CN103656600B - A kind of preparation method of 12 WENJING WAN and application - Google Patents
A kind of preparation method of 12 WENJING WAN and application Download PDFInfo
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- CN103656600B CN103656600B CN201310712990.7A CN201310712990A CN103656600B CN 103656600 B CN103656600 B CN 103656600B CN 201310712990 A CN201310712990 A CN 201310712990A CN 103656600 B CN103656600 B CN 103656600B
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Abstract
The invention provides a kind of preparation method of 12 WENJING WAN, by Fructus Evodiae 90g, Radix Angelicae Sinensis 60g, Rhizoma Chuanxiong (processed with wine) 60g, Radix Paeoniae Alba 60g, Colla Corii Asini bead 60g, Cortex Cinnamomi 60g, Cortex Moutan 60g, Rhizoma Zingiberis Recens 15g, Radix Codonopsis 60g, Rhizoma Pinelliae (processed) 90g, Radix Ophiopogonis 120g, Radix Glycyrrhizae (processed with honey) 60g makes as crude drug, employing supercritical extraction is prepared from, active constituent content is increased, dose reduces, and present invention also offers the application of 12 WENJING WAN in preparation suppression human melanoma cell M14 cell proliferation.
Description
Technical field
The present invention relates to technical field of traditional Chinese medicine preparation, be specifically related to a kind of preparation method and application of 12 WENJING WAN.
Background technology
12 WENJING WAN are recorded in Ministry of Public Health standard WS3-B-0005-89, prescription be Fructus Evodiae 90g, Radix Angelicae Sinensis 60g, Rhizoma Chuanxiong (processed with wine) 60g, Radix Paeoniae Alba 60g, Colla Corii Asini bead 60g, Cortex Cinnamomi 60g, Cortex Moutan 60g, Rhizoma Zingiberis Recens 15g, Radix Codonopsis 60g, Rhizoma Pinelliae (processed) 90g, Radix Ophiopogonis 120g, Radix Glycyrrhizae (processed with honey) 60g, above 12 tastes, be ground into fine powder, sieve, mixing.Every 100g powder adds refined honey 50g and appropriate water, general ball, and cold drying, to obtain final product, energy dispelling cold by warming the meridian, nourishing blood and dissolving stasis.For the cold and deficiency of CHONG and REN meridians, stagnation of blood stasis, menoxenia, or first or after, more or less, cold and pain in the lower abdomen and infertility.Promoting flow of QI and blood, relieving gastric hyperacidity to alleviate stomachache.
In prior art, not yet have 12 WENJING WAN extracting the report adopting supercritical technology in preparation, and adopt the method for beating powder, technique is coarse, backward, and impurity is many, causes patient's consumption excessive, is inconvenient to take, and has had a strong impact on this product and has applied clinically.
Summary of the invention
Goal of the invention: in order to solve the problem, the object of the present invention is to provide a kind of preparation method of 12 WENJING WAN.
Another object of the present invention is to provide a kind of 12 WENJING WAN to suppress the application in human melanoma cell M14 cell proliferation in preparation.
Technical scheme: the object of the invention is by following scheme realize:
A kind of preparation method of 12 WENJING WAN, by Fructus Evodiae 90g, Radix Angelicae Sinensis 60g, Rhizoma Chuanxiong (processed with wine) 60g, Radix Paeoniae Alba 60g, Colla Corii Asini bead 60g, Cortex Cinnamomi 60g, Cortex Moutan 60g, Rhizoma Zingiberis Recens 15g, Radix Codonopsis 60g, Rhizoma Pinelliae (processed) 90g, Radix Ophiopogonis 120g, Radix Glycyrrhizae (processed with honey) 60g makes as crude drug, it is characterized in that described method is made up of the following step: get Fructus Evodiae 90g, Radix Angelicae Sinensis 60g, Rhizoma Chuanxiong (processed with wine) 60g, Radix Paeoniae Alba 60g, Colla Corii Asini bead 60g, Cortex Cinnamomi 60g, Cortex Moutan 60g, Rhizoma Zingiberis Recens 15g, Radix Codonopsis 60g, Rhizoma Pinelliae (processed) 90g, Radix Ophiopogonis 120g, Radix Glycyrrhizae (processed with honey) 60g, 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 1-3m1/g crude drug min, extraction time 150-180min, obtains supercritical extract, is ground into fine powder, sieves, mixing, and every 100g powder adds refined honey 50g and appropriate water, general ball, and cold drying obtains 600g water pill, every heavy 0.3g of ball.
The preparation method of above-mentioned a kind of 12 WENJING WAN, described CO
2the percent by volume that supercritical extraction entrainer accounts for total extractant is 5%.
The preparation method of above-mentioned a kind of 12 WENJING WAN, described CO
2the extracting pressure 20MPa of supercritical extraction, temperature 40 DEG C, CO
2flow 2ml/g crude drug min, extraction time 160min.
Above-mentioned 12 WENJING WAN suppress the application in human melanoma cell M14 cell proliferation in preparation, 12 WENJING WAN are by Fructus Evodiae 90g, Radix Angelicae Sinensis 60g, Rhizoma Chuanxiong (processed with wine) 60g, Radix Paeoniae Alba 60g, Colla Corii Asini bead 60g, Cortex Cinnamomi 60g, Cortex Moutan 60g, Rhizoma Zingiberis Recens 15g, Radix Codonopsis 60g, Rhizoma Pinelliae (processed) 90g, Radix Ophiopogonis 120g, Radix Glycyrrhizae (processed with honey) 60g makes as crude drug, it is characterized in that described method is made up of the following step: get Fructus Evodiae 90g, Radix Angelicae Sinensis 60g, Rhizoma Chuanxiong (processed with wine) 60g, Radix Paeoniae Alba 60g, Colla Corii Asini bead 60g, Cortex Cinnamomi 60g, Cortex Moutan 60g, Rhizoma Zingiberis Recens 15g, Radix Codonopsis 60g, Rhizoma Pinelliae (processed) 90g, Radix Ophiopogonis 120g, Radix Glycyrrhizae (processed with honey) 60g, 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 1-3m1/g crude drug min, extraction time 150-180min, obtains supercritical extract, is ground into fine powder, sieves, mixing, and every 100g powder adds refined honey 50g and appropriate water, general ball, and cold drying obtains 600g water pill, every heavy 0.3g of ball.
Above-mentioned 12 WENJING WAN suppress the application in human melanoma cell M14 cell proliferation in preparation, CO described in 12 WENJING WAN preparation methoies
2the percent by volume that supercritical extraction entrainer accounts for total extractant is 5%.
Above-mentioned 12 WENJING WAN suppress the application in human melanoma cell M14 cell proliferation in preparation, CO described in 12 WENJING WAN preparation methoies
2the extracting pressure 20MPa of supercritical extraction, temperature 40 DEG C, CO
2flow 2ml/g crude drug min, extraction time 160min.
In prior art, the every ball 0.3g of 12 WENJING WAN, each 6-9g, 2 times on the one, adopt the every ball 0.3g of 12 WENJING WAN that the present invention is prepared into, but the medical material amount contained is original 2 times, therefore only need 3-4.5g at every turn, within 1st, take 2 times, under the condition with more active component, greatly reduce dose, add the compliance of medicine.
Detailed description of the invention
Form by the following examples, foregoing of the present invention is described in further detail again, 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 Fructus Evodiae 90g, Radix Angelicae Sinensis 60g, Rhizoma Chuanxiong (processed with wine) 60g, Radix Paeoniae Alba 60g, Colla Corii Asini bead 60g, Cortex Cinnamomi 60g, Cortex Moutan 60g, Rhizoma Zingiberis Recens 15g, Radix Codonopsis 60g, Rhizoma Pinelliae (processed) 90g, Radix Ophiopogonis 120g, Radix Glycyrrhizae (processed with honey) 60g, join in CO2 supercritical extraction device, ethanol is as entrainer, the percent by volume that entrainer accounts for total extractant is 4%, extracting pressure 15MPa, temperature 30 DEG C, CO2 flow 1m1/g crude drug min, extraction time 150min, obtain supercritical extract, be ground into fine powder, sieve, mixing, every 100g powder adds refined honey 50g and appropriate water, general ball, cold drying, obtain 600g water pill, every heavy 0.3g of ball.
Embodiment 2
Get Fructus Evodiae 90g, Radix Angelicae Sinensis 60g, Rhizoma Chuanxiong (processed with wine) 60g, Radix Paeoniae Alba 60g, Colla Corii Asini bead 60g, Cortex Cinnamomi 60g, Cortex Moutan 60g, Rhizoma Zingiberis Recens 15g, Radix Codonopsis 60g, Rhizoma Pinelliae (processed) 90g, Radix Ophiopogonis 120g, Radix Glycyrrhizae (processed with honey) 60g, join CO
2in supercritical extraction device, ethanol is as entrainer, and the percent by volume that entrainer accounts for total extractant is 6%, extracting pressure 30MPa, temperature 50 C, CO
2flow 3m1/g crude drug min, extraction time 180min, obtains supercritical extract, is ground into fine powder, sieves, mixing, and every 100g powder adds refined honey 50g and appropriate water, general ball, and cold drying obtains 600g water pill, every heavy 0.3g of ball.
Embodiment 3
Get Fructus Evodiae 90g, Radix Angelicae Sinensis 60g, Rhizoma Chuanxiong (processed with wine) 60g, Radix Paeoniae Alba 60g, Colla Corii Asini bead 60g, Cortex Cinnamomi 60g, Cortex Moutan 60g, Rhizoma Zingiberis Recens 15g, Radix Codonopsis 60g, Rhizoma Pinelliae (processed) 90g, Radix Ophiopogonis 120g, Radix Glycyrrhizae (processed with honey) 60g, join CO
2in supercritical extraction device, ethanol is as entrainer, and the percent by volume that entrainer accounts for total extractant is 5%, extracting pressure 20MPa, temperature 40 DEG C, CO
2flow 2m1/g crude drug min, extraction time 160min, obtains supercritical extract, is ground into fine powder, sieves, mixing, and every 100g powder adds refined honey 50g and appropriate water, general ball, and cold drying obtains 600g water pill, every heavy 0.3g of ball.
Embodiment 4: ten two WENJING WAN suppresses the experimentation data of M14 cell proliferation
1 experiment material
1.1 experiment cell strains
Human melanoma cell M14 cell, Nanjing Medical University's laboratory cell storehouse, DMEM+10%FBS cellar culture.
1.2 Experimental agents
Drugs: the present invention 12 WENJING WAN: prepare by embodiment 3 method.
Medicinal liquid liquid storage: take 100mg 12 WENJING WAN, 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); NaHCO3(Shanghai hundred million chemical reagent company limited Cat.No.11810-033Lot.No.1088387 of a specified duration); Trypsin(AMRESCO company lot number: 2010/04); EDTA(AMRESCO company lot number: 2009/10); PenicillinGSodiumSalt(AMRESCO company lot number: 2010242); StreptomycinSulfate(AMRESCO company lot number: 2010382); Dehydrated alcohol (Nanjing Chemistry Reagent Co., Ltd.'s lot number: 080310182); MTT (Biosharp lot number: 0793); PBS(laboratory autogamy);
1.4 experiment equipment
Lycra inverted microscope (German Leica model: DM1L); Visible-ultraviolet light microwell plate detector (MD company of U.S. model: SPECTRAMAX190); CO2 incubator (FORMA model: 3111); Super-clean bench (safe and sound Inc. of Su Jing group moulding number: SW-CJ-ZFD); Pure water instrument (Spring 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); 10cm culture dish (NEST company), 96 well culture plates (NEST company); Cell counting count board; Centrifuge tube, pipet, Tips are some.
2 experimental techniques
1) M14 cell DMEM+10%FBS in 37 DEG C, 5%CO2 carries 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 × 104/ml.
2) enter in 96 well culture plates by cell kind, every hole adds cell suspension 180 μ l, and culture plate puts into cell culture incubator (37 DEG C, 5%CO2) cellar culture.
3) according to cell growth status, generally grow to 50%-70%, add 12 WENJING WAN 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 dispositions
Adopt the correlation analysis in MicrosoftExcel2003 software and Studentt inspection, data represent with mean ± S.D..
4 experimental results
Statistical result showed after mtt assay experiment, compare with matched group, when dosage reaches 5mg/ml, to M14 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 12 WENJING WAN is to M14 cell inhibitory effect influence research
Note: compare with matched group,
*p<0.01;
*p<0.001
5 experiment conclusion
12 WENJING WAN can suppress M14 cell proliferation, and reduce the Growth of Cells number of M14 cell, this effect is dose dependent.
Claims (3)
1. the application of WENJING WAN in preparation suppression human melanoma cell M14 cell proliferation, it is characterized in that 12 WENJING WAN are by Fructus Evodiae 90g, Radix Angelicae Sinensis 60g, Rhizoma Chuanxiong (processed with wine) 60g, Radix Paeoniae Alba 60g, Colla Corii Asini bead 60g, Cortex Cinnamomi 60g, Cortex Moutan 60g, Rhizoma Zingiberis Recens 15g, Radix Codonopsis 60g, Rhizoma Pinelliae Preparata 90g, Radix Ophiopogonis 120g, Radix Glycyrrhizae (processed with honey) 60g makes as crude drug, it is characterized in that described method is made up of the following step: get Fructus Evodiae 90g, Radix Angelicae Sinensis 60g, Rhizoma Chuanxiong (processed with wine) 60g, Radix Paeoniae Alba 60g, Colla Corii Asini bead 60g, Cortex Cinnamomi 60g, Cortex Moutan 60g, Rhizoma Zingiberis Recens 15g, Radix Codonopsis 60g, Rhizoma Pinelliae Preparata 90g, Radix Ophiopogonis 120g, Radix Glycyrrhizae (processed with honey) 60g, 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 1-3m1/g crude drug min, extraction time 150-180min, obtains supercritical extract, is ground into fine powder, sieves, mixing, and every 100g powder adds refined honey 50g and appropriate water, general ball, and cold drying obtains 600g water pill, every heavy 0.3g of ball.
2. a kind of 12 WENJING WAN suppress the application in human melanoma cell M14 cell proliferation in preparation according to claim 1, it is characterized in that CO described in 12 WENJING WAN preparation methoies
2the percent by volume that supercritical extraction entrainer accounts for total extractant is 5%.
3. a kind of 12 WENJING WAN suppress the application in human melanoma cell M14 cell proliferation in preparation according to claim 1, it is characterized in that CO described in 12 WENJING WAN preparation methoies
2the extracting pressure 20MPa of supercritical extraction, temperature 40 DEG C, CO
2flow 2ml/g crude drug min, extraction time 160min.
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| CN112426491A (en) * | 2020-08-15 | 2021-03-02 | 仲崇允 | Traditional Chinese medicine composition for treating malignant melanoma |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102988665A (en) * | 2012-10-08 | 2013-03-27 | 卞毓平 | Method for preparing gynecological menstruation regulating tablets and application |
| CN102988489A (en) * | 2012-10-08 | 2013-03-27 | 卞毓平 | Preparation method and application of nose comforting tablet |
| CN102988667A (en) * | 2012-10-15 | 2013-03-27 | 李正梅 | Preparation method and application of tablet for regulating menstruation and activating blood |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102988665A (en) * | 2012-10-08 | 2013-03-27 | 卞毓平 | Method for preparing gynecological menstruation regulating tablets and application |
| CN102988489A (en) * | 2012-10-08 | 2013-03-27 | 卞毓平 | Preparation method and application of nose comforting tablet |
| CN102988667A (en) * | 2012-10-15 | 2013-03-27 | 李正梅 | Preparation method and application of tablet for regulating menstruation and activating blood |
Non-Patent Citations (2)
| Title |
|---|
| "十二温经丸";医学教育网;《网址：http://www.med66.com/new/40a187a2009/2009123zhangz203134.shtml》;20091203;1 * |
| "继发性闭经中医药治疗概况";蒋铃 等;《辽宁中医药大学学报》;20100731;第12卷(第7期);205-208 * |
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