CN104133025A - Detection method for Zhechong wound healing preparation - Google Patents
Detection method for Zhechong wound healing preparation Download PDFInfo
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- CN104133025A CN104133025A CN201410066024.7A CN201410066024A CN104133025A CN 104133025 A CN104133025 A CN 104133025A CN 201410066024 A CN201410066024 A CN 201410066024A CN 104133025 A CN104133025 A CN 104133025A
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Abstract
The invention provides a detection method for a Zhechong wound healing preparation. The detection method is characterized by comprising one or more selected from the following processes: qualitative identification of red ginseng, Himalayan teasel root and fortune's drynaria rhizome in the Zhechong wound healing preparation by using thin-layer chromatography; and determination of the content of Asperosaponin VI in the Himalayan teasel root in the Zhechong wound healing preparation by using high performance liquid chromatography. With the detection method, effective detection can be carried out on each component in the Zhechong wound healing preparation, so product quality can be better controlled.
Description
Technical field
The present invention relates to the more detection method of preparation of Yi Zhong female ground beetle wound.
Background technology
Female ground beetle wound capsule for curing is medical institutions' preparation, ground bettle, teasel root, the rhizome of davallia, calcined Dragon's bone, calcined oyster shell, red ginseng, Morinda officinalis, Radix Angelicae Sinensis eight taste medicines, consists of.Have stimulate the circulation of the blood and cause the muscles and joints to relax, the effect of promoting blood circulation and removing blood stasis, swelling and pain relieving, be mainly used in the soft tissue bruise due to traumatic injury and closed fracture, can alleviate the acute stage pain of traumatic injury.
Medical institutions' preparation refer to medical institutions according to our unit clinically need to prepare through approval, personal fixed prescription preparation.The same with other medicines, the preparation of medical institutions' preparation must carry out quality inspection with standard preparation according to the rules, underproof, must not use.According to relevant laws and regulations, medical institutions' preparation only can be sold in preparation hospital, must not on market, sell or in a disguised form sell.Or adjust and use in other medical institutions through approval.
Ground bettle: record in one 18 pages of Pharmacopoeia of People's Republic of China versions in 2010.This product is the dry body of the female worm of Corydiidae insect eupolyphoge sinensis Eupolyphaga Sinesis Walker or Ji eupolyphoge sinensis Steleophaga plancyi (Boleny).After seizure, put in boiling water and scald extremely, dry or dry.
Teasel root: record in one 309 pages of Pharmacopoeia of People's Republic of China versions in 2010.This product is the dry root of Dipsacaceae plant teasel Dipsacus asper Wall.exHenry.Excavate autumn, removes root and fibrous root, is dried to half-driedly with low baking temperature, banks up " sweating " when the inner virescence, then dry.
The rhizome of davallia: record in one 239 pages of Pharmacopoeia of People's Republic of China versions in 2010.This product is the dry rhizome of Plants of Polypodiaceae Mongolian oak fern Drynaria fortunei (Kunze) J.Sm..All can excavate the whole year, removes silt, dry, or burn remove floss (scale) again.
Calcined Dragon's bone: medicinal material Os Draconis(Fossilia Ossia Mastodi) former mineral Fossilia Ossis Mastrodi; Come from < < herbal classic > > etc.Keel be ancient times mammal as if the fossil of the bone of class, rhinoceros class, Hippocampal cortex etc.Distribution Sichuan, Shanxi, Shandong, Hebei, the Inner Mongol, Henan, Shaanxi, Gansu, Qinghai.After digging out, remove earth and impurity.Principal ingredient is calcium carbonate, calcium phosphate, also iron content, potassium, sodium, chlorine, sulfate radical etc.Property sweet puckery, flat.Enter the heart, liver, kidney, large intestine channel.Transquilization with heavy material tranquillizing the mind by relieving convulsion, arrest sweating controlling nocturnal emission with astringent drugs, the puckery intestines that stop blooding, regenerating tissue to heal wond.Control frightened epilepsy demented, palpitation is forgetful, insomnia and dreamful sleep, and spontaneous sweating, seminal emission stranguria with turbid discharge, tells nosebleed and has blood in stool, and under uterine bleeding band, rushes down dysentery prolapse of the anus, and ulcer does not close up for a long time.Calcined Dragon's bone is exactly the keel of getting outwash, on smokeless stove fire or in crucible, forges and is popular in, and takes out, and cools, and pulverizes.
Calcined oyster shell: record in one 161 pages of Pharmacopoeia of People's Republic of China versions in 2010.This product is the shell of the long oyster Ostrea of Ostreidae animal gigas Thunberg, ostrea talienwhanensis Crosse Ostrae talienwhanensis Crosse or Crassostrea rivularis Ostrea rivularis Gould.All can fish for the whole year, and fleshing is cleaned.Dry.< < Chinese Pharmacopoeia > > nineteen ninety-five version is " get clean oyster, according to method of direct calcination, be calcined to crisp " to the process of preparing Chinese medicine regulation of oyster.It is strong that calcined oyster shell can be restrained the effect of astringent or styptic treatment for spontaneous sweating deacidification, and treatment has a stomach-ache, hydrochloric acid in gastric juice etc.
Red ginseng: record in one 143 pages of Pharmacopoeia of People's Republic of China versions in 2010.This product is the cultivation product of Araliaceae ginseng Panax ginseng C.A.Mey. dry root and the rhizomes after steaming.Excavate autumn, cleans, after steaming, dry.
Morinda officinalis: record in one 75 pages of Pharmacopoeia of People's Republic of China versions in 2010.This product is the dry root of madder wort Morinda officinalis Morinda offinalis How.All can excavate the whole year, cleans, and removes fibrous root, shines to 6 seventy percent dry, beats gently flatly, dries.
Radix Angelicae Sinensis: record in one 124 pages of Pharmacopoeia of People's Republic of China versions in 2010.This product is the dry root of umbelliferae angelica Angelica sinensis (Oliv.) Diels..Autumn end excavates, and removes fibrous root and silt, after moisture slightly evaporates, is bundled into wisp, and upper canopy, with pyrotechnics smoke-dried beancurd slowly.
Quality inspection is the technical parameter of the drug quality characteristic of reflection, and index is clearly stipulated, and forms technological document, and regulation drug quality specification and the method for inspection, be exactly drug standard.The current Shang Weiyou female ground beetle wound quality determining method of preparation more, does not have composition in the other side to carry out the method for content detection yet.
Summary of the invention
The object of this invention is to provide the more detection method of preparation of Yi Zhong female ground beetle wound, described method favorable reproducibility, specificity are strong, effectively Dui female ground beetle wound more preparation carry out quality inspection, thereby the Shi female ground beetle wound steady quality, safe, controlled of preparation more.
The invention provides the more detection method of preparation of Yi Zhong female ground beetle wound, described method comprises one or more in following method: by red ginseng, teasel root, the rhizome of davallia in thin layer chromatography Dui female ground beetle wound more preparation, carry out qualitative discriminating; Content by the asperosaponin VI in the teasel root of high performance liquid chromatography Dui female ground beetle wound more preparation is measured.
Preferably, the qualitative discriminating Shi of described red ginseng is after the female ground beetle wound more ultrasonic extraction of preparation, and the methenyl choloride-ethyl acetate-methanol-water of take carries out thin-layer chromatography at 10 ℃ of following lower floor's solution of placing as developping agent; Preferably, described ultrasonic extraction Shi adds female ground beetle wound more preparation after the ultrasonic extraction of methenyl choloride, then adds the ultrasonic extraction of normal butyl alcohol, and filtrate first adds ammoniacal liquor, then adds methyl alcohol to dissolve, and obtains ultrasonic extraction solution; More preferably, the mass ratio of described methenyl choloride, ethyl acetate, methyl alcohol, water is 3:8:4.5:2.
Further, the qualitative discriminating of described red ginseng is to get more preparation content 1.5-7.5 weight portion of Suo Shu female ground beetle wound, adds methenyl choloride 10-60 parts by volume, ultrasonic 10min, discards methenyl choloride liquid, and the dregs of a decoction volatilize, add water 0.1-1.0 parts by volume wetting, add water-saturated n-butanol 5-20 parts by volume, ultrasonic processing 10-30 minute, filter, filtrate adds 1-5 times of volume ammonia solution, shakes up, place layering, get upper strata liquid evaporate to dryness, residue adds methyl alcohol 1-5 parts by volume and dissolves, as need testing solution, separately get red ginseng control medicinal material powder 0.1-1.0 weight portion, be made in the same way of control medicinal material solution, in prescription ratio and preparation technology, preparation does not contain the negative sample of red ginseng, and by the compound method of above-mentioned need testing solution, makes the negative sample solution that does not contain red ginseng, according to thin-layered chromatography (appendix VIB of < < Chinese Pharmacopoeia > > version in 2010), test, draw above-mentioned each 0.001-0.02 parts by volume of three kinds of solution, put respectively on same silica gel g thin-layer plate, with methenyl choloride-ethyl acetate-methanol-water, volume ratio 1～10:5～15:2.5～10:1.0～5.0, 10 ℃ of following lower floor's solution of placing are developping agent, launch, take out, dry, spray is with 1%～20% ethanol solution of sulfuric acid, it is clear that hot blast blows to spot colour developing, put under uviol lamp (365nm) and inspect, in test sample chromatogram, with control medicinal material chromatogram relevant position on aobvious same color spot, and negative noiseless.
Preferably, the qualitative discriminating Shi of described teasel root is Jiang female ground beetle wound more preparation with after the ultrasonic extraction of methyl alcohol, and the upper solution of normal butyl alcohol-glacial acetic acid-water of take is carried out thin-layer chromatography as developping agent; Preferably, the mass ratio of described normal butyl alcohol, glacial acetic acid, water is 4:1:5.
Further, the qualitative discriminating of described teasel root is to get more preparation content 0.1-5 weight portion of Suo Shu female ground beetle wound, adds methyl alcohol 5-20 parts by volume, ultrasonic processing 10-50 minute, 4000r/min, 5min centrifuging and taking supernatant, evaporate to dryness, residue adds methyl alcohol 0.5-5 parts by volume to be made to dissolve, as need testing solution; Separately get teasel root control medicinal material powder 0.05-1.0 weight portion, be made in the same way of control medicinal material solution; In prescription ratio and preparation technology, preparation does not contain the negative sample of teasel root, and by the compound method of above-mentioned need testing solution, makes the negative sample solution that does not contain teasel root; According to thin-layered chromatography (appendix VIB of < < Chinese Pharmacopoeia > > version in 2010), test, draw above-mentioned each 0.001-0.02 parts by volume of three kinds of solution, put respectively on same silica gel g thin-layer plate, normal butyl alcohol-glacial acetic acid-water, the upper solution of volume ratio 1～10:0.5～5:2.0～10 is developping agent, launch, exhibition is apart from about 1-10cm, take out, dry, spray is with 1%～20% ethanol solution of sulfuric acid, and it is clear that hot blast blows to spot colour developing, puts under daylight and inspect; In test sample chromatogram, with control medicinal material chromatogram relevant position on aobvious same color spot, and negative noiseless.
Preferably, the qualitative discriminating Shi of the described rhizome of davallia is Jiang female ground beetle wound more preparation with after the ultrasonic extraction of methyl alcohol, and the upper solution of toluene-ethyl acetate-formic acid-water of take is carried out thin-layer chromatography as developping agent; Preferably, the mass ratio of described toluene, ethyl acetate, formic acid, water is 1:12:2.5:3.
Further, the qualitative discriminating of the described rhizome of davallia is to get more preparation content 0.1-5 weight portion of Suo Shu female ground beetle wound, adds methyl alcohol 10-50 parts by volume, ultrasonic 5-30 minute, 4000r/min, 5min centrifuging and taking supernatant, evaporate to dryness, residue adds methyl alcohol 0.5-5 parts by volume to be made to dissolve, as need testing solution; Separately get aurantiin reference substance appropriate, add methyl alcohol and make every 1 parts by volume containing the solution of 0.05-1.00 weight portion, in contrast product solution; In prescription ratio and preparation technology, preparation does not contain the negative sample of the rhizome of davallia, and by the compound method of above-mentioned need testing solution, makes the negative sample solution that does not contain the rhizome of davallia; According to thin-layered chromatography (appendix VIB of < < Chinese Pharmacopoeia > > version in 2010), test, draw above-mentioned each 0.001-0.03 parts by volume of three kinds of solution, put respectively on same silica gel g thin-layer plate, with toluene-ethyl acetate-formic acid-water, the upper solution of volume ratio 0.1～5:5～25:1.0～10:1.0～5 is developping agent, launch, take out, dry, spray is with 1%-20% vanillic aldehyde ethanol solution of sulfuric acid, it is clear that hot blast blows to spot colour developing, puts under daylight and inspect; In test sample chromatogram, with reference substance chromatogram relevant position on aobvious same color spot, and negative noiseless.
Preferably, the assay method Shi of the content of described asperosaponin VI with after the ultrasonic processing of methyl alcohol 10min, be take acetonitrile and water as mobile phase Jiang female ground beetle wound more preparation, in liquid chromatograph, measures.
Further, the assay method Shi Qu female ground beetle of the content of described asperosaponin VI wound is the about 0.1-5.0 weight portion of preparation content powder more, accurately weighed, put in 1-20 parts by volume volumetric flask, add the ultrasonic processing of methyl alcohol 5-30 minute, dissolve and be settled to scale, as need testing solution; Separately get asperosaponin VI reference substance appropriate, accurately weighed, add methyl alcohol and make every 1 parts by volume containing the asperosaponin VI reference substance solution of asperosaponin VI 0.1-10.0 weight portion; In prescription ratio and preparation technology, preparation does not contain the negative sample of teasel root, and by the compound method of above-mentioned need testing solution, makes the negative sample solution that does not contain teasel root; According to 2010 editions one appendix VI D test of high performance liquid chromatography < < Chinese Pharmacopoeia > >, take octadecylsilane chemically bonded silica as filling agent; Water-acetonitrile that the volume ratio of take is 10 ~ 50:10 ~ 50 is mobile phase; At 212 ± 2nm detection wavelength, column temperature, be to measure at 25 ~ 40 ℃, number of theoretical plate calculates and should be not less than 3000 by asperosaponin VI peak; Precision is drawn control medicinal material solution and each 0.005 ~ 0.05 parts by volume of need testing solution respectively, and injection liquid chromatography, measures, and every 1 weight portion of preparation in asperosaponin VI, must not be less than 0.0027 weight portion containing teasel root.
The Shang Shu female ground beetle wound more formulation of preparation is powder, capsule, granule, tablet or pill.
The applicant Dui female ground beetle is created the discrimination method of each component in the preparation of healing and is tested as follows:
one, the thin layer of red ginseng is differentiated
(1) instrument
Mortar, electronic scales, tool plug conical flask, transfer pipet, water-bath, ultrasound wave extraction apparatus, funnel, filter paper, evaporating dish, sample applicator, silica G plate, chromatography cylinder, ultraviolet point sample analyser.
(2) control medicinal material
Red ginseng (lot number: 121045-201105), identify institute purchased from Chinese pharmaceutical biological product.
(3) reagent
Methenyl choloride, methyl alcohol, ethyl acetate, water, normal butyl alcohol, ammoniacal liquor, sulfuric acid, ethanol, acetone.
(4) method of inspection:
Extract solvent: normal butyl alcohol, ammoniacal liquor, methenyl choloride, ethyl acetate, acetone.
Extracting method: jolting and ultrasonic.
Developping agent: 10 ℃ of following lower floor's solution, methenyl choloride-ethyl acetate-methanol-waters (volume ratio 3:8:4.5:2) of placing of methenyl choloride-ethyl acetate-methanol-water (volume ratio 3:8:4.5:2).
Select respectively above solvent, extracting method and the developping agent of extracting to test, result is referring to table 1.
Table 1
。
Repetition test in the above conditions, finally determined that the specificity thin-layer identification method of red ginseng is as follows:
Qu female ground beetle wound is preparation content 1.5g more, adds methenyl choloride 40ml, ultrasonic 10min, discard methenyl choloride liquid, the dregs of a decoction volatilize, and add water 0.5ml wetting, add water-saturated n-butanol 10ml, ultrasonic processing 20min, filters, filtrate adds 3 times of ammonia solutions, shake up, place layering, get upper strata liquid evaporate to dryness, residue adds methyl alcohol 3ml and dissolves, as need testing solution.Separately get red ginseng control medicinal material powder 0.5g, be made in the same way of control medicinal material solution.Get the negative sample 1.5g that lacks red ginseng medicinal material, be made in the same way of negative control solution.According to appendix VIB test of thin-layered chromatography < < Chinese Pharmacopoeia > > version in 2010, draw each 2 μ L of above-mentioned three kinds of solution, put respectively on same silica gel g thin-layer plate, the 10 ℃ of following lower floor's solution placed of methenyl choloride-ethyl acetate-methanol-water (volume ratio 3:8:4.5:2) of take are developping agent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, it is clear that hot blast blows to spot colour developing, puts under uviol lamp (365nm) and inspect.In test sample chromatogram, with control medicinal material chromatogram relevant position on aobvious same color spot, and negative noiseless.
two, the thin layer of teasel root is differentiated
(1) instrument
Mortar, electronic scales, graduated cylinder, flat bottom flask, condenser pipe, rubber tube, water-bath, ultrasound wave extraction apparatus, funnel, filter paper, evaporating dish, separating funnel, absorbent cotton, sample applicator, silica G plate, chromatography cylinder, ultraviolet point sample analyser, hydro-extractor.
(2) control medicinal material
Teasel root control medicinal material (lot number: 121033-201110), identify institute purchased from Chinese pharmaceutical biological product.
(3) reagent
Normal butyl alcohol, water, ethyl acetate, glacial acetic acid, methyl alcohol, distilled water, acetone, sulfuric acid.
(4) method of inspection:
Extract solvent: methyl alcohol, ethyl acetate, ethanol, acetone are for extracting solvent.
Extracting method: ultrasonic and add hot reflux.
Developping agent: the upper solution of normal butyl alcohol-glacial acetic acid-water (volume ratio 4:1:5), normal butyl alcohol-glacial acetic acid-water (volume ratio 4:1:5).
Select respectively above solvent, extracting method and the developping agent of extracting to test, result is referring to table 2.
Table 2
。
Repetition test in the above conditions, finally determined that the specificity thin-layer identification method of teasel root is as follows:
Qu female ground beetle wound is preparation content 1g more, adds methyl alcohol 15ml, ultrasonic processing 30min, and centrifugal (4000r/min, 5min) gets supernatant, evaporate to dryness, residue adds methyl alcohol 2ml to be made to dissolve, as need testing solution.Separately get teasel root control medicinal material powder 0.2g, be made in the same way of control medicinal material solution.Get the negative sample 1g that lacks teasel root medicinal material, be made in the same way of negative control solution.According to appendix VIB test of thin-layered chromatography < < Chinese Pharmacopoeia > > version in 2010, draw each 2 μ L of above-mentioned three kinds of solution, put on same silica gel g thin-layer plate respectively, the upper solution of normal butyl alcohol-glacial acetic acid-water (volume ratio 4:1:5) of take is developping agent, launches, exhibition is apart from about 6cm, take out, dry, spray is with 10% ethanol solution of sulfuric acid, it is clear that hot blast blows to spot colour developing, puts under daylight and inspect.In test sample chromatogram, with control medicinal material chromatogram relevant position on aobvious same color spot, and negative noiseless.
three, the thin layer of the rhizome of davallia is differentiated
(1) instrument
Electronic balance, graduated cylinder, flat bottom flask, water-bath, ultrasound wave extraction apparatus, evaporating dish, sample applicator, silica G plate, baking oven, chromatography cylinder, spray bottle, ultraviolet point sample analyser.
(2) reference substance
Aurantiin reference substance (lot number: 110722-201111), all purchased from Nat'l Pharmaceutical & Biological Products Control Institute.
(3) reagent
Methenyl choloride, acetone, formic acid, methyl alcohol, ethanol, sulfuric acid.
(4) method of inspection:
Extract solvent: methyl alcohol, methenyl choloride, acetone, ethanol are for extracting solvent.
Extracting method: ultrasonic and cold soaking extracts.
Developping agent: the upper solution of toluene-ethyl acetate-formic acid-water (volume ratio 1:12:2.5:3), toluene-ethyl acetate-formic acid-water (volume ratio 1:12:2.5:3).
Select respectively above solvent, extracting method and the developping agent of extracting to test, result is referring to table 3.
Table 3
。
Repetition test in the above conditions, finally determined that the specificity thin-layer identification method of the rhizome of davallia is as follows:
Qu female ground beetle wound is preparation content 2g more, adds methyl alcohol 30ml, ultrasonic 10min, and centrifugal (4000r/min, 5min) gets supernatant, evaporate to dryness, residue adds methyl alcohol 2ml to be made to dissolve, as need testing solution.Separately get aurantiin reference substance appropriate, add methyl alcohol and make every 1ml containing the solution of 0.5mg, in contrast product solution.Get the negative sample 2g that lacks rhizome of davallia medicinal material, be made in the same way of negative control solution.According to appendix VIB test of thin-layered chromatography < < Chinese Pharmacopoeia > > version in 2010, draw each 3 μ L of above-mentioned three kinds of solution, put respectively on same silica gel g thin-layer plate, the upper solution of toluene-ethyl acetate-formic acid-water (volume ratio 1:12:2.5:3) of take is developping agent, launches, and takes out, dry, spray is with 10% vanillic aldehyde ethanol solution of sulfuric acid, and it is clear that hot blast blows to spot colour developing, puts under daylight and inspect.In test sample chromatogram, with reference substance chromatogram relevant position on aobvious same color spot, and negative noiseless.
four, the contained asperosaponin VI assay of preparation
(1) instrument
Electronic balance, conical flask, volumetric flask, 10ml transfer pipet, 50ml transfer pipet, syringe, beaker, funnel, filter paper, filter, high performance liquid chromatograph.
(2) reference substance
Asperosaponin VI (lot number: 111685-201304), identify institute purchased from Chinese pharmaceutical biological product.
(3) reagent
Methyl alcohol, acetonitrile, water, ethanol.
(4) method of inspection:
Extract solvent: the methyl alcohol of methyl alcohol, ethanol, percent by volume 50%
Extraction time: ultrasonic 5 minutes, 10 minutes, 20 minutes.
Mobile phase: methanol-water (volume ratio 28:72), water-acetonitrile (volume ratio 70:30)
Select respectively above solvent, extraction time and the mobile phase of extracting to test, result is referring to table 4.
Table 4
。
Repetition test in the above conditions, finally determined that the method for the contained asperosaponin VI assay of preparation is as follows:
Qu female ground beetle wound is the about 1g of preparation content powder more, accurately weighed, puts in 10mL volumetric flask, adds the ultrasonic processing of methyl alcohol 10min, dissolves and is settled to scale, as need testing solution; Get asperosaponin VI reference substance appropriate, accurately weighed, add methyl alcohol and make every 1mL containing the solution of asperosaponin VI 2.08mg product solution in contrast; The formula preparation of pressing said preparation does not contain the negative sample of teasel root, and makes teasel root negative sample solution by the compound method of above-mentioned need testing solution; According to high performance liquid chromatography (2010 editions one appendix VI D of < < Chinese Pharmacopoeia > >), test, precision is drawn reference substance solution and each 20 μ l of need testing solution respectively, injection liquid chromatography, measure, as shown in Figure 4, the every 1g of preparation in asperosaponin VI, must not be less than 2.70mg containing teasel root.
wu, female ground beetle wound is the contained asperosaponin VI assay-methodological study of preparation more
(1) instrument and reagent
Waters high performance liquid chromatograph: photodiode array detector, Empower chromatographic work station.
KH-500DE ultrasonic cleaner (Kunshan He Chuan ultrasonic instrument company limited).
Acetonitrile (chromatographically pure); Methyl alcohol (analyzing pure).
Asperosaponin VI reference substance (lot number: 111685-201304), identify institute purchased from Chinese pharmaceutical biological product.
Female ground beetle wound more sample (lot number: 20130311,20130319,20130324), preparation in Lanzhou lung section Hospital.
(2) chromatographic condition
Octadecyl silane is filling agent; Chromatographic column: Waters symmetry C
18post (150mm * 4.6mm, 5 μ m);
Mobile phase: acetonitrile-water (volume ratio 30:70);
Flow velocity: 1.0ml/min;
Detect wavelength: 212nm;
Column temperature: 30 ℃.
(3) chromatographic system employment and suitability test (E & ST)
Under above chromatographic condition, asperosaponin VI and all the other impurity peaks separating effects are better, and theoretical cam curve is calculated and should be not less than 3000 by asperosaponin VI.
(4) preparation of reference substance solution and need testing solution
The preparation of I, reference substance solution: it is appropriate that precision takes asperosaponin VI reference substance, accurately weighed, adds methyl alcohol and makes every 1mL containing the solution of asperosaponin VI 2.08mg, shakes up, and obtains.
The preparation: Qu female ground beetle wound of II, need testing solution is the about 1g of preparation content powder more, accurately weighed, puts in 10mL volumetric flask, adds the ultrasonic processing of methyl alcohol 10min, dissolves and is settled to scale, obtains.
The preparation of III, negative sample solution: press the formula preparation of said preparation not containing the negative sample of teasel root, be made in the same way of negative sample solution, standby.
(6) negative interference test: accurate absorption in each 20 μ l injection liquid chromatographies of need testing solution, negative sample solution and reference substance solution, from chromatogram, with the corresponding position of reference substance solution chromatographic peak retention time on, the chromatographic peak that need testing solution has identical retention time occurs, and negative sample solution at this wavelength without absorption, noiseless to the assay of asperosaponin VI in this product.See A in Fig. 4, B, C.
(7) investigation of linear relationship
It is appropriate that precision takes asperosaponin VI reference substance, accurately weighed, adds methyl alcohol and make every 1mL containing the solution of asperosaponin VI 2.08mg, shakes up, as storing solution.Accurate above-mentioned stock solution 0.1,0.2,0.5,0.8, the 1mL of drawing, is settled in 1mL volumetric flask respectively, and accurate each the 20 μ L injection liquid chromatographies of above-mentioned serial solution of drawing, record peak area.Take peak area Y as ordinate, concentration X(μ g) be horizontal ordinate, drawing standard curve, obtains its equation of linear regression Y=4 * 106 x-343365(r=0.9999).Result shows that asperosaponin VI is good linear relationship in the scope of 4.16-41.6 μ g.
(8) precision test
It is appropriate that precision takes asperosaponin VI reference substance, accurately weighed, adds methyl alcohol and make every 1mL containing the solution of asperosaponin VI 2.08mg, shakes up, in contrast product solution.The accurate reference substance solution 10 μ L that draw, under above-mentioned chromatographic condition, continuous sample introduction is 6 times, measures peak area, obtains asperosaponin VI peak area RSD(n=6) be 0.2%, show that the method precision is good.
(9) stability test
Accurate absorption with a need testing solution 20 μ L, measures peak area integrated value respectively at 0,1,2,4,6,8h sample introduction, and the RSD of result asperosaponin VI peak area is 1.6%, shows that need testing solution is good at 8h internal stability.
(10) replica test
Get with 6 parts, a collection of test sample (20130311) powder, accurately weighed, preparation method's preparation of pressing respectively test sample, under above-mentioned chromatographic condition, measure, recording its average content is 3.98mg/g, RSD(n=6) be 0.8%, show that the method repeatability is good.
(11) recovery test
Recovery test refers to that average recovery is to add medicine in concentration known sample, comes and typical curve ratio, and typical curve is also in matrix, to add medicine.
Precision takes the more about 0.5g of formulation samples powder of known content female ground beetle wound, and totally 9 parts, accurately weighed, precision adds a certain amount of reference substance respectively, by the method preparation of test sample, under above-mentioned chromatographic condition, measures, and calculates the recovery of asperosaponin VI.The average recovery rate of result asperosaponin VI is that 97.40%, RSD value is 1.29%, and result shows that the method accurately and reliably.In Table 5.
Table 5 asperosaponin VI average recovery measurement result
。
(12) mensuration of sample
Get respectively three crowdes of about 1g of test sample content powder, accurately weighed, by the method for test sample, prepare need testing solution, under above-mentioned chromatographic condition, measure, and calculate the content of asperosaponin VI.Result is referring to table 6.
Asperosaponin VI assay result in table 6 sample
。
Known according to the measurement result of above-mentioned multiple batches of sample, the every 1g of this product contains the amount of asperosaponin VI between 3.9713～4.0025mg, press minimum content 70% as content lower limit, i.e. 3.9713mg/g * 70%=2.7799mg/g ≈ 2.78mg/g, tentative this product containing teasel root with asperosaponin VI (C
17h
76o
18) amount meter, every 1g must not be less than 2.70mg.
This invention She, Ji female ground beetle is created the more detection method of preparation, comprises one or more of following discriminating and/or detection method: by thin-layer chromatography, the red ginseng in preparation, teasel root, the rhizome of davallia are carried out to qualitative discriminating; By high performance liquid chromatography, the content of contained asperosaponin VI in preparation is measured.By above-mentioned detection method, each component that Ke Yi Dui female ground beetle wound heals in preparation effectively detects, thereby better controls the quality of product.
Accompanying drawing explanation
Fig. 1 is that red ginseng of the present invention is identified thin-layer chromatogram, and wherein, 1 is red ginseng control medicinal material, and 2-4 is red ginseng test sample, and 5 is not contain the negative sample of red ginseng;
Fig. 2 is that teasel root of the present invention is identified thin-layer chromatogram, and wherein, 1 is teasel root control medicinal material, and 2-4 is teasel root test sample, and 5 is not contain the negative sample of teasel root;
Fig. 3 is that the rhizome of davallia of the present invention is identified thin-layer chromatogram, and wherein, 1 is aurantiin reference substance, and 2-4 is rhizome of davallia test sample, and 5 is not contain the negative sample of the rhizome of davallia;
Fig. 4 is the contained asperosaponin VI chromatogram of preparation of the present invention, and wherein, A-C is respectively asperosaponin VI reference substance, teasel root test sample, does not contain the negative sample of teasel root.
Embodiment
Following embodiment is convenient to understand better the present invention, but does not limit the present invention.Experimental technique in following embodiment, if no special instructions, is conventional method.Test material used in following embodiment, if no special instructions, is and purchases available from routine biochemistry reagent shop.Quantitative test in following examples, all arranges and repeats experiment, results averaged for three times." h " in this patent all represents hour." min " in this patent all represents minute.
the looser discriminating of embodiment 1 female ground beetle wound
The preparation that female ground beetle wound is looser: ground bettle 15g, teasel root 90g, rhizome of davallia 30g, calcined Dragon's bone 15g, calcined oyster shell 15g, red ginseng 20g, Morinda officinalis 20g, Radix Angelicae Sinensis 30g.
Above eight tastes, red ginseng, calcined Dragon's bone, calcined oyster shell three tastes are ground into fine powder; All the other five tastes water extractions such as ground bettle, teasel root, extract concentrate drying becomes extract powder, above red ginseng, calcined Dragon's bone, the fine powder of calcined oyster shell three tastes and the dry extract of all the other five tastes such as ground bettle, teasel root is mixed, by pharmacy conventional method, add appropriate amount of auxiliary materials to mix, obtain.
A. female ground beetle is created the thin layer discriminating of red ginseng in the powder preparation of healing
Take the looser content 1.5g of Gong Shi Pin female ground beetle wound, add methenyl choloride 40ml, ultrasonic 10min, discard methenyl choloride liquid, the dregs of a decoction volatilize, and add water 0.5ml wetting, add water-saturated n-butanol 10ml, ultrasonic processing 20min, filters, filtrate adds 3 times of volume ammonia solutions, shake up, place layering, get upper strata liquid evaporate to dryness, residue adds methyl alcohol 3ml and dissolves, as need testing solution.Separately get red ginseng control medicinal material powder 0.5g, be made in the same way of control medicinal material solution.Get the negative sample 1.5g that lacks red ginseng medicinal material, be made in the same way of negative control solution.According to appendix VIB test of thin-layered chromatography < < Chinese Pharmacopoeia > > version in 2010, draw each 2 μ L of above-mentioned three kinds of solution, put respectively on same silica gel g thin-layer plate, take methenyl choloride-ethyl acetate-methanol-water (volume ratio: 3:8:4.5:2) 10 ℃ of following lower floor's solution of placing are developping agent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, it is clear that hot blast blows to spot colour developing, puts under uviol lamp (365nm) and inspect.In test sample chromatogram, with control medicinal material chromatogram relevant position on aobvious same color spot, and negative noiseless.
B. female ground beetle is created the thin layer discriminating of teasel root in the powder preparation of healing
Take the looser content 1g of Gong Shi Pin female ground beetle wound, add methyl alcohol 15ml, ultrasonic processing 30min, centrifugal (4000r/min, 5min) gets supernatant, evaporate to dryness, residue adds methyl alcohol 2ml to be made to dissolve, as need testing solution.Separately get teasel root control medicinal material powder 0.2g, be made in the same way of control medicinal material solution.Get the negative sample 1g that lacks teasel root medicinal material, be made in the same way of negative control solution.According to thin-layered chromatography (appendix VI B), test, draw each 2 μ L of above-mentioned three kinds of solution, put on same silica gel g thin-layer plate respectively, the upper solution of normal butyl alcohol-glacial acetic acid-water (volume ratio 4:1:5) of take is developping agent, launches, exhibition is apart from about 6cm, take out, dry, spray is with 10% ethanol solution of sulfuric acid, it is clear that hot blast blows to spot colour developing, puts under daylight and inspect.In test sample chromatogram, with control medicinal material chromatogram relevant position on aobvious same color spot, and negative noiseless.
C. female ground beetle is created the thin layer discriminating of the rhizome of davallia in the powder preparation of healing
Take the looser content 2g of Gong Shi Pin female ground beetle wound, add methyl alcohol 30ml, ultrasonic 10min, centrifugal (4000r/min, 5min) gets supernatant, evaporate to dryness, residue adds methyl alcohol 2ml to be made to dissolve, as need testing solution.Separately get aurantiin reference substance appropriate, add methyl alcohol and make every 1ml containing the solution of 0.5mg aurantiin, in contrast product solution.Get the negative sample 2g that lacks rhizome of davallia medicinal material, be made in the same way of negative control solution.According to appendix VIB test of thin-layered chromatography < < Chinese Pharmacopoeia > > version in 2010, draw each 3 μ L of above-mentioned three kinds of solution, put respectively on same silica gel g thin-layer plate, the upper solution of toluene-ethyl acetate-formic acid-water (volume ratio 1:12:2.5:3) of take is developping agent, launches, and takes out, dry, spray is with 10% vanillic aldehyde ethanol solution of sulfuric acid, and it is clear that hot blast blows to spot colour developing, puts under daylight and inspect.In test sample chromatogram, with reference substance chromatogram relevant position on aobvious same color spot, and negative noiseless.
D. female ground beetle is created the more contained asperosaponin VI assay of powder preparation
According to high performance liquid chromatography (2010 editions one appendix VI D of < < Chinese Pharmacopoeia > >)
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; The acetonitrile-water (volume ratio 30:70) of take is mobile phase; Detection wavelength is 212nm, 30 ℃ of column temperatures.Number of theoretical plate calculates and should be not less than 3000 by asperosaponin VI peak.
The preparation of reference substance solution: it is appropriate that precision takes asperosaponin VI reference substance, accurately weighed, adds methyl alcohol and makes every 1mL containing the solution of asperosaponin VI 2.08mg, obtains.
The looser about 1g of content powder of preparation: Qu female ground beetle wound of need testing solution, accurately weighed, put in 10mL volumetric flask, add the ultrasonic processing of methyl alcohol 10min, dissolve and be settled to scale, obtain.
Determination method: precision is drawn reference substance solution and each 20 μ l of need testing solution respectively, and injection liquid chromatography, measures, and obtains.
The looser every 1g of female ground beetle wound contains teasel root with asperosaponin VI (C
17h
76o
18) meter, every 1g must not be less than 2.70mg.
embodiment 2
The difference of the present embodiment and embodiment 1 is: the content in the present embodiment Shi Dui female ground beetle wound capsule for curing is tested, and all the other checking procedures are all identical with embodiment 1 with test stone.
The preparation method of female ground beetle wound capsule for curing is: ground bettle 15g, teasel root 90g, rhizome of davallia 30g, calcined Dragon's bone 15g, calcined oyster shell 15g, red ginseng 20g, Morinda officinalis 20g, Radix Angelicae Sinensis 30g.Above eight tastes, red ginseng, calcined Dragon's bone, calcined oyster shell three tastes are ground into fine powder; All the other five tastes water extractions such as ground bettle, teasel root, extract concentrate drying becomes extract powder, above red ginseng, calcined Dragon's bone, the fine powder of calcined oyster shell three tastes and the dry extract of all the other five tastes such as ground bettle, teasel root is mixed, by pharmacy conventional method, add appropriate amount of auxiliary materials to mix, obtain.
embodiment 3
The difference of the present embodiment and embodiment 1 is: the present embodiment Shi Dui female ground beetle wound more particle is tested, and all the other checking procedures are all identical with embodiment 1 with test stone.
The female ground beetle wound more preparation method of particle is: ground bettle 15g, teasel root 90g, rhizome of davallia 30g, calcined Dragon's bone 15g, calcined oyster shell 15g, red ginseng 20g, Morinda officinalis 20g, Radix Angelicae Sinensis 30g.Above eight tastes, red ginseng, calcined Dragon's bone, calcined oyster shell three tastes are ground into fine powder; All the other five tastes water extractions such as ground bettle, teasel root, extract concentrate drying becomes extract powder, above red ginseng, calcined Dragon's bone, the fine powder of calcined oyster shell three tastes and the dry extract of all the other five tastes such as ground bettle, teasel root is mixed, by pharmacy conventional method, add appropriate amount of auxiliary materials to mix softwood processed, granulate and get final product.
embodiment 4
The difference of the present embodiment and embodiment 1 is: the present embodiment Shi Dui female ground beetle wound more sheet is tested, and all the other checking procedures are all identical with embodiment 1 with test stone.
The female ground beetle wound more preparation method of sheet is: ground bettle 15g, teasel root 90g, rhizome of davallia 30g, calcined Dragon's bone 15g, calcined oyster shell 15g, red ginseng 20g, Morinda officinalis 20g, Radix Angelicae Sinensis 30g.Above eight tastes, red ginseng, calcined Dragon's bone, calcined oyster shell three tastes are ground into fine powder; All the other five tastes water extractions such as ground bettle, teasel root, extract concentrate drying becomes extract powder, above red ginseng, calcined Dragon's bone, the fine powder of calcined oyster shell three tastes and the dry extract of all the other five tastes such as ground bettle, teasel root is mixed, by pharmacy conventional method, add appropriate amount of auxiliary materials to mix, compressing tablet and get final product.
embodiment 5
The difference of the present embodiment and embodiment 1 is: the present embodiment Shi Dui female ground beetle wound more pill is tested, and all the other checking procedures are all identical with embodiment 1 with test stone.
The female ground beetle wound more preparation method of pill is: ground bettle 15g, teasel root 90g, rhizome of davallia 30g, calcined Dragon's bone 15g, calcined oyster shell 15g, red ginseng 20g, Morinda officinalis 20g, Radix Angelicae Sinensis 30g.Above eight tastes, red ginseng, calcined Dragon's bone, calcined oyster shell three tastes are ground into fine powder; All the other five tastes water extractions such as ground bettle, teasel root, extract concentrate drying becomes extract powder, above red ginseng, calcined Dragon's bone, the fine powder of calcined oyster shell three tastes and the dry extract of all the other five tastes such as ground bettle, teasel root is mixed, by pharmacy conventional method, add appropriate amount of auxiliary materials to mix, pill and get final product.
the looser discriminating of embodiment 6 female ground beetle wound
The preparation method that the present embodiment Zhong female ground beetle wound is looser is identical with embodiment 1.
A. female ground beetle is created the thin layer discriminating of red ginseng in the powder preparation of healing
Take the looser content 7.5g of Gong Shi Pin female ground beetle wound, add methenyl choloride 60ml, ultrasonic 10min, discard methenyl choloride liquid, the dregs of a decoction volatilize, and add water 1.0ml wetting, add water-saturated n-butanol 20ml, ultrasonic processing 30min, filters, filtrate adds 5 times of volume ammonia solutions, shake up, place layering, get upper strata liquid evaporate to dryness, residue adds methyl alcohol 5ml and dissolves, as need testing solution.Separately get red ginseng control medicinal material powder 1.0g, be made in the same way of control medicinal material solution.Get the negative sample 1.5g that lacks red ginseng medicinal material, be made in the same way of negative control solution.According to appendix VIB test of thin-layered chromatography < < Chinese Pharmacopoeia > > version in 2010, draw each 20 μ L of above-mentioned three kinds of solution, put respectively on same silica gel g thin-layer plate, the 10 ℃ of following lower floor's solution placed of methenyl choloride-ethyl acetate-methanol-water (volume ratio 10:15:10:5) of take are developping agent, launch, take out, dry, spray is with 20% ethanol solution of sulfuric acid, it is clear that hot blast blows to spot colour developing, puts under uviol lamp (365nm) and inspect.In test sample chromatogram, with control medicinal material chromatogram relevant position on aobvious same color spot, and negative noiseless.
B. female ground beetle is created the thin layer discriminating of teasel root in the powder preparation of healing
Take the looser content 5g of Gong Shi Pin female ground beetle wound, add methyl alcohol 20ml, ultrasonic processing 50min, centrifugal (4000r/min, 5min) gets supernatant, evaporate to dryness, residue adds methyl alcohol 5ml to be made to dissolve, as need testing solution.Separately get teasel root control medicinal material powder 1.0g, be made in the same way of control medicinal material solution.Get the negative sample 1g that lacks teasel root medicinal material, be made in the same way of negative control solution.According to thin-layered chromatography (appendix VI B), test, draw each 20 μ L of above-mentioned three kinds of solution, put on same silica gel g thin-layer plate respectively, the upper solution of normal butyl alcohol-glacial acetic acid-water (volume ratio 10:5:10) of take is developping agent, launches, exhibition is apart from about 10cm, take out, dry, spray is with 20% ethanol solution of sulfuric acid, it is clear that hot blast blows to spot colour developing, puts under daylight and inspect.In test sample chromatogram, with control medicinal material chromatogram relevant position on aobvious same color spot, and negative noiseless.
C. female ground beetle is created the thin layer discriminating of the rhizome of davallia in the powder preparation of healing
Take the looser content 5g of Gong Shi Pin female ground beetle wound, add methyl alcohol 50ml, ultrasonic 30min, centrifugal (4000r/min, 5min) gets supernatant, evaporate to dryness, residue adds methyl alcohol 5ml to be made to dissolve, as need testing solution.Separately get aurantiin reference substance appropriate, add methyl alcohol and make every 1ml containing the solution of 0.05mg aurantiin, in contrast product solution.Get the negative sample 2g that lacks rhizome of davallia medicinal material, be made in the same way of negative control solution.According to appendix VIB test of thin-layered chromatography < < Chinese Pharmacopoeia > > version in 2010, draw each 3 μ L of above-mentioned three kinds of solution, put respectively on same silica gel g thin-layer plate, the upper solution of toluene-ethyl acetate-formic acid-water (volume ratio 5:25:10:5) of take is developping agent, launches, and takes out, dry, spray is with 20% vanillic aldehyde ethanol solution of sulfuric acid, and it is clear that hot blast blows to spot colour developing, puts under daylight and inspect.In test sample chromatogram, with reference substance chromatogram relevant position on aobvious same color spot, and negative noiseless.
D. female ground beetle is created the more contained asperosaponin VI assay of powder preparation
According to high performance liquid chromatography (2010 editions one appendix VI D of < < Chinese Pharmacopoeia > >)
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; The acetonitrile-water (volume ratio 50:50) of take is mobile phase; Detection wavelength is 210nm, 40 ℃ of column temperatures.Number of theoretical plate calculates and should be not less than 3000 by asperosaponin VI peak.
The preparation of reference substance solution: it is appropriate that precision takes asperosaponin VI reference substance, accurately weighed, adds methyl alcohol and makes every 1mL containing the solution of asperosaponin VI 0.1mg, obtains.
The looser about 5g of content powder of preparation: Qu female ground beetle wound of need testing solution, accurately weighed, put in 20mL volumetric flask, add the ultrasonic processing of methyl alcohol 30min, dissolve and be settled to scale, obtain.
Determination method: precision is drawn reference substance solution and each 50 μ l of need testing solution respectively, and injection liquid chromatography, measures, and obtains.
The looser every 1g of female ground beetle wound contains teasel root with asperosaponin VI (C
17h
76o
18) meter, every 1g must not be less than 2.70mg.
the looser discriminating of embodiment 7 female ground beetle wound
The preparation method that the present embodiment Zhong female ground beetle wound is looser is identical with embodiment 1.
A. female ground beetle is created the thin layer discriminating of red ginseng in the powder preparation of healing
Take the looser content 1.5g of Gong Shi Pin female ground beetle wound, add methenyl choloride 10ml, ultrasonic 10min, discard methenyl choloride liquid, the dregs of a decoction volatilize, and add water 0.1ml wetting, add water-saturated n-butanol 5ml, ultrasonic processing 10min, filters, filtrate adds 1 times of volume ammonia solution, shake up, place layering, get upper strata liquid evaporate to dryness, residue adds methyl alcohol 1ml and dissolves, as need testing solution.Separately get red ginseng control medicinal material powder 0.1g, be made in the same way of control medicinal material solution.Get the negative sample 1.5g that lacks red ginseng medicinal material, be made in the same way of negative control solution.According to appendix VIB test of thin-layered chromatography < < Chinese Pharmacopoeia > > version in 2010, draw each 1 μ L of above-mentioned three kinds of solution, put respectively on same silica gel g thin-layer plate, the 10 ℃ of following lower floor's solution placed of methenyl choloride-ethyl acetate-methanol-water (volume ratio 1:5:2.5:1.0) of take are developping agent, launch, take out, dry, spray is with 1% ethanol solution of sulfuric acid, it is clear that hot blast blows to spot colour developing, puts under uviol lamp (365nm) and inspect.In test sample chromatogram, with control medicinal material chromatogram relevant position on aobvious same color spot, and negative noiseless.
B. female ground beetle is created the thin layer discriminating of teasel root in the powder preparation of healing
Take the looser content 0.1g of Gong Shi Pin female ground beetle wound, add methyl alcohol 5ml, ultrasonic processing 10min, centrifugal (4000r/min, 5min) gets supernatant, evaporate to dryness, residue adds methyl alcohol 0.5ml to be made to dissolve, as need testing solution.Separately get teasel root control medicinal material powder 0.05g, be made in the same way of control medicinal material solution.Get the negative sample 1g that lacks teasel root medicinal material, be made in the same way of negative control solution.According to thin-layered chromatography (appendix VI B), test, draw each 1 μ L of above-mentioned three kinds of solution, put on same silica gel g thin-layer plate respectively, the upper solution of normal butyl alcohol-glacial acetic acid-water (volume ratio 1:0.5:2.0) of take is developping agent, launches, exhibition is apart from about 1cm, take out, dry, spray is with 1% ethanol solution of sulfuric acid, it is clear that hot blast blows to spot colour developing, puts under daylight and inspect.In test sample chromatogram, with control medicinal material chromatogram relevant position on aobvious same color spot, and negative noiseless.
C. female ground beetle is created the thin layer discriminating of the rhizome of davallia in the powder preparation of healing
Take the looser content 0.1g of Gong Shi Pin female ground beetle wound, add methyl alcohol 10ml, ultrasonic 5min, centrifugal (4000r/min, 5min) gets supernatant, evaporate to dryness, residue adds methyl alcohol 0.5ml to be made to dissolve, as need testing solution.Separately get aurantiin reference substance appropriate, add methyl alcohol and make every 1ml containing the solution of 1.00mg aurantiin, in contrast product solution.Get the negative sample 2g that lacks rhizome of davallia medicinal material, be made in the same way of negative control solution.According to appendix VIB test of thin-layered chromatography < < Chinese Pharmacopoeia > > version in 2010, draw each 1 μ L of above-mentioned three kinds of solution, put respectively on same silica gel g thin-layer plate, the upper solution of toluene-ethyl acetate-formic acid-water (volume ratio 0.1:5:1.0:1.0) of take is developping agent, launches, and takes out, dry, spray is with 1% vanillic aldehyde ethanol solution of sulfuric acid, and it is clear that hot blast blows to spot colour developing, puts under daylight and inspect.In test sample chromatogram, with reference substance chromatogram relevant position on aobvious same color spot, and negative noiseless.
D. female ground beetle is created the more contained asperosaponin VI assay of powder preparation
According to high performance liquid chromatography (2010 editions one appendix VI D of < < Chinese Pharmacopoeia > >)
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; The acetonitrile-water (30:70) of take is mobile phase; Detection wavelength is 214nm, 25 ℃ of column temperatures.Number of theoretical plate calculates and should be not less than 3000 by asperosaponin VI peak.
The preparation of reference substance solution: it is appropriate that precision takes asperosaponin VI reference substance, accurately weighed, adds methyl alcohol and makes every 1mL containing the solution of asperosaponin VI 10.0mg, obtains.
The looser about 5g of content powder of preparation: Qu female ground beetle wound of need testing solution, accurately weighed, put in 20mL volumetric flask, add the ultrasonic processing of methyl alcohol 30min, dissolve and be settled to scale, obtain.
Determination method: precision is drawn reference substance solution and each 5 μ l of need testing solution respectively, and injection liquid chromatography, measures, and obtains.
The looser every 1g of female ground beetle wound contains teasel root with asperosaponin VI (C
17h
76o
18) meter, every 1g must not be less than 2.70mg.
Finally it should be noted that: the foregoing is only the preferred embodiments of the present invention, be not limited to the present invention, although the present invention is had been described in detail with reference to previous embodiment, for a person skilled in the art, its technical scheme that still can record aforementioned each embodiment is modified, or part technical characterictic is wherein equal to replacement.Within the spirit and principles in the present invention all, any modification of doing, be equal to replacement, improvement etc., within all should being included in protection scope of the present invention.
Claims (10)
1. Yi Zhong female ground beetle is created the more detection method of preparation, it is characterized in that: described method comprises one or more in following method: by red ginseng, teasel root, the rhizome of davallia in thin layer chromatography Dui female ground beetle wound more preparation, carry out qualitative discriminating; Content by the asperosaponin VI in the teasel root of high performance liquid chromatography Dui female ground beetle wound more preparation is measured.
2. detection method according to claim 1, it is characterized in that: the qualitative discriminating Shi of described red ginseng is after the female ground beetle wound more ultrasonic extraction of preparation, the methenyl choloride-ethyl acetate-methanol-water of take carries out thin-layer chromatography at 10 ℃ of following lower floor's solution of placing as developping agent; Preferably, described ultrasonic extraction Shi adds female ground beetle wound more preparation after the ultrasonic extraction of methenyl choloride, then adds the ultrasonic extraction of normal butyl alcohol, and filtrate first adds ammoniacal liquor, then adds methyl alcohol to dissolve, and obtains ultrasonic extraction solution; More preferably, the mass ratio of described methenyl choloride, ethyl acetate, methyl alcohol, water is 3:8:4.5:2.
3. detection method according to claim 2, it is characterized in that: the qualitative discriminating of described red ginseng is to get more preparation content 1.5-7.5 weight portion of Suo Shu female ground beetle wound, add methenyl choloride 10-60 parts by volume, ultrasonic 10min, discard methenyl choloride liquid, the dregs of a decoction volatilize, and add water 0.1-1.0 parts by volume wetting, add water-saturated n-butanol 5-20 parts by volume, ultrasonic processing 10-30 minute, filter, filtrate adds 1-5 times of volume ammonia solution, shakes up, place layering, get upper strata liquid evaporate to dryness, residue adds methyl alcohol 1-5 parts by volume and dissolves, as need testing solution, separately get red ginseng control medicinal material powder 0.1-1.0 weight portion, be made in the same way of control medicinal material solution, in prescription ratio and preparation technology, preparation does not contain the negative sample of red ginseng, and by the compound method of above-mentioned need testing solution, makes the negative sample solution that does not contain red ginseng, according to thin-layered chromatography (appendix VIB of < < Chinese Pharmacopoeia > > version in 2010), test, draw above-mentioned each 0.001-0.02 parts by volume of three kinds of solution, put respectively on same silica gel g thin-layer plate, with methenyl choloride-ethyl acetate-methanol-water, volume ratio 1～10:5～15:2.5～10:1.0～5.0, 10 ℃ of following lower floor's solution of placing are developping agent, launch, take out, dry, spray is with 1%～20% ethanol solution of sulfuric acid, it is clear that hot blast blows to spot colour developing, put under uviol lamp (365nm) and inspect, in test sample chromatogram, with control medicinal material chromatogram relevant position on aobvious same color spot, and negative noiseless.
4. detection method according to claim 1, is characterized in that: the qualitative discriminating Shi of described teasel root is Jiang female ground beetle wound more preparation with after the ultrasonic extraction of methyl alcohol, and the upper solution of normal butyl alcohol-glacial acetic acid-water of take is carried out thin-layer chromatography as developping agent; Preferably, the mass ratio of described normal butyl alcohol, glacial acetic acid, water is 4:1:5.
5. detection method according to claim 4, it is characterized in that: the qualitative discriminating of described teasel root is to get more preparation content 0.1-5 weight portion of Suo Shu female ground beetle wound, add methyl alcohol 5-20 parts by volume, ultrasonic processing 10-50 minute, 4000r/min, 5min centrifuging and taking supernatant, evaporate to dryness, residue adds methyl alcohol 0.5-5 parts by volume to be made to dissolve, as need testing solution; Separately get teasel root control medicinal material powder 0.05-1.0 weight portion, be made in the same way of control medicinal material solution; In prescription ratio and preparation technology, preparation does not contain the negative sample of teasel root, and by the compound method of above-mentioned need testing solution, makes the negative sample solution that does not contain teasel root; According to thin-layered chromatography (appendix VIB of < < Chinese Pharmacopoeia > > version in 2010), test, draw above-mentioned each 0.001-0.02 parts by volume of three kinds of solution, put respectively on same silica gel g thin-layer plate, normal butyl alcohol-glacial acetic acid-water, the upper solution of volume ratio 1～10:0.5～5:2.0～10 is developping agent, launch, exhibition is apart from about 1-10cm, take out, dry, spray is with 1%～20% ethanol solution of sulfuric acid, and it is clear that hot blast blows to spot colour developing, puts under daylight and inspect; In test sample chromatogram, with control medicinal material chromatogram relevant position on aobvious same color spot, and negative noiseless.
6. detection method according to claim 1, is characterized in that: the qualitative discriminating Shi of the described rhizome of davallia is Jiang female ground beetle wound more preparation with after the ultrasonic extraction of methyl alcohol, and the upper solution of toluene-ethyl acetate-formic acid-water of take is carried out thin-layer chromatography as developping agent; Preferably, the mass ratio of described toluene, ethyl acetate, formic acid, water is 1:12:2.5:3.
7. detection method according to claim 6, it is characterized in that: the qualitative discriminating of the described rhizome of davallia is to get more preparation content 0.1-5 weight portion of Suo Shu female ground beetle wound, add methyl alcohol 10-50 parts by volume, ultrasonic 5-30 minute, 4000r/min, 5min centrifuging and taking supernatant, evaporate to dryness, residue adds methyl alcohol 0.5-5 parts by volume to be made to dissolve, as need testing solution; Separately get aurantiin reference substance appropriate, add methyl alcohol and make every 1 parts by volume containing the solution of 0.05-1.00 weight portion, in contrast product solution; In prescription ratio and preparation technology, preparation does not contain the negative sample of the rhizome of davallia, and by the compound method of above-mentioned need testing solution, makes the negative sample solution that does not contain the rhizome of davallia; According to thin-layered chromatography (appendix VIB of < < Chinese Pharmacopoeia > > version in 2010), test, draw above-mentioned each 0.001-0.03 parts by volume of three kinds of solution, put respectively on same silica gel g thin-layer plate, with toluene-ethyl acetate-formic acid-water, the upper solution of volume ratio 0.1～5:5～25:1.0～10:1.0～5 is developping agent, launch, take out, dry, spray is with 1%-20% vanillic aldehyde ethanol solution of sulfuric acid, it is clear that hot blast blows to spot colour developing, puts under daylight and inspect; In test sample chromatogram, with reference substance chromatogram relevant position on aobvious same color spot, and negative noiseless.
8. detection method according to claim 1, is characterized in that: the assay method Shi of the content of described asperosaponin VI with after the ultrasonic processing of methyl alcohol 10min, be take acetonitrile and water as mobile phase Jiang female ground beetle wound more preparation, in liquid chromatograph, measures.
9. detection method according to claim 8, it is characterized in that: the assay method Shi Qu female ground beetle wound of the content of described asperosaponin VI is the about 0.1-5.0 weight portion of preparation content powder more, accurately weighed, put in 1-20 parts by volume volumetric flask, add the ultrasonic processing of methyl alcohol 5-30 minute, dissolving is settled to scale, as need testing solution; Separately get asperosaponin VI reference substance appropriate, accurately weighed, add methyl alcohol and make every 1 parts by volume containing the asperosaponin VI reference substance solution of asperosaponin VI 0.1-10.0 weight portion; In prescription ratio and preparation technology, preparation does not contain the negative sample of teasel root, and by the compound method of above-mentioned need testing solution, makes the negative sample solution that does not contain teasel root; According to 2010 editions one appendix VI D test of high performance liquid chromatography < < Chinese Pharmacopoeia > >, take octadecylsilane chemically bonded silica as filling agent; Water-acetonitrile that the volume ratio of take is 10 ~ 50:10 ~ 50 is mobile phase; At 212 ± 2nm detection wavelength, column temperature, be to measure at 25 ~ 40 ℃, number of theoretical plate calculates and should be not less than 3000 by asperosaponin VI peak; Precision is drawn control medicinal material solution and each 0.005 ~ 0.05 parts by volume of need testing solution respectively, and injection liquid chromatography, measures, and every 1 weight portion of preparation in asperosaponin VI, must not be less than 0.0027 weight portion containing teasel root.
10. according to the arbitrary described detection method of claim 1-9, it is characterized in that it is powder, capsule, granule, tablet or pill that: Suo Shu female ground beetle is created the more formulation of preparation.
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