CN102138950A - Quality control method for Siberian cocklebur grass - Google Patents
Quality control method for Siberian cocklebur grass Download PDFInfo
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- CN102138950A CN102138950A CN 201110028620 CN201110028620A CN102138950A CN 102138950 A CN102138950 A CN 102138950A CN 201110028620 CN201110028620 CN 201110028620 CN 201110028620 A CN201110028620 A CN 201110028620A CN 102138950 A CN102138950 A CN 102138950A
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Abstract
The invention discloses a quality control method for Siberian cocklebur grass, which provides an identification method and a method for measuring chlorogenic acid content in medicaments by high efficiency liquid chromatography. By the improved quality standard, the quality of medicines can be better controlled. The quality control method has high specificity and excellent repeatability, and medicines are safe and effective and controllable in quality really.
Description
Technical field
The present invention relates to the Chinese medicine preparation field of quality control, be specifically related to a kind of method of quality control of Herba Xanthii grass.
Background technology
The plant resources of Herba Xanthii is very abundant, and applicating history is long, determined curative effect, and action range has unique therapeutical effect especially to many difficult miscellaneous syndromes, is rare good medicine.But pharmacopeia is not recorded this medicine, lacks the dark people's research to its quality standard, effective ingredient and pharmacological action at present yet, and limited research report is also just fragmentary.And do not see as yet for the report that the component content of the Herba Xanthii grass of difference growth collection period changes, multiple saying and rules of thumb are also arranged the collection period of Herba Xanthii grass.No matter from inheriting and develop the viewpoint of motherland's medicine, still from the needs of current people's medication, all be necessary the dark people's research of Herba Xanthii grass medicinal standard, chemical constituent and the mechanism of action and inquire into, so that development and use.
Chinese medicine for a long time, difficult quality is effectively controlled, the quality of effectively controlling tcm product is a great problem, the method for available technology adopting is imperfect, the minority specific aim is not strong, uses these methods to be difficult to reach the purpose of real control of quality.Above Herba Xanthii grass is owing to lack method of quality control well, be difficult to control the quality of Herba Xanthii grass, brought difficulty for normal production and operation, utilize more known technology to be difficult to the method for quality control that the Herba Xanthii grass product is provided, the present invention is directed to the deficiencies in the prior art, adopt new means that the quality of Herba Xanthii grass is controlled, particularly adopt with chlorogenic acid contents in this medicine of high effective liquid chromatography for measuring, solve the difficult problem of Herba Xanthii grass quality control, the present invention is directed to the method for quality control that Herba Xanthii grass provides a kind of practicality.
The present invention has proposed method of quality control to Herba Xanthii grass, this method provides the method for quality control such as content of chlorogenic acid composition in discriminating and the high effective liquid chromatography for measuring medicine, quality standard after the raising can be controlled the quality of medicine better, method of quality control of the present invention, have stronger specificity and good repeatability, real embodiment drug safety is effective, quality controllable.
Summary of the invention
Goal of the invention:, the object of the present invention is to provide a kind of method of quality control of Herba Xanthii grass in order to address the above problem.In order to reach this order ground, adopt following technical scheme:
Technical scheme:
Discrimination method is:
A: perusal Herba Xanthii grass powder is a yellow green, and 40 power microscopes are observed down, sees that the epidermis cell anticlinal wall is arranged is more straight or be shallow wavy and polygon; The pore infinitive, visible more glandular hair and nonglandular hair; The fiber bunchy, the visible pit that has, diameter 15-40 μ m; Needle-like calcium oxalate crystal diameter 5-50 μ m and calcium oxalate cluster crystal diameter 10-30 μ um; Secretory canal schizogenesis formula contains light brown secretions, diameter 20-80 μ m; Can see screw thread, bordered pit vessel, diameter 10-80 μ m;
B: get Herba Xanthii grass powder 2g, add 10% sodium hydroxide solution 50ml, reflux 30min, put coldly, filter, filtrate is used ethyl acetate extraction, each 50ml, 2 times, combined ethyl acetate liquid washes with water to neutrality, acetic acid ethyl fluid is concentrated into about 2ml, as need testing solution, other gets Fructus Xanthii control medicinal material 2g, shines medical material solution in pairs with legal system, draw each 10 μ l of above-mentioned two kinds of solution respectively, point is that toluene-ethyl acetate-glacial acetic acid of 16: 4: 1 is developing solvent with volume ratio on same silica gel g thin-layer plate, launches, take out, dry, inspect under the daylight, place under the 365nm ultraviolet light and inspect; Spray 105 ℃ of heating colour developings of 10% sulfuric acid solution, sample and Fructus Xanthii control medicinal material are equipped with same blob in identical bits;
C: get Herba Xanthii grass powder 5g, 50ml adds diethyl ether, reflux 1h, put cold, filter, filtrate low temperature is waved to 2ml, as need testing solution, other gets Herba Xanthii booth reference substance, make the 1mg/ml diethyl ether solution and compare liquid, draw above-mentioned two kinds of each 5ul of solution respectively, put respectively on same silica gel g thin-layer plate, with volume ratio is that cyclohexane extraction-ethyl acetate-formic acid of 10: 5: 0.2 is developing solvent, launch, take out, dry, put under the daylight respectively and spray 105 ℃ of heating of 10% sulfuric acid solution and develop the color, sample and Herba Xanthii booth standard substance are equipped with same blob in identical bits;
Described chlorogenic acid contents assay method is:
The chromatographic condition chromatographic column: diameter and length are 4.6mm * 250mm, the Alltima of 5 μ m
TMC18, mobile phase: the methanol of equal-volume ratio and 0.1% phosphoric acid mixed aqueous solution, flow velocity: 0.8ml/min, column temperature: 25 ℃, detect wavelength: 326nm, sample size: 20 μ l, sampling time: 60min;
It is an amount of that the preparation precision of reference substance solution takes by weighing the chlorogenic acid reference substance, adds 50% methanol and make the solution that concentration is 0.4mg/ml, as the chlorogenic acid reference substance solution;
The preparation of need testing solution is carried out uniform sampling according to 2005 editions appendix high performance liquid chromatography tests of Chinese Pharmacopoeia to sample, claims sample, precision takes by weighing mistake 50 each 500mg of mesh sieve Herba Xanthii grass medicinal powder that gather in 5-10 month, puts in the 250ml round-bottomed flask, adds 25ml50% methanol, weigh, 85 ℃ of reflux 1h place room temperature, supply original weight with 50% methanol, pipette supernatant after centrifugal, cross 0.45 μ m filter membrane, place brown sample bottle, as need testing solution;
Accurate reference substance and the need testing solution 20 μ l of drawing of assay inject chromatograph of liquid, measure, promptly;
According to chlorogenic acid content meter in the Herba Xanthii grass dry product, be no less than setting value for qualified.
The method of quality control of above-mentioned Herba Xanthii grass, chlorogenic acid content meter in the Herba Xanthii grass dry product, setting value is 0.30%.
Beneficial effect:
The present invention has proposed method of quality control to Herba Xanthii grass, this method provides the content of chlorogenic acid composition in discriminating and the high effective liquid chromatography for measuring medicine, quality standard after the raising can be controlled the quality of medicine better, method of quality control of the present invention, have stronger specificity and good repeatability, real embodiment drug safety is effective, quality controllable, determines the Herba Xanthii grass the best of gathering May.
Description of drawings
Fig. 1 is the chlorogenic acid high-efficient liquid phase chromatogram, and wherein a is a sample solution high-efficient liquid phase chromatogram in May; B is a chlorogenic acid reference substance solution high-efficient liquid phase chromatogram.Fig. 2 investigates figure for linear relationship.Fig. 3 is a Different Harvesting Time chlorogenic acid content comparison diagram.
The specific embodiment
In conjunction with the specific embodiment, the present invention is further described as follows:
1, discrimination method is:
1.1 medical material, instrument and reagent
Crude drug source: the Herba Xanthii grass medical material picks up from the Xuzhou, is accredited as the dry aerial parts of feverfew Herba Xanthii Xantliiurn sihiricum Patr. through professor Chen Jianwei of pharmaceutical college of Nanjing University of Traditional Chinese Medicine.It is standby to pulverize 100 mesh sieves.Medical material situation and collecting time see Table 1-1:
Table 1-1 sample ordinary circumstance
The automatic uv analyzer of ZF-20C camera bellows formula (the Shanghai Golconda turns round and look at village electric light instrument plant);
(lot number: 121168-200604) purchase in Nat'l Pharmaceutical ﹠ Biological Products Control Institute, Herba Xanthii booth reference substance is provided by the Zhang Shujun professor of chemical engineering institute of Qiqihar University the Fructus Xanthii control medicinal material, tlc silica gel G plate Haiyang Chemical Plant, Qingdao;
Sodium hydroxide is analytical pure (the sincere chemical reagent company limited in Shanghai, a lot number: 20090227);
Ethyl acetate is analytical pure (Chemical Reagent Co., Ltd., Sinopharm Group, a lot number: T20090306);
Toluene is analytical pure (Nanjing Chemistry Reagent Co., Ltd., lot number: 080910759);
Glacial acetic acid is analytical pure (Chemical Reagent Co., Ltd., Sinopharm Group, a lot number: T20090105);
Concentrated sulphuric acid is analytical pure (Shanghai hundred million chemical reagent company limiteies of a specified duration, a lot number: 091020);
Ethanol is analytical pure (Nanjing Chemistry Reagent Co., Ltd., lot number: 09081310771);
Ether is analytical pure (Shanghai pilot scale chemical corp, a lot number: 20100315);
Cyclohexane extraction is analytical pure (Shishewei Chemical Co., Ltd., Shanghai, a lot number: 0904101);
Formic acid is analytical pure (Shanghai Ling Feng chemical reagent company limited, lot number: 041101);
Water is distilled water.
1.2 experimental technique
A: No. 1 sample powder of perusal is a yellow green, and 40 power microscopes are observed down, sees that the epidermis cell anticlinal wall is arranged is more straight or be shallow wavy and polygon; The pore infinitive, visible more glandular hair and nonglandular hair; The fiber bunchy, the visible pit that has, diameter 15-40 μ m; Needle-like calcium oxalate crystal diameter 5-50 μ m and calcium oxalate cluster crystal diameter 10-30 μ um; Secretory canal schizogenesis formula contains light brown secretions, diameter 20-80 μ m; Can see screw thread, bordered pit vessel, diameter 10-80 μ m;
B: get sample powder 2g No. 1, add 10% sodium hydroxide solution 50ml, reflux 30min, put coldly, filter, filtrate is used ethyl acetate extraction, each 50ml, 2 times, combined ethyl acetate liquid washes with water to neutrality, acetic acid ethyl fluid is concentrated into about 2ml, as need testing solution, other gets Fructus Xanthii control medicinal material 2g, shines medical material solution in pairs with legal system, draw each 10 μ l of above-mentioned two kinds of solution respectively, point is that toluene-ethyl acetate-glacial acetic acid of 16: 4: 1 is developing solvent with volume ratio on same silica gel g thin-layer plate, launches, take out, dry, inspect under the daylight, place under the 365nm ultraviolet light and inspect; Spray 105 ℃ of heating colour developings of 10% sulfuric acid solution, sample and Fructus Xanthii control medicinal material are equipped with same blob in identical bits;
C: get sample powder 5g No. 1,50ml adds diethyl ether, reflux 1h, put cold, filter, filtrate low temperature is waved to 2ml, as need testing solution, other gets Herba Xanthii booth reference substance, make the 1mg/ml diethyl ether solution and compare liquid, draw above-mentioned two kinds of each 5ul of solution respectively, put respectively on same silica gel g thin-layer plate, with volume ratio is that cyclohexane extraction-ethyl acetate-formic acid of 10: 5: 0.2 is developing solvent, launch, take out, dry, put under the daylight respectively and spray 105 ℃ of heating of 10% sulfuric acid solution and develop the color, sample and Herba Xanthii booth standard substance are equipped with same blob in identical bits.
2. the chlorogenic acid contents assay method is:
2.1 medical material, instrument and reagent
Crude drug source
The Herba Xanthii grass medical material picks up from the Xuzhou, is accredited as the dry aerial parts of feverfew Herba Xanthii Xantliiurn sihiricum Patr. through professor Chen Jianwei of pharmaceutical college of Nanjing University of Traditional Chinese Medicine.It is standby to pulverize 100 mesh sieves.Medical material situation and collecting time see Table 1-1:
Table 1-1 sample ordinary circumstance
Agilent1100 type high performance liquid chromatograph (comprising online degasser, quaternary gradient pump, automatic sampler, column oven, diode array detector), Mettler Toledo AG285 100,000/balance, medical supersonic washer (KQ-500E type, Kunshan ultrasonic instrument company limited);
Chromatograph methanol (production of U.S. Tedia company), water is the WAHAHA pure water; Analyze methanol (Nanjing Chemistry Reagent Co., Ltd., lot number: 09020310079), (1Guanghua Chemical Plant Co., Ltd., Guangdong, lot number: 20071101), the chlorogenic acid reference substance is purchased the (lot number: 110753-200413) in Nat'l Pharmaceutical ﹠ Biological Products Control Institute to analyze phosphoric acid.
2.2 method and result
2.2.1 measure the selection of wavelength
It is an amount of to get the chlorogenic acid reference substance, adds 50% methanol and makes the solution that concentration is 0.443mg/ml, and in the scanning of 200～400nm wave-length coverage, the maximum absorption wavelength λ max that records chlorogenic acid is 326nm.
2.2.2 chromatographic condition
Chromatographic column: diameter and length are 4.6mm * 250mm, the Alltima of 5 μ m
TMC18, mobile phase: the methanol of equal-volume ratio and 0.1% phosphoric acid mixed aqueous solution, flow velocity: 0.8ml/min, column temperature: 25 ℃, detect wavelength: 326nm, sample size: 20 μ l, sampling time: 60min;
2.2.3 the preparation of reference substance solution
Precision takes by weighing chlorogenic acid reference substance 4.43mg, adds 50% methanol and makes the solution that concentration is 0.443mg/ml, as the chlorogenic acid reference substance solution.
2.2.4 the preparation of need testing solution
According to 2005 editions appendix high performance liquid chromatography tests of Chinese Pharmacopoeia, sample is carried out uniform sampling, claim sample.Precision takes by weighing each 500mg of Herba Xanthii grass medicinal powder (crossing 50 mesh sieves) that gathers in above-mentioned 5-10 month, puts in the 250ml round-bottomed flask, adds 25ml50% methanol, weighs, and 85 ℃ of reflux 1h place room temperature, supply original weight with 50% methanol.Pipette supernatant after centrifugal, cross 0.45 μ m filter membrane, place brown sample bottle, as need testing solution.
2.2.5 linear relationship is investigated
Precision takes by weighing the chlorogenic acid reference substance solution, and accurate respectively 0.025ml, 0.05ml, 0.1ml, 0.2ml, 0.5ml, 1.0ml, the 2.0ml of drawing places the 10ml measuring bottle, add 50% methanol to scale, shake up, sample introduction 20 μ l, the record chromatogram sees accompanying drawing 1 for details, and linear relationship the results are shown in accompanying drawing 2.
By last figure as seen, chlorogenic acid is in 1.1075～88.6 μ g/ml scopes, and its chromatographic peak area (A) is linear with concentration (C), and regression equation is A=42.509C-19.445 (R
2=0.9999).
2.2.6 precision test
Precision takes by weighing medicinal powder 100mg No. 2, presses the preparation method preparation of need testing solution, presses above-mentioned chromatographic condition continuous sample introduction 6 times, the peak area of asking, and calculating RSD is 0.81%, illustrates that the precision of instrument is good, the precision data see Table 4-2.
Table 4-2 precision data (n=6)
2.2.7 stability test
Precision takes by weighing medicinal powder 100mg No. 2, press the preparation method preparation of need testing solution, press above-mentioned chromatographic condition sample introduction at 0h, 2h, 4h, 6h, 8h, 10h, 12h, 24h respectively, RSD is 1.47%, illustrate that test sample is basicly stable in 24h, concrete stability data sees Table 4-3.
Table 4-3 stability data (n=8)
2.2.8 replica test
Get medicinal powder No. 2, the preparation method of pressing need testing solution, 5 parts of need testing solutions of system, accurate sample introduction 20.0 μ l measure according to said method respectively, and calculating RSD is 1.73%, illustrates that this method repeatability is good, and concrete data see Table 4-4.
Table 4-4 repeatability data (n=5)
2.2.9 average recovery
Precision takes by weighing medicinal powder 500mg No. 2, totally 6 parts, place the 250ml round-bottomed flask, the accurate 50% methanol 25ml that adds adds 44.3 μ g/ml chlorogenic acid contrast solution 1.0ml, presses the operation of need testing solution preparation method, measure in the 326nm place, average recovery rate is 97.33% as a result, and RSD is 1.76% (n=6), the results are shown in Table 4-5.
Table 4-5 chlorogenic acid average recovery experiment (n=6)
2.2.10 sample size is measured
Precision takes by weighing each 500mg of medicinal powder 1～No. 6, and 3 parts, be respectively 1-1,1-2,1-3, surplus and the like, press the operation of need testing solution preparation method, sample introduction under above chromatographic condition the results are shown in Table 4-6, and the Different Harvesting Time chlorogenic acid content is relatively seen accompanying drawing 3.
Table 4-6 determination of chlorogenic acid (n=3)
Claims (2)
1. the method for quality control of a Herba Xanthii grass comprises and differentiating and the chlorogenic acid contents assay method, it is characterized in that described discrimination method is:
A: perusal Herba Xanthii grass powder is a yellow green, and 40 power microscopes are observed down, sees that the epidermis cell anticlinal wall is arranged is more straight or be shallow wavy and polygon; The pore infinitive, visible more glandular hair and nonglandular hair; The fiber bunchy, the visible pit that has, diameter 15-40 μ m; Needle-like calcium oxalate crystal diameter 5-50 μ m and calcium oxalate cluster crystal diameter 10-30 μ um; Secretory canal schizogenesis formula contains light brown secretions, diameter 20-80 μ m; Can see screw thread, bordered pit vessel, diameter 10-80 μ m;
B: get Herba Xanthii grass powder 2g, add 10% sodium hydroxide solution 50ml, reflux 30min, put coldly, filter, filtrate is used ethyl acetate extraction, each 50ml, 2 times, combined ethyl acetate liquid washes with water to neutrality, acetic acid ethyl fluid is concentrated into about 2ml, as need testing solution, other gets Fructus Xanthii control medicinal material 2g, shines medical material solution in pairs with legal system, draw each 10 μ l of above-mentioned two kinds of solution respectively, point is that toluene-ethyl acetate-glacial acetic acid of 16: 4: 1 is developing solvent with volume ratio on same silica gel g thin-layer plate, launches, take out, dry, inspect under the daylight, place under the 365nm ultraviolet light and inspect; Spray 105 ℃ of heating colour developings of 10% sulfuric acid solution, sample and Fructus Xanthii control medicinal material are equipped with same blob in identical bits;
C: get Herba Xanthii grass powder 5g, 50ml adds diethyl ether, reflux 1h, put cold, filter, filtrate low temperature is waved to 2ml, as need testing solution, other gets Herba Xanthii booth reference substance, make the 1mg/ml diethyl ether solution and compare liquid, draw above-mentioned two kinds of each 5ul of solution respectively, put respectively on same silica gel g thin-layer plate, with volume ratio is that cyclohexane extraction-ethyl acetate-formic acid of 10: 5: 0.2 is developing solvent, launch, take out, dry, put under the daylight respectively and spray 105 ℃ of heating of 10% sulfuric acid solution and develop the color, sample and Herba Xanthii booth standard substance are equipped with same blob in identical bits;
Described chlorogenic acid contents assay method is:
The chromatographic condition chromatographic column: diameter and length are 4.6mm * 250mm, the Alltima of 5 μ m
TMC18, mobile phase: the methanol of equal-volume ratio and 0.1% phosphoric acid mixed aqueous solution, flow velocity: 0.8ml/min, column temperature: 25 ℃, detect wavelength: 326nm, sample size: 20 μ l, sampling time: 60min;
It is an amount of that the preparation precision of reference substance solution takes by weighing the chlorogenic acid reference substance, adds 50% methanol and make the solution that concentration is 0.4mg/ml, as the chlorogenic acid reference substance solution;
The preparation of need testing solution is carried out uniform sampling according to 2005 editions appendix high performance liquid chromatography tests of Chinese Pharmacopoeia to sample, claims sample, precision took by weighing 50 each 500mg of mesh sieve Herba Xanthii grass medicinal powder, put in the 250ml round-bottomed flask, added 25ml50% methanol, weigh, 85 ℃ of reflux 1h place room temperature, supply original weight with 50% methanol, pipette supernatant after centrifugal, cross 0.45 μ m filter membrane, place brown sample bottle, as need testing solution;
Accurate reference substance and the need testing solution 20 μ l of drawing of assay inject chromatograph of liquid, measure, promptly;
According to chlorogenic acid content meter in the Herba Xanthii grass dry product, be no less than setting value for qualified.
2. according to the method for quality control of the described Herba Xanthii grass of claim 1, it is characterized in that according to chlorogenic acid content meter in the Herba Xanthii grass dry product, setting value is 0.30%.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103193742A (en) * | 2013-04-12 | 2013-07-10 | 南京中医药大学 | Xanthatin derivative and medicine use thereof |
| CN108918721A (en) * | 2018-08-01 | 2018-11-30 | 山西大学 | A kind of measuring method of tussilago Content of Chlorogenic Acid |
| CN114199818A (en) * | 2021-12-02 | 2022-03-18 | 广东一方制药有限公司 | Construction method of near-infrared quantitative detection model of cocklebur fruit traditional Chinese medicine formula particles and quantitative detection method thereof |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1053364A (en) * | 1991-03-02 | 1991-07-31 | 张玉昆 | The preparation method of xanthium sibiricum volatile oil pharmaceutical |
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2011
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Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1053364A (en) * | 1991-03-02 | 1991-07-31 | 张玉昆 | The preparation method of xanthium sibiricum volatile oil pharmaceutical |
Non-Patent Citations (2)
| Title |
|---|
| 《徐州医学院学报》 20100930 徐圣秋等 UV法测定不同采收期苍耳草中总酚酸含量 第574-576页 1-2 第30卷, 第9期 2 * |
| 《江西中医学院学报》 19940228 陈根顺等 苍耳草的生药组织鉴定 第40-41页 1-2 第6卷, 第2期 2 * |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103193742A (en) * | 2013-04-12 | 2013-07-10 | 南京中医药大学 | Xanthatin derivative and medicine use thereof |
| CN103193742B (en) * | 2013-04-12 | 2014-08-13 | 南京中医药大学 | Xanthatin derivative and medicine use thereof |
| CN108918721A (en) * | 2018-08-01 | 2018-11-30 | 山西大学 | A kind of measuring method of tussilago Content of Chlorogenic Acid |
| CN114199818A (en) * | 2021-12-02 | 2022-03-18 | 广东一方制药有限公司 | Construction method of near-infrared quantitative detection model of cocklebur fruit traditional Chinese medicine formula particles and quantitative detection method thereof |
| CN114199818B (en) * | 2021-12-02 | 2024-05-14 | 广东一方制药有限公司 | Construction method of near infrared quantitative detection model of fructus xanthil traditional Chinese medicine formula particles and quantitative detection method thereof |
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