CN100998617A - Quality control method for Xindakang preparation - Google Patents
Quality control method for Xindakang preparation Download PDFInfo
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- CN100998617A CN100998617A CN 200610022691 CN200610022691A CN100998617A CN 100998617 A CN100998617 A CN 100998617A CN 200610022691 CN200610022691 CN 200610022691 CN 200610022691 A CN200610022691 A CN 200610022691A CN 100998617 A CN100998617 A CN 100998617A
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Abstract
A quality control method for Chinese medicine 'Xindakang' includes such steps as testing its characters, examining its appearance, thin-layer chromatography discrimination of quercetin and isorhamnetin, and measuring the contents of quercetin, isorhamnetin and kaempferol.
Description
Technical field
The present invention relates to a kind of method of quality control of Xindakang preparation, belong to the technical field of medicine being carried out quality control.
Background technology
Xindakang preparation is that the effective site Fructus Hippophae total flavones and the adjuvant that extract from plant Fructus Hippophae (Hippophae L.) are prepared from.Fructus Hippophae total flavones is used for the treatment of cardiovascular and cerebrovascular disease and has obtained universally acknowledged, modern pharmacology research and clinical practice show: Fructus Hippophae total flavones has coronary artery dilator, increase the heart and brain tissues amount of blood supply, reduce cardiac load, anticoagulant, microcirculation improvement prevents thrombosis and atherosclerotic generation, suppress lipid peroxidation, effects such as slow down aging." XINDAKANG PIAN " wherein and " Flavonihippophae capsule " are published respectively on the 14 in Chinese ministry standard Chinese traditional patent formulation preparation and new drug are become a full member 33 of standards, and use for many years clinically, obtaining than satisfactory therapeutic effects aspect the treatment cardiovascular and cerebrovascular disease.But find after deliberation, existing Xindakang preparation exists the irrational shortcoming of method of quality control, as the XINDAKANG PIAN on the present market, comprise coated tablet and Film coated tablets, because its method for making, prescription are in fact the same, are the coating difference, (one is WS3-B-2666-97 and carry out two standards, one is WS3-B-2666-2002), wasted standard resource; In version Chinese Pharmacopoeia appendix XVG reference substance in 2005, control medicinal material, reference extract, do not recorded in addition in the existing standard still at the standard substance isorhamnetin glucoside that uses, this brings certain degree of difficulty for quality control of product; Existing Xindakang preparation exists quality control standard simple, the uppity shortcoming of product quality, be embodied in: be the general name of flavone material because of Fructus Hippophae flavone, and its discriminating and content control are all weighed with single isorhamnetin, so can not effectively control the quality of said preparation, thereby influence its clinical efficacy.
Summary of the invention
Purpose of the present invention provides a kind of method of quality control of Xindakang preparation.It is simple to the present invention is directed to original Xindakang preparation quality control standard, the uppity shortcoming of product quality, method of quality control to this preparation is studied, worked out the thin layer chromatography discriminating of Quercetin, isorhamnetin in the preparation, and increased mensuration to the total content of Quercetin, isorhamnetin, kaempferol in the preparation, improved the quality control standard of Xindakang preparation, control method in this quality standard can be controlled the quality of Xindakang preparation effectively, thereby has guaranteed the clinical efficacy of said preparation.
Xindakang preparation of the present invention is to be prepared from by Fructus Hippophae total flavones (Fructus Hippophae flavone) and adjuvant.Its method for making be Fructus Hippophae through extract Fructus Hippophae flavone, get Fructus Hippophae flavone 50g (containing total flavones) in isorhamnetin 5g, be ground into fine powder, add appropriate amount of auxiliary materials and make the different preparations that comprise tablet (comprising buccal tablet, dispersible tablet, chewable tablet), capsule, soft capsule, drop pill according to conventional method.
Method of quality control of the present invention mainly comprise in character, inspection, discriminating and the assay project partly or entirely; Wherein differentiate and comprise that with Quercetin reference substance, isorhamnetin reference substance be the thin layer chromatography discriminating that flavone aglycone in the preparation is differentiated in contrast; Assay comprises the assay to Quercetin, isorhamnetin, kaempferol total amount in the preparation.
The discrimination method of Quercetin, isorhamnetin is to be contrast with Quercetin reference substance and isorhamnetin reference substance respectively, and with toluene: ethyl acetate: formic acid=2～10: 1～8: 0.1～1.5 solution is the thin layer chromatography of developing solvent.
Described discrimination method comprises following:
Get this preparation or its content, porphyrize adds ethanol and makes dissolving, filters, and filtrate is as need testing solution; Other gets Quercetin and isorhamnetin reference substance, adds ethanol and makes mixed solution, in contrast product solution; Test according to the Chinese Pharmacopoeia thin layer chromatography, draw above-mentioned need testing solution, reference substance solution, put respectively on same silica gel g thin-layer plate, with toluene: ethyl acetate: formic acid=2～10: 1～8: 0.1～1.5 is developing solvent, launch, take out, dry, spray is put under the ultra-violet lamp and is inspected with 1～5% aluminum chloride alcoholic solution; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.
Discrimination method comprises following more specifically:
Get this preparation or its content, porphyrize is got fine powder 2～20mg, adds dehydrated alcohol 2～20ml, and supersound process 1～15 minute filters, and filtrate is as need testing solution; Other gets Quercetin and isorhamnetin reference substance, adds dehydrated alcohol and makes the mixed solution that every 1ml contains 1mg～10mg, in contrast product solution; Test according to an appendix VIB of Chinese Pharmacopoeia thin layer chromatography, draw each 1～20ul of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene: ethyl acetate: formic acid=2～10: 1～8: 0.1～1.5 is developing solvent, launch, take out, dry, spray is put under the 365nm ultra-violet lamp and is inspected with 1～5% aluminum chloride alcoholic solution; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.
Quercetin, isorhamnetin, the total quantity measuring method of kaempferol are to be contrast with isorhamnetin reference substance, Quercetin reference substance, kaempferol reference substance in the Xindakang preparation, and with methanol: 0.4% phosphoric acid=30～60: 30～60 is the high performance liquid chromatography of mobile phase.
Described content assaying method comprises following:
Quercetin, isorhamnetin, kaempferol are according to the Chinese Pharmacopoeia high effective liquid chromatography for measuring: chromatographic column is the C18 post, and with methanol: 0.4% phosphoric acid=30～60: 30～60 is mobile phase; The detection wavelength is 370nm; Theoretical cam curve is calculated by the isorhamnetin peak should be not less than 3000; Precision takes by weighing isorhamnetin, kaempferol, Quercetin reference substance, adds dissolve with methanol and makes mixed solution, gets reference substance solution; Get this preparation or its content, the accurate title, decide, and puts in the tool plug conical flask, the accurate methanol that adds, claim decide weight, supersound extraction is put coldly, and weight decided in title again, supply the weight that subtracts mistake with methanol, shake up, filter, precision is measured subsequent filtrate and is put in the tool plug conical flask, add methanol, hydrochloric acid, put reflux in the water-bath, cooling, in the dislocation measuring bottle, be diluted to scale, shake up with methanol, filter, get subsequent filtrate, get need testing solution; Accurate reference substance solution and the need testing solution drawn injects chromatograph of liquid, measures, promptly;
When this preparation contained total flavones (in isorhamnetin) and is 5mg, each dosage form contained Quercetin (C
15H
10O
7), isorhamnetin (C
16H
12O
7), kaempferol (C
15H
10O
6) total amount: every in tablet should be no less than labelled amount 80%, capsule and every every of soft capsule should be no less than labelled amount 80%, the every ball of drop pill should be no less than 80% of labelled amount.
When this preparation contained total flavones (in isorhamnetin) and is 10mg, each dosage form contained Quercetin (C
15H
10O
7), isorhamnetin (C
16H
12O
7), kaempferol (C
15H
10O
6) total amount: every in tablet should be no less than labelled amount 80%, capsule and every every of soft capsule should be no less than labelled amount 80%, the every ball of drop pill should be no less than 80% of labelled amount.
Content assaying method comprises following more specifically:
Quercetin, isorhamnetin, kaempferol shine an appendix VID of Chinese Pharmacopoeia high effective liquid chromatography for measuring: chromatographic column is the C18 post; With methanol: 0.4% phosphoric acid=30～60: 30～60 is mobile phase; The detection wavelength is 370nm; Theoretical cam curve is calculated by the isorhamnetin peak should be not less than 3000; Precision takes by weighing puts P
2O
5Dry 10～48 hours isorhamnetin, kaempferol, 4 hours Quercetin reference substance of 120 ℃ of dryings add methanol and make the mixed solution that every 1ml contains isorhamnetin 20ug, kaempferol 2ug, Quercetin 25ug respectively in the vacuum drying apparatus, promptly get reference substance solution; Get that this preparation or its content 100～200mg are accurate to be claimed surely, porphyrize is got and is equivalent to total flavones 10mg sample, the accurate title, decide, and puts in the tool plug conical flask, the accurate methanol 25ml that adds, claim to decide weight, at power 135W, supersound process is 30 minutes under the frequency 59kHz, puts cold, claim again to decide weight, supply the weight that subtracts mistake, shake up with methanol, filter, discard filtrate just, precision is measured subsequent filtrate 5ml, put in the tool plug conical flask, add methanol 15ml, hydrochloric acid (1 → 2) 5ml, put in 80～100 ℃ of water-baths reflux 30 minutes, cooling in the transposition 50ml measuring bottle, is diluted to scale with methanol rapidly, shake up, filter with the 0.45um microporous filter membrane, get subsequent filtrate, promptly get need testing solution; Accurate respectively reference substance solution and each 10ul of need testing solution of drawing injects chromatograph of liquid, measures, promptly;
When this preparation contained total flavones (in isorhamnetin) and is 5mg, each dosage form contained Quercetin (C
15H
10O
7), isorhamnetin (C
16H
12O
7), kaempferol (C
15H
10O
6) total amount: every in tablet should be no less than labelled amount 80%, capsule and every every of soft capsule should be no less than labelled amount 80%, the every ball of drop pill should be no less than 80% of labelled amount.
When this preparation contained total flavones (in isorhamnetin) and is 10mg, each dosage form contained Quercetin (C
15H
10O
7), isorhamnetin (C
16H
12O
7), kaempferol (C
15H
10O
6) total amount: every in tablet should be no less than labelled amount 80%, capsule and every every of soft capsule should be no less than labelled amount 80%, the every ball of drop pill should be no less than 80% of labelled amount.
Described method of quality control comprises:
Character:
For tablet: content be pale brown color to brown, mildly bitter flavor;
For capsule: content is that fawn is to dun granule and powder; Feeble QI, mildly bitter flavor;
For soft capsule: content is the yellow oily liquid that contains a small amount of suspended solid extractum; Feeble QI, mildly bitter flavor;
For drop pill: this product is brown drop pill; Feeble QI, mildly bitter flavor;
Differentiate:
Get this preparation or its content, porphyrize adds ethanol and makes dissolving, filters, and filtrate is as need testing solution; Other gets Quercetin and isorhamnetin reference substance, adds ethanol and makes mixed solution, in contrast product solution; Test according to the Chinese Pharmacopoeia thin layer chromatography, draw above-mentioned need testing solution, reference substance solution, put respectively on same silica gel g thin-layer plate, with toluene: ethyl acetate: formic acid=2～10: 1～8: 0.1～1.5 is developing solvent, launch, take out, dry, spray is put under the ultra-violet lamp and is inspected with 1～5% aluminum chloride alcoholic solution; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.
Check: this tablet, capsule, soft capsule or drop pill should meet pertinent regulations under Chinese Pharmacopoeia tablet, capsule, soft capsule or the drop pill item respectively.
Assay
(1) total flavones
The preparation of reference substance solution: precision takes by weighing the isorhamnetin reference substance, adds dehydrated alcohol and makes the solution that every 1ml contains isorhamnetin 20ug, promptly;
The preparation of standard curve: precision is measured reference substance solution 1,2,3,4,5,6ml, put respectively in the 10ml measuring bottle, add 1～5% aluminum chloride ethanol solution 1.0ml, be diluted to scale with dehydrated alcohol, shake up, placing 10 minutes, is blank with the reagent corresponding, according to ultraviolet visible spectrophotometry (appendix VA of Chinese Pharmacopoeia), wavelength place at 430nm measures absorbance, with the absorbance is vertical coordinate, and concentration is abscissa, the drawing standard curve;
Algoscopy: this preparation tablet is got 20 and is removed coating, 10～20 of capsule or drop pills, it is fixed to get the accurate title of its content, porphyrize, mixing, get 0.1g, the accurate title, decide, and puts in the tool plug conical flask, it is an amount of to add dehydrated alcohol, and reflux is 15 minutes in 80 ℃～100 ℃ baths, puts cold, in the dislocation 100ml measuring bottle, be diluted to scale, shake up with dehydrated alcohol, filter, discard filtrate just, precision is measured subsequent filtrate 10ml, puts in the 25ml measuring bottle, be diluted to scale with dehydrated alcohol, shake up, precision is measured 3ml, puts in the 10ml measuring bottle, add 1～5% aluminum chloride ethanol solution 1.0ml, be diluted to scale with dehydrated alcohol, shake up, as need testing solution; Other gets the same amount need testing solution that does not add developer and does blank, method under the sighting target directrix curve preparation, from " placing 10 minutes ", according to ultraviolet visible spectrophotometry (appendix VA of Chinese Pharmacopoeia), wavelength place at 430nm measures absorbance, from standard curve, read the weight (ug) of isorhamnetin in the test sample, calculate, promptly;
When this preparation contains total flavones (in isorhamnetin) and is 5mg, contain Fructus Hippophae flavone with isorhamnetin (C
16H
12O
7) meter, every in tablet should be labelled amount 90.0～110.0%, capsule and every of soft capsule should be labelled amount 90.0～110.0%, the every ball of drop pill should be 90.0～110.0% of labelled amount.
When this preparation contains total flavones (in isorhamnetin) and is 10mg, contain Fructus Hippophae flavone with isorhamnetin (C
16H
12O
7) meter, every in tablet should be labelled amount 90.0～110.0%, capsule and every of soft capsule should be labelled amount 90.0～110.0%, the every ball of drop pill should be labelled amount 90.0～110.0%,
(2) Quercetin, isorhamnetin, kaempferol are according to an appendix VID of Chinese Pharmacopoeia high effective liquid chromatography for measuring: chromatographic column is the C18 post, and with methanol: 0.4% phosphoric acid=30～60: 30～60 is mobile phase; The detection wavelength is 370nm; Theoretical cam curve is calculated by the isorhamnetin peak should be not less than 3000; Precision takes by weighing isorhamnetin, kaempferol, Quercetin reference substance, adds dissolve with methanol and makes mixed solution, gets reference substance solution; Get this preparation or its content, the accurate title, decide, and puts in the tool plug conical flask, the accurate methanol that adds, claim decide weight, supersound extraction is put coldly, and weight decided in title again, supply the weight that subtracts mistake with methanol, shake up, filter, precision is measured subsequent filtrate and is put in the tool plug conical flask, add methanol, hydrochloric acid, put reflux in the water-bath, cooling, in the dislocation measuring bottle, be diluted to scale, shake up with methanol, filter, get subsequent filtrate, get need testing solution; Accurate reference substance solution and the need testing solution drawn injects chromatograph of liquid, measures, promptly;
When this preparation contained total flavones (in isorhamnetin) and is 5mg, each dosage form contained Quercetin (C
15H
10O
7), isorhamnetin (C
16H
12O
7), kaempferol (C
15H
10O
6) total amount: every in tablet should be no less than labelled amount 80%, capsule and every every of soft capsule should be no less than labelled amount 80%, the every ball of drop pill should be no less than 80% of labelled amount.
When this preparation contained total flavones (in isorhamnetin) and is 10mg, each dosage form contained Quercetin (C
15H
10O
7), isorhamnetin (C
16H
12O
7), kaempferol (C
15H
10O
6) total amount: every in tablet should be no less than labelled amount 80%, capsule and every every of soft capsule should be no less than labelled amount 80%, the every ball of drop pill should be no less than 80% of labelled amount.
The applicant finds after deliberation, adopts the quality of following method of quality control with this preparation of easier control, more helps guaranteeing the clinical efficacy of preparation, so described method of quality control also can comprise:
Character:
For tablet: content be pale brown color to brown, mildly bitter flavor;
For capsule: content is that fawn is to dun granule and powder; Feeble QI, mildly bitter flavor;
For soft capsule: content is the yellow oily liquid that contains a small amount of suspended solid extractum; Feeble QI, mildly bitter flavor;
For drop pill: this product is brown drop pill; Feeble QI, mildly bitter flavor;
Differentiate:
Get 1 in this preparation or 1, porphyrize adds ethanol 5ml or 10ml, and supersound process 5 minutes is made the solution of 1mg/ml, filters, and filtrate is as need testing solution; Other gets Quercetin and isorhamnetin reference substance, adds ethanol and makes the mixed solution that every 1ml contains 1mg, in contrast product solution; Test according to an appendix VIB of Chinese Pharmacopoeia thin layer chromatography, draw each 1～2ul of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene: ethyl acetate: formic acid=5: 4: 0.5 is developing solvent, launch, take out, dry, spray is put under the 365nm ultra-violet lamp and is inspected with 1～3% aluminum chloride alcoholic solution; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.
Check:
Should meet in appendix of Chinese Pharmacopoeia each pertinent regulations under tablet, capsule, soft capsule or the drop pill item.
Assay
(1) total flavones
The preparation of reference substance solution: precision takes by weighing and adopts P
2O
5Dry isorhamnetin reference substance more than 12 hours adds dehydrated alcohol and makes the solution that every 1ml contains isorhamnetin 20ug, promptly;
The preparation of standard curve: precision is measured reference substance solution 1,2,3,4,5,6ml, put respectively in the 10ml measuring bottle, add 1～3% aluminum chloride ethanol solution 1.0ml, be diluted to scale with dehydrated alcohol, shake up, placing 10 minutes, is blank with the reagent corresponding, according to ultraviolet visible spectrophotometry (appendix VA of Chinese Pharmacopoeia), wavelength place at 430nm measures absorbance, with the absorbance is vertical coordinate, and concentration is abscissa, the drawing standard curve;
Algoscopy: this preparation is got 20 in tablet, get 10～20 seed lac wafers, the drop pill content, the accurate title, decided the porphyrize mixing, get about 0.1g, the accurate title, decide, and puts in the tool plug conical flask, it is an amount of to add dehydrated alcohol, and reflux is 15 minutes in 80 ℃ of water-baths, puts cold, in the dislocation 100ml measuring bottle, be diluted to scale, shake up with dehydrated alcohol, filter, discard filtrate just, precision is measured subsequent filtrate 10ml, puts in the 25ml measuring bottle, be diluted to scale with dehydrated alcohol, shake up, precision is measured 3ml, puts in the 10ml measuring bottle, add 1～% aluminum chloride ethanol solution 1.0ml, be diluted to scale with dehydrated alcohol, shake up, as need testing solution; Other gets the same amount need testing solution that does not add developer and does blank, according to the method under the above-mentioned standard curve preparation, from " placing 10 minutes ", according to ultraviolet visible spectrophotometry (appendix VA of Chinese Pharmacopoeia), wavelength place at 430nm measures absorbance, from standard curve, read the weight (ug) of isorhamnetin in the test sample, calculate, promptly;
When this preparation contains total flavones (in isorhamnetin) and is 5mg, contain Fructus Hippophae flavone with isorhamnetin (C
16H
12O
7) meter, every in tablet should be labelled amount 90.0～110.0%, capsule and every of soft capsule should be labelled amount 90.0～110.0%, the every ball of drop pill should be 90.0～110.0% of labelled amount.
When this preparation contains total flavones (in isorhamnetin) and is 10mg, contain Fructus Hippophae flavone with isorhamnetin (C
16H
12O
7) meter, every in tablet should be labelled amount 90.0～110.0%, capsule and every of soft capsule should be labelled amount 90.0～110.0%, the every ball of drop pill should be labelled amount 90.0～110.0%,
(2) Quercetin, isorhamnetin, kaempferol adopt high effective liquid chromatography for measuring: chromatographic column is the C18 post; With methanol: 0.4% phosphoric acid=50: 50 is mobile phase; The detection wavelength is 370nm; Theoretical cam curve is calculated by the isorhamnetin peak should be not less than 3000; Precision takes by weighing with P
2O
5For desiccant drying under reduced pressure isorhamnetin more than 12 hours, kaempferol, 4 hours Quercetin reference substance of 120 ℃ of dryings an amount of, add methanol and make the mixed solution that every 1ml contains isorhamnetin 20ug, kaempferol 2ug, Quercetin 25ug respectively, promptly get reference substance solution; This preparation is got 20 in tablet and is removed coating, get 10 seed lac wafers, the drop pill content, the accurate title, decided the porphyrize mixing, get and be equivalent to total flavones 10mg, accurately claim surely, put in the tool plug conical flask, the accurate methanol 25ml that adds claims to decide weight, supersound process (power 135W, frequency 59kHz) 30 minutes, put coldly, claim to decide weight again, supply the weight that subtracts mistake with methanol, shake up, filter, discard filtrate just, precision is measured subsequent filtrate 5ml, put in the tool plug conical flask, add methanol 15ml, hydrochloric acid (1 → 2) 5ml, put in the water-bath reflux 30 minutes, cooling is transferred in the 50ml measuring bottle rapidly, is diluted to scale with methanol, shake up, filter with the 0.45um microporous filter membrane, get subsequent filtrate, promptly get need testing solution; Accurate respectively reference substance solution and each 10ul of need testing solution of drawing injects chromatograph of liquid, measures, promptly;
When this preparation contained total flavones (in isorhamnetin) and is 5mg, each dosage form contained Quercetin (C
15H
10O
7), isorhamnetin (C
16H
12O
7), kaempferol (C
15H
10O
6) total amount: every in tablet should be no less than labelled amount 80%, capsule and every every of soft capsule should be no less than labelled amount 80%, the every ball of drop pill should be no less than 80% of labelled amount.
When this preparation contained total flavones (in isorhamnetin) and is 10mg, each dosage form contained Quercetin (C
15H
10O
7), isorhamnetin (C
16H
12O
7), kaempferol (C
15H
10O
6) total amount: every in tablet should be no less than labelled amount 80%, capsule and every every of soft capsule should be no less than labelled amount 80%, the every ball of drop pill should be no less than 80% of labelled amount.
This method of quality control is the improvement that the method for quality control that has the Xindakang preparation product now is carried out, and it is improved one's methods and comprises: will have two kinds of product standards of XINDAKANG PIAN coated tablet and Film coated tablets now and merge; Being combined the product standard character adjusts; The chemical reaction colour developing of a former total flavones of differentiating is differentiated that the specificity that deletion replaces Quercetin in the Fructus Hippophae flavone, isorhamnetin differentiates; The reference substance isorhamnetin adopts P by the pharmacopeia new demand in the assay item
2O
5Seasoning; Increase with high effective liquid chromatography for measuring Quercetin, isorhamnetin, kaempferol total content.
Because the present invention compared with prior art, the total content of Quercetin, isorhamnetin, kaempferol in the high effective liquid chromatography for measuring preparation is differentiated and adopted to the thin layer chromatography that has increased Quercetin, isorhamnetin, the being specific to property of method that is adopted is strong, precision is high, favorable reproducibility, response rate height, improve the quality control standard of former Xindakang preparation, thereby guaranteed the quality and the clinical efficacy of said preparation.
In order to ensure method of quality control science of the present invention, reasonable, feasible, the applicant studies discriminating in the method and content assaying method, and concrete testing data is as follows:
[discriminating] adopts Quercetin and isorhamnetin to compare the discriminating Xindakang preparation through test, and its feminine gender is noiseless, clear spot.The point sample amount adopts 10ul, and speckle is hangover a bit, and adjusting the point sample amount is 1～2ul.
Adopt the silica gel g thin-layer plate of different manufacturers, launch, all can obtain satisfied separating resulting with above-mentioned thin layer chromatography condition.
[inspection] should meet Chinese Pharmacopoeia tablet (appendix ID of Chinese Pharmacopoeia), capsule, soft capsule (appendix IL of Chinese Pharmacopoeia) or drop pill (appendix IK of Chinese Pharmacopoeia) item pertinent regulations down.
[assay] total flavones is only studied the drying mode of isorhamnetin reference substance with the primary standard method, the results are shown in Table 1.
Table 1
| The reference substance title | Isorhamnetin | ||
| The reference substance lot number | 110860-200407 | ||
| 100 ℃ are used for test sample general flavone content (mg/ sheet) after being dried to constant weight | 4.94 | 4.95 | 4.93 |
| Use P 2O 5After dry 12h is above, test sample general flavone content (mg/ sheet) | 4.95 | 4.93 | 4.95 |
(XINDAKANG PIAN sample lot number: 060301, specification: the 5mg/ sheet)
Above result shows: after adopting the dry isorhamnetin reference substance of dual mode, survey the content of total flavone no significant difference, adopt P so the isorhamnetin reference substance changes into
2O
5More than the dry 12h, more to meet the requirement of pharmacopeia to the reference substance drying condition.
The content assaying method of the Quercetin in the employing high effective liquid chromatography for measuring XINDAKANG PIAN that Quercetin, isorhamnetin, kaempferol increase newly, isorhamnetin, kaempferol total amount.Method is carried out assay with reference to the method for the high effective liquid chromatography for measuring isorhamnetin that records under Fructus Hippophae item of Chinese Pharmacopoeia to Quercetin, isorhamnetin, kaempferol in the Xindakang preparation.
1. maximum absorption wavelength is selected
Get Quercetin, isorhamnetin, kaempferol reference substance, make reference substance solution with methanol respectively, in the interscan of 200-600nm wave-length coverage, Quercetin has maximum absorption wavelength at the 371nm place, isorhamnetin has maximum absorption wavelength at the 369.5nm place, and kaempferol has maximum absorption wavelength at the 366.5nm place.With reference to the detection wavelength of isorhamnetin under Fructus Hippophae item of Chinese Pharmacopoeia version in 2005,, Quercetin, isorhamnetin, kaempferide be chosen as 370nm so being detected wavelength.
2. the selection of mobile phase
The assay item current downflow of isorhamnetin is methanol mutually under Fructus Hippophae item of Chinese Pharmacopoeia version in 2005: 0.4% phosphoric acid=58: 42, the assay item current downflow of Quercetin, isorhamnetin, kaempferol is methanol mutually in the Folium Ginkgo: 0.4% phosphoric acid=50: 50, this product is the single medicinal material preparation, so, determine that mobile phase the best is methanol: 0.4% phosphoric acid=50: 50 with reference to Chinese Pharmacopoeia.
3. chromatographic condition and system suitability test
Mobile phase: methanol: 0.4% phosphoric acid=50: 50
Chromatographic column: (5,um2 50 * 4.6mm) for the DiamonsilC18 of Di Ma company
Flow velocity: 1ml/s, 35 ℃ of column temperatures;
Under this condition in the sample Quercetin, isorhamnetin, kaempferol can reach baseline separation with other component, meet the requirements with adjacent chromatographic peak separating degree.Theoretical cam curve is calculated by the isorhamnetin peak should be not less than 3000.
4. the preparation method research of need testing solution
The main component of Xindakang preparation is a Fructus Hippophae total flavones, form by isorhamnetin-3-rutinoside, isorhamnetin-3-rhamnoside, Quercetin-3-glucoside, kaempferol-multiple flavonoid glycoside compounds such as 3-glucoside, described composition is difficult directly to be measured, and its aglycon is mainly Quercetin, isorhamnetin, kaempferol, so carry out assay after adopting acid hydrolysis to make flavonoid glycoside change into flavone aglycone.
Because Quercetin, isorhamnetin, kaempferol all can be dissolved in methanol,, serve as to extract solvent to extract with methanol 25ml with reference to the preparation method of need testing solution in the Flavonihippophae capsule quality standard.Get 20 of this product, remove coating, the accurate title, decide, porphyrize is got an amount of (being equivalent to total flavones 10mg), and accurate the title decides, put in the tool plug conical flask, the accurate methanol 25ml that adds claims to decide weight, supersound process (power 135W, frequency 59kHz) is put cold, claim again to decide weight, supply the weight that subtracts mistake, shake up with methanol, filter, discard filtrate just, precision is measured subsequent filtrate 5ml, put in the tool plug conical flask, add methanol 15ml, hydrochloric acid, put reflux in the water-bath, cooling is transferred in the 50ml measuring bottle rapidly, be diluted to scale with methanol, shake up, filter with microporous filter membrane (0.45um), get subsequent filtrate, promptly.Adopt above-mentioned chromatographic condition, measure.
4.1 extraction time research 20 (30 or 40) minute,
Add hydrochloric acid (1 → 2) 3 (5 or 7) ml 4.2 add the research of acid amount,
4.3 reflux 20 in the investigation water-bath of hydrolysis time (30 or 40) minute,
Experimental result sees Table 2.
Table 2
| The investigation project | Ultrasonic time | Add the acid amount | Hydrolysis time | ||||||
| The investigation condition | 20min | 30min | 40min | 3ml | 5ml | 7ml | 20min | 30min | 40min |
| Quercetin, isorhamnetin, kaempferide total content (mg/ sheet) | 4.42 | 4.45 | 4.45 | 4.58 | 4.64 | 4.56 | 4.62 | 4.65 | 4.61 |
Result: by test relatively, determine that the test liquid preparation method is: get 20 of this product, remove coating, the accurate title, decide, and porphyrize is got an amount of (being equivalent to total flavones 10mg), the accurate title, decide, and puts in the tool plug conical flask, the accurate methanol 25ml that adds, claim to decide weight, supersound process (power 135W, frequency 59kHz) 30 minutes, put coldly, claim again to decide weight, supply the weight that subtracts mistake with methanol, shake up, filter, discard filtrate just, precision is measured subsequent filtrate 5ml, put in the tool plug conical flask, add methanol 15ml, hydrochloric acid (1 → 2) 5ml, put in the water-bath reflux 30 minutes, cooling is transferred in the 50ml measuring bottle rapidly, is diluted to scale with methanol, shake up, filter with microporous filter membrane (0.45um), get subsequent filtrate, promptly.
5. specificity test
With the XINDAKANG PIAN negative sample that lacks Fructus Hippophae flavone, press the test sample disposal methods after, measure, the result is noiseless on the position at Quercetin, isorhamnetin, kaempferol peak.
6. the preparation of standard curve
Precision takes by weighing Quercetin reference substance 20.69mg, puts in the 50ml measuring bottle, adds dissolve with methanol and is diluted to scale, shakes up, in contrast the product stock solution.Precision takes by weighing kaempferol reference substance 3.23mg, puts in the 10ml measuring bottle, adds dissolve with methanol and is diluted to scale, shakes up, in contrast the product stock solution.Precision takes by weighing isorhamnetin reference substance 8.46mg, puts in the 25ml measuring bottle, adds dissolve with methanol and is diluted to scale, shakes up, in contrast the product stock solution.Precision is measured Quercetin reference substance stock solution 10ml, kaempferol reference substance stock solution 1ml, isorhamnetin reference substance stock solution 10ml in same 50ml measuring bottle respectively, is diluted to scale with methanol, shakes up, as mixing reference substance solution.Precision is measured mixing reference substance solution 1,2,4,6,8,10ml in the 10ml measuring bottle respectively again, is diluted to scale with methanol, shakes up, and draws the 10ul sample introduction.With the sample size is abscissa, and peak area is a vertical coordinate drawing standard curve.Calculate regression equation.
Quercetin regression equation: A=4636757.285C-18655.16986 (r
2=0.999989135)
Kaempferol regression equation: A=4025498.961C-4063.736986 (r
2=0.999377663)
Isorhamnetin regression equation: A=4551361C-22470.2 (r
2=0.999995)
The result shows: under above-mentioned chromatographic condition, Quercetin is between 0.08276-0.8276ug, sample feeding amount and peak area linear relationship are good, kaempferol is between 0.00646-0.0646ug, sample feeding amount and peak area linear relationship are good, isorhamnetin is between 0.06768-0.6768ug, and sample feeding amount and peak area linear relationship are good.
7. precision test
7.1 accurate Quercetin, kaempferol, each 10ul of isorhamnetin mixing reference substance solution of drawing of instrument precision repeats sample introduction 5 times under above-mentioned chromatographic condition.The results are shown in Table 3.
The test of table 3 instrument precision
| The sample introduction number of times | Quercetin | Kaempferide | Isorhamnetin |
| 1 | 1486544 | 118686 | 767727 |
| 2 | 1451610 | 113661 | 746763 |
| 3 | 1483973 | 120058 | 768565 |
| 4 | 1483038 | 119708 | 762612 |
| 5 | 1485164 | 120285 | 766082 |
| RSD(％) | 1.0 | 2.3 | 1.2 |
7.2 replica test
Parallel getting with 6 parts in batch XINDAKANG PIAN sample by " assay " test liquid preparation method test down, adopted above-mentioned chromatographic condition, measures Quercetin, kaempferol, isorhamnetin content.The results are shown in Table 4.
Table 4 replica test result
| Test number (TN) | 1 | 2 | 3 | 4 | 5 | 6 | RSD(％) |
| Quercetin, kaempferide, isorhamnetin total content (mg/ sheet) | 4.82 | 4.83 | 4.82 | 4.82 | 4.80 | 4.82 | 1.1 |
Above result shows: Quercetin, and kaempferide, the disorderly content of different Fructus rhamni (Rhamnus davurica Pall.) has repeatability preferably
8, accuracy (average recovery test)
Precision is measured Quercetin reference substance solution (concentration 0.9348mg/ml) 3ml, nimbecetin reference substance solution (concentration 0.2196mg/ml) 1ml, isorhamnetin reference substance solution (concentration 0.54mg/ml) 3ml, totally 6 parts, put respectively in 6 tool plug conical flasks, solvent evaporated in the water-bath, standby.Get the known sample that contains Quercetin 19.07mg/g, kaempferol 1.40mg/g, isorhamnetin 11.60mg/g, the accurate title, decide, put in the above-mentioned tool plug conical flask, measure Quercetin, kaempferol, the total metering method of isorhamnetin and carry out application of sample and reclaim experiment, the results are shown in Table 5, table 6, table 7 by drafting.
Table 5 Quercetin average recovery result of the test
| Numbering | Sampling amount (g) | Quercetin amount (mg) in the sample | Quercetin addition (mg) | Record Quercetin amount (mg) | Response rate % | Average recovery rate % | RSD value (%) |
| 1 | 0.1607 | 3.17 | 2.8044 | 5.78 | 93.1 | 92.7 | 0.82 |
| 2 | 0.1550 | 3.05 | 2.8044 | 5.64 | 92.4 | ||
| 3 | 0.1496 | 2.95 | 2.8044 | 5.68 | 93.8 | ||
| 4 | 0.1413 | 2.78 | 2.8044 | 5.37 | 92.4 | ||
| 5 | 0.1444 | 2.84 | 2.8044 | 5.45 | 93.1 | ||
| 6 | 0.1463 | 2.88 | 2.8044 | 5.45 | 91.6 |
Table 6 kaempferol average recovery result of the test
| Numbering | Sampling amount (g) | Kaempferide amount (mg) in the sample | Kaempferide addition (mg) | Record kaempferide amount (mg) | Response rate % | Average recovery rate % | RSD value (%) |
| 1 | 0.1607 | 0.22 | 0.2196 | 0.42 | 91.1 | 90.3 | 2.08 |
| 2 | 0.1550 | 0.22 | 0.2196 | 0.42 | 91.1 | ||
| 3 | 0.1496 | 0.21 | 0.2196 | 0.41 | 91.1 | ||
| 4 | 0.1413 | 0.20 | 0.2196 | 0.39 | 86.5 | ||
| 5 | 0.1444 | 0.20 | 0.2196 | 0.40 | 91.1 | ||
| 6 | 0.1463 | 0.20 | 0.2196 | 0.40 | 91.1 |
Table 7 isorhamnetin average recovery result of the test
| Numbering | Sampling amount (g) | Isorhamnetin amount (mg) in the sample | Isorhamnetin addition (mg) | Record isorhamnetin amount (mg) | Response rate % | Average recovery rate % | RSD value (%) |
| 1 | 0.1607 | 1.86 | 1.62 | 3.36 | 92.6 | 91.7 | 1.13 |
| 2 | 0.1550 | 1.80 | 1.62 | 3.29 | 92.0 | ||
| 3 | 0.1496 | 1.74 | 1.62 | 3.24 | 92.6 | ||
| 4 | 0.1413 | 1.64 | 1.62 | 3.11 | 90.7 | ||
| 5 | 0.1444 | 1.68 | 1.62 | 3.17 | 92.0 | ||
| 6 | 0.1463 | 1.70 | 1.62 | 3.16 | 90.1 |
Above result's test shows: this method has the response rate preferably, and accuracy is good.
9, ruggedness experiment
9.1 stability test is got need testing solution and measured content in the preparation back after 0,1,3,6,12,24 hour, it the results are shown in Table 8.
Table 8 stability test result
| Standing time | 0 | 1 | 3 | 6 | 12 | 24 | RSD(％) |
| Quercetin | 1547874 | 1540112 | 1543021 | 1532368 | 1485664 | 1566521 | 1.8 |
| Kaempferol | 124117 | 122975 | 125646 | 125481 | 119476 | 126826 | 2.1 |
| Isorhamnetin | 801384 | 797535 | 800284 | 793973 | 765418 | 816134 | 2.1 |
Above result shows: test sample is better at 24 hours internal stabilities.
9.2 the test that the mobile phase ratio changes
Sample thief prepares need testing solution by the preparation method of above-mentioned definite need testing solution, and mobile phase is respectively: 1. methanol-0.4% phosphoric acid (45: 55)); 2. methanol-0.4% phosphoric acid (50: 50); 3. methanol-0.4% phosphoric acid (55: 45).Constant at other chromatographic conditions, only change under the situation of mobile phase, measurement result is investigated.Measure by sample introduction under the above-mentioned chromatographic condition, measurement result sees Table 9.
The investigation that table 9 mobile phase ratio changes
| Methanol-0.4% phosphoric acid (45: 55) | Methanol-0.4% phosphoric acid (50: 50) | Methanol-0.4% phosphoric acid (55: 45) | |
| Quercetin, kaempferide, isorhamnetin total content (mg/ sheet) | 4.49 | 4.58 | 4.47 |
According to above measurement result, RSD is 1.3%, and the less variation of mobile phase ratio does not have influence to measurement result.
9.3 the investigation of different brands chromatographic column
Get same batch sample, prepare need testing solution by the preparation method of above-mentioned definite need testing solution, chromatographic column is respectively: (1) Kromasil.200 * 4.6mmID, 5um; (2) Dickma 250 * 4.mmID 5um; (3) phenomemex 150 * 4.6 ID 5um.Constant in mobile phase and other chromatographic conditions, only change employed chromatographic column, measurement result is investigated.Measure by sample introduction under the above-mentioned chromatographic condition, measurement result sees Table 10.
The investigation of table 10 different brands chromatographic column
| Kromasil | Dickma | phenomemex | |
| Quercetin, the disorderly element in mountain, isorhamnetin total content (mg/ sheet) | 4.47 | 4.52 | 4.55 |
According to above measurement result, RSD is 1.09%, and the chromatographic column of different brands does not have obvious influence to measurement result.
According to above result of the test as can be known, the ruggedness of this method is better.
10, sample size is measured
By the Xindakang preparation Quercetin of drafting, kaempferide, isorhamnetin content assaying method, former XINDAKANG PIAN that keeps sample for a long time and commercially available prod have been carried out assay, the results are shown in Table 11, table 12, table 13.
Five batches of XINDAKANG PIAN of table 11 (specification: 5mg) assay result
| Lot number | Quercetin (mg) | Nimbecetin (mg) | Isorhamnetin (mg) | Total amount (mg/ sheet) | Account for labelled amount (%) |
| 060103 | 2.88 | 0.20 | 1.67 | 4.75 | 95.0 |
| 060202 | 2.78 | 0.20 | 1.62 | 4.60 | 92.0 |
| 060203 | 2.91 | 0.20 | 1.71 | 4.83 | 96.6 |
| 060301 | 2.96 | 0.21 | 1.73 | 4.90 | 98.0 |
| 060402 | 2.96 | 0.21 | 1.73 | 4.90 | 98.0 |
Three batches of XINDAKANG PIAN of table 12 (specification: 10mg) assay result
| Lot number | Quercetin (mg) | Nimbecetin (mg) | Isorhamnetin (mg) | Total amount (mg/ sheet) | Account for labelled amount (%) |
| 060101 | 6.14 | 0.25 | 3.27 | 9.66 | 96.6 |
| 060201 | 5.76 | 0.34 | 3.27 | 9.37 | 93.7 |
| 060401 | 5.47 | 0.37 | 3.22 | 9.06 | 90.6 |
Three batches of XINDAKANG PIAN of table 13 (specification: 5mg) assay result
| Manufacturer | Lot number | Quercetin (mg) | Nimbecetin (mg) | Isorhamnetin (mg) | Total amount (mg/ sheet) | Account for labelled amount (%) |
| Shanghai Tiancifu Biological Engineering Co., Ltd. | 051102 | 1.12 | 0.37 | 2.59 | 4.08 | 81.6 |
| Chuanda Huaxi Pharmaceutical Industry Co., Ltd. Sichuan Prov. | 050401 | 0.99 | 0.30 | 2.92 | 4.21 | 84.2 |
| The refined Pharmaceutical limited company that reaches in Sichuan | 050610 | 1.01 | 0.22 | 3.02 | 4.25 | 85 |
As can be seen from the above results, Quercetin, kaempferol, isorhamnetin total content are equivalent to 81.6%～98.0% of its total flavones labelled amount in the employing high effective liquid chromatography for measuring XINDAKANG PIAN.This standard of considering is just carried out assay to existing procucts, therefore is its limit standard with the minimum data that records temporarily.Promptly adopt Quercetin in the high effective liquid chromatography for measuring XINDAKANG PIAN, kaempferol, isorhamnetin total amount, should be not less than 80.0% of labelled amount.
The specific embodiment
Further specify the present invention by the following examples, but not as limitation of the present invention, method of quality control of the present invention also can be used for the quality control of following other Flavonihippophae dosage form medicine.
Embodiment one: described tablet quality control method comprises
Character: this product is coated tablet or Film coated tablets, removes to show pale brown color behind the coating to brown, mildly bitter flavor.
Differentiate: get 1 of this product, remove coating, porphyrize adds every 5ml of ethanol (5mg) or 10ml (10mg), and supersound process 5 minutes filters, and filtrate is as need testing solution; Other gets Quercetin and isorhamnetin reference substance, adds ethanol and makes the mixed solution that every 1ml contains 1mg, in contrast product solution; Test according to thin layer chromatography (appendix VIB of Chinese Pharmacopoeia), draw each 1～2ul of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene: ethyl acetate: formic acid=5: 4: 0.5 is developing solvent, launch, take out, dry, spray is put under the 365nm ultra-violet lamp and is inspected with 1% aluminum chloride alcoholic solution; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.
Check: should meet each relevant under tablet item regulation (appendix ID of Chinese Pharmacopoeia).
Assay
(1) total flavones
The preparation of reference substance solution: precision takes by weighing the isorhamnetin reference substance, adds dehydrated alcohol and makes the solution that every 1ml contains isorhamnetin 20ug, promptly.
The preparation of standard curve: precision is measured reference substance solution 1,2,3,4,5,6ml, put respectively in the 10ml measuring bottle, add 1% aluminum chloride ethanol solution 1.0ml, be diluted to scale with dehydrated alcohol, shake up, placing 10 minutes, is blank with the reagent corresponding, according to ultraviolet visible spectrophotometry (appendix VA of Chinese Pharmacopoeia version in 2005), wavelength place at 430nm measures absorbance, with the absorbance is vertical coordinate, and concentration is abscissa, the drawing standard curve.
Algoscopy: get 20 in this tablet, remove coating, the accurate title, decide, porphyrize is got 0.1g, and accurate the title decides, put in the tool plug conical flask, it is an amount of to add dehydrated alcohol, and reflux is 15 minutes in 80 ℃ of water-baths, put coldly, in the dislocation 100ml measuring bottle, be diluted to scale with dehydrated alcohol, shake up, filter, discard filtrate just, precision is measured subsequent filtrate 10ml, put in the 25ml measuring bottle, be diluted to scale, shake up with dehydrated alcohol, precision is measured 3ml, put in the 10ml measuring bottle, add 1% aluminum chloride ethanol solution 1.0ml, be diluted to scale with dehydrated alcohol, shake up, as need testing solution; Other gets the same amount need testing solution that does not add developer and makes blank, and the method under the sighting target directrix curve preparation from " placing 10 minutes ", is measured absorbance in accordance with the law, reads the weight (ug) of isorhamnetin in the test sample from standard curve, calculates, promptly.
Every in this tablet contains Fructus Hippophae flavone with isorhamnetin (C
16H
12O
7) meter, should be 90.0%～110.0% of labelled amount.
Every in this tablet contains total flavones (in isorhamnetin) 1. 5mg, 2. 10mg.
(2) Quercetin, isorhamnetin, kaempferol are measured according to high performance liquid chromatography (appendix VID of Chinese Pharmacopoeia version in 2005):
Chromatographic column is the C18 post, and flow velocity is 1ml/s, and column temperature is 35 ℃, and with methanol: 0.4% phosphoric acid=50: 50 is mobile phase; The detection wavelength is 370nm; Theoretical cam curve is calculated by the isorhamnetin peak should be not less than 3000.
The preparation of reference substance solution: precision takes by weighing with P
2O
5An amount of for desiccant drying under reduced pressure isorhamnetin more than 12 hours, kaempferol, 4 hours Quercetin reference substance of 120 ℃ of dryings, add methanol and make the mixed solution that every 1ml contains isorhamnetin 20ug, kaempferol 2ug, Quercetin 25ug respectively, promptly.
The preparation of need testing solution: get 20 of this product, remove coating, the accurate title, decide, porphyrize is got an amount of (being equivalent to total flavones 10mg), and accurate the title decides, put in the tool plug conical flask, the accurate methanol 25ml that adds claims to decide weight, supersound process (power 135W, frequency 59kHz) 30 minutes is put cold, claim again to decide weight, supply the weight that subtracts mistake, shake up with methanol, filter, discard filtrate just, precision is measured subsequent filtrate 5ml, put in the tool plug conical flask, add methanol 15ml, hydrochloric acid (1 → 2) 5ml, put in the water-bath reflux 30 minutes, cooling is transferred in the 50ml measuring bottle rapidly, be diluted to scale with methanol, shake up, filter with microporous filter membrane (0.45um), get subsequent filtrate, promptly.
Algoscopy: accurate respectively reference substance solution and each 10ul of need testing solution of drawing, inject chromatograph of liquid, measure, promptly.
Every in this tablet contains Quercetin (C
15H
10O
7), isorhamnetin (C
16H
12O
7), kaempferol (C
15H
10O
6) total amount be no less than 80.0% of total flavones labelled amount;
[function with cure mainly] reinforce functional activities of the heart, blood stasis dispelling is promoted blood circulation, the expectorant activating the spleen.Be used for a little less than the insufficiency of heart-QI, heart arteries and veins stasis of blood resistance, phlegm-damp is stranded nervous, cardiopalmus, pained due to the spleen, and it is uncomfortable in chest to breathe hard, and blood vessels are not smooth, cough the tired disease of waiting.
[usage and consumption] is oral, a 10mg, and 3 times on the one, three months is a course of treatment.
[specification] every contains 1. 2. 10mg of 5mg of total flavones (in isorhamnetin)
Dry place is put in [storage a surname] sealing.
Embodiment two: described capsule method of quality control comprises
Character: content is that fawn is to dun granule and powder; Feeble QI, mildly bitter flavor.
Differentiate: get this capsule content 5mg or 10mg, add ethanol 5ml or 10ml, supersound process 5 minutes is made the solution of 1mg/ml, filters, and filtrate is as need testing solution; Other gets Quercetin and isorhamnetin reference substance, adds ethanol and makes the mixed solution that every 1ml contains 1mg, in contrast product solution; Test according to an appendix VIB of Chinese Pharmacopoeia version in 2005 thin layer chromatography, draw each 1～2ul of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene: ethyl acetate: formic acid=5: 4: 0.5 is developing solvent, launch, take out, dry, spray is put under the 365nm ultra-violet lamp and is inspected with 3% aluminum chloride alcoholic solution; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.
Check: should meet pertinent regulations under the capsule item (appendix IL of Chinese Pharmacopoeia version in 2005).。
Assay
(1) total flavones
The preparation of reference substance solution: precision takes by weighing the isorhamnetin reference substance, adds dehydrated alcohol and makes the solution that every 1ml contains isorhamnetin 20ug, promptly;
The preparation of standard curve: precision is measured reference substance solution 1,2,3,4,5,6ml, put respectively in the 10ml measuring bottle, add 3% aluminum chloride ethanol solution 1.0ml, be diluted to scale with dehydrated alcohol, shake up, placing 10 minutes, is blank with the reagent corresponding, according to ultraviolet visible spectrophotometry (appendix VA of Chinese Pharmacopoeia version in 2005), wavelength place at 430nm measures absorbance, with the absorbance is vertical coordinate, and concentration is abscissa, the drawing standard curve.
Algoscopy: it is tolerant to get this capsule 10～20 intragranulars, and accurate the title decided porphyrize, mixing is got 0.1g, and accurate the title decides, put in the tool plug conical flask, it is an amount of to add dehydrated alcohol, and reflux is 15 minutes in 80 ℃ of water-baths, put coldly, in the dislocation 100ml measuring bottle, be diluted to scale with dehydrated alcohol, shake up, filter, discard filtrate just, precision is measured subsequent filtrate 10ml, put in the 25ml measuring bottle, be diluted to scale, shake up with dehydrated alcohol, precision is measured 3ml, put in the 10ml measuring bottle, add 3% aluminum chloride ethanol solution 1.0ml, be diluted to scale with dehydrated alcohol, shake up, as need testing solution; Other gets the same amount need testing solution that does not add developer and makes blank, and the method under the sighting target directrix curve preparation from " placing 10 minutes ", is measured absorbance in accordance with the law, reads the weight (uG) of isorhamnetin in the test sample from standard curve, calculates, promptly;
This capsule contains Fructus Hippophae flavone with isorhamnetin (C
16H
12O
7) meter, should be 90.0%～110.0% of labelled amount.
Every of this capsule contains flavone (in isorhamnetin) 1. 5mg, 2. 10mg.
(2) Quercetin, isorhamnetin, kaempferol are measured according to high performance liquid chromatography (appendix VID of Chinese Pharmacopoeia version in 2005).
Chromatographic column is the C18 post, and flow velocity is 1ml/s, and column temperature is 35 ℃, and with methanol: 0.4% phosphoric acid=50: 50 is mobile phase; The detection wavelength is 370nm.Theoretical cam curve is calculated by the isorhamnetin peak should be not less than 3000.
The preparation of reference substance solution: precision takes by weighing with P
2O
5An amount of for desiccant drying under reduced pressure isorhamnetin more than 12 hours, kaempferol, 4 hours Quercetin reference substance of 120 ℃ of dryings, add methanol and make the mixed solution that every 1ml contains isorhamnetin 20ug, kaempferol 2ug, Quercetin 25ug respectively, promptly;
The preparation of need testing solution: get content an amount of (being equivalent to total flavones 10mg) under the determination of total flavonoids item, the accurate title, decide, and puts in the tool plug conical flask, the accurate methanol 25ml that adds claims to decide weight, supersound process (power 135W, frequency 59kHz) 30 minutes, put coldly, claim to decide weight again, supply the weight that subtracts mistake with methanol, shake up, filter, discard filtrate just, precision is measured subsequent filtrate 5ml, puts in the tool plug conical flask, add methanol 15ml, hydrochloric acid (1 → 2) 5ml put in the water-bath reflux 30 minutes, rapidly cooling, be transferred in the 50ml measuring bottle, be diluted to scale with methanol, shake up, filter with microporous filter membrane (0.45um), get subsequent filtrate, promptly.
Accurate respectively reference substance solution and each 10ul of need testing solution of drawing of algoscopy injects chromatograph of liquid, measures, promptly.
Every of this capsule contains Quercetin (C
15H
10O
7), isorhamnetin (C
16H
12O
7), kaempferol (C
15H
10O
6) total amount be no less than 80.0% of total flavones labelled amount.
Embodiment three: this soft capsule method of quality control comprises
Character: content is the yellow oily liquid that contains a small amount of suspended solid extractum; Feeble QI, mildly bitter flavor.
Differentiate: get this capsule content 5mg or 10mg, add ethanol 5ml or 10ml, supersound process 5 minutes is made the solution of 1mg/ml, filters, and filtrate is as need testing solution; Other gets Quercetin and isorhamnetin reference substance, adds ethanol and makes the mixed solution that every 1ml contains 1mg, in contrast product solution; Test according to an appendix VIB of Chinese Pharmacopoeia version in 2005 thin layer chromatography, draw each 1～2ul of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene: ethyl acetate: formic acid=5: 4: 0.5 is developing solvent, launch, take out, dry, spray is put under the 365nm ultra-violet lamp and is inspected with 3% aluminum chloride alcoholic solution; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.
Check: should meet pertinent regulations under the capsule item (appendix IL of Chinese Pharmacopoeia version in 2005).
Assay
(1) total flavones
The preparation of reference substance solution: precision takes by weighing the isorhamnetin reference substance, adds dehydrated alcohol and makes the solution that every 1ml contains isorhamnetin 20ug, promptly;
The preparation of standard curve: precision is measured reference substance solution 1,2,3,4,5,6ml, put respectively in the 10ml measuring bottle, add 3% aluminum chloride ethanol solution 1.0ml, be diluted to scale with dehydrated alcohol, shake up, placing 10 minutes, is blank with the reagent corresponding, according to ultraviolet visible spectrophotometry (appendix VA of Chinese Pharmacopoeia version in 2005), wavelength place at 430nm measures absorbance, with the absorbance is vertical coordinate, and concentration is abscissa, the drawing standard curve.
Algoscopy: it is tolerant to get this preparation 10～20 intragranulars, and accurate the title decided mixing, get 0.1g, the accurate title, decide, and puts in the tool plug conical flask, it is an amount of to add dehydrated alcohol, and reflux is 15 minutes in 80 ℃ of water-baths, puts cold, in the dislocation 100ml measuring bottle, be diluted to scale, shake up with dehydrated alcohol, filter, discard filtrate just, precision is measured subsequent filtrate 10ml, puts in the 25ml measuring bottle, be diluted to scale with dehydrated alcohol, shake up, precision is measured 3ml, puts in the 10ml measuring bottle, add 3% aluminum chloride ethanol solution 1.0ml, be diluted to scale with dehydrated alcohol, shake up, as need testing solution; Other gets the same amount need testing solution that does not add developer and makes blank, and the method under the sighting target directrix curve preparation from " placing 10 minutes ", is measured absorbance in accordance with the law, reads the weight (ug) of isorhamnetin in the test sample from standard curve, calculates, promptly;
This soft capsule contains Fructus Hippophae flavone with isorhamnetin (C
16H
12O
7) meter, should be 90.0%～110.0% of labelled amount.
Every of this soft capsule contains flavone (in isorhamnetin) 1. 5mg, 2. 10mg.
(2) Quercetin, isorhamnetin, kaempferide are measured according to high performance liquid chromatography (appendix VID of Chinese Pharmacopoeia version in 2005).
Chromatographic column is the C18 post, and flow velocity is 1ml/s, and column temperature is 35 ℃, and with methanol: 0.4% phosphoric acid=50: 50 is mobile phase; The detection wavelength is 370nm; Theoretical cam curve is calculated by the isorhamnetin peak should be not less than 3000.
The preparation of reference substance solution: precision takes by weighing with P
2O
5An amount of for desiccant drying under reduced pressure isorhamnetin more than 12 hours, kaempferol, 4 hours Quercetin reference substance of 120 ℃ of dryings, add methanol and make the mixed solution that every 1ml contains isorhamnetin 20ug, kaempferol 2ug, Quercetin 25ug respectively, promptly;
The preparation of need testing solution: get total flavones and measure item content an amount of (being equivalent to total flavones 10mg) down, the accurate title, decide, and puts in the tool plug conical flask, the accurate methanol 25ml that adds claims to decide weight, supersound process (power 135W, frequency 59kHz) 30 minutes, put coldly, claim to decide weight again, supply the weight that subtracts mistake with methanol, shake up, filter, discard filtrate just, precision is measured subsequent filtrate 5ml, puts in the tool plug conical flask, add methanol 15ml, hydrochloric acid (1 → 2) 5ml put in the water-bath reflux 30 minutes, rapidly cooling, be transferred in the 50ml measuring bottle, be diluted to scale with methanol, shake up, filter with microporous filter membrane (0.45um), get subsequent filtrate, promptly.
Algoscopy: accurate respectively reference substance solution and each 10ul of need testing solution of drawing, inject chromatograph of liquid, measure, promptly.
Every of this soft capsule contains Quercetin (C
15H
10O
7), isorhamnetin (C
16H
12O
7), kaempferol (C
15H
10O
6) total amount be no less than 80.0% of total flavones labelled amount.
Embodiment four: this drop pill method of quality control comprises
Character: content is that fawn is to dun granule and powder; Feeble QI, mildly bitter flavor.
Differentiate: get this drop pill content 5mg or 10mg, add ethanol 5ml or 10ml, supersound process 5 minutes is made the solution of 1mg/ml, filters, and filtrate is as need testing solution; Other gets Quercetin and isorhamnetin reference substance, adds ethanol and makes the mixed solution that every 1ml contains 1mg, in contrast product solution; Test according to an appendix VIB of Chinese Pharmacopoeia version in 2005 thin layer chromatography, draw each 1～2ul of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene: ethyl acetate: formic acid=5: 4: 0.5 is developing solvent, launch, take out, dry, spray is put under the 365nm ultra-violet lamp and is inspected with 1% aluminum chloride alcoholic solution; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.
Check: should meet pertinent regulations under the drop pill item (appendix IK of Chinese Pharmacopoeia version in 2005).
Assay
(1) total flavones
The preparation of reference substance solution: precision takes by weighing the isorhamnetin reference substance, adds dehydrated alcohol and makes the solution that every 1ml contains isorhamnetin 20ug, promptly;
The preparation of standard curve: precision is measured reference substance solution 1,2,3,4,5,6ml, put respectively in the 10ml measuring bottle, add 1% aluminum chloride ethanol solution 1.0ml, be diluted to scale with dehydrated alcohol, shake up, placing 10 minutes, is blank with the reagent corresponding, according to ultraviolet visible spectrophotometry (appendix VA of Chinese Pharmacopoeia version in 2005), wavelength place at 430nm measures absorbance, with the absorbance is vertical coordinate, and concentration is abscissa, the drawing standard curve.
Algoscopy: this preparation 10～20 intragranulars are tolerant, and accurate the title decided mixing, get 0.1g, the accurate title, decide, and puts in the tool plug conical flask, it is an amount of to add dehydrated alcohol, and reflux is 15 minutes in 80 ℃ of water-baths, puts cold, in the dislocation 100ml measuring bottle, be diluted to scale, shake up with dehydrated alcohol, filter, discard filtrate just, precision is measured subsequent filtrate 10ml, puts in the 25ml measuring bottle, be diluted to scale with dehydrated alcohol, shake up, precision is measured 3ml, puts in the 10ml measuring bottle, add 1% aluminum chloride ethanol solution 1.0ml, be diluted to scale with dehydrated alcohol, shake up, as need testing solution; Other gets the same amount need testing solution that does not add developer and makes blank, and the method under the sighting target directrix curve preparation from " placing 10 minutes ", is measured absorbance in accordance with the law, reads the weight (ug) of isorhamnetin in the test sample from standard curve, calculates, promptly;
This drop pill contains Fructus Hippophae flavone with isorhamnetin (C
16H
12O
7) meter, should be 90.0%～110.0% of labelled amount.
The every ball of this drop pill contains flavone (in isorhamnetin) 1. 5mg, 2. 10mg.
(2) Quercetin, isorhamnetin, kaempferol are measured according to high performance liquid chromatography (appendix VID of Chinese Pharmacopoeia).
Chromatographic column is the C18 post, and flow velocity is 1ml/s, and column temperature is 35 ℃, and with methanol: 0.4% phosphoric acid=50: 50 is mobile phase; The detection wavelength is 370nm.Theoretical cam curve is calculated by the isorhamnetin peak should be not less than 3000.
The preparation of reference substance solution: precision takes by weighing with P
2O
5An amount of for desiccant drying under reduced pressure isorhamnetin more than 12 hours, kaempferol, 4 hours Quercetin reference substance of 120 ℃ of dryings, add methanol and make the mixed solution that every 1ml contains isorhamnetin 20ug, kaempferol 2ug, Quercetin 25ug respectively, promptly;
The preparation of need testing solution: get the content an amount of (being equivalent to total flavones 10mg) under the total flavones mensuration item, the accurate title, decide, and puts in the tool plug conical flask, the accurate methanol 25ml that adds claims to decide weight, supersound process (power 135W, frequency 59kHz) 30 minutes, put coldly, claim to decide weight again, supply the weight that subtracts mistake with methanol, shake up, filter, discard filtrate just, precision is measured subsequent filtrate 5ml, puts in the tool plug conical flask, add methanol 15ml, hydrochloric acid (1 → 2) 5ml put in the water-bath reflux 30 minutes, rapidly cooling, be transferred in the 50ml measuring bottle, be diluted to scale with methanol, shake up, filter with microporous filter membrane (0.45um), get subsequent filtrate, promptly.
Accurate respectively reference substance solution and each 10ul of need testing solution of drawing of algoscopy injects chromatograph of liquid, measures, promptly.
The every ball of this drop pill contains Quercetin (C
15H
10O
7), isorhamnetin (C
16H
12O
7), kaempferol (C
15H
10O
6) total amount be no less than 80.0% of total flavones labelled amount.
Claims (9)
1. the method for quality control of an Xindakang preparation, described preparation comprises tablet, capsule, soft capsule, drop pill, it is characterized in that: described method of quality control mainly comprise in character, inspection, discriminating, the assay project partly or entirely; Wherein differentiate and comprise that with Quercetin reference substance, isorhamnetin reference substance be the thin layer chromatography discriminating that flavone aglycone in the preparation is differentiated in contrast; Assay comprises the assay to Quercetin, isorhamnetin, kaempferol total amount in the preparation.
2. according to the method for quality control of the described Xindakang preparation of claim 1, it is characterized in that: discrimination method is to be contrast with Quercetin reference substance and isorhamnetin reference substance, and with toluene: ethyl acetate: formic acid=2～10: 1～8: 0.1～1.5 solution is the tlc identification method of developing solvent.
3. according to the method for quality control of claim 1 or 2 described Xindakang preparations, it is following to it is characterized in that described discrimination method comprises:
Get this preparation or its content, porphyrize adds ethanol and makes dissolving, filters, and filtrate is as need testing solution; Other gets Quercetin and isorhamnetin reference substance, adds ethanol and makes mixed solution, in contrast product solution; Test according to the Chinese Pharmacopoeia thin layer chromatography, draw above-mentioned need testing solution, reference substance solution, put respectively on same silica gel g thin-layer plate, with toluene: ethyl acetate: formic acid=2～10: 1～8: 0.1～1.5 is developing solvent, launch, take out, dry, spray is put under the ultra-violet lamp and is inspected with 1～5% aluminum chloride alcoholic solution; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.
4. according to the method for quality control of the described Xindakang preparation of claim 3, it is characterized in that: discrimination method comprises following more specifically:
Get this preparation or its content, porphyrize is got fine powder 2～20mg, adds dehydrated alcohol 2～20ml, and supersound process 1～15 minute filters, and filtrate is as need testing solution; Other gets Quercetin and isorhamnetin reference substance, adds dehydrated alcohol and makes the mixed solution that every 1ml contains 1mg～10mg, in contrast product solution; Test according to an appendix VIB of Chinese Pharmacopoeia thin layer chromatography, draw each 1～20ul of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene: ethyl acetate: formic acid=2～10: 1～8: 0.1～1.5 is developing solvent, launch, take out, dry, spray is put under the 365nm ultra-violet lamp and is inspected with 1～5% aluminum chloride alcoholic solution; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.
5. according to the method for quality control of the described Xindakang preparation of claim 1, it is characterized in that: the assay method of Quercetin, isorhamnetin, kaempferol total content is to be contrast with isorhamnetin, Quercetin, kaempferol reference substance in the preparation, and with methanol: 0.4% phosphoric acid=30～60: 30～60 is the high performance liquid chromatography of mobile phase.
6. according to the method for quality control of claim 1 or 5 described Xindakang preparations, it is characterized in that: described content assaying method comprises following:
Quercetin, isorhamnetin, kaempferol are according to the Chinese Pharmacopoeia high effective liquid chromatography for measuring: chromatographic column is the C18 post, and with methanol: 0.4% phosphoric acid=30～60: 30～60 is mobile phase; The detection wavelength is 370nm; Theoretical cam curve is calculated by the isorhamnetin peak should be not less than 3000; Precision takes by weighing isorhamnetin, kaempferol, Quercetin reference substance, adds dissolve with methanol and makes mixed solution, gets reference substance solution; Get this preparation or its content, the accurate title, decide, and puts in the tool plug conical flask, the accurate methanol that adds, claim decide weight, supersound extraction is put coldly, and weight decided in title again, supply the weight that subtracts mistake with methanol, shake up, filter, precision is measured subsequent filtrate and is put in the tool plug conical flask, add methanol, hydrochloric acid, put reflux in the water-bath, cooling, in the dislocation measuring bottle, be diluted to scale, shake up with methanol, filter, get subsequent filtrate, get need testing solution; Accurate reference substance solution and the need testing solution drawn injects chromatograph of liquid, measures, promptly;
This preparation contains total flavones when counting 5mg with isorhamnetin, and each dosage form contains the total amount of Quercetin, isorhamnetin, kaempferol: every in tablet should be no less than labelled amount 80%, capsule and every of soft capsule should be no less than labelled amount 80%, the every ball of drop pill should be no less than 80% of labelled amount;
This preparation contains total flavones when counting 10mg with isorhamnetin, and each dosage form contains the total amount of Quercetin, isorhamnetin, kaempferol: every in tablet should be no less than labelled amount 80%, capsule and every of soft capsule should be no less than labelled amount 80%, the every ball of drop pill should be no less than 80% of labelled amount.
7. according to the method for quality control of the described Xindakang preparation of claim 6, it is characterized in that: content assaying method comprises following more specifically:
Quercetin, isorhamnetin, kaempferol shine an appendix VID of Chinese Pharmacopoeia high effective liquid chromatography for measuring: chromatographic column is the C18 post; With methanol: 0.4% phosphoric acid=30～60: 30～60 is mobile phase; The detection wavelength is 370nm; Theoretical cam curve is calculated by the isorhamnetin peak should be not less than 3000; Precision takes by weighing puts P
2O
5Dry 10～48 hours isorhamnetin, kaempferol, 4 hours Quercetin reference substance of 120 ℃ of dryings add methanol and make the mixed solution that every 1ml contains isorhamnetin 20ug, kaempferol 2ug, Quercetin 25ug respectively in the vacuum drying apparatus, promptly get reference substance solution; Get that this preparation or its content 100～200mg are accurate to be claimed surely, porphyrize is got and is equivalent to total flavones 10mg sample, the accurate title, decide, and puts in the tool plug conical flask, the accurate methanol 25ml that adds, claim to decide weight, at power 135W, supersound process is 30 minutes under the frequency 59kHz, puts cold, claim again to decide weight, supply the weight that subtracts mistake, shake up with methanol, filter, discard filtrate just, precision is measured subsequent filtrate 5ml, put in the tool plug conical flask, add methanol 15ml, hydrochloric acid (1 → 2) 5ml, put in 80～100 ℃ of water-baths reflux 30 minutes, cooling in the transposition 50ml measuring bottle, is diluted to scale with methanol rapidly, shake up, filter with the 0.45um microporous filter membrane, get subsequent filtrate, promptly get need testing solution; Accurate respectively reference substance solution and each 10ul of need testing solution of drawing injects chromatograph of liquid, measures, promptly;
This preparation contains total flavones when counting 5mg with isorhamnetin, and each dosage form contains the total amount of Quercetin, isorhamnetin, kaempferol: every in tablet should be no less than labelled amount 80%, capsule and every of soft capsule should be no less than labelled amount 80%, the every ball of drop pill should be no less than 80% of labelled amount;
This preparation contains total flavones when counting 10mg with isorhamnetin, and each dosage form contains the total amount of Quercetin, isorhamnetin, kaempferol: every in tablet should be no less than labelled amount 80%, capsule and every of soft capsule should be no less than labelled amount 80%, the every ball of drop pill should be no less than 80% of labelled amount.
8. according to the method for quality control of the described Xindakang preparation of claim 1, it is characterized in that described method of quality control comprises:
Character:
For tablet: content be pale brown color to brown, mildly bitter flavor;
For capsule: content is that fawn is to dun granule and powder; Feeble QI, mildly bitter flavor;
For soft capsule: content is the yellow oily liquid that contains a small amount of suspended solid extractum; Feeble QI, mildly bitter flavor;
For drop pill: this product is brown drop pill; Feeble QI, mildly bitter flavor;
Differentiate:
Get this preparation or its content, porphyrize adds ethanol and makes dissolving, filters, and filtrate is as need testing solution; Other gets Quercetin and isorhamnetin reference substance, adds ethanol and makes mixed solution, in contrast product solution; Test according to the Chinese Pharmacopoeia thin layer chromatography, draw above-mentioned need testing solution, reference substance solution, put respectively on same silica gel g thin-layer plate, with toluene: ethyl acetate: formic acid=2～10: 1～8: 0.1～1.5 is developing solvent, launch, take out, dry, spray is put under the ultra-violet lamp and is inspected with 1～5% aluminum chloride alcoholic solution; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color;
Check: this tablet, capsule, soft capsule or drop pill should meet pertinent regulations under Chinese Pharmacopoeia tablet, capsule, soft capsule or the drop pill item respectively;
Assay
(1) total flavones
The preparation of reference substance solution: precision takes by weighing the isorhamnetin reference substance, adds dehydrated alcohol and makes the solution that every 1ml contains isorhamnetin 20ug, promptly;
The preparation of standard curve: precision is measured reference substance solution 1,2,3,4,5,6ml, put respectively in the 10ml measuring bottle, add 1～5% aluminum chloride ethanol solution 1.0ml, be diluted to scale with dehydrated alcohol, shake up, placing 10 minutes, is blank with the reagent corresponding, according to an appendix VA of Chinese Pharmacopoeia ultraviolet visible spectrophotometry, wavelength place at 430nm measures absorbance, with the absorbance is vertical coordinate, and concentration is abscissa, the drawing standard curve;
Algoscopy: this preparation tablet is got 20 and is removed coating, 10～20 of capsule or drop pills, it is fixed to get the accurate title of its content, porphyrize, mixing, get 0.1g, the accurate title, decide, and puts in the tool plug conical flask, it is an amount of to add dehydrated alcohol, and reflux is 15 minutes in 80～100 ℃ of baths, puts cold, in the dislocation 100ml measuring bottle, be diluted to scale, shake up with dehydrated alcohol, filter, discard filtrate just, precision is measured subsequent filtrate 10ml, puts in the 25ml measuring bottle, be diluted to scale with dehydrated alcohol, shake up, precision is measured 3ml, puts in the 10ml measuring bottle, add 1～5% aluminum chloride ethanol solution 1.0ml, be diluted to scale with dehydrated alcohol, shake up, as need testing solution; Other gets the same amount need testing solution that does not add developer and does blank, method under the sighting target directrix curve preparation, from " placing 10 minutes ", according to an appendix V of Chinese Pharmacopoeia A ultraviolet visible spectrophotometry, wavelength place at 430nm measures absorbance, from standard curve, read the weight (ug) of isorhamnetin in the test sample, calculate, promptly;
This preparation contains total flavones when counting 5mg with isorhamnetin, contain Fructus Hippophae flavone in isorhamnetin, every in tablet should be labelled amount 90.0～110.0%, capsule and every of soft capsule should be labelled amount 90.0～110.0%, the every ball of drop pill should be 90.0～110.0% of labelled amount;
This preparation contains total flavones when counting 10mg with isorhamnetin, contain Fructus Hippophae flavone in isorhamnetin, every in tablet should be labelled amount 90.0～110.0%, capsule and every of soft capsule should be labelled amount 90.0～110.0%, the every ball of drop pill should be 90.0～110.0% of labelled amount;
(2) Quercetin, isorhamnetin, kaempferol adopt high effective liquid chromatography for measuring:
Chromatographic column is the C18 post, and with methanol: 0.4% phosphoric acid=30～60: 30～60 is mobile phase; The detection wavelength is 370nm; Theoretical cam curve is calculated by the isorhamnetin peak should be not less than 3000; Precision takes by weighing isorhamnetin, kaempferol, Quercetin reference substance, adds dissolve with methanol and makes mixed solution, gets reference substance solution; Get this preparation or its content, the accurate title, decide, and puts in the tool plug conical flask, the accurate methanol that adds, claim decide weight, supersound extraction is put coldly, and weight decided in title again, supply the weight that subtracts mistake with methanol, shake up, filter, precision is measured subsequent filtrate and is put in the tool plug conical flask, add methanol, hydrochloric acid, put reflux in the water-bath, cooling, in the dislocation measuring bottle, be diluted to scale, shake up with methanol, filter, get subsequent filtrate, get need testing solution; Accurate reference substance solution and the need testing solution drawn injects chromatograph of liquid, measures, promptly;
This preparation contains total flavones when counting 5mg with isorhamnetin, and each dosage form contains the total amount of Quercetin, isorhamnetin, kaempferol: every in tablet should be no less than labelled amount 80%, capsule and every of soft capsule should be no less than labelled amount 80%, the every ball of drop pill should be no less than 80% of labelled amount;
This preparation contains total flavones when counting 10mg with isorhamnetin, and each dosage form contains the total amount of Quercetin, isorhamnetin, kaempferol: every in tablet should be no less than labelled amount 80%, capsule and every of soft capsule should be no less than labelled amount 80%, the every ball of drop pill should be no less than 80% of labelled amount.
9. the method for quality control of described Xindakang preparation according to Claim 8 is characterized in that described method of quality control further comprises:
Character:
For tablet: content be pale brown color to brown, mildly bitter flavor;
For capsule: content is that fawn is to dun granule and powder; Feeble QI, mildly bitter flavor;
For soft capsule: content is the yellow oily liquid that contains a small amount of suspended solid extractum; Feeble QI, mildly bitter flavor;
For drop pill: this product is brown drop pill; Feeble QI, mildly bitter flavor;
Differentiate:
Get 1 in this preparation or 1, porphyrize adds ethanol 5ml or 10ml, and supersound process 5 minutes is made the solution of 1mg/ml, filters, and filtrate is as need testing solution; Other gets Quercetin and isorhamnetin reference substance, adds ethanol and makes the mixed solution that every 1ml contains 1mg, in contrast product solution; Test according to an appendix VIB of Chinese Pharmacopoeia thin layer chromatography, draw each 1～2ul of above-mentioned two kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene: ethyl acetate: formic acid=5: 4: 0.5 is developing solvent, launch, take out, dry, spray is put under the 365nm ultra-violet lamp and is inspected with 1～3% aluminum chloride alcoholic solution; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color;
Check: should meet in appendix of Chinese Pharmacopoeia each pertinent regulations under tablet, capsule, soft capsule or the drop pill item;
Assay
(1) total flavones
The preparation of reference substance solution: precision takes by weighing and adopts P
2O
5Dry isorhamnetin reference substance more than 12 hours adds dehydrated alcohol and makes the solution that every 1ml contains isorhamnetin 20ug, promptly;
The preparation of standard curve: precision is measured reference substance solution 1,2,3,4,5,6ml, put respectively in the 10ml measuring bottle, add 1～3% aluminum chloride ethanol solution 1.0ml, be diluted to scale with dehydrated alcohol, shake up, placing 10 minutes, is blank with the reagent corresponding, according to an appendix V of Chinese Pharmacopoeia A ultraviolet visible spectrophotometry, wavelength place at 430nm measures absorbance, with the absorbance is vertical coordinate, and concentration is abscissa, the drawing standard curve;
Algoscopy: this preparation is got 20 in tablet, get 10-20 seed lac wafer, the drop pill content, the accurate title, decided the porphyrize mixing, get about 0.1g, the accurate title, decide, and puts in the tool plug conical flask, it is an amount of to add dehydrated alcohol, and reflux is 15 minutes in 80 ℃ of water-baths, puts cold, in the dislocation 100ml measuring bottle, be diluted to scale, shake up with dehydrated alcohol, filter, discard filtrate just, precision is measured subsequent filtrate 10ml, puts in the 25ml measuring bottle, be diluted to scale with dehydrated alcohol, shake up, precision is measured 3ml, puts in the 10ml measuring bottle, add 1～3% aluminum chloride ethanol solution 1.0ml, be diluted to scale with dehydrated alcohol, shake up, as need testing solution; Other gets the same amount need testing solution that does not add developer and does blank, according to the method under the above-mentioned standard curve preparation, from " placing 10 minutes ", according to an appendix V of Chinese Pharmacopoeia A ultraviolet visible spectrophotometry, wavelength place at 430nm measures absorbance, from standard curve, read the weight (ug) of isorhamnetin in the test sample, calculate, promptly;
This preparation contains total flavones when counting 5mg with isorhamnetin, contain Fructus Hippophae flavone in isorhamnetin, every in tablet should be labelled amount 90.0～110.0%, capsule and every of soft capsule should be labelled amount 90.0～110.0%, the every ball of drop pill should be 90.0～110.0% of labelled amount;
This preparation contains total flavones when counting 10mg with isorhamnetin, contain Fructus Hippophae flavone in isorhamnetin, every in tablet should be labelled amount 90.0～110.0%, capsule and every of soft capsule should be labelled amount 90.0～110.0%, the every ball of drop pill should be 90.0～110.0% of labelled amount;
(2) Quercetin, isorhamnetin, kaempferol shine the Chinese Pharmacopoeia high effective liquid chromatography for measuring:
Chromatographic column is the C18 post; With methanol: 0.4% phosphoric acid=50: 50 is mobile phase; The detection wavelength is 370nm; Theoretical cam curve is calculated by the isorhamnetin peak should be not less than 3000; Precision takes by weighing with P
2O
5For desiccant drying under reduced pressure isorhamnetin more than 12 hours, kaempferol, 4 hours Quercetin reference substance of 120 ℃ of dryings an amount of, add methanol and make the mixed solution that every 1ml contains isorhamnetin 20ug, kaempferol 2ug, Quercetin 25ug respectively, promptly get reference substance solution; This preparation is got 20 in tablet and is removed coating, get 10-20 seed lac wafer, the drop pill content, the accurate title, decided the porphyrize mixing, get and be equivalent to total flavones 10mg, accurately claim surely, put in the tool plug conical flask, the accurate methanol 25ml that adds claims to decide weight, at power 135W, supersound process is 30 minutes under the frequency 59kHz, put cold, claim again to decide weight, supply the weight that subtracts mistake, shake up with methanol, filter, discard filtrate just, precision is measured subsequent filtrate 5ml, puts in the tool plug conical flask, add methanol 15ml, hydrochloric acid (1 → 2) 5ml put in the water-bath reflux 30 minutes, rapidly cooling, be transferred in the 50ml measuring bottle, be diluted to scale with methanol, shake up, filter with the 0.45um microporous filter membrane, get subsequent filtrate, promptly get need testing solution; Accurate respectively reference substance solution and each 10ul of need testing solution of drawing injects chromatograph of liquid, measures, promptly;
This preparation contains total flavones when counting 5mg with isorhamnetin, and each dosage form contains the total amount of Quercetin, isorhamnetin, kaempferol: every in tablet should be no less than labelled amount 80%, capsule and every of soft capsule should be no less than labelled amount 80%, the every ball of drop pill should be no less than 80% of labelled amount;
This preparation contains total flavones when counting 10mg with isorhamnetin, and each dosage form contains the total amount of Quercetin, isorhamnetin, kaempferol: every in tablet should be no less than labelled amount 80%, capsule and every of soft capsule should be no less than labelled amount 80%, the every ball of drop pill should be no less than 80% of labelled amount.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101140267B (en) * | 2007-10-10 | 2010-09-08 | 广西万寿堂药业有限公司 | Quality detecting method of mountain green tea blood pressure reducing preparations |
| CN105758980A (en) * | 2016-04-27 | 2016-07-13 | 广西壮族自治区梧州食品药品检验所 | Method for measuring total flavones in wild chrysanthemum flowers |
| CN107281233A (en) * | 2016-06-13 | 2017-10-24 | 中国科学院西北高原生物研究所 | Protection purposes of the total seabuckthorn flavone to blood vessel endothelium |
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2006
- 2006-12-29 CN CN200610022691A patent/CN100998617B/en active Active
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101140267B (en) * | 2007-10-10 | 2010-09-08 | 广西万寿堂药业有限公司 | Quality detecting method of mountain green tea blood pressure reducing preparations |
| CN105758980A (en) * | 2016-04-27 | 2016-07-13 | 广西壮族自治区梧州食品药品检验所 | Method for measuring total flavones in wild chrysanthemum flowers |
| CN107281233A (en) * | 2016-06-13 | 2017-10-24 | 中国科学院西北高原生物研究所 | Protection purposes of the total seabuckthorn flavone to blood vessel endothelium |
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