CN103638450B - A kind of preparation method of Muxiangliqi Tablet and application - Google Patents
A kind of preparation method of Muxiangliqi Tablet and application Download PDFInfo
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Abstract
The invention belongs to technical field of Chinese medicines, be specifically related to a kind of preparation method and application of Muxiangliqi Tablet.The preparation method of Muxiangliqi Tablet of the present invention, be made up as crude drug of Radix Astragali 280g, fiber crops rare 80g, Radix Angelicae Sinensis 190g, Herba Ecliptae 220g, Rhizoma Atractylodis 190g, Flos Carthami 150g, Radix Paeoniae Rubra 140g, Semen Persicae 140g, Radix Achyranthis Bidentatae 150g, Rhizoma Chuanxiong 120g, Fructus Aurantii 70g, Radix Platycodonis 100g, adopt supercritical extraction and microwave extracting to be prepared from, content of baicalin is improved a lot.Present invention also offers the application of Muxiangliqi Tablet in preparation suppression mouse mammary tumor cells C127 cell proliferation.
Description
Technical field
The invention belongs to technical field of Chinese medicines, be specifically related to a kind of preparation method and application of Muxiangliqi Tablet.
Background technology
Muxiangliqi Tablet standard No. WS-11113(ZD-1113)-2002, be recorded in national standard for traditional Chinese medicines compilation internal medicine taste fascicle.Boil Rhizoma Curcumae 24g, Fructus Crataegi 96g by Radix Aucklandiae 48g, Rhizoma Cyperi (processed with vinegar) 96g, Radix Linderae 96g, Pericarpium Citri Reticulatae Viride (processed with vinegar) 48g, Pericarpium Citri Reticulatae 96g, Fructus Aurantii Immaturus 96g, Fructus Aurantii 96g, stir-frying with ginger juice Cortex Magnoliae Officinalis 96g, vinegar Rhizoma Sparganii 24g, vinegar, Semen Arecae 120g, Fructus Evodiae (processed) 12g, Cortex Cinnamomi 12g, Radix Et Rhizoma Nardostachyos 15g, Radix Platycodonis 48g, Radix Scutellariae 96g, Radix Et Rhizoma Rhei 48g, Semen Pharbitidis (parched) 48g make as crude drug, there is promoting the circulation of QI to alleviate the stagnation in middle-JIAO, change effect of stagnant relieving constipation.For depression of QI, stop eating and cut off the water, breast side of body painful abdominal mass is vexed, distension and fullness in the abdomen, nausea and vomiting, falls full noisy, constipation.
In prior art, Muxiangliqi Tablet is not yet had to extract the report adopting supercritical and microwave technology in preparation, and Chinese medicine directly beats powder, volatile oil is drawn, the method of soak by water, technique is coarse, backward, and impurity is many, causes patient's consumption excessive, be inconvenient to take, had a strong impact on this product and applied clinically.
Summary of the invention
In order to solve above-mentioned technical problem, the invention provides a kind of preparation method of Muxiangliqi Tablet.
Another object of the present invention is to provide a kind of Muxiangliqi Tablet to suppress the application in mouse mammary tumor cells C127 cell proliferation in preparation.
The object of the invention is by following scheme realize:
A kind of preparation method of Muxiangliqi Tablet, by Radix Aucklandiae 48g, Rhizoma Cyperi (processed with vinegar) 96g, Radix Linderae 96g, Pericarpium Citri Reticulatae Viride (processed with vinegar) 48g, Pericarpium Citri Reticulatae 96g, Fructus Aurantii Immaturus 96g, Fructus Aurantii 96g, stir-frying with ginger juice Cortex Magnoliae Officinalis 96g, vinegar Rhizoma Sparganii 24g, vinegar boils Rhizoma Curcumae 24g, Fructus Crataegi 96g, Semen Arecae 120g, Fructus Evodiae (processed) 12g, Cortex Cinnamomi 12g, Radix Et Rhizoma Nardostachyos 15g, Radix Platycodonis 48g, Radix Scutellariae 96g, Radix Et Rhizoma Rhei 48g, Semen Pharbitidis (parched) 48g makes as crude drug, described preparation method is made up of the following step: get the Radix Aucklandiae, Radix Et Rhizoma Rhei, Pericarpium Citri Reticulatae, Fructus Aurantii Immaturus, Fructus Aurantii, Radix Et Rhizoma Nardostachyos, join in CO2 supercritical extraction device, ethanol is as entrainer, the percent by volume that entrainer accounts for total extractant is 4-6%, extracting pressure 15-30MPa, temperature 30-50 DEG C, CO2 flow l-3ml/g crude drug min, extraction time 180-220min, obtain supercritical extract, for subsequent use, get all the other Chinese medicines, pulverize, add 50% ethanol of 1.6L, drop in microwave extracting apparatus and carry out microwave extracting, extraction power 600-800W, extracts 2 times, each 5-10 minute, combining extraction liquid, concentrated, be added on Dl01 macroporous adsorptive resins, 50% ethanol elution, collect 5 times amount column volume eluents, decompression recycling ethanol, concentrated and dry, obtain microwave extraction thing, for subsequent use, by above-mentioned supercritical extract and the mixing of microwave extraction thing, add starch, 70% ethanol granule, dry, tabletting, makes 500, the heavy 0.5g of every sheet.
In the preparation method of above-mentioned Muxiangliqi Tablet, described CO
2in supercritical extraction, entrainer accounts for the percent by volume of total extractant is 5%.
In the preparation method of above-mentioned Muxiangliqi Tablet, described CO
2in supercritical extraction, extracting pressure is 25MPa.
In the preparation method of above-mentioned Muxiangliqi Tablet, described CO
2in supercritical extraction, extraction temperature is 60 DEG C.
In the preparation method of above-mentioned Muxiangliqi Tablet, described CO
2cO in supercritical extraction
2flow 2ml/g crude drug min.
In the preparation method of above-mentioned Muxiangliqi Tablet, described CO
2in supercritical extraction, extraction time is 200min.
In the preparation method of above-mentioned Muxiangliqi Tablet, described microwave extracting power is 700W.
In the preparation method of above-mentioned Muxiangliqi Tablet, each extraction time of described microwave extracting is 8 minutes.
Above-mentioned Muxiangliqi Tablet suppresses the application in mouse mammary tumor cells C127 cell proliferation in preparation.
In prior art, Muxiangliqi Tablet is oral, one time 4 ~ 8,2 times on the one.Muxiangliqi Tablet dosage is large.The heavy 0.5g of the every sheet of the Muxiangliqi Tablet adopting the inventive method to be prepared into, only needs 2-4 sheet at every turn, within 1st, takes 2 times.Dose is greatly reduced under the condition with more active component.This conclusion can be proved by following test.
The comparison of content of baicalin in Muxiangliqi Tablet prepared by test one, distinct methods
L, instrument and reagent Muxiangliqi Tablet of the present invention: by the preparation of embodiment 1 method, use 1215g crude drug, makes 500 through extracting, the heavy 0.5g of every sheet.Former Muxiangliqi Tablet, according to WS-11113(ZD-1113)-2002 standard method preparations, in contrast.Agilent1200 high performance liquid chromatograph; METTLERAE240 electronic analytical balance; Baicalin reference substance (Nat'l Pharmaceutical & Biological Products Control Institute).
2, method
Measure according to high performance liquid chromatography (Chinese Pharmacopoeia version in 2010 annex VD).
Chromatographic condition and system suitability octadecylsilane chemically bonded silica are filler; Methanol-water-glacial acetic acid (35: 61: 4) is mobile phase; Determined wavelength is 283nm.Number of theoretical plate presses the calculating of baicalin peak should lower than 2500.
The preparation precision of reference substance solution takes at 60 DEG C of drying under reduced pressure baicalin reference substance of 4 hours appropriate, adds methanol and makes the solution of every 1ml containing 0.04mg, to obtain final product.
The preparation of product need testing solution of the present invention gets Muxiangliqi Tablet of the present invention, porphyrize, gets 1g, accurately weighed, puts in 100ml measuring bottle, add water 30ml, and put in water-bath and heat 30 minutes, add methanol 50ml, supersound process 30 minutes, lets cool, add methanol to scale, shake up, filter, get subsequent filtrate, to obtain final product.
The Muxiangliqi Tablet of contrast, porphyrize are got in the preparation of reference product need testing solution, get 1g, accurately weighed, put in 100ml measuring bottle, add water 30ml, and put in water-bath and heat 30 minutes, add methanol 50ml, supersound process 30 minutes, lets cool, add methanol to scale, shake up, filter, get subsequent filtrate, to obtain final product.
Algoscopy is accurate respectively draws reference substance solution and each 10 μ L of need testing solution, injection liquid chromatography, measures, to obtain final product.
3, result
Result shows, in Muxiangliqi Tablet of the present invention, the content of baicalin is 4-7mg/ sheet; And the content of baicalin is 0.9mg/ sheet in former Muxiangliqi Tablet, every sheet content of baicalin is equivalent to the 4-8 of former tablet content doubly, and when dose reduces, content of baicalin improves a lot.
Above-mentioned research shows, adopt Muxiangliqi Tablet prepared by preparation method of the present invention, active constituent content is far away higher than WS-11113(ZD-1113) Muxiangliqi Tablet prepared of method recorded of-2002 standards.
Detailed description of the invention
Below in conjunction with specific embodiment, the present invention is further described, so that those skilled in the art more understands the present invention, but this should be interpreted as that the scope of the above-mentioned theme of the present invention is only limitted to following example, all technology realized based on foregoing of the present invention all belong to scope of the present invention.
Embodiment 1
Get Radix Aucklandiae 48g, Rhizoma Cyperi (processed with vinegar) 96g, Radix Linderae 96g, Pericarpium Citri Reticulatae Viride (processed with vinegar) 48g, Pericarpium Citri Reticulatae 96g, Fructus Aurantii Immaturus 96g, Fructus Aurantii 96g, stir-frying with ginger juice Cortex Magnoliae Officinalis 96g, vinegar Rhizoma Sparganii 24g, vinegar boils Rhizoma Curcumae 24g, Fructus Crataegi 96g, Semen Arecae 120g, Fructus Evodiae (processed) 12g, Cortex Cinnamomi 12g, Radix Et Rhizoma Nardostachyos 15g, Radix Platycodonis 48g, Radix Scutellariae 96g, Radix Et Rhizoma Rhei 48g, Semen Pharbitidis (parched) 48g, the Radix Aucklandiae, Radix Et Rhizoma Rhei, Pericarpium Citri Reticulatae, Fructus Aurantii Immaturus, Fructus Aurantii, Radix Et Rhizoma Nardostachyos are joined CO
2in supercritical extraction device, ethanol is as entrainer, and the percent by volume that entrainer accounts for total extractant is 5%, extracting pressure 25MPa, temperature 40 DEG C, CO
2flow 2ml/g crude drug min, extraction time 200min, obtains supercritical extract, for subsequent use; Get all the other Chinese medicines, pulverize, add 50% ethanol of 1.6L, drop in microwave extracting apparatus and carry out microwave extracting, extraction power 700W, extracts 2 times, each 8 minutes, combining extraction liquid, concentrated, be added on Dl01 macroporous adsorptive resins, 50% ethanol elution, collect 5 times amount column volume eluents, decompression recycling ethanol, concentrated and dry, obtain microwave extraction thing, for subsequent use; By above-mentioned supercritical extract and the mixing of microwave extraction thing, add starch, 70% ethanol granule, dry, tabletting, makes 500, the heavy 0.5g of every sheet.
After testing, in finished product, the content of baicalin is 6.22mg/ sheet.
Embodiment 2
Get Radix Aucklandiae 48g, Rhizoma Cyperi (processed with vinegar) 96g, Radix Linderae 96g, Pericarpium Citri Reticulatae Viride (processed with vinegar) 48g, Pericarpium Citri Reticulatae 96g, Fructus Aurantii Immaturus 96g, Fructus Aurantii 96g, stir-frying with ginger juice Cortex Magnoliae Officinalis 96g, vinegar Rhizoma Sparganii 24g, vinegar boils Rhizoma Curcumae 24g, Fructus Crataegi 96g, Semen Arecae 120g, Fructus Evodiae (processed) 12g, Cortex Cinnamomi 12g, Radix Et Rhizoma Nardostachyos 15g, Radix Platycodonis 48g, Radix Scutellariae 96g, Radix Et Rhizoma Rhei 48g, Semen Pharbitidis (parched) 48g, the Radix Aucklandiae, Radix Et Rhizoma Rhei, Pericarpium Citri Reticulatae, Fructus Aurantii Immaturus, Fructus Aurantii, Radix Et Rhizoma Nardostachyos are joined CO
2in supercritical extraction device, ethanol is as entrainer, and the percent by volume that entrainer accounts for total extractant is 4%, extracting pressure 30MPa, temperature 50 C, CO
2flow 3ml/g crude drug min, extraction time 180min, obtains supercritical extract, for subsequent use; Get all the other Chinese medicines, pulverize, add 50% ethanol of 1.6L, drop in microwave extracting apparatus and carry out microwave extracting, extraction power 600W, extracts 2 times, each 5 minutes, combining extraction liquid, concentrated, be added on Dl01 macroporous adsorptive resins, 50% ethanol elution, collect 5 times amount column volume eluents, decompression recycling ethanol, concentrated and dry, obtain microwave extraction thing, for subsequent use; By above-mentioned supercritical extract and the mixing of microwave extraction thing, add starch, 70% ethanol granule, dry, tabletting, makes 500, the heavy 0.5g of every sheet.
After testing, in finished product, the content of baicalin is 5.78mg/ sheet.
Embodiment 3
Get Radix Aucklandiae 48g, Rhizoma Cyperi (processed with vinegar) 96g, Radix Linderae 96g, Pericarpium Citri Reticulatae Viride (processed with vinegar) 48g, Pericarpium Citri Reticulatae 96g, Fructus Aurantii Immaturus 96g, Fructus Aurantii 96g, stir-frying with ginger juice Cortex Magnoliae Officinalis 96g, vinegar Rhizoma Sparganii 24g, vinegar boils Rhizoma Curcumae 24g, Fructus Crataegi 96g, Semen Arecae 120g, Fructus Evodiae (processed) 12g, Cortex Cinnamomi 12g, Radix Et Rhizoma Nardostachyos 15g, Radix Platycodonis 48g, Radix Scutellariae 96g, Radix Et Rhizoma Rhei 48g, Semen Pharbitidis (parched) 48g, the Radix Aucklandiae, Radix Et Rhizoma Rhei, Pericarpium Citri Reticulatae, Fructus Aurantii Immaturus, Fructus Aurantii, Radix Et Rhizoma Nardostachyos are joined CO
2in supercritical extraction device, ethanol is as entrainer, and the percent by volume that entrainer accounts for total extractant is 6%, extracting pressure 15MPa, temperature 30 DEG C, CO
2flow 1ml/g crude drug min, extraction time 220min, obtains supercritical extract, for subsequent use; Get all the other Chinese medicines, pulverize, add 50% ethanol of 1.6L, drop in microwave extracting apparatus and carry out microwave extracting, extraction power 800W, extracts 2 times, each 10 minutes, combining extraction liquid, concentrated, be added on Dl01 macroporous adsorptive resins, 50% ethanol elution, collect 5 times amount column volume eluents, decompression recycling ethanol, concentrated and dry, obtain microwave extraction thing, for subsequent use; By above-mentioned supercritical extract and the mixing of microwave extraction thing, add starch, 70% ethanol granule, dry, tabletting, makes 500, the heavy 0.5g of every sheet.
After testing, in finished product, the content of baicalin is 4.96mg/ sheet.
Embodiment 4: Muxiangliqi Tablet suppresses the experimentation data of mouse mammary tumor cells C127 cell proliferation
1. experiment material
1.1 experiment cell strains
Mouse mammary tumor cells C127 cell, Shandong University's laboratory cell storehouse, DMEM+10%FBS cellar culture.
1.2 Experimental agents
Drugs: Muxiangliqi Tablet of the present invention: prepare by embodiment 1 method.
Medicinal liquid liquid storage: take 100mg Muxiangliqi Tablet, is dissolved in 5ml dehydrated alcohol, 0.2 μm of frit, 500 μ ldoff pipe subpackages ,-20 DEG C of storages, and simultaneously 0.2 μm of frit dehydrated alcohol is in order to the use of matched group.
1.3 experiment reagent
DMEM (GIBCO company Cat.No.12100-061Lot.No.758137); Hyclone (Tian Hang bio tech ltd, Zhejiang Lot.No.100419); NaHC0
3(Shanghai hundred million chemical reagent company limited Cat.No.11810-033Lot.No.1088387 of a specified duration); Trypsin(AMRESCO company); EDTA(AMRESCO company); PenicillinGSodiumSalt(AMRESCO company 1); StreptomycinSulfate (AMRESCO); Dehydrated alcohol (Zibo Ya Dulan Trade Co., Ltd.); MTT (Biosharp lot number: 0793): PBS(laboratory autogamy);
1.4 experiment equipment
Lycra inverted microscope (German Leica model: DMIL); Visible-ultraviolet light microwell plate detector (MD company of U.S. model: SPECTRAMAX190); C0
2incubator (FORMA model: 3111); Super-clean bench (safe and sound Inc. of Su Jing group moulding number: SW-CJ-ZFD); Pure water instrument (Sprlng company of U.S. model: S/N020579); Accurate pipettor (French Gilson Inc model: P2); Electronic balance (German Sai Duolisi company limited model: BT323S); Full-automatic high-pressure autoclave (Japanese SANYO company model: MLS-3020); Table electrothermal air dry oven (the accurate experimental facilities company in Shanghai model: DHG9123A); Refrigerator (Siemens Company's model: KG18V21TI); Liquid nitrogen container (CBS model: 2001); Low speed centrifuge (Anting Scientific Instrument Factory, Shanghai's model: KA-1000); 0.2 μm of filter (MILLIPORE model: SLGP033RB); 1cm culture dish (NEST company), 96 well culture plates (NEST company); Cell counting count board; Centrifuge tube, pipet, Tips are some.
2. experimental technique
1) C127 cell DMEM+10%FBS is in 37 DEG C, 5%C0
2carry out cellar culture (10cm culture dish), when Growth of Cells is to logarithmic (log) phase, collecting cell, discards culture fluid, PBS fine laundering 3 times, add 3ml0.25% trypsin-0.04%EDTA, after 37 DEG C of digestion 2min, add 5ml complete medium neutralization reaction wherein, proceeded in centrifuge tube after piping and druming cell, the centrifugal 5min of 1000rpm, adjustment concentration of cell suspension 3 × 10
4individual/ml.
2) enter in 96 well culture plates by cell kind, every hole adds cell suspension 180 μ l, culture plate put into cell culture incubator (37 DEG C, 5%C0
2) cellar culture.
3) according to cell growth status, generally grow to 50%-70%, add Muxiangliqi Tablet solution, continue to cultivate 24h.
4) add 20 μ lMTT solution (5mg/ml, i.e. 0.5%MTT) after 24h, continue to cultivate 4h.
5) after 4h, buckle method removes supernatant, pats dry gently with absorbent paper, and every hole adds 200 μ l dimethyl sulfoxide, puts low-speed oscillation 10min on shaking table, crystal is fully dissolved.The light absorption value in each hole is measured at enzyme-linked immunosorbent assay instrument 490nm place.
6) background (do not add cell, only add culture fluid) is set, control wells (the medicine dissolution medium of cell, same concentrations, culture fluid, MTT, dimethyl sulfoxide) simultaneously, often organizes the multiple hole of setting 6.
7) result represents with the suppression ratio of medicine to cell: cell proliferation suppression ratio (%)=(control wells OD value-dosing holes OD value), control wells OD value × 100%.Experiment repetition 3 times.
3. statistical disposition
Adopt the correlation analysis in MicrosoftExcel2007 software and Studentt inspection, data represent with mean ± S.D..
4. experimental result
Statistical result showed after mtt assay experiment, compare with matched group, when dosage reaches 5mg/ml, to C127 cell inhibitory effect variant (P<0.05), dosage this difference when 10mg/ml has significance (P<0.01), has pole significant difference (P<0.001) when dosage reaches 15-20mg/ml.
Table 1 Muxiangliqi Tablet is to C127 cell inhibitory effect influence research (X ± SD)
| Group | Drug level (mg/ml) | Suppression ratio (%) |
| Matched group | 0 | 0 |
| 1 | 5 | 9.24±8.91 |
| 2 | 10 | 24.6±15.88* |
| 3 | 15 | 34.9±17.92** |
| 4 | 20 | 45.7±18.98** |
Note: compare with matched group, * P<O.01; * P<0.001.
5. experiment conclusion
Muxiangliqi Tablet can suppress C127 cell proliferation, and reduce the Growth of Cells number of C127 cell, this effect is dose dependent.
Claims (7)
1. the application of Muxiangliqi Tablet in preparation suppression mouse mammary tumor cells C127 cell proliferation, it is characterized in that, described Muxiangliqi Tablet is by Radix Aucklandiae 48g, Rhizoma Cyperi (processed with vinegar) 96g, Radix Linderae 96g, Pericarpium Citri Reticulatae Viride (processed with vinegar) 48g, Pericarpium Citri Reticulatae 96g, Fructus Aurantii Immaturus 96g, Fructus Aurantii 96g, stir-frying with ginger juice Cortex Magnoliae Officinalis 96g, vinegar Rhizoma Sparganii 24g, vinegar boils Rhizoma Curcumae 24g, Fructus Crataegi 96g, Semen Arecae 120g, Fructus Evodiae (processed) 12g, Cortex Cinnamomi 12g, Radix Et Rhizoma Nardostachyos 15g, Radix Platycodonis 48g, Radix Scutellariae 96g, Radix Et Rhizoma Rhei 48g, Semen Pharbitidis (parched) 48g makes as crude drug, described Muxiangliqi piece preparation method is made up of the following step: get the Radix Aucklandiae, Radix Et Rhizoma Rhei, Pericarpium Citri Reticulatae, Fructus Aurantii Immaturus, Fructus Aurantii, Radix Et Rhizoma Nardostachyos, join CO
2in supercritical extraction device, ethanol is as entrainer, and the percent by volume that entrainer accounts for total extractant is 4-6%, extracting pressure 15-30MPa, temperature 30-50 DEG C, CO
2flow l-3ml/g crude drug min, extraction time 180-220min, obtains supercritical extract, for subsequent use, get all the other Chinese medicines, pulverize, add 50% ethanol of 1.6L, drop in microwave extracting apparatus and carry out microwave extracting, extraction power 600-800W, extracts 2 times, each 5-10 minute, combining extraction liquid, concentrated, be added on Dl01 macroporous adsorptive resins, 50% ethanol elution, collect 5 times amount column volume eluents, decompression recycling ethanol, concentrated and dry, obtain microwave extraction thing, for subsequent use, by above-mentioned supercritical extract and the mixing of microwave extraction thing, add starch, 70% ethanol granule, dry, tabletting, makes 500, the heavy 0.5g of every sheet.
2. the Muxiangliqi Tablet according to claim l suppresses the application in mouse mammary tumor cells C127 cell proliferation in preparation, it is characterized in that, described CO
2in supercritical extraction, entrainer accounts for the percent by volume of total extractant is 5%.
3. the Muxiangliqi Tablet according to claim l suppresses the application in mouse mammary tumor cells C127 cell proliferation in preparation, it is characterized in that, described CO
2in supercritical extraction, extracting pressure is 25MPa.
4. the Muxiangliqi Tablet according to claim l suppresses the application in mouse mammary tumor cells C127 cell proliferation in preparation, it is characterized in that, described CO
2cO in supercritical extraction
2flow 2ml/g crude drug min.
5. the Muxiangliqi Tablet according to claim l suppresses the application in mouse mammary tumor cells C127 cell proliferation in preparation, it is characterized in that, described CO
2in supercritical extraction, extraction time is 200min.
6. the Muxiangliqi Tablet according to claim l suppresses the application in mouse mammary tumor cells C127 cell proliferation in preparation, and it is characterized in that, described microwave extracting power is 700W.
7. the Muxiangliqi Tablet according to claim l suppresses the application in mouse mammary tumor cells C127 cell proliferation in preparation, and it is characterized in that, each extraction time of described microwave extracting is 8 minutes.
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