identifies and derives from medical jurisprudence that everyone fingerprint is had nothing in common with each other in small detail.According to these difference, by " comparison " mode, can determine to differentiate everyone feature.Dna fingerprinting has appearred in the development along with modern biotechnology.By the dna fingerprinting analysis, can differentiate evaluation to biosomes such as people, animal, plants.
Autonomic drug uses finger-print (especially chromatographic fingerprinting) to be set something afoot the seventies in 20th century aspect quality control.Current traditional Chinese medicine fingerprint (fringerprinting) is used the dna fingerprinting development, is a kind of comprehensive, quantifiable chromatogram identification of means.What grow up at first is chemical composition of Chinese materia medica chromatographic fingerprinting, particularly high performance liquid chromatography (HPLC) finger-print.HPLC has very high degree of separation, can separate the chemical constitution of complexity and form the different peak of height and form a chromatogram spy, the height of these chromatographic peaks and peak area have represented various different chemical compositions and its content and drug efficacy study result contact respectively will produce Chinese medicine spectrum effect.
A lot of Chinese patent drugs are made up of multiple flavour of a drug, for example colleague's White Phoenix Bolus of Black-bone Chicken is made up of 19 flavor medicinal materials, medicinal material involves a wide range of knowledge, complex chemical composition, only depend on its tens active components as its quality control index, be difficult to control the inherent quality situation of Chinese patent drug ball comprehensively, more can not make judgement to its true and false quality.Therefore, utilize and to set up chromatographic fingerprinting to control its quality more objective more comprehensively than conventional criterias such as thin layer discriminating, assays.
Summary of the invention
At the defective of prior art in the existence of traditional Chinese medicine ingredients context of detection.The inventor is through a large amount of experiments, studied the detection method of differentiating the multiple medicinal material of prescription in a kind of Chinese medicine preparation simultaneously, the result proves, compare with traditional thin-layer chromatography discrimination method, method of the present invention avoids existing method need prepare need testing solution and control medicinal material solution respectively, the shortcoming that adopts different developping agent systems to test respectively again, qualification result is also more objective and accurate.
Contribution of the present invention is: the inventor is applied to finger-print holistic approach and the evaluation of Chinese medicine preparation quality.Be example with colleague's White Phoenix Bolus of Black-bone Chicken, adopt the HPLC method to set up colleague's White Phoenix Bolus of Black-bone Chicken (water-honeyed pill) preparation finger, for the traditional Chinese medicine fingerprint analysis provides new thinking.
The purpose of this invention is to provide a kind of detection method that contains the Chinese patent drug of at least two kinds of compositions in the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii and the Radix Angelicae Sinensis, can obtain result accurately by this method, and avoided other impurity to disturb as far as possible.
The invention provides a kind of detection method that contains the Chinese patent drug of at least two kinds of compositions in the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii and the Radix Angelicae Sinensis, this method comprises:
(1). the preparation of need testing solution, take by weighing test sample, add and be equivalent to the methyl alcohol that test sample 5-10 doubly measures, dissolving, filter, filtrate is reclaimed solvent and is concentrated into driedly, and the water that residue adds with above-mentioned methyl alcohol equivalent makes dissolving, put cold, by the macroporous resin adsorption post, the 2-10 water elution doubly with above-mentioned quantity of methyl alcohol discards water liquid, use 15-45%wt and ethanol elution above-mentioned methyl alcohol equivalent again, discard eluent, continue with the 2-6 ethanol elution doubly for above-mentioned quantity of methyl alcohol of 75-95%wt, collect eluent, evaporate to dryness, residue is with the methyl alcohol dissolving and move to measuring bottle, adds methyl alcohol to scale, crosses miillpore filter, get subsequent filtrate, make need testing solution;
(2). the preparation of control medicinal material solution, get at least two kinds of each 1g of control medicinal material in the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii and the Radix Angelicae Sinensis respectively, shine medicinal material solution with need testing solution in pairs with legal system;
(3). chromatographic condition is mobile phase A with the acetonitrile, is Mobile phase B with water, according to the A of different proportion: the B wash-out; The detection wavelength is 201-290nm;
(4). measure, draw each 10 μ L of above-mentioned solution, inject liquid chromatograph, measure;
(5). the result detects, in the chromatogram of test sample, with the position of the identical retention time of control medicinal material chromatogram on, inspect has corresponding chromatographic peak whether respectively.
The miillpore filter that filters usefulness in the above-mentioned steps (1) is preferably the miillpore filter of 0.2～0.75 μ m, the more preferably miillpore filter of 0.4～0.5 μ m.
Detection method of the present invention adopts sample to use the detection method of high performance liquid chromatography earlier again through pre-treatment, changed usually containing the sample treatment of at least two kinds of medicinal materials in the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii and the Radix Angelicae Sinensis in the liquid chromatography, successfully eliminated the assorted peak in the liquid phase collection of illustrative plates beyond the composition to be measured in the Chinese patent medicine preparation, the result is more precise and stable.
Above-mentioned a kind of detection method that contains the Chinese patent drug of at least two kinds of compositions in the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii and the Radix Angelicae Sinensis provided by the invention, concrete steps comprise:
1. the preparation of need testing solution: precision takes by weighing test sample (for example 6g), add and to be equivalent to methyl alcohol that test sample 5-10 doubly measures (preferably with respect to the 6g test sample, quantity of methyl alcohol is about 50mL), dissolving, preferred ultrasonic processing (power 300W, frequency 50kHz) 30 minute, filter, filtrate is reclaimed solvent and is concentrated into dried, the water that residue adds with above-mentioned methyl alcohol equivalent makes dissolving, put cold, by D101 type macroporous resin adsorption post (the about 60g of resin of dress post, internal diameter 2cm, long 12cm), approximately being that (preferred 5-7 is doubly to methyl alcohol for the 2-10 water doubly of above-mentioned quantity of methyl alcohol, the about 50mL of quantity of methyl alcohol for example, the about 300mL of water then) wash-out, discard water liquid, use 15-45%wt (preferred 30%wt) and ethanol (for example 300mL) wash-out above-mentioned methyl alcohol equivalent again, discard eluent, continue with the 2-6 that is about above-mentioned quantity of methyl alcohol (preferred 3-5 is doubly to methyl alcohol) ethanol (for example about 50mL of quantity of methyl alcohol, the then about 200mL of this ethanol) wash-out doubly of 75-95%wt (preferred 80%wt), collect eluent, evaporate to dryness, residue is with the methyl alcohol dissolving and be transferred to (for example test sample is 6g, and then preferred measuring bottle is 4-6mL, more preferably the 5mL measuring bottle) in the measuring bottle, add methyl alcohol to scale, cross miillpore filter (miillpore filter of preferred 0.45 μ m), get subsequent filtrate, make need testing solution;
2. the preparation of control medicinal material solution: get at least two kinds of each 1g of control medicinal material in the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii, the Radix Angelicae Sinensis respectively, shine medicinal material solution with need testing solution in pairs with legal system;
3. chromatographic condition and system suitability test: be filling agent with the octadecylsilane chemically bonded silica; Being mobile phase A with the acetonitrile, is Mobile phase B with water, and the amount of carrying out reducing successively according to the B by 100%wt B increases the amount of A until the gradient elution of the order of the A of 100%wt; The time of gradient elution is 100-150 minute; The detection wavelength is 201-290nm, and preferred 203-180nm for example detects wavelength and is about 203nm (when detecting the root of herbaceous peony, genseng, Radix Glycyrrhizae and/or sweet wormwood), detects preferably about 280nm (when the detection red sage root, Ligusticum wallichii and/or Radix Angelicae Sinensis) of wavelength;
4. measure: according to high performance liquid chromatography (appendix VI D) test, accurate each the 10 μ L of above-mentioned solution that draw inject liquid chromatograph, measure;
5. the result detects: in the chromatogram of test sample, with the position of the identical retention time of control medicinal material chromatogram on, inspect has corresponding chromatographic peak whether respectively, and ultraviolet spectrum is consistent.
If in the test sample chromatogram, on the position of identical with reference substance chromatogram retention time with control medicinal material, corresponding chromatographic peak is arranged respectively, and ultraviolet spectrum is consistent, then can judges the medicinal material or the medicinal ingredient that contain reference substance in this test sample (sample); Further can convert according to the area at peak and contain the medicinal material of reference substance or the content of medicinal ingredient in this test sample (sample).
Test sample involved in the present invention refers to contain the Chinese patent drug (preparation) of at least two kinds of compositions in the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii and the Radix Angelicae Sinensis.
In the above-mentioned detection method of the present invention, the step that also preferably comprises the preparation of another contrast solution, it is the step of the preparation of reference substance solution, the preparation of this reference substance solution specifically comprises: take by weighing the reference substance corresponding with control medicinal material, add methyl alcohol and make reference substance solution, cross 0.45 μ m miillpore filter, get subsequent filtrate and make reference substance solution again; This reference substance is selected from Paeoniflorin, liquiritin, ginsenoside Rb
1, at least a among the ginsenoside Rf, tanshinone IIA, Astragaloside IV, onocerin.
When above-mentioned detection method when detecting the method for the Chinese patent drug contain the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii and Radix Angelicae Sinensis, the preparation of described reference substance solution comprises that adding methyl alcohol makes every 1mL and contain Paeoniflorin 0.06mg/mL, liquiritin 0.02mg/mL, ginsenoside Rb respectively
10.2mg/mL, the solution of Tanshinone I I A0.016mg/mL product solution in contrast; Reference substance solution is carried out the detection of described step (3) and (4), obtain the finger-print of reference substance, then in the result of described step (5) detects, in the chromatogram of test sample, with the position of control medicinal material and the identical retention time of reference substance chromatogram on, inspect has corresponding chromatographic peak whether respectively.
Be example with the White Phoenix Bolus of Black-bone Chicken, the White Phoenix Bolus of Black-bone Chicken (water-honeyed pill) that commodity are produced available from company of Tongrentang; The detection method of effective constituent comprises in this White Phoenix Bolus of Black-bone Chicken (water-honeyed pill) Chinese patent drug:
1. the preparation of need testing solution: precision takes by weighing White Phoenix Bolus of Black-bone Chicken (for example 6g), add the methyl alcohol (preferred 50mL) that is equivalent to test sample 5-10 and doubly measures, dissolving, preferred ultrasonic processing, filter, filtrate is reclaimed solvent and is concentrated into driedly, and the water that residue adds with above-mentioned methyl alcohol equivalent makes dissolving, put cold, by D101 type macroporous resin adsorption post, the 2-10 water elution doubly to be about above-mentioned quantity of methyl alcohol discards water liquid, use the ethanol elution of the about and above-mentioned methyl alcohol equivalent of 15-45%wt again, discard eluent, continue with the 2-6 that the is about above-mentioned quantity of methyl alcohol ethanol elution doubly of 75-95%wt, collect eluent, evaporate to dryness, residue is with the methyl alcohol dissolving and be transferred to (for example preferred measuring bottle is 4-6mL) in the measuring bottle, adds methyl alcohol to scale, crosses miillpore filter (miillpore filter of 0.25-0.5 μ m), get subsequent filtrate, make need testing solution;
2. the preparation of control medicinal material solution: get the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii, each 1g of Radix Angelicae Sinensis control medicinal material respectively, shine medicinal material solution with need testing solution in pairs with legal system;
3. chromatographic condition and system suitability test: be filling agent with the octadecylsilane chemically bonded silica; Being mobile phase A with the acetonitrile, is Mobile phase B with water, and the amount of carrying out reducing successively according to the B by 100%wt B increases the amount of A until the gradient elution of the order of the A of 100%wt; The time of gradient elution is 100-150 minute; The detection wavelength is 201-290nm, and preferred 203-180nm for example detects wavelength and is about 203nm (when detecting the root of herbaceous peony, genseng, Radix Glycyrrhizae and sweet wormwood), detects preferably about 280nm (when the detection red sage root, Ligusticum wallichii and Radix Angelicae Sinensis) of wavelength;
4. measure: according to high performance liquid chromatography (appendix VI D) test, accurate each the 10 μ L of above-mentioned solution that draw inject liquid chromatograph, measure;
5. the result detects: in the chromatogram of test sample, with the position of control medicinal material and the identical retention time of reference substance chromatogram on, inspect has corresponding chromatographic peak whether respectively, and ultraviolet spectrum is consistent.
If in the test sample chromatogram, on the position of identical retention time with the control medicinal material chromatogram, corresponding chromatographic peak is arranged respectively, and ultraviolet spectrum is consistent, then can judges the medicinal material or the medicinal ingredient that contain reference substance in this test sample (sample); Further can convert according to the area at peak and contain the medicinal material of reference substance or the content of medicinal ingredient in this test sample (sample).
In the step 1 of above-mentioned detection method, it is the preparation of need testing solution, passing through D101 type macroporous resin adsorption post, with water elution, contain the carbohydrate content that contains in the test sample in the water liquid, this carbohydrate content can cause interference to the collection of illustrative plates peak of HPLC, so the present invention selects to discard water liquid, use the ethanol elution of 15-45%wt again, trial test of the present invention shows, this part composition that elutes overlaps with the finger-print of the flow point of 80%wt ethanol elution, therefore unavoidably also will cause interference to testing result, so the present invention's selection discards the eluent of the ethanol elution of 15-45%wt, continue with the ethanol elution of 75-95%wt, repeatedly the result of trial test shows, the chromatographic peak of this part eluent is maximum, so the present invention collects the eluent of the ethanol elution of 75-95%wt.
Be example with the White Phoenix Bolus of Black-bone Chicken, the detection method of effective constituent is the same substantially in this White Phoenix Bolus of Black-bone Chicken (water-honeyed pill) Chinese patent drug, the Chinese patent drug that can detect test sample equally be the medicinal material that whether contains reference substance in the White Phoenix Bolus of Black-bone Chicken (water-bindered pill) with and component content in Chinese patent drug.
The present invention has mainly adopted the reference substance medicinal material of effective constituent in the Chinese patent drug and reference substance (being equivalent to pure product) to make finger-print with the method for efficient liquid phase, compare with the efficient liquid phase collection of illustrative plates of the Chinese patent medicine preparation that contains effective constituent again, can contrast whether contain above-mentioned effective constituent in this Chinese patent medicine preparation.
In a preferred embodiment of the invention, be example with White Phoenix Bolus of Black-bone Chicken (water-honeyed pill), the detection method of effective constituent comprises in this White Phoenix Bolus of Black-bone Chicken (water-honeyed pill) Chinese patent drug:
1. the preparation of need testing solution: get White Phoenix Bolus of Black-bone Chicken, porphyrize, precision takes by weighing 6g, add methyl alcohol 50mL, ultrasonic processing (power 300W, frequency 50kHz) 30 minutes, put coldly, filter, filtrate is reclaimed solvent and also is concentrated into dried, residue adds water 50mL makes dissolving, put cold, by the about 60g of D101 type macroporous resin adsorption post (internal diameter 2cm, long 12cm), with water 300mL wash-out, discard water liquid, use 30% ethanol 300mL wash-out again, discard eluent, continue with 80% ethanol 200mL wash-out, collect eluent, evaporate to dryness, residue is with the methyl alcohol dissolving and be transferred to the 5mL measuring bottle, adds methyl alcohol to scale, shake up, cross 0.45 μ m miillpore filter, get subsequent filtrate, make need testing solution;
2. the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii, each 1g of Radix Angelicae Sinensis control medicinal material are got in the preparation of control medicinal material solution respectively, shine medicinal material solution in pairs with legal system;
The preparation that also can prepare reference substance solution simultaneously: it is an amount of to get reference substance respectively, adds methyl alcohol and makes every 1mL and contain Paeoniflorin 0.06mg/mL, liquiritin 0.02mg/mL, ginsenoside Rb respectively
10.2mg/mL, the solution of Tanshinone I I A 0.016mg/mL, product solution in contrast;
For example get reference substance Paeoniflorin, liquiritin, ginsenoside Rb respectively
1, tanshinone IIA, add methyl alcohol and make every 1mL and contain Paeoniflorin 0.06mg/mL, liquiritin 0.02mg/mL, ginsenoside Rb respectively
10.2mg/mL, the solution of Tanshinone I I A 0.016mg/mL, product solution in contrast;
3. chromatographic condition and system suitability test: be filling agent with the octadecylsilane chemically bonded silica; Being mobile phase A with the acetonitrile, is Mobile phase B with water, and the time of gradient elution is 100-150 minute; According to the form below carries out gradient elution; The detection wavelength is 201-205nm (preferred 203nm), main medicinal material or the medicinal ingredients such as the root of herbaceous peony, genseng, Radix Glycyrrhizae, sweet wormwood of detecting under this wavelength, and wavelength 255-290nm, preferred 270-285nm, main medicinal material or the medicinal ingredients such as the red sage root, Ligusticum wallichii, Radix Angelicae Sinensis of detecting under this wavelength;
Table HPLC eluent gradient
4. determination method: according to high performance liquid chromatography (appendix VI D) test, accurate each the 10 μ L of above-mentioned solution that draw inject liquid chromatograph, measure.
5. the result detects: in the test sample chromatogram, with control medicinal material (or simultaneously and reference substance) the identical retention time of chromatogram position on, corresponding chromatographic peak is arranged respectively, and ultraviolet spectrum is consistent.
The detection method of effective constituent is that the inventor gropes to set up with the basis of garbled data in a large amount of experiments in the Chinese medicine preparation that contains two or more medicinal ingredient in the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii, the Radix Angelicae Sinensis simultaneously of the present invention, and described experiment is groped and the work of garbled data roughly following (be example with White Phoenix Bolus of Black-bone Chicken (water-honeyed pill that Tongrentang produces)):
1, the investigation of the foundation of analytical approach and influence factor.
1.1 the selection of sample extraction condition
Different types of extraction solvent such as methyl alcohol, ethanol, water have been investigated in experiment respectively; The extraction solvent of different proportion such as 30%wt methyl alcohol, 50%wt methyl alcohol, 70%wt methyl alcohol; Different extracting method such as ultrasonic, hot reflux, Soxhlet is extracted.The result shows that methyl alcohol extraction method gained chromatographic peak is more in the chromatogram that the different proportion methanol solvate extracts.Investigated the need testing solution that extracting n-butyl alcohol and methyl alcohol extract simultaneously, measure finger-print under above-mentioned chromatographic condition, the chromatogram of two kinds of methods does not have the difference of essence.So the methyl alcohol extraction method is adopted in this experiment, guarantees that the multiple composition in colleague's White Phoenix Bolus of Black-bone Chicken can be embodied in the finger-print.Colleague's White Phoenix Bolus of Black-bone Chicken is made up of 19 flavor medicinal materials, it is the compound preparation of a complexity, methanolic extract chromatogram peak is many but separation is not good, have nothing in common with each other according to each main effectively compositional polarity in colleague's White Phoenix Bolus of Black-bone Chicken finished product, so investigated by high polarity to low polarity and crossed macroporous resin column, collect different proportion ethanol elution thing and separate each polar fraction and adopt different solvents (chloroform, ethyl acetate, normal butyl alcohol, methyl alcohol) Soxhlet to extract by the paramount polarity of low polarity, collect two kinds of separation methods of different parts extract.Chromatographic peak is overlapping serious in the different parts Soxhlet extract gained chromatogram, and the baseline injustice, and chromatographic peak can not obtain fine separation.Adopted macroporous resin column method gained chromatographic peak many and separate good.After the chromatographic peak feature of having investigated different proportion ethanol elution thing, finally determined the ethanol elution liquid proportional.
1.2 the selection of macroreticular resin type
Adopt D-101 type macroreticular resin and AB-8 type macroreticular resin sample separation.The latter separates saponins chemical constitution poor effect, and the test sample saponin active ingredient is more, so select D-101 type macroreticular resin.
Reach the optimization of type of elution mutually 1.3 flow
Colleague's White Phoenix Bolus of Black-bone Chicken ingredient is more, and nature difference is bigger, adopt the mode of isocratic elution be difficult at short notice with its effective constituent wash-out with separate, thereby adopt the gradient elution program to separate composition in colleague's White Phoenix Bolus of Black-bone Chicken.In the selection of flow phase system, according to raw medicinal material effective constituent classification and proportion, mostly be acetonitrile-water, acetonitrile-acid system.So investigated acetonitrile-water, acetonitrile-0.5% glacial acetic acid, acetonitrile-0.1% glacial acetic acid, acetonitrile-0.1% formic acid gradient elution chromatography (GEC) system respectively.The result shows that in acetonitrile-acid system, the noise jamming of chromatographic peak baseline is serious; In the acetonitrile-water system, chromatographic peak is separated preferably, and baseline is steady.So adopt the acetonitrile-water system as the phase that flows.
1.4 detect the selection of wavelength
Under the above-mentioned chromatographic condition, in 200～400nm scope need testing solution is carried out spectral scan, 3D chromatogram per sample is to choose the detection wavelength that can provide maximum chromatographic peak information.And through testing all the main fingerprint peakses of chromatographic fingerprinting that are difficult to keep this compound under the discovery single wavelength.Many, comparatively concentrated at 203nm and 280nm place chromatographic peak information, baseline is more steady, and collection of illustrative plates has kept the main fingerprint peaks of other wavelength under these two wavelength.So optimum is chosen 203nm and 280nm wavelength as detecting wavelength, also according to kind, batch difference of Chinese patent drug, and difference in experiment conditions, also can select the wavelength of 201-290nm as detecting wavelength.
1.5 the selection of object of reference
The chromatographic peak of a composition under 280nm in the red sage root separates well with other chromatographic peaks, and retention time is suitable, and the peak type is stable; The peak purity inspection shows that this chromatographic peak is one-component, so select this peak as the reference peak under the 280nm, all the other each chromatographic peaks all relatively calculate relative retention time and relative peak area ratio with it.The chromatographic peak of a composition under 203nm in the Radix Glycyrrhizae separates well with other chromatographic peaks, and the peak type is stable; The peak purity inspection shows that this chromatographic peak is one-component, so select this peak as the reference peak under the 203nm, all the other each chromatographic peaks all relatively calculate relative retention time and relative peak area ratio with it.
When not containing the red sage root and/or Radix Glycyrrhizae in the test sample, also can be single according to component, separate well, the peak type is stable etc., and standard is carried out examination, picks out suitable chromatographic peak as the reference peak.
1.6 assay method
Draw need testing solution 10 μ L, inject the HPLC chromatograph, other draws methanol solvate 10 μ L and injects HPLC as blank solution, and the result shows under 203nm solvent peak is arranged but do not disturb the chromatographic peak ownership, and solvent-free peak disturbs under the 280nm.
2, methodological study
2.1 precision experiment:
Get colleague's White Phoenix Bolus of Black-bone Chicken (water-honeyed pill, lot number 4030220), the preparation need testing solution, continuous sample introduction 6 times records each total peak relative retention time and relative peak area.
Each total peak relative retention time is stable, and the RSD of each total peak relative peak area is less than 3%.The relevant regulations that meet " technical requirement (provisional) of traditional Chinese medicine finger-print research " precision test.
2.2 replica test:
Get 6 parts of same lot number (4030220) colleague's White Phoenix Bolus of Black-bone Chickens (water-honeyed pill), the accurate title, decide, and is equipped with need testing solution by " 2 " below legal system, and sample introduction records each total peak relative retention time and relative peak area respectively.
Each total peak relative retention time is stable, and the RSD of each total peak relative peak area meets the relevant regulations of " technical requirement (provisional) of traditional Chinese medicine finger-print research " replica test less than 3%.
2.3 same need testing solution (the same) is got in stability test, respectively at 0,4,8,12,16,20,24h sample introduction, records each total peak relative retention time and relative peak area.
By colleague's White Phoenix Bolus of Black-bone Chicken (water-honeyed pill) sample HPLC finger-print be studies show that it is feasible setting up colleague's White Phoenix Bolus of Black-bone Chicken (water-honeyed pill) HPLC fingerprint spectrum method.This method is stable, favorable reproducibility, can be used as one of standard of colleague's White Phoenix Bolus of Black-bone Chicken quality control.
3, the drafting of colleague's White Phoenix Bolus of Black-bone Chicken HPLC finger-print and feature chromatographic peak ultraviolet spectrum are pointed out
3.1 the drafting of finger-print.
Accurate colleague's White Phoenix Bolus of Black-bone Chicken need testing solution 10 μ L that draw inject the HPLC instrument, measure the record chromatogram by above-mentioned condition.The results are shown in Figure 1 and Fig. 2.
The finger-print of other control medicinal materials is owed to give temporarily.
Except special mark and explanation, the concentration of other solvents (comprising percent concentration %) is mass concentration (%wt) among the present invention.
The Chinese patent drug that the present invention detects is to contain in the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii and the Radix Angelicae Sinensis at least two kinds Chinese patent medicine preparation, be preferably the Chinese patent medicine preparation that contains the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii and Radix Angelicae Sinensis, comprise the Chinese medicine preparation of White Phoenix Bolus of Black-bone Chicken and/or the dosage changing form on the White Phoenix Bolus of Black-bone Chicken basis, the Chinese medicine preparation of this dosage changing form comprises the preparation of the Chinese prescription that contains White Phoenix Bolus of Black-bone Chicken.For example the Chinese patent drug that detects of the present invention comprises the water-honeyed pill of White Phoenix Bolus of Black-bone Chicken, the water-bindered pill, Wuji Baifeng capsule and/or the WUJI BAIFENG KOUFUYE of White Phoenix Bolus of Black-bone Chicken.
Detection method of the present invention adopts sample to use the detection method of high performance liquid chromatography earlier again through pre-treatment, changed usually containing the sample treatment of the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii and/or Radix Angelicae Sinensis medicinal material in the liquid chromatography, successfully eliminated the assorted peak in the liquid phase collection of illustrative plates beyond the composition to be measured in the preparation, novelty technology of the present invention especially is successfully to have developed carries out one-time detection to the Chinese patent medicine preparation that contains multidigit Chinese medicine and can contrast the technology that obtains testing result, and makes testing result more precise and stable.
Description of drawings
Fig. 1: the HPLC finger-print of colleague's White Phoenix Bolus of Black-bone Chicken (water-honeyed pill) (detecting wavelength 203nm).
Fig. 2: the HPLC finger-print of colleague's White Phoenix Bolus of Black-bone Chicken (water-honeyed pill) (detecting wavelength 280nm).
Wherein, marked label on Fig. 1 and Fig. 2 and be totally 19 peaks of 1～19, this 19 peaks and relatively and the relative retention time at No. 17 peaks formed the HPLC finger-print of colleague's White Phoenix Bolus of Black-bone Chicken (water-honeyed pill) jointly.
Embodiment
Describe the present invention in detail below in conjunction with drawings and Examples, but do not limit practical range of the present invention.
Embodiment one. the quality determining method of colleague's White Phoenix Bolus of Black-bone Chicken (big honeyed bolus, Tongrentang's pharmacy)
Chromatographic condition and system suitability test: be filling agent with the octadecylsilane chemically bonded silica; Being mobile phase A with the acetonitrile, is Mobile phase B with water, and according to the form below carries out gradient elution; The detection wavelength is 203nm (root of herbaceous peony, genseng, Radix Glycyrrhizae, sweet wormwood medicinal material), 280nm (red sage root, Ligusticum wallichii, Radix Angelicae Sinensis medicinal material).
Table HPLC eluent gradient
The preparation of need testing solution: it is an amount of to get this product, shreds, and adds equivalent zeyssatite and grinds well, precision takes by weighing 12g, adds methyl alcohol 50mL, ultrasonic processing (power 300W, frequency 50kHz) 30 minutes, put coldly, filter, filtrate is reclaimed solvent and is concentrated into dried, residue adds water 50mL makes dissolving, put cold, by the about 60g of D101 type macroporous resin adsorption post (internal diameter 2cm, long 12cm), with water 300mL wash-out, discard water liquid, use 30% ethanol 300mL wash-out again, discard eluent, continue with 80% ethanol 200mL wash-out, collect eluent, evaporate to dryness, residue is with the methyl alcohol dissolving and be transferred to the 5mL measuring bottle, add methyl alcohol to scale, shake up, cross 0.45 μ m miillpore filter, get subsequent filtrate, make need testing solution.
The preparation of control medicinal material solution: get the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii, each 1g of Radix Angelicae Sinensis control medicinal material respectively, shine medicinal material solution in pairs with legal system.
Determination method: according to high performance liquid chromatography (appendix VI D) test, accurate each the 10 μ L of above-mentioned solution that draw inject liquid chromatograph, measure the record chromatogram.
In the test sample chromatogram, on the position of identical retention time with the control medicinal material chromatogram, corresponding chromatographic peak is arranged respectively, and ultraviolet spectrum is consistent.
The drafting of finger-print: the HPLC finger-print of colleague's White Phoenix Bolus of Black-bone Chicken (detecting wavelength 203nm) is seen Fig. 1; The HPLC finger-print of colleague's White Phoenix Bolus of Black-bone Chicken (detecting wavelength 280nm) is seen Fig. 2.
Wherein, marked label on Fig. 1 and Fig. 2 and be totally 19 peaks of 1～19, this 19 peaks and relatively and the relative retention time at No. 17 peaks formed the HPLC finger-print of colleague's White Phoenix Bolus of Black-bone Chicken (water-honeyed pill) jointly.No. 17 peaks and No. 6 peaks are with reference to the peak.
Embodiment two. the detection method of effective constituent in the White Phoenix Bolus of Black-bone Chicken (water-bindered pill, the pharmacy of remittance benevolence)
1. the preparation of need testing solution: precision takes by weighing White Phoenix Bolus of Black-bone Chicken (water-bindered pill 6g), add the methyl alcohol that is equivalent to the about 8 times of amounts of test sample, dissolving, preferred ultrasonic processing 20-40 minute filters, and filtrate is reclaimed solvent and is concentrated into dried, residue adds about 50mL water makes dissolving, by D101 type macroporous resin adsorption post (the about 60g of resin of dress post, internal diameter 2cm, long 12cm), water elution with about 300mL, discard water liquid, use the ethanol elution of the 50mL of 25-35%wt again, discard eluent, the ethanol elution of about 200mL of the usefulness that continues 75-85%wt (preferred 80%wt), collect eluent, evaporate to dryness, residue is with the methyl alcohol dissolving and be transferred to (preferred 5mL measuring bottle) in the measuring bottle, add methyl alcohol to scale, cross miillpore filter (miillpore filter of preferred 0.45 μ m), get subsequent filtrate, make need testing solution;
2. the preparation of control medicinal material solution: get the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii, each 1g of Radix Angelicae Sinensis control medicinal material respectively, shine medicinal material solution with need testing solution in pairs with legal system;
3. chromatographic condition and system suitability test: be filling agent with the octadecylsilane chemically bonded silica; Being mobile phase A with the acetonitrile, is Mobile phase B with water, and the amount of carrying out reducing successively according to the B by 100%wt B increases the amount of A until the gradient elution of the order of the A of 100%wt; The time of gradient elution is 100-150 minute; The detection wavelength is 201-290nm, preferred 203-180nm, and for example detecting wavelength is 203nm (detecting the root of herbaceous peony, genseng, Radix Glycyrrhizae and/or sweet wormwood), detects wavelength 280nm (detecting the red sage root, Ligusticum wallichii and/or Radix Angelicae Sinensis);
4. measure: according to high performance liquid chromatography (appendix VI D) test, accurate each the 10 μ L of above-mentioned solution that draw inject liquid chromatograph, measure;
5. the result detects: in the chromatogram of test sample, with the position of the identical retention time of control medicinal material chromatogram on, inspect has corresponding chromatographic peak whether respectively, and ultraviolet spectrum is consistent.
The drafting of finger-print: the HPLC finger-print of colleague's White Phoenix Bolus of Black-bone Chicken is seen Fig. 1 and Fig. 2.Marked 19 peaks on Fig. 1 and Fig. 2, the relative retention time at these 19 peaks and relative and No. 17 peaks has been formed the HPLC finger-print of White Phoenix Bolus of Black-bone Chicken (water-honeyed pill) jointly, can be used as the finger-print of the contrast usefulness of White Phoenix Bolus of Black-bone Chicken (water-bindered pill) equally.
In the test sample chromatogram, on the position of identical retention time with the control medicinal material chromatogram, corresponding chromatographic peak is arranged respectively, and ultraviolet spectrum is consistent, then can judges and contain the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii, Radix Angelicae Sinensis control medicinal material or medicinal ingredient in this test sample (sample).
Embodiment three. and other contain the detection method of effective constituent in the Chinese medicine preparation of two or more medicinal ingredient in the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii, the Radix Angelicae Sinensis
1. the preparation of need testing solution: precision takes by weighing test sample (should contain the root of herbaceous peony in this test sample, genseng, the red sage root, 6g), add the methyl alcohol that is equivalent to the about 8 times of amounts of test sample, dissolving, filter, filtrate recovery solvent is concentrated into dried, and residue adds about 50mL water makes dissolving, by D101 type macroporous resin adsorption post (the about 60g of resin of dress post), water elution with about 300mL, discard water liquid, use the ethanol elution of the 50mL of 25-35%wt again, discard eluent, the ethanol elution of about 200mL of the usefulness that continues 75%wt, collect eluent, evaporate to dryness, residue is with the methyl alcohol dissolving and be transferred to (5mL) in the measuring bottle, add methyl alcohol to scale, cross miillpore filter (miillpore filter of preferred 0.45 μ m), get subsequent filtrate, make need testing solution;
2. the preparation of control medicinal material solution: get the root of herbaceous peony, genseng, each 1g of red sage root control medicinal material respectively, shine medicinal material solution with need testing solution in pairs with legal system;
3. chromatographic condition and system suitability test: be filling agent with the octadecylsilane chemically bonded silica; Being mobile phase A with the acetonitrile, is Mobile phase B with water, and the amount of carrying out reducing successively according to the B by 100%wt B increases the amount of A until the gradient elution of the order of the A of 100%wt; The time of gradient elution is 100-150 minute; The detection wavelength is 201-290nm, and preferred 203-180nm for example detects wavelength and is about 203nm (when detecting the root of herbaceous peony, genseng), detects preferably about 280nm (when detecting the red sage root) of wavelength;
4. measure: according to high performance liquid chromatography (appendix VI D) test, accurate each the 10 μ L of above-mentioned solution that draw inject liquid chromatograph, measure;
5. the result detects: in the chromatogram of test sample, with the position of control medicinal material and the identical retention time of reference substance chromatogram on, inspect has corresponding chromatographic peak whether respectively, and ultraviolet spectrum is consistent.
In the test sample chromatogram, on the position of identical retention time with the control medicinal material chromatogram, corresponding chromatographic peak is arranged respectively, and ultraviolet spectrum is consistent, then judges in this test sample (sample) and contain the root of herbaceous peony, genseng, the red sage root.
If when control medicinal material and reference substance solution increase control medicinal material such as sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii, Radix Angelicae Sinensis or medicinal ingredient, can from collection of illustrative plates, find out, do not contain sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii, Radix Angelicae Sinensis medicinal material or medicinal ingredient in this test sample.
Embodiment four. the detection method of the effective constituent of WUJI BAIFENG KOUFUYE (river, Jiangxi is strange)
1, instrument and reagent
Waters Alliance2695 high performance liquid chromatograph; Waters 2996 diode array detector; Waters Empower 2 data processing software systems.
B ü CHI Rotary Evaporators (Rotavapor R-114; Waterbath B-480); Lion ancient cooking vessel SHB-III circulation ability of swimming is used vacuum pump (Zhengzhou Greatwall Scientific Industrial ﹠ Trading Co., Ltd.) more; The medical numerical control supersonic washer of KQ-250DE type (Kunshan Ultrasonic Instruments Co., Ltd.); FW100 type high speed Universalpulverizer (Tianjin Tai Site Instr Ltd.).D-101 type macroreticular resin (Anhui Samsung resin Science and Technology Ltd.).
Methyl alcohol, ethanol are analytical reagent; HPLC is chromatographically pure with acetonitrile (J.T.Baker company).Water is pure water, crosses Millipore 0.22 μ m filter membrane.
Paeoniflorin, liquiritin, ginsenoside Rb1, ginsenoside Rf, tanshinone IIA, Astragaloside IV, onocerin reference substance, the root of herbaceous peony, Radix Glycyrrhizae, sweet wormwood, the red sage root, genseng, Ligusticum wallichii, Radix Angelicae Sinensis control medicinal material are provided by Nat'l Pharmaceutical ﹠ Biological Products Control Institute.Colleague's White Phoenix Bolus of Black-bone Chicken (water-honeyed pill), colleague's White Phoenix Bolus of Black-bone Chicken feminine gender (lacking flavor) are provided by Beijing TongrenTang Co., Ltd Tongrentang Pharmaceutical Factory, Beijing.The red sage root, genseng, the root of herbaceous peony, Radix Glycyrrhizae, Radix Angelicae Sinensis, Ligusticum wallichii, sweet wormwood, rhizoma cyperi medicinal material sample are provided by Beijing TongrenTang Co., Ltd Tongrentang Pharmaceutical Factory, Beijing, open the associate professor of pharmacy that continues through Nat'l Pharmaceutical ﹠ Biological Products Control Institute and identify.
2, the preparation of test solution product: get WUJI BAIFENG KOUFUYE 20mL, by the D101 type macroporous resin adsorption post (internal diameter 2cm, long 12cm) that pre-service is good, with water 300mL wash-out, discard water liquid, use 30% ethanol 300mL wash-out again, discard eluent, continue with 80% ethanol 200mL wash-out, collect eluent, evaporate to dryness with methyl alcohol dissolving and be transferred to the 5mL measuring bottle, adds methyl alcohol to scale, shake up, cross 0.45 μ m miillpore filter, get subsequent filtrate, namely.
3, the preparation of reference substance solution: get Paeoniflorin, liquiritin, ginsenoside Rb respectively
1, ginsenoside Rf, Tanshinone I I A, Astragaloside IV, onocerin reference substance be an amount of, adds methyl alcohol and make reference substance solution, crosses 0.45 μ m miillpore filter, gets subsequent filtrate, namely.
4, HPLC chromatographic condition:
Chromatographic column: Alltech Alltima C
18Post (4.6mm * 250mm, 5 μ m); Phase flows: see the following form; Flow velocity: 1mLmin
-1Detect wavelength: 203nm, 280nm; Column temperature: 25 ℃; Sample size: 10 μ L.Chromatogram writing time is 126 minutes.
Table HPLC eluent gradient
Embodiment five. result's contrast of Chinese patent drug reference examples
The Chinese patent drug reference examples is to have deleted the finished product that one or several effective constituents obtain on the prescription basis of former Chinese patent drug.Present embodiment is to have deleted the root of herbaceous peony and genseng on the prescription basis of White Phoenix Bolus of Black-bone Chicken.
1. instrument and reagent: with embodiment one.
2. chromatographiccondition: with embodiment one.
3. the preparation of test sample (reference examples) solution: with embodiment one.
4. measure: precision is drawn reference substance solution and each 10ul of test sample (reference examples) solution respectively, injects liquid chromatograph, measures, namely.
5. contrast collection of illustrative plates: the result does not contain the root of herbaceous peony and genseng in the test sample (reference examples) as can be seen, and other compositions for example red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii and Radix Angelicae Sinensis then are present in the test sample (reference examples).
Result's proof is consistent with the actual composition of reference examples.
Embodiment six. the detection method of effective constituent in the Wuji Baifeng capsule (Beijing Tongrentang)
1. the preparation of need testing solution: precision takes by weighing Wuji Baifeng capsule 's content 6g, add the methyl alcohol that is equivalent to the about 8 times of amounts of test sample, dissolving, preferred ultrasonic processing 20-40 minute filters, and filtrate is reclaimed solvent and is concentrated into dried, residue adds about 50mL water makes dissolving, by D101 type macroporous resin adsorption post (the about 60g of resin of dress post, internal diameter 2cm, long 12cm), water elution with about 300mL, discard water liquid, use the ethanol elution of the 50mL of 25-35%wt again, discard eluent, the ethanol elution of about 200mL of the usefulness that continues 75-85%wt (preferred 80%wt), collect eluent, evaporate to dryness, residue is with the methyl alcohol dissolving and be transferred to (preferred 5mL measuring bottle) in the measuring bottle, add methyl alcohol to scale, cross miillpore filter (miillpore filter of preferred 0.45 μ m), get subsequent filtrate, make need testing solution;
2. the preparation of control medicinal material solution: get the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii, each 1g of Radix Angelicae Sinensis control medicinal material respectively, shine medicinal material solution with need testing solution in pairs with legal system;
3. chromatographic condition and system suitability test: be filling agent with the octadecylsilane chemically bonded silica; Being mobile phase A with the acetonitrile, is Mobile phase B with water, and the amount of carrying out reducing successively according to the B by 100%wt B increases the amount of A until the gradient elution of the order of the A of 100%wt; The time of gradient elution is 100-150 minute; The detection wavelength is 201-290nm, preferred 203-180nm, and for example detecting wavelength is 203nm (detecting the root of herbaceous peony, genseng, Radix Glycyrrhizae and/or sweet wormwood), detects wavelength 280nm (detecting the red sage root, Ligusticum wallichii and/or Radix Angelicae Sinensis);
4. measure: according to high performance liquid chromatography (appendix VI D) test, accurate each the 10 μ L of above-mentioned solution that draw inject liquid chromatograph, measure;
5. the result detects: in the chromatogram of test sample, with the position of the identical retention time of control medicinal material chromatogram on, inspect has corresponding chromatographic peak whether respectively, and ultraviolet spectrum is consistent.
In the test sample chromatogram, on the position of identical retention time with the control medicinal material chromatogram, corresponding chromatographic peak is arranged respectively, and ultraviolet spectrum is consistent, then can judges and contain the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii, Radix Angelicae Sinensis control medicinal material or medicinal ingredient in this test sample (sample).
Claims (7)
1. the detection method of effective constituent in the Chinese patent drug that contains the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii and Radix Angelicae Sinensis, this Chinese patent drug is colleague's White Phoenix Bolus of Black-bone Chicken, this method comprises:
(1). the preparation of need testing solution, precision takes by weighing test sample, adds to be equivalent to the methyl alcohol that test sample 5-10 doubly measures, dissolving, filter, filtrate is reclaimed solvent and is concentrated into driedly, and the water that residue adds with above-mentioned methyl alcohol equivalent makes dissolving, put cold, by the macroporous resin adsorption post, the 2-10 water elution doubly with above-mentioned quantity of methyl alcohol discards water liquid, use 15-45%wt and ethanol elution above-mentioned methyl alcohol equivalent again, discard eluent, continue with the 2-6 ethanol elution doubly for above-mentioned quantity of methyl alcohol of 75-95%wt, collect eluent, evaporate to dryness, residue dissolves with methyl alcohol and is transferred in the measuring bottle, adds methyl alcohol to scale, crosses miillpore filter, get subsequent filtrate, make need testing solution;
(2). the preparation of control medicinal material solution, get the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii and each 1g of Radix Angelicae Sinensis control medicinal material respectively, shine medicinal material solution with need testing solution in pairs with legal system;
(3). chromatographic condition is mobile phase A with the acetonitrile, is Mobile phase B with water, according to the A of different proportion: the B wash-out; The detection wavelength is 201-290nm;
(4). measure, draw each 10 μ L of above-mentioned solution, inject liquid chromatograph, measure;
(5). the result detects, in the chromatogram of test sample, with the position of the identical retention time of control medicinal material chromatogram on, inspect has corresponding chromatographic peak whether respectively.
2. detection method as claimed in claim 1, wherein, the concentration of described need testing solution is that every ml is equivalent to test sample 0.1-10g.
3. detection method as claimed in claim 1, wherein, the described detection wavelength of step (3) is 203-280nm.
4. detection method as claimed in claim 1, wherein, step (3) is described to flow mutually for being mobile phase A with the acetonitrile, is Mobile phase B with water, and the amount that reduces B successively according to the B by 100%wt increases the amount of A and carries out gradient elution until the order of the A of 100%wt.
5. detection method as claimed in claim 1, wherein, this detection method also comprises the step of the preparation of reference substance solution, being prepared as of this reference substance solution takes by weighing the reference substance corresponding with control medicinal material, add methyl alcohol and make reference substance solution, cross 0.45 μ m miillpore filter, get subsequent filtrate and make reference substance solution again; This reference substance is Paeoniflorin, liquiritin, ginsenoside Rb
1, ginsenoside Rf, tanshinone IIA, Astragaloside IV and onocerin.
6. detection method as claimed in claim 5, it is the detection method that contains the Chinese patent drug of the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii and Radix Angelicae Sinensis, wherein, in the preparation of described reference substance solution, comprise that adding methyl alcohol makes every 1mL and contain Paeoniflorin 0.06mg, liquiritin 0.02mg, ginsenoside Rb respectively
10.2mg, the solution of tanshinone IIA 0.016mg product solution in contrast; Reference substance solution is carried out the detection of described step (3) and (4), obtain the finger-print of reference substance, then in described step (5), in the chromatogram of test sample, with the position of control medicinal material and the identical retention time of reference substance chromatogram on, inspect has corresponding chromatographic peak whether respectively.
7. as claim 1,2,4,5,6 each described detection methods, it is the detection method that contains the Chinese patent drug of the root of herbaceous peony, genseng, the red sage root, sweet wormwood, Radix Glycyrrhizae, Ligusticum wallichii and Radix Angelicae Sinensis, and wherein, step (3) is to be filling agent with the octadecylsilane chemically bonded silica; Being mobile phase A with the acetonitrile, is Mobile phase B with water, and the time of gradient elution is 100-150 minute; Detect wavelength and under 201-205nm, detect the root of herbaceous peony, genseng, Radix Glycyrrhizae, sweet wormwood, detect wavelength and detect the red sage root, Ligusticum wallichii, Radix Angelicae Sinensis at 255-290nm.
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| CN102507838B (en) * | 2011-10-31 | 2013-12-25 | 哈尔滨乐泰药业有限公司 | Method for detecting quality of traditional Chinese medicine capsules for regulating collateral channels and soothing liver |
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| CN104749265B (en) * | 2013-12-27 | 2017-05-17 | 天津中新药业集团股份有限公司乐仁堂制药厂 | Detection method for effective components in silky fowl tablet |
| CN103698464A (en) * | 2014-01-13 | 2014-04-02 | 吉林省通化博祥药业股份有限公司 | Anti-cerebral-thrombosis tablet quality standard |
| CN108459129B (en) * | 2017-02-17 | 2020-06-05 | 华润三九医药股份有限公司 | Quality control method of radix Stephaniae Tetrandrae and Poria decoction composition |
| CN108459091B (en) * | 2017-02-17 | 2020-08-07 | 华润三九医药股份有限公司 | Method for establishing fingerprint spectrum of angelica sinensis Sini decoction composition |
| CN108459128B (en) * | 2017-02-17 | 2020-06-05 | 华润三九医药股份有限公司 | Quality control method of angelica sinensis Sini decoction composition |
| CN109932437B (en) * | 2017-12-19 | 2022-03-04 | 四川济生堂药业有限公司 | Detection method of pharmaceutical composition |
| CN109507310B (en) * | 2018-10-22 | 2021-08-03 | 广州市香雪制药股份有限公司 | Fingerprint spectrum construction method and detection method of essence-nourishing jade-planting prescription |
| CN111337580A (en) * | 2018-12-19 | 2020-06-26 | 神威药业集团有限公司 | HPLC (high Performance liquid chromatography) characteristic spectrum of four-component preparation as well as construction method and application thereof |
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| US20060292246A1 (en) * | 2005-06-20 | 2006-12-28 | Rey-Yuh Wu | Characteristic mass spectral fingerprint setting method and rapid identification method for Chinese herbal medicines and prescriptions |
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| US20060292246A1 (en) * | 2005-06-20 | 2006-12-28 | Rey-Yuh Wu | Characteristic mass spectral fingerprint setting method and rapid identification method for Chinese herbal medicines and prescriptions |
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