CN101732405B - Common lamiophlomis extract, medicine composition containing same and quality control method - Google Patents
Common lamiophlomis extract, medicine composition containing same and quality control method Download PDFInfo
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- CN101732405B CN101732405B CN 200910225156 CN200910225156A CN101732405B CN 101732405 B CN101732405 B CN 101732405B CN 200910225156 CN200910225156 CN 200910225156 CN 200910225156 A CN200910225156 A CN 200910225156A CN 101732405 B CN101732405 B CN 101732405B
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Abstract
The invention provides a common lamiophlomis extract which contains 1.5-30mg/g of shanzhiside methylester. The invention also provides a medicine composition containing the common lamiophlomis extract and a method for controlling the quality of the common lamiophlomis extract or the medicine composition containing the common lamiophlomis extract. The invention has stable medicine effect of the extract, definite medicine effect and strong controllability and controls the quality of the extract by taking the shanzhiside methylester as an index component. The quality control method is accurate, reliable, stable and controllable, is used for measuring the content of the shanzhiside methylester in a common lamiophlomis medicinal material or a preparation and can effectively monitor the quality of a common lamiophlomis medicine.
Description
Technical field
The present invention relates to a kind of common lamiophlomis root extract and contain pharmaceutical composition and the method for quality control of this extract, belong to drug world.
Background technology
Lamiophlomis rotata is Tibetan's conventional crude drugs, for the root of labiate lamiophlomis rotata Lamio-phlomis rotata (Benth.) Kudo and root-like stock, herb, grows on height above sea level the 3000m above plateau or high mountain.The lamiophlomis rotata cold nature, bitter, slightly poisonous; Effect: invigorate blood circulation, the row stagnation resolvation is swollen, pain relieving; Cure mainly and get injured by a fall muscles and bones, sprain one's back and frustrate gas, yellow water is amassed in the joint.Its chemical composition has: flavones ingredient, iridoid, phenylethyl alcohol glycoside etc., the pharmacologically active of report has, analgesic activity, anastalsis, antiinflammatory action, antitumor, impact (seeing Zhang Feng etc., the chemical composition of lamiophlomis rotata and pharmacological action, pharmacy practice magazine the 3rd phase of the 26th volume in 2008) on bone marrow granulocyte CFU-GM (CFU-D).
About the method for quality control of lamiophlomis rotata and preparation, " Chinese Pharmacopoeia 2005 version one one (the 184th and 541 page) has been recorded lamiophlomis rotata medicinal material and Radix Lamiophlomidis Rotatae capsule, has stipulated the test item of flavones ingredient (cyanidenon and/or general flavone).Many all institutes, the compound that obtains pure useful value from plant is very difficult, and cost is very high, simultaneously also for fear of the spinoff that is difficult to expect, the general effective pure compound that does not use in the plant, and use medicinal plant extract direct effect medicinal raw material, cost is low and use easy.But a defective of plant extracts is, because the plant collecting season fluctuations that all can cause active constituent content different from the place of production, simultaneously, if main effective constituent and content thereof are unclear, very large difficulty is caused to the quality control of medicine in the capital, therefore, in order to keep the steady quality of autonomic drug, clearly know in the plant that each basic effective component and content thereof are very important.Because flavones ingredient is not to be one of main active in the common lamiophlomis root preparation, therefore only by the quality control to flavones ingredient, still can not monitor the quality of common lamiophlomis root preparation fully.
Mountain Cape jasmine glycosides methyl esters is one of iridoid in the lamiophlomis rotata, and the people such as Zhang Chengzhong (iridoid glycoside in the Tibet medicine lamivphlomis root. Chinese herbal medicine .1992 the 10th phase the 509th of the 23rd volume, 510 and 560 pages) from Tibet medicine lamivphlomis root, obtain first mountain Cape jasmine glycosides methyl esters.Because complex chemical composition in the lamiophlomis rotata, can reflect by certain composition wherein the quality of lamiophlomis rotata medicinal material or extract, for a person skilled in the art, be difficult to predict.The relevant report that also mountain Cape jasmine glycosides methyl esters is not used at present quality control index.
Summary of the invention
Technical scheme of the present invention has provided a kind of common lamiophlomis root extract, and another technical scheme of the present invention has provided pharmaceutical composition and the method for quality control thereof that contains this extract.
The invention provides a kind of common lamiophlomis root extract, it contains mountain Cape jasmine glycosides methyl esters 1.5-30mg/g.Preferably, it contains mountain Cape jasmine glycosides methyl esters 12.5-30mg/g.Further preferably, it contains mountain Cape jasmine glycosides methyl esters 18.1-30mg/g.
Wherein, described lamiophlomis rotata derives from the root of labiate lamiophlomis rotata Lamiophlomis rotata (Benth.) Kudo and root-like stock, herb.
Common lamiophlomis root extract of the present invention is water or the extractive with organic solvent of lamiophlomis rotata medicinal material.Described organic solvent is 5-100% ethanol.
The present invention also provides a kind of pharmaceutical composition, and it contains described common lamiophlomis root extract is active component, adds the medicament that pharmaceutically acceptable auxiliary material or complementary composition are prepared from.
Wherein, described medicament is capsule, tablet, granule, pill, oral liquid, soft capsule or pill.
The present invention also provides a kind of method of controlling lamiophlomis rotata medicinal material or extract or containing the drug regimen amount of common lamiophlomis root extract, be to carry out quality control take mountain Cape jasmine glycosides methyl esters as index components, the content that wherein contains mountain Cape jasmine glycosides methyl esters in the common lamiophlomis root extract is 1.5-30mg/g; Wherein preferably, the content of mountain Cape jasmine glycosides methyl esters is 12.5-30mg/g; Still more preferably, the content of mountain Cape jasmine glycosides methyl esters is 18.1-30mg/g.
It is take mountain Cape jasmine glycosides methyl esters as index components, carries out quality control, comprises the steps:
A, by high effective liquid chromatography for measuring, adopt reverse-phase chromatographic column, mobile phase is comprised of organic phase and water, it is 220-250nm that ultraviolet detects wavelength;
B, get mountain Cape jasmine glycosides methyl esters reference substance, add Methanol and become reference substance solution;
C, get lamiophlomis rotata medicinal material or preparation, make need testing solution;
D, accurate reference substance solution and the need testing solution injection liquid chromatography drawn of difference are measured, and be get final product.
Wherein, the described reverse-phase chromatographic column of step a is take octadecylsilane chemically bonded silica as filling agent; Described organic phase is methyl alcohol or acetonitrile, and described water is pure water, phosphoric acid solution or acetum; Described detection wavelength is 235nm; Reference substance solution described in the step b is the solution that every 1ml contains 0.02-0.40mg mountain Cape jasmine glycosides methyl esters; The preparation of the described need testing solution of step c may further comprise the steps: get lamiophlomis rotata medicinal material, extract or preparation 0.05-1.0g, and accurately weighed, put in the tool plug conical flask, accurate 50-100% methanol solution 25ml, close plug, the weighed weight of adding, add hot reflux 30-90 minute, let cool, weighed weight is again supplied the weight of less loss with the 50-100% methanol solution, shake up, filter, get subsequent filtrate, and get final product.
Wherein, the described methyl alcohol of step a and described pure water volume ratio are 10: 90-30: 70.
Wherein, the described methyl alcohol of step a and described pure water volume ratio are 20: 80.
Wherein, the described concentration of methanol solution of step c is 70%.
Extract efficacy stability of the present invention, controllability is strong.By mountain Cape jasmine glycosides methyl esters is controlled the quality of extract as index components, drug effect is clear and definite, method of quality control of the present invention accurately and reliably, stablize controlledly, the content that uses the method to measure mountain Cape jasmine glycosides methyl esters in lamiophlomis rotata medicinal material or the preparation can be monitored the quality of lamiophlomis rotata medicine effectively.
Below the present invention is described in further detail by embodiment, but do not limit the present invention, those skilled in the art can make according to the present invention various changes and distortion, only otherwise break away from spirit of the present invention, all should belong to the scope of claims of the present invention.
Description of drawings
Fig. 1 reference substance mountain Cape jasmine glycosides methyl esters HPLC collection of illustrative plates
Fig. 2 lamiophlomis rotata medicinal material HPLC collection of illustrative plates
Fig. 3 mountain Cape jasmine glycosides methyl esters canonical plotting
Fig. 4 Radix Lamiophlomidis Rotatae capsule HPLC of the present invention collection of illustrative plates
Embodiment
The preparation of embodiment 1 extract of the present invention
Get lamiophlomis rotata medicinal material 1000g, pulverize boiling 3 times, the 1st time 2 hours, 2nd, be respectively for 3 times 1 hour, each amount of water is 8 times, and collecting decoction filters, the clear cream of filtrate simmer down to relative density 1.30 (85 ℃) dry below 80 ℃, is pulverized, and gets cream.
The preparation of embodiment 2 extracts of the present invention
Get lamiophlomis rotata medicinal material 1000g, pulverize, add 20 times of water rettings 24 hours, water temperature is 85 ℃, filter, and the clear cream of filtrate simmer down to relative density 1.30 (85 ℃), drying is pulverized, and gets cream.
The preparation of embodiment 3 extracts of the present invention
Get lamiophlomis rotata medicinal material 1000g, pulverize, in the pressure extractor, add the water pressurised extraction 3 times, the 1st time 2 hours, be respectively 1 hour for the 2nd, 3 time, each amount of water is 8 times, and temperature is 120 ℃, merges extract, filters, and is concentrated, and drying is pulverized, and gets cream.
The preparation of embodiment 4 extracts of the present invention
Get lamiophlomis rotata medicinal material 1000g, pulverize, boiling 3 times, the 1st time 2 hours, be respectively 1 hour for the 2nd, 3 time, each amount of water is 8 times, collecting decoction filters, and is concentrated, and adding ethanol to solution, to contain the alcohol amount be 30%, left standstill 12 hours, get supernatant, concentrated, drying is pulverized, and gets cream.
The preparation of embodiment 5 extracts of the present invention
Get lamiophlomis rotata medicinal material 1000g, pulverize, add 20 times of amount 70% ethanol percolations and extract, the extract Recycled ethanol, concentrated, drying is pulverized, and gets cream.
The preparation of embodiment 6 extracts of the present invention
Get lamiophlomis rotata medicinal material 1000g, pulverize, add 20 times of amount 30% ethanol percolations and extract, the extract Recycled ethanol, concentrated, drying is pulverized, and gets cream.
The preparation of embodiment 7 extracts of the present invention
Get lamiophlomis rotata medicinal material 1000g, pulverize, add 8 times of amount 70% alcohol refluxs and extract 3 times, the extract Recycled ethanol, concentrated, drying is pulverized, and gets cream.
The preparation of embodiment 8 medicine capsules of the present invention
With the medicinal extract of embodiment 1 preparation, dry below 80 ℃, add an amount of starch, granulation, drying incapsulates, and makes 1000, and get final product.
The preparation of embodiment 9 medicinal soft capsule agent of the present invention
Get soybean oil 300g and be heated to 80 degree, add beeswax 30g, it is dissolved, cool to 40 ℃ of adding embodiment 2 medicinal extract fine powders, mixing is pressed into soft capsule, is drying to obtain 1000 of Duyiwei soft capsules.
The selection of chromatographic condition:
1. detect the selection of wavelength
Mountain Cape jasmine glycosides methyl esters reference substance solution is carried out absorbing wavelength scanning, and the result: mountain Cape jasmine glycosides methyl esters has absorption maximum at 235nm wavelength place, and test selects 235nm for detecting wavelength.
2. the selection of mobile phase
The test ginseng was once selected 1. methyl alcohol-0.4% phosphoric acid solution (20: 80) of mobile phase; 2. acetonitrile-water (11: 89); 3. methanol-water (18: 82), 4. methanol-water (20: 80) carries out assay to the mountain Cape jasmine glycosides methyl esters in lamiophlomis rotata medicinal material and the Radix Lamiophlomidis Rotatae capsule, all obtains the symmetrical chromatographic peak that mountain Cape jasmine glycosides methyl esters peak base separates in the sample.Yet, consider that long-term use acid solution can reduce chromatographic column post effect, it is high that acetonitrile pollutes large cost, therefore methodological study is take methanol aqueous solution as mobile phase, 3. 4. appearance time is late than condition for condition, and selected peak shape 4. methanol-water (20: 80) good, that cost is low, appearance time is suitable is that mobile phase is for further study at last.
3. the investigation of chromatographic column
Test was once selected: 1. Dikma platisil C18 (250mm * 4.6mm, 5um), 2. Phenomenex Luna C18 (150mm * 4.6mm, 5um), 3. Alltima C18 (250mm * 4.6mm, 5 μ m) chromatographic column of three brands is carried out assay to the mountain Cape jasmine glycosides methyl esters in the lamiophlomis rotata medicinal material, all obtain the symmetrical chromatographic peak that mountain Cape jasmine glycosides methyl esters peak base separates in the sample, degree of separation is greater than 1.5, theoretical cam curve is calculated greater than 3000, therefore the method scope of application is wider by Cape jasmine glycosides methyl esters peak, mountain.Methodological study is take Dikma platisil C18 (250mm * 4.6mm, 5um) as analytical column.As shown in Figure 1.
4. the preparation of need testing solution
This test is studied the preparation method of need testing solution, and extracting method, the factors such as solvent, extraction time of extracting are investigated, and test method and result are as follows.The medicinal material lot number is: 0821101.
4.1 extracting method is selected
Investigated the lab analysis conventional method: add hot reflux and ultrasonic extracting method.
1. heating and refluxing extraction is got approximately 0.3g of this product powder (crossing sieve No. three), and is accurately weighed, puts in the tool plug conical flask accurate methyl alcohol 25ml, the close plug of adding, weighed weight added hot reflux 60 minutes, let cool, and weighed weight is again supplied the weight of less loss with methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.
2. approximately 0.3g of this product powder (crossing sieve No. three) is got in ultrasonic extraction, and is accurately weighed, puts in the tool plug conical flask, accurate methyl alcohol 25ml, close plug, the weighed weight of adding, ultrasonic processing (power 250W, frequency 50kHz) 60 minutes lets cool, weighed weight again, supply the weight of less loss with methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.Measure by above-mentioned chromatographic condition, measurement result is as follows.
Table 1 extracting method is investigated test findings
| Extracting method | Added hot reflux 60 minutes | Ultrasonic processing 60 minutes |
| Mountain Cape jasmine glycosides methyl esters content (mg/g) | 3.34 | 3.27 |
[0061]Therefore heating and refluxing extraction slightly is better than ultrasonic processing to the extraction of mountain Cape jasmine glycosides methyl esters in the lamiophlomis rotata medicinal material, therefore heating and refluxing extraction is selected in test.
4.2 extracting solvent selects
Investigated conventional reagent: the methanol aqueous solution of methyl alcohol and variable concentrations.
1. methyl alcohol is got approximately 0.3g of this product powder (crossing sieve No. three), and is accurately weighed, puts in the tool plug conical flask accurate methyl alcohol 25ml, the close plug of adding, weighed weight added hot reflux 60 minutes, let cool, and weighed weight is again supplied the weight of less loss with methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.
2. 70% methanol solution is got approximately 0.3g of this product powder (crossing sieve No. three), and is accurately weighed, puts in the tool plug conical flask, accurate 70% methanol solution 25ml, close plug, the weighed weight of adding, added hot reflux 60 minutes, let cool, weighed weight is again supplied the weight of less loss with 70% methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.
3. 30% methanol solution is got approximately 0.3g of this product powder (crossing sieve No. three), and is accurately weighed, puts in the tool plug conical flask, accurate 30% methanol solution 25ml, close plug, the weighed weight of adding, added hot reflux 60 minutes, let cool, weighed weight is again supplied the weight of less loss with 30% methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.Measure by above-mentioned chromatographic condition, measurement result is as follows.
Table 2 extracts solvent and investigates test findings
| Extract solvent | Methyl alcohol | 70% methyl alcohol | 30% methyl alcohol |
| Mountain Cape jasmine glycosides methyl esters content (mg/g) | 3.34 | 3.36 | 3.13 |
Therefore, best to the extraction efficiency of mountain Cape jasmine glycosides methyl esters in the lamiophlomis rotata medicinal material with 70% methanol solution in three kinds of extraction solvents, therefore 70% methanol solution is selected in test.
4.3 extraction time is selected
Under the condition of determining extracting method, extraction solvent, extraction time is investigated.
1. get approximately 0.3g of this product powder (crossing sieve No. three), accurately weighed, put in the tool plug conical flask accurate 70% methanol solution 25ml, the close plug of adding, weighed weight added hot reflux 30 minutes, let cool, and weighed weight is again supplied the weight of less loss with 70% methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.
2. get approximately 0.3g of this product powder (crossing sieve No. three), accurately weighed, put in the tool plug conical flask accurate 70% methanol solution 25ml, the close plug of adding, weighed weight added hot reflux 60 minutes, let cool, and weighed weight is again supplied the weight of less loss with 70% methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.
3. get approximately 0.3g of this product powder (crossing sieve No. three), accurately weighed, put in the tool plug conical flask accurate 70% methanol solution 25ml, the close plug of adding, weighed weight added hot reflux 90 minutes, let cool, and weighed weight is again supplied the weight of less loss with 70% methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.Measure by above-mentioned chromatographic condition, measurement result is as follows.
Table 3 extraction time investigation test findings
| Extraction time (min) | 30 | 60 | 90 |
| Mountain Cape jasmine glycosides methyl esters content (mg/g) | 3.14 | 3.36 | 3.34 |
Therefore, extract after 60 minutes in the sample content of mountain Cape jasmine glycosides methyl esters substantially unchanged, historical facts or anecdotes is tested and is selected to add hot reflux 60 minutes.
In sum, the preparation method of need testing solution is: get approximately 0.3g of this product powder (crossing sieve No. three), and accurately weighed, put in the tool plug conical flask accurate 70% methanol solution 25ml, the close plug of adding, weighed weight added hot reflux 60 minutes, let cool, weighed weight again, supply the weight of less loss with 70% methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.
The content results of measuring according to the method described above Cape jasmine glycosides methyl esters in mountain in 08070546,08080358,08,090,943 3 batch of medicinal material is as follows, collection of illustrative plates as shown in Figure 2:
| The medicinal material lot number | 08070546 | 08080358 | 08090943 |
| Mountain Cape jasmine glycosides methyl esters content (mg/g) | 8.7 | 6.8 | 5.9 |
The assay of mountain Cape jasmine glycosides methyl esters in embodiment 11 common lamiophlomis root extracts
1, instrument and reagent
Instrument: Agilent 1100 high performance liquid chromatographs and auxiliary work station
Sartorius CP225D electronic balance Sartorius BS124S electronic balance
Mountain Cape jasmine glycosides methyl esters reference substance: self-control, purity is 97.7%;
Reagent: chromatographic grade methyl alcohol (available from U.S. Fisher chemical reagents corporation), water is double distilled water (self-control); It is pure that other reagent is analysis.
2, chromatographic condition
Detect wavelength: 235nm
Mobile phase: methanol-water solution (20: 80)
Chromatographic column: take octadecylsilane chemically bonded silica as filling agent.
3, assay
Measure according to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 D).
Chromatographic condition and system suitability: octadecylsilane chemically bonded silica is filling agent, and take methanol-water (20: 80) as mobile phase, it is 235nm that ultraviolet detects wavelength, and number of theoretical plate calculates by Cape jasmine glycosides methyl esters peak, mountain should be not less than 3000.
The preparation of reference substance solution: it is an amount of that precision takes by weighing mountain Cape jasmine glycosides methyl esters reference substance, adds methyl alcohol and make the solution that every 1ml contains 0.1mg, shakes up, and get final product.
The preparation of need testing solution: get 08070546,08080358,08,090,943 3 batch of medicinal material, adopt respectively the described 7 kinds of extracting method of above embodiment 1-7 to extract, obtain medicinal extract sample 1-21, get respectively 0.15g measures mountain Cape jasmine glycosides methyl esters by the method for embodiment 10 content, the every batch of replicate determination 2 times, test findings such as table 4.
The content of mountain Cape jasmine glycosides methyl esters in table 4 common lamiophlomis root extract
The assay of mountain Cape jasmine glycosides methyl esters in embodiment 12 Radix Lamiophlomidis Rotatae capsules
1 instrument and reagent
Instrument: Agilent 1100 high performance liquid chromatographs and auxiliary work station; LC-2010 high performance liquid chromatograph and auxiliary work station.
Sartorius CP225D electronic balance Sartorius BS124S electronic balance
Mountain Cape jasmine glycosides methyl esters reference substance: self-control or commercially available;
Reagent: chromatographic grade methyl alcohol, acetonitrile are chromatographically pure (available from U.S. Fisher chemical reagents corporation), and water is double distilled water (self-control); It is pure that other reagent is analysis.
2, chromatographic condition
2.1 detect the selection of wavelength
Mountain Cape jasmine glycosides methyl esters reference substance solution is carried out absorbing wavelength scanning, and the result: mountain Cape jasmine glycosides methyl esters has absorption maximum at 235nm wavelength place, and test selects 235nm for detecting wavelength.
2.2 the selection of mobile phase
1. methyl alcohol-0.4% phosphoric acid solution (20: 80) of mobile phase is selected in test; 2. acetonitrile-water (11: 89); 3. methanol-water (18: 82), 4. methanol-water (20: 80) carries out assay to the mountain Cape jasmine glycosides methyl esters in the Radix Lamiophlomidis Rotatae capsule, obtains the symmetrical chromatographic peak that mountain Cape jasmine glycosides methyl esters peak base separates in the sample.Methodological study take mobile phase 1. methanol-water (20: 80) as mobile phase.
2.3 the investigation of chromatographic column
Test ginseng was once selected: 1. Dikma platisil C18 (250mm * 4.6mm, 5 μ m), 2. Phenomenex Luna C18 (150mm * 4.6mm, 5 μ m), 3. Alltima C18 (250mm * 4.6mm, 5 μ m) chromatographic column of three brands is carried out assay to the mountain Cape jasmine glycosides methyl esters in the Radix Lamiophlomidis Rotatae capsule, all obtain the symmetrical chromatographic peak that mountain Cape jasmine glycosides methyl esters peak base separates in the sample, degree of separation is greater than 1.5, theoretical cam curve is calculated greater than 3000, therefore the method scope of application is wider by Cape jasmine glycosides methyl esters peak, mountain.
Methodological study is analytical column with Dikma platisil C18 (250mm * 4.6mm, 5 μ m).
Reference substance solution preparation method: it is an amount of that precision takes by weighing mountain Cape jasmine glycosides methyl esters reference substance, adds methyl alcohol and make the solution that every 1ml contains 0.1mg, and get final product.
3, the preparation of need testing solution
This test is studied the preparation method of test sample (Radix Lamiophlomidis Rotatae capsule lot number: by the preparation of embodiment 8 methods) solution, and extracting method, the factors such as solvent, extraction time of extracting are investigated, and test method and result are as follows.
3.1 extracting method is selected
Investigated the lab analysis conventional method: add hot reflux and ultrasonic extracting method.
1. heating and refluxing extraction is got this product content, and porphyrize is got approximately 0.2g, and is accurately weighed, puts in the tool plug conical flask, the accurate methyl alcohol 25ml that adds, close plug, weighed weight added hot reflux 60 minutes, let cool, more weighed weight, supply the weight of less loss with methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.
2. this product content is got in ultrasonic extraction, and porphyrize is got approximately 0.2g, accurately weighed, put in the tool plug conical flask the accurate methyl alcohol 25ml that adds, close plug, weighed weight, ultrasonic processing (power 250W, frequency 50kHz) 60 minutes, lets cool, more weighed weight, supply the weight of less loss with methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.Measure by above-mentioned chromatographic condition, measurement result is as follows.
Table 5 extracting method is investigated test findings
| Extracting method | Added hot reflux 60 minutes | Ultrasonic processing 60 minutes |
| Mountain Cape jasmine glycosides methyl esters content (mg/g) | 11.73 | 10.92 |
Therefore heating and refluxing extraction slightly is better than ultrasonic processing to the extraction of mountain Cape jasmine glycosides methyl esters in the Radix Lamiophlomidis Rotatae capsule, therefore heating and refluxing extraction is selected in test.
3.2 extracting solvent selects
Investigated conventional reagent: the methanol aqueous solution of methyl alcohol and variable concentrations.
1. methyl alcohol is got this product content, and porphyrize is got approximately 0.2g, and is accurately weighed, puts in the tool plug conical flask, the accurate methyl alcohol 25ml that adds, close plug, weighed weight added hot reflux 60 minutes, let cool, more weighed weight, supply the weight of less loss with methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.
2. 70% methyl alcohol is got this product content, and porphyrize is got approximately 0.2g, and is accurately weighed, puts in the tool plug conical flask, the accurate 70% methyl alcohol 25ml that adds, close plug, weighed weight added hot reflux 60 minutes, let cool, more weighed weight, supply the weight of less loss with 70% methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.
3. 30% methyl alcohol is got this product content, and porphyrize is got approximately 0.2g, and is accurately weighed, puts in the tool plug conical flask, the accurate 30% methyl alcohol 25ml that adds, close plug, weighed weight added hot reflux 60 minutes, let cool, more weighed weight, supply the weight of less loss with 30% methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.Measure by above-mentioned chromatographic condition, measurement result is as follows.
Table 6 extracts solvent and investigates test findings
| Extract solvent | Methyl alcohol | 70% methyl alcohol | 30% methyl alcohol |
| Mountain Cape jasmine glycosides methyl esters content (mg/g) | 10.82 | 11.73 | 11.54 |
Therefore, best to the extraction efficiency of mountain Cape jasmine glycosides methyl esters in the Radix Lamiophlomidis Rotatae capsule with 70% methyl alcohol in three kinds of extraction solvents, therefore 70% methyl alcohol is selected in test.
3.3 extraction time is selected
Under the condition of determining extracting method, extraction solvent, extraction time is investigated.
1. get this product content, porphyrize is got approximately 0.2g, and is accurately weighed, puts in the tool plug conical flask, the accurate 70% methyl alcohol 25ml that adds, close plug, weighed weight added hot reflux 30 minutes, let cool, more weighed weight, supply the weight of less loss with 70% methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.
2. get this product content, porphyrize is got approximately 0.2g, and is accurately weighed, puts in the tool plug conical flask, the accurate 70% methyl alcohol 25ml that adds, close plug, weighed weight added hot reflux 60 minutes, let cool, more weighed weight, supply the weight of less loss with 70% methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.
3. get this product content, porphyrize is got approximately 0.2g, and is accurately weighed, puts in the tool plug conical flask, the accurate 70% methyl alcohol 25ml that adds, close plug, weighed weight added hot reflux 90 minutes, let cool, more weighed weight, supply the weight of less loss with 70% methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.Measure by above-mentioned chromatographic condition, measurement result is as follows.
Table 7 extraction time investigation test findings
| Extraction time (min) | 30 | 60 | 90 |
| Mountain Cape jasmine glycosides methyl esters content (mg/g) | 11.12 | 11.74 | 11.51 |
Therefore, extract after 60 minutes in the sample content of mountain Cape jasmine glycosides methyl esters substantially unchanged, historical facts or anecdotes is tested and is selected to add hot reflux 60 minutes.
In sum, the preparation method of need testing solution: get this product content, porphyrize, get approximately 0.2g, accurately weighed, put in the tool plug conical flask, accurate 70% methyl alcohol 25ml, close plug, the weighed weight of adding, added hot reflux 60 minutes, and let cool, more weighed weight, supply the weight of less loss with 70% methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.
4, blank test
Said preparation is the preparation that lamiophlomis rotata single medicinal material is made after extracting, and does not add other in the preparation process mountain Cape jasmine glycosides methyl esters is measured noisy auxiliary material, therefore blank the interference do not examined or check in this research.
5, linear relationship is investigated
Precision is drawn mountain Cape jasmine glycosides methyl esters reference substance solution (0.250mg/ml) 0.25,0.5,1,2,4,8 μ l respectively, the injection liquid chromatography, measure peak area according to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 D), experimental result is as follows.Take peak area (A) as ordinate (Y), mountain Cape jasmine glycosides methyl esters reference substance sample size (ug) is horizontal ordinate (X), the drawing standard curve.
Table 8 mountain Cape jasmine glycosides methyl esters reference substance standard curve determination result
| Mountain Cape jasmine glycosides methyl esters sample size (μ g) | 0.0625 | 0.125 | 0.250 | 0.500 | 1.000 | 2.000 |
| Peak area | 86.4 | 172.4 | 370 | 703 | 1457.6 | 2817.6 |
Calculate regression equation: typical curve Y=1413.4X+6.9, related coefficient 0.9998.Therefore:
Mountain Cape jasmine glycosides methyl esters sample size is the good linear relation with chromatographic peak area in 0.0625～2.000ug scope.Canonical plotting is seen Fig. 3.
6, stability test
Get Radix Lamiophlomidis Rotatae capsule (lot number: 0808036337), prepare need testing solution by method for selecting, preserve under the room temperature, respectively at 0,2,4,8 hours points after the configuration, accurate 5 μ l, the injection liquid chromatography drawn, measure by the text condition determination, test findings is as follows.
Table 9 study on the stability test findings
| Standing time (hour) | 0 | 2 | 4 | 8 | Mean value | RSD(％) |
| The sample peak area | 339.2 | 343.1 | 342.6 | 328.6 | 338.4 | 1.99 |
Therefore need testing solution is measured in rear 8 hours of preparation, and the result is stable.
7, precision test
The accurate lower need testing solution of stability of drawing repeats sample introduction 5 times, and each 5ul measures mountain Cape jasmine glycosides methyl esters peak area, and test findings is as follows.
Table 10 precision is investigated test findings
| Numbering | 1 | 2 | 3 | 4 | 5 | Mean value | RSD(％) |
| The sample peak area | 339.2 | 341.7 | 352.8 | 346.7 | 343.1 | 344.7 | 1.53 |
Therefore test sample peak area mean value is that 344.7, RSD is 1.53%, shows that precision is better.
8, replica test
Get same batch of Radix Lamiophlomidis Rotatae capsule content, porphyrize is got approximately 0.2g, parallel 5 parts, accurately weighed, put in the tool plug conical flask, precision adds 70% methyl alcohol 25ml, close plug, weighed weight respectively, added hot reflux 60 minutes, and let cool, more weighed weight, supply the weight of less loss with 70% methyl alcohol, shake up, filter, get subsequent filtrate, and get final product.
Measure by the assay method under the quality standard text assay item, experimental result is as follows.
Table 11 reappearance is investigated test findings
Therefore content mean value is 11.42mg/g, and RSD=1.30% shows that reappearance is good.
9, recovery test
Adopt the application of sample absorption method, the Radix Lamiophlomidis Rotatae capsule content (mountain Cape jasmine glycosides methyl esters content is 11.42mg/g) of getting known content is 0.1g approximately, parallel 6 parts, accurately weighed, put in the tool plug conical flask, wherein 1., 2. a part difference precision adds mountain Cape jasmine glycosides methyl esters reference substance solution (C=0.2263mg/ml) 4.0ml, replenishes 70% methyl alcohol to 25ml; 3., 4. a part difference precision adds mountain Cape jasmine glycosides methyl esters reference substance solution (C=0.2263mg/ml) 5.0ml, replenishes 70% methyl alcohol to 25ml; 5., 6. a part difference precision adds mountain Cape jasmine glycosides methyl esters reference substance solution (C=0.2263mg/ml) 6.0ml, replenishes methyl alcohol to 25ml; Close plug, weighed weight added hot reflux 60 minutes, let cool, and weighed weight is again supplied the weight of less loss with 70% methyl alcohol, shake up, and filters, and gets subsequent filtrate, and get final product.
Measure by the assay method under the quality standard text assay item, be calculated as follows the recovery, test findings is as follows.
Table 12 average recovery is investigated test findings
Therefore: adopting the application of sample recovery test averaging of income recovery is that 98.4%, RSD is 0.91%, shows that the recovery is better.
10, sample determination
Get the Radix Lamiophlomidis Rotatae capsule content 0.2g of embodiment 8, measure the content of 10 mountain Cape jasmine glycosides methyl esters, every replicate determination 2 times, test findings such as table 13, as shown in Figure 4
Mountain Cape jasmine glycosides methyl esters assay result in table 13 Radix Lamiophlomidis Rotatae capsule
| The sample sequence number | Mountain Cape jasmine glycosides methyl esters content (mg/g) |
| 1 | 7.3 |
| 2 | 7.7 |
| 3 | 8.2 |
| 4 | 8.2 |
| 5 | 8.8 |
| 6 | 7.6 |
| 7 | 7.9 |
| 8 | 7.8 |
| 9 | 8.5 |
| 10 | 8.0 |
| Mean value | 8.0 |
The assay of mountain Cape jasmine glycosides methyl esters in the embodiment 13 lamiophlomis rotata medicinal materials
Measure according to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 D).
Chromatographic condition and system suitability: octadecylsilane chemically bonded silica is filling agent, and take methanol-water (20: 80) as mobile phase, it is 235nm that ultraviolet detects wavelength, and number of theoretical plate calculates by Cape jasmine glycosides methyl esters peak, mountain should be not less than 3000.
The preparation of reference substance solution: it is an amount of that precision takes by weighing mountain Cape jasmine glycosides methyl esters reference substance, adds methyl alcohol and make the solution that every 1ml contains 0.1mg, shakes up, and get final product.
The preparation of need testing solution: get lamiophlomis rotata medicinal material aerial part powder 0.3g, accurately weighed, put in the tool plug conical flask, accurate 70% methanol solution 25ml, close plug, the weighed weight of adding, added hot reflux 60 minutes, let cool, weighed weight is again supplied the weight of less loss with 70% methyl alcohol, shake up, filter, get subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution and each 5 μ l injection liquid chromatography of need testing solution respectively, measures, and get final product.
According to the method described above the content of Cape jasmine glycosides methyl esters in mountain in the different batches lamiophlomis rotata medicinal material is measured the every batch of replicate determination 2 times, test findings such as table 14.
Mountain Cape jasmine glycosides methyl esters assay result in the table 14 lamiophlomis rotata medicinal material
| The sample lot number | Mountain Cape jasmine glycosides methyl esters content (mg/g) |
| 0804126 | 5.2 |
| 0879780 | 6.0 |
| 0806709 | 5.5 |
| 0804721 | 2.9 |
| 0816758 | 5.8 |
| 0864962 | 12.2 |
| 0882995 | 8.9 |
| 0821101 | 3.9 |
| 0805968 | 6.5 |
| Average | 6.3 |
The results show: the selected method with mountain Cape jasmine glycosides methyl esters in high effective liquid chromatography for measuring lamiophlomis rotata medicinal material and the preparation is easy and simple to handle, mensuration is accurate, and precision is good, good reproducibility, is suitable for the quality control of lamiophlomis rotata medicinal material and preparation.
Carry out following pharmacodynamics test with above-mentioned by embodiment 1-7 gained common lamiophlomis root extract sample 1-21.
Test example 1 common lamiophlomis root extract hemostasis of the present invention, analgesic test
1. experiment purpose:
Adopt acetic acid to cause mouse writhing method and mouse docking method, analgesia, the anastalsis of comparative sample 1-21 common lamiophlomis root extract.
2. experiment material
(1) medicine: sample 1-21: common lamiophlomis root extract to sepia medicinal extract, by this laboratory self-control, all is configured to debita spissitudo with physiological saline for subsequent use for brown before use; Compound Paracetamol Tablets: white tablet, specification: the 0.3g/ sheet, Jilin Province Bai Kang medicine company Ltd produces, lot number: 20060102, be mixed with debita spissitudo with physiological saline before use, for subsequent use; Yunnan Baiyao: white powder, specification: the 4g/ bottle, Yunnan Baiyao Group Co.,Ltd produces, lot number: 071054, be mixed with debita spissitudo with physiological saline before use, for subsequent use.
(2) animal: Kunming mouse, body weight 18～22g.Provided animal occupancy permit number by the Traditional Chinese Medicine Research Institute, Sichuan Province: SCXK (river) 2008-10.Animal rank: three grades.
(3) reagent: glacial acetic acid: specification: the 500ml/ bottle, analyze pure, by Chinese Chengdu chemical reagent factory product, lot number: 20070503, draw glacial acetic acid 0.3ml and add physiological saline to 50ml, be made into 0.6% acetum for testing.Physiological saline: specification: the 500ml/ bottle, colourless clear liquid is produced lot number: B070414 by Kelun Pharm Ind Co., Ltd., Sichuan.
(4) instrument: FA1004 type electronic balance, range: 100g, precision: 0.1mg, the flat instrument and meter of upper current chart company limited produces.Stopwatch: Shanghai stopwatch factory product.
3. experimental technique and result
3.1 Dichlorodiphenyl Acetate causes the impact of mice pain reaction: get 230 of Kunming mouses, male and female half and half are divided into 10 groups at random by the body weight sex, and 10 every group, male and female half and half.The every mouse oral administration gavage of blank group physiological saline 10ml/kg body weight, positive controls gavages Aminodyne Compound 0.23g/kg (according to the content meter of aspirin in the prescription) body weight, and each group of administration gavages respectively sample 1-21 group 3g (crude drug)/kg body weight.Administration every day 1 time, successive administration 3 days, 30min after the last administration, each is organized mouse peritoneal and injects 0.6% acetum 0.2ml/ mouse, each mouse of observed and recorded writhing number of times in the 15min behind lumbar injection acetic acid, its significance difference opposite sex relatively between organizing.The results are shown in Table 15.
Table 15 Dichlorodiphenyl Acetate causes the impact of mice pain reaction
Compare with the blank group: P
*＜0.05, P
*＜0.01, P
* *＜0.001
Table 15 is the result show, compares with the blank group, and sample 1-21 all can suppress acetic acid induced mice pain.Except 9,16 and 18, all the other samples all have significant difference, wherein the P value less than 0.05 sample have 2,11,12,13,14,17,19,20 and 21, P value less than 0.01 sample have 4,5,6 and 10, P value have 1,3,7,8 and 15 less than 0.001 sample.
3.2 the impact on the mouse docking bleeding time: get 230 of Kunming mouses, be divided at random 10 groups by the body weight sex, 10 every group, male and female half and half.The every mouse oral administration gavage of negative control group physiological saline 10ml/kg body weight, positive controls gavages Yunnan Baiyao 1.2g (crude drug)/kg body weight, and each group of administration gavages respectively sample 1-21 group 3.0g (crude drug)/kg body weight.Administration every day 1 time, successive administration 3 days, 1h after the last administration, cut with profit Mouse Tail-tip 3mm place is cut off, put into 37 ℃ of physiological saline, begin timing when allowing blood overflow voluntarily, until hemorrhagely stop voluntarily, calculate the bleeding time, its significance difference opposite sex relatively between organizing.
3.3 experimental result the results are shown in Table 16.
Table 16 is on the mouse impact in docking bleeding time
| Group | Dosage (g/kg) | Number of animals (only) | Bleeding time (s, x ± s) |
| The blank group | NS | 10 | 516.3±119.8 |
| Positive controls | 1.2 | 10 | 313.4±100.6*** |
| Sample 1 | 3g crude drug/kg | 10 | 308.3±104.4*** |
| Sample 2 | 3g crude drug/kg | 10 | 329.5±118.9* |
| Sample 3 | 3g crude drug/kg | 10 | 314.8±115.2*** |
| Sample 4 | 3g crude drug/kg | 10 | 315.6±106.6** |
| Sample 5 | 3g crude drug/kg | 10 | 310.4±110.8** |
| Sample 6 | 3g crude drug/kg | 10 | 310.5±116.1** |
| Sample 7 | 3g crude drug/kg | 10 | 318.1±111.5*** |
| Sample 8 | 3g crude drug/kg | 10 | 310.2±94.5*** |
| Sample 9 | 3g crude drug/kg | 10 | 433.7±122.8 |
| Sample 10 | 3g crude drug/kg | 10 | 320.3±95.9** |
| Sample 11 | 3g crude drug/kg | 10 | 379.5±108.2* |
| Sample 12 | 3g crude drug/kg | 10 | 353.8±120.6* |
| Sample 13 | 3g crude drug/kg | 10 | 345.6±117.5* |
| Sample 14 | 3g crude drug/kg | 10 | 354.4±110.8* |
| Sample 15 | 3g crude drug/kg | 10 | 318.0±101.9*** |
| Sample 16 | 3g crude drug/kg | 10 | 441.8±144.5 |
| Sample 17 | 3g crude drug/kg | 10 | 395.0±106.2* |
| Sample 18 | 3g crude drug/kg | 10 | 457.9±130.2 |
| Sample 19 | 3g crude drug/kg | 10 | 406.6±100.1* |
| Sample 20 | 3g crude drug/kg | 10 | 363.1±112.9* |
| Sample 21 | 3g crude drug/kg | 10 | 361.8±99.5* |
Compare with the blank group: P
*＜0.05, P
*＜0.01, P
* *＜0.001
Table 16 is the result show, compares with the blank group, and sample 1-21 all can shorten the bleeding time of docking mouse.Except 9,16 and 18, all the other samples all have significant difference, wherein the P value less than 0.05 have sample 2,11,12,13,14,17,19,20 and 21, P value less than 0.01 have sample 4,5,6 and 10, P value less than 0.001 sample 1,3,7,8 and 15 arranged.
4 conclusions
Show by analgesia and two results of pharmacodynamic test of stopping blooding, each batch common lamiophlomis root extract that the mountain Cape jasmine glycosides methyl esters content of different medicinal materials, different extraction process gained does not wait all has pain easing and hemostasis effect in various degree, and what of its content show certain dose-effect trend with the conspicuousness of hemostatic analgesia effect.See the following form
| Sample number into spectrum | Mountain Cape jasmine glycosides methyl esters content (mg/g) | The writhing number of times (inferior, x ± s) | Bleeding time (s, x ± s) |
| The blank group | - | 53.9±11.0 | 516.3±119.8 |
| Positive controls | - | 16.7±10.0*** | 313.4±100.6*** |
| 16 | 0.4 | 48.9±19.2 | 441.8±144.5 |
| 9 | 0.7 | 45.7±16.2 | 433.7±122.8 |
| 18 | 1.2 | 40.8±12.5 | 457.9±130.2 |
| 19 | 1.5 | 35.9±10.5* | 406.6±100.1* |
| 20 | 2.6 | 35.2±8.0* | 363.1±112.9* |
| 17 | 3.1 | 31.9±10.2* | 395.0±106.2* |
| 11 | 3.9 | 28.9±10.6* | 379.5±108.2* |
| 2 | 5.8 | 29.7±12.1* | 329.5±118.9* |
| 21 | 6.7 | 28.4±10.5* | 361.8±99.5* |
| 13 | 7.3 | 32.3±9.0* | 345.6±117.5* |
| 12 | 8.5 | 30.8±8.8* | 353.8±120.6* |
| 14 | 8.9 | 27.9±13.6* | 354.4±110.8* |
| 4 | 12.5 | 23.9±11.8** | 315.6±106.6** |
| 5 | 13.4 | 24.2±10.2** | 310.4±110.8** |
| 10 | 14.6 | 26.3±9.9** | 320.3±95.9** |
| 6 | 15.9 | 26.1±9.5** | 310.5±116.1** |
| 15 | 18.1 | 17.3±9.9*** | 318.0±101.9*** |
| 3 | 19.0 | 17.6±9.9*** | 314.8±115.2*** |
| 7 | 21.3 | 17.2±10.1*** | 318.1±111.5*** |
| 8 | 22.5 | 19.5±8.0*** | 310.2±94.5*** |
| 1 | 30.0 | 19.1±9.0*** | 308.3±104.4*** |
[0208]Above-mentioned evidence, take mountain Cape jasmine glycosides methyl esters as index components, the mountain Cape jasmine glycosides methyl esters content of different medicinal materials, different extraction process gained, in the scope of mountain Cape jasmine glycosides methyl esters content at 1.5-30mg/g, all have obvious drug effect, and efficacy stability, again take mountain Cape jasmine glycosides methyl esters content as 12.5-30mg/g as good, still more preferably, containing mountain Cape jasmine glycosides methyl esters 18.1-30mg/g is optimal selection.
In addition, above-mentioned test also illustrates, the height of the content of mountain Cape jasmine glycosides methyl esters has determined the quality of common lamiophlomis root extract in the medicinal material, lamiophlomis rotata medicinal material mountain Cape jasmine glycosides methyl esters content is 8.7mg/g among the sample 1-7, no matter adopts which kind of technique, can make the mountain Cape jasmine glycosides methyl esters of common lamiophlomis root extract within the scope of limiting the quantity of that the present invention requires, become effective active substance, and extraction process is more simplified, and has saved operation steps, convenient greatly production; When quality of medicinal material not good, when mountain Cape jasmine glycosides methyl esters content is low in the medicinal material, the content of mountain Cape jasmine glycosides methyl esters might not reach scope of the presently claimed invention in the common lamiophlomis root extract, can not still can adopt refining, method for concentration as effective active matter, improve the content of mountain Cape jasmine glycosides methyl esters, make it to reach the scope of limiting the quantity of of the present invention, also can use as effective active matter, enlarge the adaptability to medicinal material, for large production provides wider selection.
This evidence, common lamiophlomis root extract efficacy stability of the present invention, controllability is strong, and which kind of extraction process no matter the content by direct control mountain Cape jasmine glycosides methyl esters adopt, and all can reach the purpose of control drug quality of the present invention.
The pharmacodynamics test that test example 2 adopts general flavone to carry out quality control
In order further to prove beneficial effect of the present invention, by " quality control standard of Chinese Pharmacopoeia 2005 version lamiophlomis rotata adopts general flavone to control the quality of common lamiophlomis root extract.
Extracting method:
Method 1: get the lamiophlomis rotata medicinal material, be ground into meal, put in the percolator, after 12 hours, with 5ml/min speed diacolation, obtain the lamiophlomis rotata percolate with 20 times of 70% ethanol with 70% alcohol immersion; The lamiophlomis rotata percolate is added water left standstill 36 hours, get supernatant concentration, drying.
Method 2: get the lamiophlomis rotata medicinal material, be ground into meal, add 30% alcohol reflux and extract 2 times, add 8 times of amounts of ethanol for the first time, refluxed 1 hour, add 10 times of amounts of ethanol, refluxed 1 hour for the 2nd time; Merge phegma, reduced pressure concentration, Recycled ethanol is to without the alcohol flavor, and is concentrated again, drying.
Method 3: get the lamiophlomis rotata medicinal material, be ground into meal, add the ultrasonic extraction of water 1 hour of 10 times of amounts of water, extract is concentrated, drying.
" extracting method of Chinese Pharmacopoeia 2005 version lamiophlomis rotata: get lamiophlomis rotata, pulverize, boiling 3 times, each 1 hour, collecting decoction filtered, and the clear cream of filtrate simmer down to relative density 1.30 is dry below 80 ℃.
General flavone content in the common lamiophlomis root extract, the result is as follows:
Sample 1-12 is carried out common lamiophlomis root extract hemostasis, analgesic test by the method for test example 1, and test findings sees the following form 17:
Table 17 Dichlorodiphenyl Acetate causes the impact of mice pain reaction
| Group | Dosage | Number of animals (only) | The writhing number of times (inferior, x ± s) | Inhibiting rate (%) | |
| The | NS | 10 | 48.2±12.2 | - | |
| Positive controls | 0.23mg/ |
10 | 15.1±8.2*** | 68.7 | |
| |
3g crude drug/ |
10 | 23.3±9.4** | 51.6 | |
| |
3g crude drug/ |
10 | 28.5±11.8* | 40.9 | |
| Sample 3 | 3g crude drug/ |
10 | 38.2±12.6 | 20.7 | |
| Sample 4 | 3g crude drug/ |
10 | 18.1±9.6*** | 62.4 | |
| |
3g crude drug/ |
10 | 24.2±8.8** | 49.8 | |
| Sample 6 | 3g crude drug/ |
10 | 32.1±9.4* | 33.4 | |
| Sample 7 | 3g crude drug/ |
10 | 39.8±14.0 | 17.4 | |
| |
3g crude drug/ |
10 | 17.9±10.2*** | 62.9 | |
| Sample 9 | 3g crude drug/ |
10 | 40.2±14.8 | 16.6 | |
| |
3g crude drug/ |
10 | 21.3±10.7*** | 43.3 | |
| Sample 11 | 3g crude drug/ |
10 | 36.1±9.9* | 25.1 | |
| Sample 12 | 3g crude drug/ |
10 | 18.4±11.1*** | 61.8 |
Compare with the blank group: P
*＜0.05, P
*＜0.01, P
* *＜0.001
Table 18 is on the mouse impact in docking bleeding time
| Group | Dosage (g/kg) | Number of animals (only) | Bleeding time (s, x ± s) | |
| The | NS | 10 | 516.3±119.8 | |
| Positive controls | 1.2 | 10 | 305.9±110.6*** | |
| |
3g crude drug/ |
10 | 313.9±95.8*** | |
| |
3g crude drug/ |
10 | 368.9±105.6** | |
| Sample 3 | 3g crude drug/ |
10 | 455.9±132.1 | |
| Sample 4 | 3g crude drug/ |
10 | 323±100.8*** | |
| |
3g crude drug/ |
10 | 366.2±100.8* | |
| Sample 6 | 3g crude drug/ |
10 | 333.9±116.1*** | |
| Sample 7 | 3g crude drug/ |
10 | 360.2±110.6** | |
| |
3g crude drug/ |
10 | 356.0±119.2** | |
| Sample 9 | 3g crude drug/ |
10 | 448.7±140.2 | |
| |
3g crude drug/ |
10 | 410.7±104.5* | |
| Sample 11 | 3g crude drug/ |
10 | 389.2±114.5* | |
| Sample 12 | 3g crude drug/ |
10 | 312.2±100.1*** |
Compare with the blank group: P*＜0.05, P**＜0.01, P***＜0.001
By evidence, adopt " the extracting method of Chinese Pharmacopoeia 2005 version lamiophlomis rotata, wherein general flavone content is in the scope of pharmacopeia regulation, efficacy stability, and the common lamiophlomis root extract that adopts other extraction process to obtain, wherein the content of general flavone is unstable, the drug effect of extract is also unstable, such as sample 9 and sample 11, under identical general flavone content, drug effect is not identical, its general flavone content does not show corresponding dose-effect relationship with analgesia, hemostatic healing efficacy, therefore, only adopt general flavone as index, can not effectively monitor the quality of common lamiophlomis root extract.See the following form
By above-mentioned evidence, common lamiophlomis root extract of the present invention is used for analgesia, hemostasis, efficacy stability, and controllability is strong.By the content of control mountain Cape jasmine glycosides methyl esters, can effectively control the quality of extract, improve the stability of drug effect and medicine.
Claims (2)
1. the method for quality that detects lamiophlomis rotata medicinal material and extract thereof or contain the pharmaceutical composition of common lamiophlomis root extract is characterized in that: be to carry out quality testing take mountain Cape jasmine glycosides methyl esters as index components;
Comprise the steps:
A, by high effective liquid chromatography for measuring, adopt reverse-phase chromatographic column, mobile phase is comprised of organic phase and water, it is 220-250nm that ultraviolet detects wavelength;
B, get mountain Cape jasmine glycosides methyl esters reference substance, add Methanol and become reference substance solution;
C, get lamiophlomis rotata medicinal material, extract or preparation, make need testing solution;
D, accurate reference substance solution and the need testing solution injection liquid chromatography drawn of difference are measured, and be get final product;
Wherein, the described reverse-phase chromatographic column of step a is take octadecylsilane chemically bonded silica as filling agent; Described organic phase is methyl alcohol or acetonitrile, and described water is pure water, phosphoric acid solution or acetum; Described detection wavelength is 235nm; Reference substance solution described in the step b is the solution that every 1ml contains 0.02-0.40mg mountain Cape jasmine glycosides methyl esters; The preparation of the described need testing solution of step c may further comprise the steps: get lamiophlomis rotata medicinal material, extract or preparation 0.05-1.0g, and accurately weighed, put in the tool plug conical flask, accurate 50-100% methanol solution 25ml, close plug, the weighed weight of adding, add hot reflux 30-90 minute, let cool, weighed weight is again supplied the weight of less loss with the 50-100% methanol solution, shake up, filter, get subsequent filtrate, and get final product;
Wherein, the described methyl alcohol of step a and described pure water volume ratio are 10:90-30:70.
2. quality determining method according to claim 1 is characterized in that: contain mountain Cape jasmine glycosides methyl esters 1.5-30mg/g in the described common lamiophlomis root extract.
3, quality determining method according to claim 2 is characterized in that: the content that contains mountain Cape jasmine glycosides methyl esters in the common lamiophlomis root extract is 12.5-30mg/g.
4, quality determining method according to claim 3 is characterized in that: the content that contains mountain Cape jasmine glycosides methyl esters in the common lamiophlomis root extract is 18.1-30mg/g.
5, quality determining method according to claim 1 is characterized in that: the described methyl alcohol of step a and described pure water volume ratio are 20:80.
6, quality determining method according to claim 1 is characterized in that: the described concentration of methanol solution of step c is 70%.
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