CN103308644A - Quality detection method for miscarriage-preventing leonurus preparation - Google Patents
Quality detection method for miscarriage-preventing leonurus preparation Download PDFInfo
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Abstract
The invention relates to a quality detection method for a traditional Chinese medicine preparation, and in particular relates to a quality detection method for a miscarriage-preventing leonurus preparation. In the quality detection method, stachydrine hydrochloride, rhizoma cyperi, a-cyperone, angelica sinensis, ligusticum wallichii, baicalin and hesperidin are identified by thin-layer chromatography, and content determination is performed on paeoniflorin by high-performance liquid chromatography. The quality detection method disclosed by the invention is scientific and advanced, good in specificity and reproducibility, and capable of greatly controlling the quality of the miscarriage-preventing leonurus preparation.
Description
Technical field
The present invention relates to a kind of quality determining method of Chinese medicine preparation, be specifically related to the quality determining method of abortion preventing Yimu preparation.
Background technology
The abortion preventing Yimu ball records in the health drug standards promulgated by the ministries or commissions of the Central Government (the 12 55 pages of Traditional Chinese medicine historical preparations), and standard code is WS3-B-2318-97, has menstruation regulating, invigorates blood circulation, and is antiabortive.Be used for qi-blood deficiency, irregular menstruation, fetal irritability.Its prescription is: motherwort 100g, rhizoma cyperi (vinegar system) 40g, Ligusticum wallichii 40g, Radix Angelicae Sinensis 40g, teasel root 30g, tarragon 30g, root of herbaceous peony 30g, bighead atractylodes rhizome 30g, the bark of eucommia (salt solution system) 30g, Radix Codonopsis 30g, Poria cocos 30g, fructus amomi 20g, donkey-hide gelatin (stir-fry) 20g, root of large-flowered skullcap 20g, dried orange peel 20g, prepared rhizome of rehmannia 100g, Radix Glycyrrhizae 10g.Without differentiating and the assay item, need to formulate truly feasible quality determining method to guarantee product quality and clinical efficacy in the abortion preventing Yimu ball quality standard.The report that has a small amount of this variety and quality to detect in the document, as having put down in writing the detection method of content of Paeoniflorin in the abortion preventing Yimu ball in the 34th phase of the 49th volume " contemporary Chinese doctor " Dec in 2011 " the HPLC method is measured the content of Paeoniflorin in the abortion preventing Yimu ball " literary composition, but we are when using the method to test, find that it has weak point, be necessary the method is improved.
Summary of the invention
The object of the invention is to provide the quality determining method of a kind of quality determining method of Chinese medicine preparation, particularly abortion preventing Yimu preparation.
The quality determining method of abortion preventing Yimu preparation comprises the one or more of following detection: 1. thin-layered chromatography is differentiated stachydrine hydrochloride; 2. thin-layered chromatography is differentiated rhizoma cyperi and a-cyperolone; 3. thin-layered chromatography is differentiated Radix Angelicae Sinensis and Ligusticum wallichii; 4. thin-layered chromatography is differentiated scutelloside; 5. thin-layered chromatography is differentiated aurantiamarin; 6. the content of high effective liquid chromatography for measuring Paeoniflorin.
The quality determining method of abortion preventing Yimu preparation comprises the one or more of following detection:
1. thin-layered chromatography is differentiated stachydrine hydrochloride
This product porphyrize is got in the preparation of solid pharmaceutical preparation need testing solution, gets in right amount, adds 20%～90% methanol solution, 10～100ml, heating and refluxing extraction 0.5～4 hour filters, and filtrate is steamed to nothing alcohol flavor, regulate pH value to 1～2 with watery hydrochloric acid, by strong acid cation exchange resin column, wash with water to efflux closely colourless, discard eluent, use again strong ammonia solution ethanolic solution (19 → 100) wash-out, collect eluent, evaporate to dryness, residue makes dissolving with ethanol 1ml, as need testing solution.
This product mixing is got in the preparation of liquid preparation need testing solution, get an amount of, regulate pH value to 1～2 with watery hydrochloric acid, by strong acid cation exchange resin column, wash with water to efflux closely colourless, discard eluent, use again strong ammonia solution ethanolic solution (19 → 100) wash-out, collect eluent, evaporate to dryness, residue makes dissolving with ethanol 1ml, as need testing solution.
The stachydrine hydrochloride reference substance is got in the preparation of reference substance solution, adds ethanol and makes the solution that every 1ml contains 1～2mg, in contrast product solution.
Absorption need testing solution, reference substance solution are put respectively on same silica gel g thin-layer plate, take n-butyl alcohol-hydrochloric acid-ethyl acetate 6～10: 2～4: 1 as developping agent, launch, take out, dry, 90～120 ℃ were heated 2～20 minutes, and let cool, spray dries up with 1% ferric trichloride ethanolic solution-1: 10 mixed solution of rare bismuth potassium iodide test solution.In the test sample chromatogram, with reference substance chromatogram relevant position on, the spot of aobvious same color.
2. thin-layered chromatography is differentiated rhizoma cyperi and a-cyperolone
Get this product porphyrize or mixing, get an amount of, put in the round-bottomed flask, add water an amount of, connect volatile oil determination apparatus, add water to scale from the analyzer upper end, and overflow enters till the flask, adds sherwood oil (60～90 ℃) 1～5ml again, adds hot reflux 0.5～4 hour, let cool, get petroleum ether layer as need testing solution.Other gets rhizoma cyperi control medicinal material 0.5～2g, the 10～100ml that adds diethyl ether, and ultrasonic processing 10～60 minutes lets cool, and filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, in contrast medicinal material solution.Get again a-cyperolone reference substance, add ethyl acetate and make the solution that every 1ml contains 0.5～2mg, in contrast product solution.Draw respectively above-mentioned three kinds of solution, point is on same silica GF254 thin layer plate, take toluene-ethyl acetate 8～10: 1 as developping agent, launch, take out, dry, put under the ultraviolet lamp 254nm and inspect, in the test sample chromatogram, with control medicinal material and reference substance chromatogram relevant position on, the spot of aobvious same color.
3. thin-layered chromatography is differentiated Radix Angelicae Sinensis and Ligusticum wallichii
This product porphyrize is got in the preparation of solid pharmaceutical preparation need testing solution, gets in right amount, adds sherwood oil (30～60 ℃) 10～100ml, ultrasonic processing 10～60 minutes, get sherwood oil liquid and volatilize, residue adds sherwood oil (30～60 ℃) 1ml makes dissolving, as need testing solution.
This product mixing is got in the preparation of liquid preparation need testing solution, gets in right amount, puts in the separating funnel, add sherwood oil (30～60 ℃), jolting is extracted, and minute gets petroleum ether layer to volatilize, residue adds sherwood oil (30～60 ℃) 1ml makes dissolving, as need testing solution.
Radix Angelicae Sinensis and each 0.2～2g of Ligusticum wallichii control medicinal material are got in the preparation of control medicinal material solution, add sherwood oil (30～60 ℃) 2～10ml, and supernatant is got in ultrasonic processing 10～60 minutes, in contrast medicinal material solution.
Draw respectively mentioned solution, put on same silica gel g thin-layer plate, take normal hexane-ethyl acetate 5～8: 1 as developping agent, launches, take out, dry, put under the ultraviolet lamp (365nm) and inspect, in the test sample chromatogram, with control medicinal material chromatogram relevant position on, the spot of aobvious same color.
4. thin-layered chromatography is differentiated scutelloside
This product porphyrize is got in the preparation of solid pharmaceutical preparation need testing solution, gets in right amount, and the 20～100ml that adds diethyl ether added hot reflux 10～120 minutes, filters, and discards ether solution.The dregs of a decoction are flung to ether, add methyl alcohol 10～100ml, add hot reflux 10～300 minutes, filter, filtrate evaporate to dryness, residue add water 10～100ml makes dissolving, filter, get filtrate, with salt acid for adjusting pH value to 2, extract with the ethyl acetate jolting, get acetic acid ethyl fluid, evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution.
This product mixing is got in the preparation of liquid preparation need testing solution, gets in right amount, with salt acid for adjusting pH value to 2, extracts with the ethyl acetate jolting, gets acetic acid ethyl fluid, and evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution.
The scutelloside reference substance is got in the preparation of reference substance solution, adds methyl alcohol and makes the solution that every 1ml contains 0.5～2mg, in contrast product solution.
Draw above-mentioned two kinds of solution, put respectively in same take the carboxymethylcellulose sodium solution that contains 4% sodium acetate on the silica gel g thin-layer plate of binder, take ethyl acetate-butanone-formic acid-water 4～6: 2～4: 0.5～1.5: 0.5～1.5 as developping agent, launch, take out, dry, spray is with 5% ferric trichloride ethanolic solution.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, the spot of aobvious same color.
5. thin-layered chromatography is differentiated aurantiamarin
This product porphyrize is got in the preparation of solid pharmaceutical preparation need testing solution, gets in right amount, and the 20～100ml that adds diethyl ether added hot reflux 10～120 minutes, filters, and discards ether solution.The dregs of a decoction are flung to ether, add methyl alcohol 10～100ml, add hot reflux 10～300 minutes, filter, filtrate evaporate to dryness, residue add water 10～100ml makes dissolving, filter, get filtrate, with salt acid for adjusting pH value to 2, extract with the ethyl acetate jolting, get acetic acid ethyl fluid, evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution.
This product mixing is got in the preparation of liquid preparation need testing solution, gets in right amount, with salt acid for adjusting pH value to 2, extracts with the ethyl acetate jolting, gets acetic acid ethyl fluid, and evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution.
The aurantiamarin reference substance is got in the preparation of reference substance solution, adds methyl alcohol and makes saturated solution, in contrast product solution.
Draw above-mentioned need testing solution and reference substance solution, put respectively in same with the silica gel g thin-layer plate that contains 1% NaOH on, take methenyl choloride-methanol-water 30～35: lower floor's solution of 15～20: 4～6 is as developping agent, launch, take out, dry, spray is put under the ultraviolet lamp (365nm) and is inspected with the aluminium choride test solution.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, the fluorescence spot of aobvious same color.
6. the content of high effective liquid chromatography for measuring Paeoniflorin
The chromatographic condition octadecylsilane chemically bonded silica is filling agent; Acetonitrile-water is mobile phase at 15: 85; The detection wavelength is 230nm.
It is an amount of that the preparation precision of reference substance solution takes by weighing the Paeoniflorin reference substance, adds 50% methyl alcohol and make the solution that contains 10～60 μ g among every 1ml, and get final product.
This product porphyrize is got in the preparation of solid pharmaceutical preparation need testing solution, gets an amount of, accurately weighed, put in the tool plug conical flask accurate 50%～90% methyl alcohol, 25～100ml, the weighed weight of adding, added hot reflux 20～200 minutes, and let cool, weighed weight, supply the weight of loss with 50%～90% methyl alcohol, shake up, filter, get subsequent filtrate 10ml, evaporate to dryness, 2～5ml makes dissolving with 50%～90% methyl alcohol, is added on the neutral alumina column, with methyl alcohol 50～200ml wash-out, collect eluent, steam near and do, add the dissolving of 50% methyl alcohol, and be settled in the 10ml measuring bottle, filter, get subsequent filtrate, and get final product.
This product mixing is got in the preparation of liquid preparation need testing solution, and precision measures in right amount, evaporate to dryness, 2～5ml makes dissolving with 50%～90% methyl alcohol, be added on the neutral alumina column, with methyl alcohol 50～200ml wash-out, collect eluent, steam near and do, add the dissolving of 50% methyl alcohol, and be settled in the 10ml measuring bottle, filter, get subsequent filtrate, and get final product.
Determination method is accurate reference substance solution, need testing solution 1～20 μ l of drawing respectively, and the injection liquid chromatography is measured, and be get final product.
Above-mentioned detection need to not carry out according to sequencing, and can also carry out other conventional sense simultaneously, as: proterties, limit test of microbe etc.
Abortion preventing Yimu preparation of the present invention, refer to the conventional formulation that is prepared from modern technology in abortion preventing Yimu ball prescription ratio, this conventional formulation comprises the clinical or pharmaceutically acceptable common formulations such as tablet, capsule, granule, mixture, syrup, pill, powder.
The sampling amount of abortion preventing Yimu preparation of the present invention refers to remove the outer packaging material of preparation, the amount of regularlying sample and deciding.
During thin-layer chromatography of the present invention was differentiated, the point sample amount of need testing solution and reference substance solution was 1～20 μ l.
Quality determining method science of the present invention, advanced person, specificity, favorable reproducibility can be controlled the quality of abortion preventing Yimu preparation well.
By reading following description and appending claims, can understand better these and other feature, aspect and advantage of the present invention.
Embodiment
Although this instructions, should be believed following explanation by particularly pointing out and clear claimed claims of the present invention are drawn a conclusion and will understand better the present invention.
Except as otherwise noted, all number percents and the ratio that relate to herein, all " the Chinese pharmacopoeia regulation was carried out by version in 2010 in the preparations of solution and test solution etc.
Word as used herein " preferably " and variant refer to can provide embodiment of the present invention of specific beneficial effect under specific environment.Yet other embodiment also can be preferred under identical or other environment.In addition, the detailed description of one or more preferred embodiments does not represent that other embodiment is useless, and is not intended to get rid of from category of the present invention other embodiment.
Experimental example 1: thin-layered chromatography is differentiated stachydrine hydrochloride
In the need testing solution preparation that abortion preventing Yimu ball stachydrine hydrochloride thin-layer chromatography is differentiated, concentration to institute's reagent adding methyl alcohol, the volume of institute's reagent adding, the heating and refluxing extraction time etc. is investigated, in order to send better beneficial effect, i.e. balance between " having clearly stachydrine hydrochloride spot in the thin-layer chromatography " and " inspection cost ":
The concentration of preferred methyl alcohol is 20%～90%, more preferably 40%～60%, also more preferably 50%.
It is 10～100ml that preferred reagent adds volume, 30～70ml more preferably, also 50ml more preferably.
The preferred heating and refluxing extraction time is 0.5～4 hour, more preferably 0.8～2 hour, and also more preferably 1 hour.
The thin-layer chromatography condition that abortion preventing Yimu ball stachydrine hydrochloride is differentiated is investigated, in order to send better beneficial effect, i.e. balance between " the stachydrine hydrochloride spot has preferably degree of separation in the test sample thin-layer chromatography " and " inspection cost ":
Developping agent is preferably n-butyl alcohol-hydrochloric acid-ethyl acetate 6～10: 2～4: 1, and more preferably n-butyl alcohol-hydrochloric acid-ethyl acetate 7～9: 2.5～3.5: 1, n-butyl alcohol-hydrochloric acid-ethyl acetate 8: 3: 1 more preferably also.
Temperature and time to thin layer plate heating colour developing is investigated, in order to send better beneficial effect, i.e. and balance between " the spot colour developing is clear " and " inspection cost ":
Preferred heating-up temperature is 90～120 ℃, more preferably 100～110 ℃, and also more preferably 105 ℃.
Be 2～20 minutes preferred heat time heating time, more preferably 5～15 minutes, and also more preferably 10 minutes.
In the experimental study, in the selection of developer, once tested the color developing effect of rare bismuth potassium iodide test solution, the result is take 1% ferric trichloride ethanolic solution-rare bismuth potassium iodide test solution (1: 10) mixed solution as best.
Carry out demonstration test by embodiment 1 stachydrine hydrochloride thin-layer identification method, as a result in the test sample chromatogram, with the corresponding position of stachydrine hydrochloride reference substance chromatogram on, the spot of aobvious same color.The thin-layer chromatography good separating effect, the spot rounding, Rf value is moderate, and reappearance and specificity are good, and negative control is noiseless.
Experimental example 2: thin-layered chromatography is differentiated rhizoma cyperi and a-cyperolone
In the need testing solution preparation of abortion preventing Yimu ball rhizoma cyperi and the discriminating of a-cyperolone thin-layer chromatography, volume to institute's reagent adding sherwood oil (60～90 ℃), heating return time etc. is investigated, in order to send better beneficial effect, i.e. balance between " having clearly corresponding spot in the thin-layer chromatography with on rhizoma cyperi and the corresponding position of a-cyperolone " and " inspection cost ":
Preferred reagent sherwood oil (60～90 ℃) adding volume is 1～5ml, 1.5～3ml more preferably, also 2ml more preferably.
The preferred heating and refluxing extraction time is 0.5～4 hour, more preferably 1～3 hour, and also more preferably 2 hours.
In rhizoma cyperi control medicinal material solution preparation, to the volume of institute's reagent adding ether, the ultrasonic processing time etc. are investigated, in order to send better beneficial effect, i.e. and balance between " control medicinal material has clearly feature spot " and " inspection cost ":
It is 10～100ml that preferred reagent ether adds volume, 20～50ml more preferably, also 30ml more preferably.
The preferred ultrasonic processing time is 10～60 minutes, more preferably 15～30 minutes, and also more preferably 20 minutes.
Thin-layer chromatography condition to abortion preventing Yimu ball rhizoma cyperi and the discriminating of a-cyperolone is investigated, in order to send better beneficial effect, i.e. balance between " the feature spot in the test sample thin-layer chromatography has suitable Rf value and degree of separation preferably " and " inspection cost ":
Developping agent is preferably toluene-ethyl acetate 8～10: 1, and more preferably toluene-ethyl acetate 8.5～9.5: 1, toluene-ethyl acetate 9: 1 more preferably also.
Carry out demonstration test by embodiment 1 rhizoma cyperi and a-cyperolone thin-layer identification method, as a result in the test sample chromatogram, with rhizoma cyperi and the corresponding position of a-cyperolone chromatogram on, the principal spot of aobvious same color.The thin-layer chromatography good separating effect, the spot rounding, Rf value is moderate, and reappearance and specificity are good, and negative control is noiseless.
In the experimental study, once prepared by the following method need testing solution: get this product 20g, porphyrize is used ether 50ml, and ultrasonic processing 30min filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, as need testing solution.Test findings show this kind extracting method with the reference substance relevant position, negative sample has one to disturb spot, specificity is bad.
Experimental example 3: thin-layered chromatography is differentiated Radix Angelicae Sinensis and Ligusticum wallichii
In the need testing solution preparation of abortion preventing Yimu ball Radix Angelicae Sinensis and the discriminating of Ligusticum wallichii thin-layer chromatography, to what add, reagent sherwood oil (30～60 ℃) volume that is used for ultrasonic extraction, ultrasonic times etc. are investigated, in order to send better beneficial effect, i.e. balance between " having clearly corresponding spot in the test sample thin-layer chromatography with on Radix Angelicae Sinensis and the corresponding position of Ligusticum wallichii " and " inspection cost ":
Preferred reagent sherwood oil (30～60 ℃) adding volume is 10～100ml, 30～70ml more preferably, also 50ml more preferably.
The preferred ultrasonic processing time is 10～60 minutes, more preferably 20～50 minutes, and also more preferably 40 minutes.
In Radix Angelicae Sinensis and the preparation of Ligusticum wallichii control medicinal material solution, to the volume of add for the reagent sherwood oil (30～60 ℃) of ultrasonic processing, the ultrasonic processing times etc. are investigated, in order to send better beneficial effect, i.e. balance between " control medicinal material has clearly feature spot " and " inspection cost ":
Preferred reagent sherwood oil (30～60 ℃) the adding volume that is used for ultrasonic processing is 2～10ml, 3～7ml more preferably, also 5ml more preferably.
The preferred ultrasonic processing time is 10～60 minutes, more preferably 20～50 minutes, and also more preferably 40 minutes.
The thin-layer chromatography condition that abortion preventing Yimu ball Radix Angelicae Sinensis and Ligusticum wallichii are differentiated is investigated, in order to send better beneficial effect, i.e. balance between " the feature spot in the thin-layer chromatography has preferably degree of separation " and " inspection cost ":
Developping agent is preferably normal hexane-ethyl acetate 5～8: 1, and more preferably normal hexane-ethyl acetate 5.5～7: 1, normal hexane-ethyl acetate 6: 1 more preferably also.
Carry out demonstration test by embodiment 1 Radix Angelicae Sinensis and Ligusticum wallichii thin-layer identification method, as a result in the test sample chromatogram, with Radix Angelicae Sinensis and the corresponding position of Ligusticum wallichii chromatogram on, the main fluorescence spot of aobvious same color.The thin-layer chromatography good separating effect, the spot rounding, Rf value is moderate, and reappearance and specificity are good, and negative control is noiseless.
Experimental example 4: thin-layered chromatography is differentiated scutelloside
In the need testing solution preparation that abortion preventing Yimu ball scutelloside thin-layer chromatography is differentiated, to the reagent ether that adds, the volume of methyl alcohol, the heating return time, ethyl acetate extraction number of times and consumption etc. are investigated, in order to send better beneficial effect, i.e. balance between " having clearly corresponding spot in the test sample thin-layer chromatography with on the corresponding position of aurantiamarin " and " inspection cost ":
Ether adds volume, is preferably 20～100ml, 30～70ml more preferably, also 50ml more preferably.
Heating return time after adding diethyl ether is preferably 10～120 minutes, and more preferably 20～50 minutes, also more preferably 30 minutes.
Methyl alcohol adds volume, is preferably 10～100ml, 30～70ml more preferably, also 50ml more preferably.
Add the heating return time behind the methyl alcohol, be preferably 10～300 minutes, more preferably 20～100 minutes, also more preferably 45 minutes.
With the extraction time that the ethyl acetate jolting is extracted, all can extract more than 1 time, be preferably more preferably 2 times 1～4 time.
The consumption of ethyl acetate can be decided routinely during jolting was extracted, and was preferably 10～100ml, more preferably 30ml.
The thin-layer chromatography condition that abortion preventing Yimu ball scutelloside is differentiated is investigated, in order to send better beneficial effect, i.e. balance between " the feature spot in the thin-layer chromatography has preferably degree of separation " and " inspection cost ":
Developping agent is preferably ethyl acetate-butanone-formic acid-water 4～6: 2～4: 0.5～1.5: 0.5～1.5, ethyl acetate-butanone-formic acid-water 4.5～5.5: 2.5～3.5: 0.8～1.2: 0.8～1.2 more preferably, also ethyl acetate-butanone-formic acid-water 5: 3: 1 more preferably: 1.
Carry out demonstration test by embodiment 1 scutelloside thin-layer identification method, as a result in the test sample chromatogram, with the corresponding position of scutelloside chromatogram on, the spot of aobvious same color.The thin-layer chromatography good separating effect, the spot rounding, Rf value is moderate, and reappearance and specificity are good, and negative control is noiseless.
Experimental example 5: thin-layered chromatography is differentiated aurantiamarin
The need testing solution preparation method that abortion preventing Yimu ball aurantiamarin thin-layer chromatography is differentiated is identical with the need testing solution preparation method in the experimental example 4.
In the experimental study, also once test: get this product 20g, pulverize, add methyl alcohol 50ml, heating and refluxing extraction 30 minutes filters, filtrate evaporate to dryness, residue add water 40ml makes dissolving, extracts 2 times with the methenyl choloride jolting, each 30ml merges methenyl choloride liquid, with ammonia solution washing 2 times, each 20ml, abandon or adopt ammonia solution, get methenyl choloride liquid evaporate to dryness, residue adds methyl alcohol 1ml as need testing solution.Other gets dried orange peel control medicinal material 0.5g, adds sherwood oil (30～60 ℃) 5ml, and dipping spends the night, and gets supernatant, in contrast medicinal material solution.Draw respectively each 5 μ 1 of above-mentioned two kinds of solution, put on same silica gel g thin-layer plate, take toluene-ethyl acetate-formic acid (5: 4: 1) as developping agent, put under the ultraviolet lamp (365nm) and inspect.It is bad that test findings shows that this kind method spot separates.
The thin-layer chromatography condition that abortion preventing Yimu ball aurantiamarin is differentiated is investigated, in order to send better beneficial effect, i.e. balance between " the feature spot in the thin-layer chromatography has preferably degree of separation " and " inspection cost ":
Developping agent is preferably methenyl choloride-methanol-water 30～35: 15～20: lower floor's solution of 4～6, more preferably methenyl choloride-methanol-water 31～33: 16～18: lower floor's solution of 4.5～5.5, also lower floor's solution of 32: 17: 5 of methenyl choloride-methanol-water more preferably.
Carry out demonstration test by embodiment 1 aurantiamarin thin-layer identification method, as a result in the test sample chromatogram, with the corresponding position of aurantiamarin chromatogram on, the spot of aobvious same color.The thin-layer chromatography good separating effect, the spot rounding, Rf value is moderate, and reappearance and specificity are good, and negative control is noiseless.
Experimental example 6: the content of high effective liquid chromatography for measuring Paeoniflorin
In the detection method of content of the 34th phase of the 49th volume " contemporary Chinese doctor " Dec in 2011 " the HPLC method is measured the content of Paeoniflorin in a abortion preventing Yimu ball " civilian abortion preventing Yimu ball Paeoniflorin, the preparation method of need testing solution is: get this product porphyrize, precision takes by weighing 4g, put in the conical flask, precision adds methyl alcohol 50ml, close plug, weighed weight, ultrasonic processing 30min, supply the weight of less loss with methyl alcohol, filter, get subsequent filtrate, through 0.45 μ m filtering with microporous membrane, and get final product.
Compare with the content assaying method of Paeoniflorin in the abortion preventing Yimu ball described in the document, the inventive method has adopted the method that adds hot reflux to extract, and extracts more fully; And adopt neutral alumina column to remove impurity, and greatly reduce the impurity content in the need testing solution, and do not affected the accuracy of mensuration, prolonged the serviceable life of chromatographic column.Through the methodology checking, measure by the inventive method, good separation, the methodological studies such as linearity, stability, repeatability, the recovery all meet the requirement of assay, can be used for controlling the quality of this preparation.
It is as follows that the content assaying method of abortion preventing Yimu ball is learned investigation:
1. the preparation of solution
It is an amount of that the preparation precision of reference substance solution takes by weighing the Paeoniflorin reference substance, adds 50% methyl alcohol and make the solution that contains 10～60 μ g among every 1ml, and get final product.
This product 20g is got in the preparation of solid pharmaceutical preparation need testing solution, and porphyrize is got 1g, accurately weighed, put in the tool plug conical flask the accurate 70% methyl alcohol 50ml that adds, weighed weight added hot reflux 30 minutes, let cool, weighed weight is supplied the weight of loss with 70% methyl alcohol, shake up, filter, get subsequent filtrate 10ml, evaporate to dryness, make dissolving with 50% methyl alcohol 2ml, be added in (100～200 orders, 2g on the neutral alumina column, internal diameter 1cm), with methyl alcohol 50～200ml wash-out, collect eluent, steam near and do, add the dissolving of 50% methyl alcohol, and be settled in the 10ml measuring bottle, filter, get subsequent filtrate, and get final product.
Take content as index, and consider cost, the consumption that adds hot reflux solvent for use and consumption, heating return time, methanol-eluted fractions etc. investigated:
Add the hot reflux solvent for use, be preferably 50%～90% methyl alcohol, 60%～80% methyl alcohol more preferably, also 70% methyl alcohol more preferably.
Add the consumption of hot reflux solvent for use, be preferably 25～100ml, more preferably 50ml.
The heating return time is preferably 20～200 minutes, and more preferably 25～40 minutes, also more preferably 30 minutes.
The preparation negative sample solution of negative sample solution is to take by weighing all the other flavour of a drug except the root of herbaceous peony in the prescription ratio, makes root of herbaceous peony negative sample by method for making, gets this negative sample does not contain white Peony Root by preparation method's preparation of need testing solution negative sample solution again.
2. chromatographic condition and system suitability
Chromatographic column: octadecylsilane chemically bonded silica chromatographic column (Amethyst C18-P post, 4.6 * 250mm); Weil-McLain dragon chromatographic work station; Acetonitrile-water (15: 85) is mobile phase; The detection wavelength is 230nm.Number of theoretical plate calculates by Paeoniflorin should be not less than 3000.
Draw respectively Paeoniflorin reference substance solution, need testing solution and do not contain each 10 μ l of negative sample solution of the root of herbaceous peony, the injection liquid chromatography detects.Paeoniflorin and other component reach baseline separation under this condition, degree of separation R＞1.5, and number of theoretical plate is pressed the Paeoniflorin peak and is calculated greater than 3000.Without absorption peak, visible feminine gender is noiseless in the relevant position for negative sample solution chromatogram.
3. reference substance purity test
Get Paeoniflorin reference substance 5.78mg, put in the 10ml measuring bottle, add 50% methyl alcohol and make dissolving and be diluted to scale, shake up, by above-mentioned chromatographic condition, sample introduction 20 μ l measure the desolventizing peak, calculate with areas of peak normalization method, content is 99.7%, meets the assay requirement, calculates by 100% during calculating.
4. methodology checking
Linear relationship is investigated: with peak area integration A Paeoniflorin sample size C (μ g) is carried out regretional analysis, sample size is good in 0.0578～1.156 μ g scope internal linear relation, and the straight-line pass initial point, and available single-point external standard method is carried out measurement and calculation.The instrument precision test: Paeoniflorin peak area mean value is that 172813, RSD is 0.46% as a result, shows that precision is better.The test liquid stability test: mean value is that 172258, RSD is 1.12% in 12 hours, shows that need testing solution is stable in 12 hours.Replica test: content mean value is 0.5526mg/g, and RSD is 1.22%, shows method repeatability better.The average recovery test: average recovery rate is that 99.98%, RSD is 0.84%, shows that the method recovery is better.
Specific embodiment is as follows:
The quality determining method of embodiment 1 pill
1. thin-layered chromatography is differentiated stachydrine hydrochloride
Get this product 8g, porphyrize adds 50% methanol solution 50ml, added hot reflux 1 hour, and let cool, filter, filtrate is steamed to without the alcohol flavor, regulates pH value to 1～2 with watery hydrochloric acid, by Na-type strong acid cation exchange resin column (internal diameter 1.5cm, the high 10cm of post) on, washes with water to efflux closely colourlessly, discard eluent, use again strong ammonia solution ethanolic solution (19 → 100) 100ml wash-out, collect eluent, evaporate to dryness, residue makes dissolving with ethanol 1ml, as need testing solution.Other gets the stachydrine hydrochloride reference substance, adds ethanol and makes the solution that every 1ml contains 2mg, in contrast product solution.Draw need testing solution 10 μ l, reference substance solution 5 μ l, put respectively on same silica gel g thin-layer plate, take n-butyl alcohol-hydrochloric acid-ethyl acetate (8: 3: 1) as developping agent, launch, take out, dry, 105 ℃ were heated approximately 10 minutes, let cool, spray dries up with cold wind with 1% ferric trichloride ethanolic solution-rare bismuth potassium iodide test solution (1: 10) mixed solution.In the test sample chromatogram, with reference substance chromatogram relevant position on, the spot of aobvious same color.
2. thin-layered chromatography is differentiated rhizoma cyperi and a-cyperolone
Get this product 20g, porphyrize is put in the 500ml round-bottomed flask, adds water 250ml, connect volatile oil determination apparatus, add water to scale from the analyzer upper end, and overflow enters till the flask, adding sherwood oil (60～90 ℃) 2ml, add hot reflux 2 hours, and let cool, get petroleum ether layer as need testing solution.Other gets rhizoma cyperi control medicinal material 1g, the 30ml that adds diethyl ether, and ultrasonic processing 20 minutes lets cool, and filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, in contrast medicinal material solution.Get again a-cyperolone reference substance, add ethyl acetate and make the solution that every 1ml contains 1mg, in contrast product solution.Draw respectively above-mentioned three kinds of each 5ul of solution, point is on same silica GF254 thin layer plate, take toluene-ethyl acetate (9: 1) as developping agent, launch, take out, dry, put under the ultraviolet lamp (254nm) and inspect, in the test sample chromatogram, with control medicinal material and reference substance chromatogram relevant position on, the spot of aobvious same color.
3. thin-layered chromatography is differentiated Radix Angelicae Sinensis and Ligusticum wallichii
Get this product 12g, porphyrize adds sherwood oil (30～60 ℃) 50ml, and ultrasonic processing 40 minutes filters, and filtrate volatilizes, and residue adds sherwood oil (30～60 ℃) 1ml makes dissolving, as need testing solution.Other gets Radix Angelicae Sinensis and each 0.5g of Ligusticum wallichii control medicinal material, adds sherwood oil (30～60 ℃) 5ml, and supernatant is got in ultrasonic processing 40 minutes, in contrast medicinal material solution.Draw respectively above-mentioned two kinds of each 5ul of solution, point is on same silica gel g thin-layer plate, take normal hexane-ethyl acetate (6: 1) as developping agent, launch, take out, dry, put under the ultraviolet lamp (365nm) and inspect, in the test sample chromatogram, with control medicinal material chromatogram relevant position on, the spot of aobvious same color.
4. thin-layered chromatography is differentiated scutelloside
Get this product 18g, porphyrize, the 50ml that adds diethyl ether added hot reflux 30 minutes, filtered, and discarded ether solution.The dregs of a decoction are flung to ether, add methyl alcohol 50ml, add hot reflux 45 minutes, filter, filtrate evaporate to dryness, residue add water 30ml makes dissolving, filter, filtrate is extracted 2 times with the ethyl acetate jolting with salt acid for adjusting pH value to 2, each 30ml, combined ethyl acetate liquid, evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution.Other gets the scutelloside reference substance, adds methyl alcohol and makes the solution that every 1ml contains 1mg, in contrast product solution.Draw each 2～5ul of above-mentioned two kinds of solution, put respectively in same take the carboxymethylcellulose sodium solution that contains 4% sodium acetate on the silica gel g thin-layer plate of binder, make into strips, take ethyl acetate-butanone-formic acid-water (5: 3: 1: 1) as developping agent, put the interior presaturation of expansion cylinder 30 minutes, launch, take out, dry, spray is with 5% ferric trichloride ethanolic solution.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, the spot of aobvious same color.
5. thin-layered chromatography is differentiated aurantiamarin
Get this product 18g, porphyrize, the 50ml that adds diethyl ether added hot reflux 30 minutes, filtered, and discarded ether solution.The dregs of a decoction are flung to ether, add methyl alcohol 50ml, add hot reflux 45 minutes, filter, filtrate evaporate to dryness, residue add water 30ml makes dissolving, filter, filtrate is extracted 2 times with the ethyl acetate jolting with salt acid for adjusting pH value to 2, each 30ml, combined ethyl acetate liquid, evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution.Get the aurantiamarin reference substance, add methyl alcohol and make saturated solution, in contrast product solution.Draw need testing solution and each 5ul of reference substance solution, put respectively in same take the carboxymethylcellulose sodium solution that contains 1% NaOH on the silica gel g thin-layer plate of binder, make into strips, take lower floor's solution of chloroform-methanol-water (32: 17: 5) as developping agent, put in the expansion cylinder presaturation 30 minutes, launch, take out, dry, spray is put under the ultraviolet lamp (365nm) and is inspected with the aluminium choride test solution.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, the fluorescence spot of aobvious same color.
6. the content of high effective liquid chromatography for measuring Paeoniflorin
The chromatographic condition octadecylsilane chemically bonded silica is filling agent; Acetonitrile-water (15: 85) is mobile phase; The detection wavelength is 230nm.
It is an amount of that the preparation precision of reference substance solution takes by weighing the Paeoniflorin reference substance, adds 50% methyl alcohol and make the solution that contains 12 μ g among every 1ml, and get final product.
This product 20g is got in the preparation of need testing solution, and porphyrize is got the approximately weight of 1g, accurately weighed, put in the tool plug conical flask the accurate 70% methyl alcohol 50ml that adds, weighed weight adds hot reflux 30min, lets cool, weighed weight is supplied the weight of loss with 70% methyl alcohol, shake up, filter, get subsequent filtrate 10ml, evaporate to dryness, make dissolving with 50% methyl alcohol 2ml, be added in neutral alumina column (100～200 orders, 2g, internal diameter 1cm) on, with methyl alcohol 80ml wash-out, collects loading efflux and eluent, steam near and do, add the dissolving of 50% methyl alcohol, and be settled in the 10ml measuring bottle, filter, get subsequent filtrate, and get final product.
Determination method is accurate reference substance solution, each 10 μ l of need testing solution of drawing respectively, and the injection liquid chromatography is measured, and be get final product.
The quality determining method of embodiment 2 tablets, capsule, granule, powder
1. thin-layered chromatography is differentiated stachydrine hydrochloride
Get this product porphyrize, get 5g, add 20% methanol solution 100ml, heating and refluxing extraction 4 hours filters, and filtrate is steamed to nothing alcohol flavor, regulate pH value to 1～2 with watery hydrochloric acid, by strong acid cation exchange resin column, wash with water to efflux closely colourless, discard eluent, use again strong ammonia solution ethanolic solution (19 → 100) wash-out, collect eluent, evaporate to dryness, residue makes dissolving with ethanol 1ml, as need testing solution.Get the stachydrine hydrochloride reference substance, add ethanol and make the solution that every 1ml contains 1～2mg, in contrast product solution.Absorption need testing solution, reference substance solution are put respectively on same silica gel g thin-layer plate, take n-butyl alcohol-hydrochloric acid-ethyl acetate 6: 4: 1 as developping agent, launch, take out, dry, 120 ℃ were heated 2 minutes, and let cool, spray dries up with 1% ferric trichloride ethanolic solution-1: 10 mixed solution of rare bismuth potassium iodide test solution.In the test sample chromatogram, with reference substance chromatogram relevant position on, the spot of aobvious same color.
2. thin-layered chromatography is differentiated rhizoma cyperi and a-cyperolone
Get this product porphyrize, get 8g, put in the round-bottomed flask, add water an amount of, connect volatile oil determination apparatus, add water to scale from the analyzer upper end, and overflow enters till the flask, add again sherwood oil (60～90 ℃) 5ml, add hot reflux 4 hours, and let cool, get petroleum ether layer as need testing solution.Other gets rhizoma cyperi control medicinal material 2g, the 100ml that adds diethyl ether, and ultrasonic processing 60 minutes lets cool, and filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, in contrast medicinal material solution.Get again a-cyperolone reference substance, add ethyl acetate and make the solution that every 1ml contains 2mg, in contrast product solution.Draw respectively above-mentioned three kinds of solution, put on same silica GF254 thin layer plate, take toluene-ethyl acetate 10: 1 as developping agent, launch, take out, dry, put under the ultraviolet lamp 254nm and inspect, in the test sample chromatogram, with control medicinal material and reference substance chromatogram relevant position on, the spot of aobvious same color.
3. thin-layered chromatography is differentiated Radix Angelicae Sinensis and Ligusticum wallichii
Get this product porphyrize, get 5g, add sherwood oil (30～60 ℃) 100ml, ultrasonic processing 60 minutes is got sherwood oil liquid and is volatilized, and residue adds sherwood oil (30～60 ℃) 1ml makes dissolving, as need testing solution.
Get Radix Angelicae Sinensis and each 2g of Ligusticum wallichii control medicinal material, add sherwood oil (30～60 ℃) 10ml, supernatant is got in ultrasonic processing 60 minutes, in contrast medicinal material solution.Draw respectively mentioned solution, put on same silica gel g thin-layer plate, take normal hexane-ethyl acetate 8: 1 as developping agent, launch, take out, dry, put under the ultraviolet lamp (365nm) and inspect, in the test sample chromatogram, with control medicinal material chromatogram relevant position on, the spot of aobvious same color.
4. thin-layered chromatography is differentiated scutelloside
Get this product porphyrize, get 5g, the 100ml that adds diethyl ether added hot reflux 120 minutes, filtered, and discarded ether solution.The dregs of a decoction are flung to ether, add methyl alcohol 100ml, add hot reflux 300 minutes, filter, filtrate evaporate to dryness, residue add water 100ml makes dissolving, filter, get filtrate, with salt acid for adjusting pH value to 2, extract 1 time with ethyl acetate 100ml jolting, get acetic acid ethyl fluid, evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution.Other gets the scutelloside reference substance, adds methyl alcohol and makes the solution that every 1ml contains 2mg, in contrast product solution.Draw above-mentioned two kinds of solution, put respectively in same take the carboxymethylcellulose sodium solution that contains 4% sodium acetate on the silica gel g thin-layer plate of binder, take ethyl acetate-butanone-formic acid-water 6: 2: 1.5: 0.5 as developping agent, launch, take out, dry, spray is with 5% ferric trichloride ethanolic solution.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, the spot of aobvious same color.
5. thin-layered chromatography is differentiated aurantiamarin
Get this product porphyrize, get 5g, the 100ml that adds diethyl ether added hot reflux 120 minutes, filtered, and discarded ether solution.The dregs of a decoction are flung to ether, add methyl alcohol 100ml, add hot reflux 300 minutes, filter, filtrate evaporate to dryness, residue add water 100ml makes dissolving, filter, get filtrate, with salt acid for adjusting pH value to 2, extract 1 time with ethyl acetate 100ml jolting, get acetic acid ethyl fluid, evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution.Get the aurantiamarin reference substance, add methyl alcohol and make saturated solution, in contrast product solution.Draw above-mentioned need testing solution and reference substance solution, put respectively in same with the silica gel g thin-layer plate that contains 1% NaOH on, take lower floor's solution of 30: 20: 6 of methenyl choloride-methanol-water as developping agent, launch, take out, dry, spray is put under the ultraviolet lamp (365nm) and is inspected with the aluminium choride test solution.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, the fluorescence spot of aobvious same color.
6. the content of high effective liquid chromatography for measuring Paeoniflorin
The chromatographic condition octadecylsilane chemically bonded silica is filling agent; Acetonitrile-water is mobile phase at 15: 85; The detection wavelength is 230nm.
It is an amount of that the preparation precision of reference substance solution takes by weighing the Paeoniflorin reference substance, adds 50% methyl alcohol and make the solution that contains 60 μ g among every 1ml, and get final product.
This product porphyrize is got in the preparation of need testing solution, gets 10g, and is accurately weighed, put in the tool plug conical flask accurate 90% methyl alcohol 100ml, the weighed weight of adding, added hot reflux 200 minutes, and let cool, weighed weight, supply the weight of loss with 90% methyl alcohol, shake up, filter, get subsequent filtrate 10ml, evaporate to dryness, 2～5ml makes dissolving with 90% methyl alcohol, is added on the neutral alumina column, with methyl alcohol 200ml wash-out, collect eluent, steam near and do, add the dissolving of 50% methyl alcohol, and be settled in the 10ml measuring bottle, filter, get subsequent filtrate, and get final product.
Determination method is accurate reference substance solution, the need testing solution 20 μ l of drawing respectively, and the injection liquid chromatography is measured, and be get final product.
The quality determining method of embodiment 3 syrups, mixture
1. thin-layered chromatography is differentiated stachydrine hydrochloride
Get this product mixing, get 2g, regulate pH value to 1～2 with watery hydrochloric acid, by strong acid cation exchange resin column, wash with water to efflux closely colourless, discard eluent, use again strong ammonia solution ethanolic solution (19 → 100) wash-out, collect eluent, evaporate to dryness, residue makes dissolving with ethanol 1ml, as need testing solution.Get the stachydrine hydrochloride reference substance, add ethanol and make the solution that every 1ml contains 1mg, in contrast product solution.Absorption need testing solution, reference substance solution are put respectively on same silica gel g thin-layer plate, take n-butyl alcohol-hydrochloric acid-ethyl acetate 10: 2: 1 as developping agent, launch, take out, dry, 90 ℃ were heated 20 minutes, and let cool, spray dries up with 1% ferric trichloride ethanolic solution-1: 10 mixed solution of rare bismuth potassium iodide test solution.In the test sample chromatogram, with reference substance chromatogram relevant position on, the spot of aobvious same color.
2. thin-layered chromatography is differentiated rhizoma cyperi and a-cyperolone
Get this product mixing, get 5g, put in the round-bottomed flask, add water an amount of, connect volatile oil determination apparatus, add water to scale from the analyzer upper end, and overflow enters till the flask, add again sherwood oil (60～90 ℃) 1ml, add hot reflux 0.5 hour, and let cool, get petroleum ether layer as need testing solution.Other gets rhizoma cyperi control medicinal material 0.5g, the 10m1 that adds diethyl ether, and ultrasonic processing 10 minutes lets cool, and filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, in contrast medicinal material solution.Get again a-cyperolone reference substance, add ethyl acetate and make the solution that every 1ml contains 0.5mg, in contrast product solution.Draw respectively above-mentioned three kinds of solution, put on same silica GF254 thin layer plate, take toluene-ethyl acetate 8: 1 as developping agent, launch, take out, dry, put under the ultraviolet lamp 254nm and inspect, in the test sample chromatogram, with control medicinal material and reference substance chromatogram relevant position on, the spot of aobvious same color.
3. thin-layered chromatography is differentiated Radix Angelicae Sinensis and Ligusticum wallichii
Get this product mixing, get 10g, put in the separating funnel, add sherwood oil (30～60 ℃) 20ml, jolting is extracted, and minute gets petroleum ether layer to volatilize, and residue adds sherwood oil (30～60 ℃) 1ml makes dissolving, as need testing solution.Get Radix Angelicae Sinensis and each 0.2g of Ligusticum wallichii control medicinal material, add sherwood oil (30～60 ℃) 2ml, supernatant is got in ultrasonic processing 10 minutes, in contrast medicinal material solution.Draw respectively mentioned solution, put on same silica gel g thin-layer plate, take normal hexane-ethyl acetate 5: 1 as developping agent, launch, take out, dry, put under the ultraviolet lamp (365nm) and inspect, in the test sample chromatogram, with control medicinal material chromatogram relevant position on, the spot of aobvious same color.
4. thin-layered chromatography is differentiated scutelloside
Get this product mixing, get 10g, with salt acid for adjusting pH value to 2, extract 3 times with the ethyl acetate jolting, each 20ml gets acetic acid ethyl fluid, and evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution.Other gets the scutelloside reference substance, adds methyl alcohol and makes the solution that every 1ml contains 0.5mg, in contrast product solution.Draw above-mentioned two kinds of solution, put respectively in same take the carboxymethylcellulose sodium solution that contains 4% sodium acetate on the silica gel g thin-layer plate of binder, take ethyl acetate-butanone-formic acid-water 4: 4: 0.5: 1.5 as developping agent, launch, take out, dry, spray is with 5% ferric trichloride ethanolic solution.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, the spot of aobvious same color.
5. thin-layered chromatography is differentiated aurantiamarin
Get this product mixing, get 10g, with salt acid for adjusting pH value to 2, extract 3 times with the ethyl acetate jolting, each 20ml gets acetic acid ethyl fluid, and evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution.Get the aurantiamarin reference substance, add methyl alcohol and make saturated solution, in contrast product solution.Draw above-mentioned need testing solution and reference substance solution, put respectively in same with the silica gel g thin-layer plate that contains 1% NaOH on, take lower floor's solution of 35: 15: 4 of methenyl choloride-methanol-water as developping agent, launch, take out, dry, spray is put under the ultraviolet lamp (365nm) and is inspected with the aluminium choride test solution.In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, the fluorescence spot of aobvious same color.
6. the content of high effective liquid chromatography for measuring Paeoniflorin
The chromatographic condition octadecylsilane chemically bonded silica is filling agent; Acetonitrile-water is mobile phase at 15: 85; The detection wavelength is 230nm.
It is an amount of that the preparation precision of reference substance solution takes by weighing the Paeoniflorin reference substance, adds 50% methyl alcohol and make the solution that contains 10 μ g among every 1ml, and get final product.
This product mixing is got in the preparation of need testing solution, and precision measures 1g, and evaporate to dryness makes dissolving with 90% methyl alcohol 2ml, is added on the neutral alumina column, with methyl alcohol 50ml wash-out, collect eluent, steam near and do, add the dissolving of 50% methyl alcohol, and be settled in the 10ml measuring bottle, filter, get subsequent filtrate, and get final product.
Determination method is accurate reference substance solution, the need testing solution 5 μ l of drawing respectively, and the injection liquid chromatography is measured, and be get final product.
Although illustrated and described specific embodiments of the present invention, it is obvious to those skilled in the art that in the situation that do not break away from the spirit and scope of the invention and can make multiple other change and modification.Therefore, in additional claims, be intended to comprise all such changes and modification in the scope of the invention.
Claims (1)
1. the quality determining method of an abortion preventing Yimu preparation is characterized in that, this quality determining method comprises the one or more of following detection:
1. thin-layered chromatography is differentiated stachydrine hydrochloride
This product porphyrize is got in the preparation of solid pharmaceutical preparation need testing solution, gets in right amount, adds 20%～90% methanol solution, 10～100ml, heating and refluxing extraction 0.5～4 hour filters, and filtrate is steamed to nothing alcohol flavor, regulate pH value to 1～2 with watery hydrochloric acid, by strong acid cation exchange resin column, wash with water to efflux closely colourless, discard eluent, use again strong ammonia solution ethanolic solution (19 → 100) wash-out, collect eluent, evaporate to dryness, residue makes dissolving with ethanol 1ml, as need testing solution;
This product mixing is got in the preparation of liquid preparation need testing solution, get an amount of, regulate pH value to 1～2 with watery hydrochloric acid, by strong acid cation exchange resin column, wash with water to efflux closely colourless, discard eluent, use again strong ammonia solution ethanolic solution (19 → 100) wash-out, collect eluent, evaporate to dryness, residue makes dissolving with ethanol 1ml, as need testing solution;
The stachydrine hydrochloride reference substance is got in the preparation of reference substance solution, adds ethanol and makes the solution that every 1ml contains 1～2mg, in contrast product solution;
Absorption need testing solution, reference substance solution are put respectively on same silica gel g thin-layer plate, take n-butyl alcohol-hydrochloric acid-ethyl acetate 6～10: 2～4: 1 as developping agent, launch, take out, dry, 90～120 ℃ were heated 2～20 minutes, and let cool, spray dries up with 1% ferric trichloride ethanolic solution-1: 10 mixed solution of rare bismuth potassium iodide test solution; In the test sample chromatogram, with reference substance chromatogram relevant position on, the spot of aobvious same color;
2. thin-layered chromatography is differentiated rhizoma cyperi and a-cyperolone
Get this product porphyrize or mixing, get an amount of, put in the round-bottomed flask, add water an amount of, connect volatile oil determination apparatus, add water to scale from the analyzer upper end, and overflow enters till the flask, adds 60～90 ℃ of sherwood oil 1～5ml again, adds hot reflux 0.5～4 hour, let cool, get petroleum ether layer as need testing solution; Other gets rhizoma cyperi control medicinal material 0.5～2g, the 10～100ml that adds diethyl ether, and ultrasonic processing 10～60 minutes lets cool, and filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, in contrast medicinal material solution; Get again a-cyperolone reference substance, add ethyl acetate and make the solution that every 1ml contains 0.5～2mg, in contrast product solution; Draw respectively above-mentioned three kinds of solution, point is on same silica GF254 thin layer plate, take toluene-ethyl acetate 8～10: 1 as developping agent, launch, take out, dry, put under the ultraviolet lamp 254nm and inspect, in the test sample chromatogram, with control medicinal material and reference substance chromatogram relevant position on, the spot of aobvious same color;
3. thin-layered chromatography is differentiated Radix Angelicae Sinensis and Ligusticum wallichii
This product porphyrize is got in the preparation of solid pharmaceutical preparation need testing solution, gets in right amount, adds 30～60 ℃ of sherwood oil 10～100ml, and ultrasonic processing 10～60 minutes is got sherwood oil liquid and volatilized, and residue adds 30～60 ℃ of sherwood oil 1ml makes dissolving, as need testing solution;
This product mixing is got in the preparation of liquid preparation need testing solution, gets in right amount, puts in the separating funnel, adds 30～60 ℃ of sherwood oils, and jolting is extracted, and minute gets petroleum ether layer to volatilize, and residue adds sherwood oil (30～60 ℃) 1ml makes dissolving, as need testing solution;
Radix Angelicae Sinensis and each 0.2～2g of Ligusticum wallichii control medicinal material are got in the preparation of control medicinal material solution, add 30～60 ℃ of sherwood oil 2～10ml, and supernatant is got in ultrasonic processing 10～60 minutes, in contrast medicinal material solution;
Draw respectively mentioned solution, put on same silica gel g thin-layer plate, take normal hexane-ethyl acetate 5～8: 1 as developping agent, launches, take out, dry, put under the ultraviolet lamp 365nm and inspect, in the test sample chromatogram, with control medicinal material chromatogram relevant position on, the spot of aobvious same color;
4. thin-layered chromatography is differentiated scutelloside
This product porphyrize is got in the preparation of solid pharmaceutical preparation need testing solution, gets in right amount, and the 20～100ml that adds diethyl ether added hot reflux 10～120 minutes, filters, and discards ether solution; The dregs of a decoction are flung to ether, add methyl alcohol 10～100ml, add hot reflux 10～300 minutes, filter, filtrate evaporate to dryness, residue add water 10～100ml makes dissolving, filter, get filtrate, with salt acid for adjusting pH value to 2, extract with the ethyl acetate jolting, get acetic acid ethyl fluid, evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution;
This product mixing is got in the preparation of liquid preparation need testing solution, gets in right amount, with salt acid for adjusting pH value to 2, extracts with the ethyl acetate jolting, gets acetic acid ethyl fluid, and evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution;
The scutelloside reference substance is got in the preparation of reference substance solution, adds methyl alcohol and makes the solution that every 1ml contains 0.5～2mg, in contrast product solution;
Draw above-mentioned two kinds of solution, put respectively in same take the carboxymethylcellulose sodium solution that contains 4% sodium acetate on the silica gel g thin-layer plate of binder, take ethyl acetate-butanone-formic acid-water 4～6: 2～4: 0.5～1.5: 0.5～1.5 as developping agent, launch, take out, dry, spray is with 5% ferric trichloride ethanolic solution; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, the spot of aobvious same color;
5. thin-layered chromatography is differentiated aurantiamarin
This product porphyrize is got in the preparation of solid pharmaceutical preparation need testing solution, gets in right amount, and the 20～100ml that adds diethyl ether added hot reflux 10～120 minutes, filters, and discards ether solution.The dregs of a decoction are flung to ether, add methyl alcohol 10～100ml, add hot reflux 10～300 minutes, filter, filtrate evaporate to dryness, residue add water 10～100ml makes dissolving, filter, get filtrate, with salt acid for adjusting pH value to 2, extract with the ethyl acetate jolting, get acetic acid ethyl fluid, evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution;
This product mixing is got in the preparation of liquid preparation need testing solution, gets in right amount, with salt acid for adjusting pH value to 2, extracts with the ethyl acetate jolting, gets acetic acid ethyl fluid, and evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution;
The aurantiamarin reference substance is got in the preparation of reference substance solution, adds methyl alcohol and makes saturated solution, in contrast product solution;
Draw above-mentioned need testing solution and reference substance solution, put respectively in same with the silica gel g thin-layer plate that contains 1% NaOH on, take methenyl choloride-methanol-water 30～35: lower floor's solution of 15～20: 4～6 is as developping agent, launch, take out, dry, spray is put under the ultraviolet lamp 365nm and is inspected with the aluminium choride test solution; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, the fluorescence spot of aobvious same color;
6. the content of high effective liquid chromatography for measuring Paeoniflorin
The chromatographic condition octadecylsilane chemically bonded silica is filling agent; Acetonitrile-water is mobile phase at 15: 85; The detection wavelength is 230nm;
It is an amount of that the preparation precision of reference substance solution takes by weighing the Paeoniflorin reference substance, adds 50% methyl alcohol and make the solution that contains 10～60 μ g among every 1ml, and get final product;
This product porphyrize is got in the preparation of solid pharmaceutical preparation need testing solution, gets an amount of, accurately weighed, put in the tool plug conical flask accurate 50%～90% methyl alcohol, 25～100ml, the weighed weight of adding, added hot reflux 20～200 minutes, and let cool, weighed weight, supply the weight of loss with 50%～90% methyl alcohol, shake up, filter, get subsequent filtrate 10ml, evaporate to dryness, 2～5ml makes dissolving with 50%～90% methyl alcohol, is added on the neutral alumina column, with methyl alcohol 50～200ml wash-out, collect eluent, steam near and do, add the dissolving of 50% methyl alcohol, and be settled in the 10ml measuring bottle, filter, get subsequent filtrate, and get final product;
This product mixing is got in the preparation of liquid preparation need testing solution, and precision measures in right amount, evaporate to dryness, 2～5ml makes dissolving with 50%～90% methyl alcohol, be added on the neutral alumina column, with methyl alcohol 50～200ml wash-out, collect eluent, steam near and do, add the dissolving of 50% methyl alcohol, and be settled in the 10ml measuring bottle, filter, get subsequent filtrate, and get final product;
Determination method is accurate reference substance solution, need testing solution 1～20 μ l of drawing respectively, and the injection liquid chromatography is measured, and be get final product.
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| CN2013102760194A CN103308644A (en) | 2013-07-03 | 2013-07-03 | Quality detection method for miscarriage-preventing leonurus preparation |
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| CN2013102760194A CN103308644A (en) | 2013-07-03 | 2013-07-03 | Quality detection method for miscarriage-preventing leonurus preparation |
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| CN2013102760194A Pending CN103308644A (en) | 2013-07-03 | 2013-07-03 | Quality detection method for miscarriage-preventing leonurus preparation |
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| CN104749312A (en) * | 2013-12-27 | 2015-07-01 | 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 |
| CN104997953A (en) * | 2014-04-24 | 2015-10-28 | 河北以岭医药研究院有限公司 | Method for determining contents of two components in traditional Chinese medicinal composition, and method for discriminating various components |
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| CN110361496A (en) * | 2019-08-01 | 2019-10-22 | 西安碑林药业股份有限公司 | The detection method of scutelloside in a kind of Chinese materia medica preparation Jinsangsanjie ball |
| CN113552246A (en) * | 2021-05-27 | 2021-10-26 | 贵州医科大学附属医院 | Quality control method of red granules |
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