CN102988577B - Preparation method and application of brain-soothing hypertension pill - Google Patents
Preparation method and application of brain-soothing hypertension pill Download PDFInfo
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- CN102988577B CN102988577B CN2012103882692A CN201210388269A CN102988577B CN 102988577 B CN102988577 B CN 102988577B CN 2012103882692 A CN2012103882692 A CN 2012103882692A CN 201210388269 A CN201210388269 A CN 201210388269A CN 102988577 B CN102988577 B CN 102988577B
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- 238000002360 preparation method Methods 0.000 title claims abstract description 22
- 206010020772 Hypertension Diseases 0.000 title abstract 4
- 239000006187 pill Substances 0.000 title abstract 4
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- 229940079593 drug Drugs 0.000 claims abstract description 13
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- IKIIZLYTISPENI-ZFORQUDYSA-N baicalin Chemical compound O1[C@H](C(O)=O)[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1OC(C(=C1O)O)=CC2=C1C(=O)C=C(C=1C=CC=CC=1)O2 IKIIZLYTISPENI-ZFORQUDYSA-N 0.000 claims abstract description 12
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- 208000028591 pheochromocytoma Diseases 0.000 claims description 3
- 238000000034 method Methods 0.000 abstract description 10
- 230000002401 inhibitory effect Effects 0.000 abstract description 3
- WTPPRJKFRFIQKT-UHFFFAOYSA-N 1,6-dimethyl-8,9-dihydronaphtho[1,2-g][1]benzofuran-10,11-dione;1-methyl-6-methylidene-8,9-dihydro-7h-naphtho[1,2-g][1]benzofuran-10,11-dione Chemical compound O=C1C(=O)C2=C3CCCC(=C)C3=CC=C2C2=C1C(C)=CO2.O=C1C(=O)C2=C3CCC=C(C)C3=CC=C2C2=C1C(C)=CO2 WTPPRJKFRFIQKT-UHFFFAOYSA-N 0.000 abstract 1
- 241000427159 Achyranthes Species 0.000 abstract 1
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- 241000050051 Chelone glabra Species 0.000 abstract 1
- 244000037364 Cinnamomum aromaticum Species 0.000 abstract 1
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- 201000003352 adrenal gland pheochromocytoma Diseases 0.000 abstract 1
- 235000005272 common selfheal Nutrition 0.000 abstract 1
- SZVJSHCCFOBDDC-UHFFFAOYSA-N iron(II,III) oxide Inorganic materials O=[Fe]O[Fe]O[Fe]=O SZVJSHCCFOBDDC-UHFFFAOYSA-N 0.000 abstract 1
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- WEEMDRWIKYCTQM-UHFFFAOYSA-N 2,6-dimethoxybenzenecarbothioamide Chemical compound COC1=CC=CC(OC)=C1C(N)=S WEEMDRWIKYCTQM-UHFFFAOYSA-N 0.000 description 1
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- 229910052757 nitrogen Inorganic materials 0.000 description 1
- YTJSFYQNRXLOIC-UHFFFAOYSA-N octadecylsilane Chemical compound CCCCCCCCCCCCCCCCCC[SiH3] YTJSFYQNRXLOIC-UHFFFAOYSA-N 0.000 description 1
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Abstract
The invention provides a preparation method of a brain-soothing hypertension pill. The brain-soothing hypertension pill is prepared by using 100g of baical skullcap roots, 60g of common selfheal fruit-spike, 60g of sophora flower bud, 60g of forged lodestone, 60g of twotooth achyranthes roots, 100g of Chinese angelica, 40g of rehmannia, 40g of danshen roots, 20g of sanguisuga, 60g of uncaria rhynchophylla, 100g of semen cassia, 20g of lumbricus and 40g of nacre as bulk drugs and adopting supercritical extraction and microwave-assisted extraction modes. By adopting the method, the content of baicalin is greatly improved; and the invention also provides application of the brain-soothing hypertension pill in preparation of drugs for inhibiting cell proliferation of rat adrenal pheochromocytoma PC-12.
Description
Technical field
The present invention relates to the Chinese medicine preparation technical field, be specifically related to a kind of preparation method and application of Qingnao Jiangyapian.
Background technology
Qingnao Jiangyapian is recorded in Ministry of Public Health standard WS3-B-0424-90, made as crude drug by Radix Scutellariae 100g, Spica Prunellae 60g, Flos Sophorae Immaturus 60g, Magnetitum (calcined) 60g, Radix Achyranthis Bidentatae 60g, Radix Angelicae Sinensis 100g, Radix Rehmanniae 40g, Radix Salviae Miltiorrhizae 40g, Hirudo 20g, Ramulus Uncariae Cum Uncis 60g, Semen Cassiae 100g, Pheretima 20g, Concha Margaritifera 40g, can brain-benefiting blood pressure-reducing.Be used for blood pressure higher, dizzy dizzy, insomnia forgetfulness, hypomnesis.
In prior art, not yet there is Qingnao Jiangyapian to adopt the report of supercritical and microwave technology aspect preparation extracting, and adopts the method that powder and decocting boil of beating, 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 used clinically.
Summary of the invention
Goal of the invention: in order to address the above problem, the object of the present invention is to provide a kind of preparation method of Qingnao Jiangyapian.
Another object of the present invention is to provide the application of a kind of Qingnao Jiangyapian in preparation inhibition Clonal Rat Pheochromocytoma tumor PC-12 cell proliferation medicine.
Technical scheme: the objective of the invention is to realize by following scheme:
A kind of preparation method of Qingnao Jiangyapian, made as crude drug by Radix Scutellariae 100g, Spica Prunellae 60g, Flos Sophorae Immaturus 60g, Magnetitum (calcined) 60g, Radix Achyranthis Bidentatae 60g, Radix Angelicae Sinensis 100g, Radix Rehmanniae 40g, Radix Salviae Miltiorrhizae 40g, Hirudo 20g, Ramulus Uncariae Cum Uncis 60g, Semen Cassiae 100g, Pheretima 20g, Concha Margaritifera 40g, described method is comprised of the following step: get Radix Angelicae Sinensis, 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 ℃, CO
2Flow 1-3m1/g crude drug min, extraction time 150-180min gets supercritical extract, and is standby; Get all the other Chinese medicines, pulverize, add 70% ethanol of 2L, drop in microwave extracting apparatus and carry out microwave extracting, extraction power 400-600W extracts 2 times, each 4-8 minute, combining extraction liquid, concentrated, be added on the D101 macroporous adsorptive resins, 50% ethanol elution is collected 5 times of amount column volume eluents, decompression recycling ethanol, concentrated and dry, get the microwave extraction thing, standby; Above-mentioned supercritical extract and microwave extraction thing are mixed, add starch, 70% ethanol granule processed, drying, tabletting is made 1000, every heavy 0.5g.
The preparation method of above-mentioned a kind of Qingnao Jiangyapian, described CO
2The percent by volume that the supercritical extraction entrainer accounts for total extractant is 5%.
The preparation method of above-mentioned a kind of Qingnao Jiangyapian, described microwave extracting power 500W extracts 6 minutes at every turn.
The preparation method of above-mentioned a kind of Qingnao Jiangyapian, described CO
2The extracting pressure 20MPa of supercritical extraction, 40 ℃ of temperature, CO
2Flow 2ml/g crude drug min, extraction time 160min.
The application of above-mentioned Qingnao Jiangyapian in preparation inhibition PC-12 cell proliferation medicine.
In prior art, every 0.5g of Qingnao Jiangyapian, each 4-6 sheet, 3 times on the one, the every 0.5g of Qingnao Jiangyapian that adopts the present invention to be prepared into only needs 3 at every turn, takes 3 times in 1st, has greatly reduced dose having under the condition of more active component.This conclusion can be by following evidence.
The comparison of content of baicalin in the Qingnao Jiangyapian of test one, distinct methods preparation
1, instrument and reagent Qingnao Jiangyapian of the present invention: press embodiment 3 method preparations, use the 665g crude drug, make 1000 through extraction, every heavy 0.5g.Former Qingnao Jiangyapian by the preparation of ministry standard method, uses the 665g crude drug, makes 1000 through extraction, every heavy 0.5g.The Agilent1200 high performance liquid chromatograph; The METTLERAE240 electronic analytical balance; Baicalin reference substance (Nat'l Pharmaceutical ﹠ Biological Products Control Institute).
2, method
Chromatographic condition and system suitability: be filler with octadecylsilane chemically bonded silica; Methanol-water-phosphoric acid (40:60:0.2) is mobile phase; The detection wavelength is 280nm.Number of theoretical plate is pressed baicalin peak calculating, should be not less than 3000.
The preparation of reference substance solution: precision takes at 60 ℃ of drying under reduced pressure baicalin reference substance of 4 hours appropriate, adds methanol and makes the solution that every 1ml contains 18 μ g, and get final product.
The preparation of need testing solution: get Qingnao Jiangyapian of the present invention and former Qingnao Jiangyapian, porphyrize, mixing is got 1g, and is accurately weighed, and precision adds 70% ethanol 20ml, close plug, supersound process 10 minutes, centrifugal, get supernatant, and get final product.
Algoscopy is accurate reference substance solution and each 20 μ l of need testing solution of drawing respectively, and the injection liquid chromatography is measured, and be get final product.
3, result
Result shows, in Qingnao Jiangyapian of the present invention, the content of baicalin is the 2.43mg/ sheet; And in former Qingnao Jiangyapian, the content of baicalin is the 0.56mg/ sheet, in the situation that dose reduces, content of baicalin improves a lot.
Above-mentioned studies show that, the Qingnao Jiangyapian that adopts the present invention to prepare, active constituent content is higher than the standby Qingnao Jiangyapian of ministry standard legal system.
The specific embodiment
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 only limits to following example, all technology that realizes based on foregoing of the present invention all belong to scope of the present invention.
Embodiment 1
Get Radix Scutellariae 100g, Spica Prunellae 60g, Flos Sophorae Immaturus 60g, Magnetitum (calcined) 60g, Radix Achyranthis Bidentatae 60g, Radix Angelicae Sinensis 100g, Radix Rehmanniae 40g, Radix Salviae Miltiorrhizae 40g, Hirudo 20g, Ramulus Uncariae Cum Uncis 60g, Semen Cassiae 100g, Pheretima 20g, Concha Margaritifera 40g, with Radix Angelicae Sinensis, join CO
2In the supercritical extraction device, ethanol is as entrainer, and the percent by volume that entrainer accounts for total extractant is 4%, extracting pressure 15MPa, and 30 ℃ of temperature, CO2 flow 1m1/g crude drug min, extraction time 150min gets supercritical extract, and is standby; Get all the other Chinese medicines, pulverize, add 70% ethanol of 2L, drop in microwave extracting apparatus and carry out microwave extracting, extraction power 400W extracts 2 times, each 4 minutes, combining extraction liquid, concentrated, be added on the D101 macroporous adsorptive resins, 50% ethanol elution is collected 5 times of amount column volume eluents, decompression recycling ethanol, concentrated and dry, get the microwave extraction thing, standby; Above-mentioned supercritical extract and microwave extraction thing are mixed, add starch, 70% ethanol granule processed, drying, tabletting is made 1000, every heavy 0.5g.
After testing, in finished product, the content of baicalin is the 2.48mg/ sheet.
Embodiment 2
Get Radix Scutellariae 100g, Spica Prunellae 60g, Flos Sophorae Immaturus 60g, Magnetitum (calcined) 60g, Radix Achyranthis Bidentatae 60g, Radix Angelicae Sinensis 100g, Radix Rehmanniae 40g, Radix Salviae Miltiorrhizae 40g, Hirudo 20g, Ramulus Uncariae Cum Uncis 60g, Semen Cassiae 100g, Pheretima 20g, Concha Margaritifera 40g, with Radix Angelicae Sinensis, join CO
2In the 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 gets supercritical extract, and is standby; Get all the other Chinese medicines, pulverize, add 70% ethanol of 2L, drop in microwave extracting apparatus and carry out microwave extracting, extraction power 600W extracts 2 times, each 8 minutes, combining extraction liquid, concentrated, be added on the D101 macroporous adsorptive resins, 50% ethanol elution is collected 5 times of amount column volume eluents, decompression recycling ethanol, concentrated and dry, get the microwave extraction thing, standby; Above-mentioned supercritical extract and microwave extraction thing are mixed, add starch, 70% ethanol granule processed, drying, tabletting is made 1000, every heavy 0.5g.
After testing, in finished product, the content of baicalin is the 2.33mg/ sheet.
Embodiment 3
Get Radix Scutellariae 100g, Spica Prunellae 60g, Flos Sophorae Immaturus 60g, Magnetitum (calcined) 60g, Radix Achyranthis Bidentatae 60g, Radix Angelicae Sinensis 100g, Radix Rehmanniae 40g, Radix Salviae Miltiorrhizae 40g, Hirudo 20g, Ramulus Uncariae Cum Uncis 60g, Semen Cassiae 100g, Pheretima 20g, Concha Margaritifera 40g, with Radix Angelicae Sinensis, join CO
2In the supercritical extraction device, ethanol is as entrainer, and the percent by volume that entrainer accounts for total extractant is 5%, extracting pressure 20MPa, 40 ℃ of temperature, CO
2Flow 2m1/g crude drug min, extraction time 160min gets supercritical extract, and is standby; Get all the other Chinese medicines, pulverize, add 70% ethanol of 2L, drop in microwave extracting apparatus and carry out microwave extracting, extraction power 500W extracts 2 times, each 6 minutes, combining extraction liquid, concentrated, be added on the D101 macroporous adsorptive resins, 50% ethanol elution is collected 5 times of amount column volume eluents, decompression recycling ethanol, concentrated and dry, get the microwave extraction thing, standby; Above-mentioned supercritical extract and microwave extraction thing are mixed, add starch, 70% ethanol granule processed, drying, tabletting is made 1000, every heavy 0.5g.
After testing, in finished product, the content of baicalin is the 2.43mg/ sheet.
Embodiment 4: Qingnao Jiangyapian suppresses the experimentation data of PC-12 cell proliferation
1 experiment material
1.1 experiment cell strain
Clonal Rat Pheochromocytoma tumor (PC-12), Nanjing Zhengkuan Pharmaceutical Technology Co., Ltd.'s laboratory cell bank, DMEM+10%FBS cellar culture.
1.2 Experimental agents
Drugs: Qingnao Jiangyapian of the present invention: press embodiment 3 method preparations.
The medicinal liquid liquid storage: take the 100mg Qingnao Jiangyapian, be dissolved in the 5ml dehydrated alcohol, 0.2 μ m filter filters, 500 μ ldoff pipe packing, and-20 ℃ of storages, 0.2 μ m filter filters dehydrated alcohol in order to the use of matched group simultaneously.
1.3 experiment reagent
The Cat.No.12100-061Lot.No.758137 of DMEM(GIBCO company); Hyclone (Hangzhoupro, sky, Zhejiang bio tech ltd 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); Penicillin G Sodium Salt(AMRESCO company lot number: 2010242); Streptomycin Sulfate(AMRESCO company lot number: 2010382); Dehydrated alcohol (Nanjing Chemistry Reagent Co., Ltd.'s lot number: 080310182); MTT (Biosharp lot number: 0793); The autogamy of PBS(laboratory); 1.4 experiment equipment
Lycra inverted microscope (German Leica model: DM1L); As seen-ultraviolet light microwell plate detector (U.S. MD company model: SPECTRAMAX190); CO2 incubator (FORMA model: 3111); (safe and sound company of Su Jing group makes model to super-clean bench: SW-CJ-ZFD); Pure water instrument (U.S. Spring company 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 (Shanghai accurate experimental facilities company 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 μ m 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) the PC-12 cell carries out cellar culture (10cm culture dish) with DMEM+10%FBS in 37 ℃, 5%CO2, when Growth of Cells during to logarithmic (log) phase, collecting cell discards culture fluid, PBS fine laundering 3 times, add 3ml0.25% trypsin-0.04%EDTA, after 37 ℃ of digestion 2min, add wherein 5ml complete medium neutralization reaction, after the piping and druming cell, it is changed in centrifuge tube, the centrifugal 5min of 1000rpm adjusts 3 * 104/ml of concentration of cell suspension.
2) the cell kind is entered in 96 well culture plates, every hole adds cell suspension 180 μ l, culture plate put into cell culture incubator (37 ℃, 5%CO2) cellar culture.
3) according to the Growth of Cells situation, generally grow to 50%-70%, add Qingnao Jiangyapian solution, continue to cultivate 24h.
4) add 20 μ l MTT solution (5mg/ml, i.e. 0.5%MTT) after 24h, continue to cultivate 4h.
5) after 4h, the buckle method is removed supernatant, pats dry gently with absorbent paper, and every hole adds 200 μ l dimethyl sulfoxide, puts low-speed oscillation 10min on shaking table, and crystal is fully dissolved.Light absorption value in each hole of measurement, enzyme-linked immunosorbent assay instrument 490nm place.6) background (do not add cell, only add culture fluid) is set simultaneously, control wells (the medicine dissolution medium of cell, same concentrations, culture fluid, MTT, dimethyl sulfoxide) is set 6 multiple holes for every group.
7) result represents with the suppression ratio of medicine to cell:
Cell increment suppression ratio (%)=(control wells OD value-dosing holes OD value)/control wells OD value * 100%.Experiment repeats 3 times.
3 statistical dispositions
Adopt correlation analysis and Student t check in Microsoft Excel2003 software, data represent with mean ± S.D..
4 experimental results
Statistical result showed after the mtt assay experiment, compare with matched group, when dosage reaches 5mg/ml, to PC-12 cell inhibitory effect variant (P＜0.05), dosage this difference when 10mg/ml has significance (P＜0.01), and utmost point significant difference (P＜0.001) is arranged when dosage reaches 15-20mg/ml.
Table 1 Qingnao Jiangyapian affects the PC-12 cell inhibitory effect and grinds
Annotate: compare * P＜0.01 with matched group; * P＜0.001
5 experiment conclusion
Qingnao Jiangyapian can suppress the PC-12 cell proliferation, reduces the Growth of Cells number of PC-12 cell, and this effect is dose dependent.
Claims (1)
1. the application of Qingnao Jiangyapian in preparation inhibition Clonal Rat Pheochromocytoma tumor PC-12 cell proliferation medicine, the preparation method of above-mentioned Qingnao Jiangyapian is for getting Radix Scutellariae 100g, Spica Prunellae 60g, Flos Sophorae Immaturus 60g, Magnetitum (calcined) 60g, Radix Achyranthis Bidentatae 60g, Radix Angelicae Sinensis 100g, Radix Rehmanniae 40g, Radix Salviae Miltiorrhizae 40g, Hirudo 20g, Ramulus Uncariae Cum Uncis 60g, Semen Cassiae 100g, Pheretima 20g, Concha Margaritifera 40g, with Radix Angelicae Sinensis, join CO
2In the supercritical extraction device, ethanol is as entrainer, and the percent by volume that entrainer accounts for total extractant is 5%, extracting pressure 20MPa, 40 ℃ of temperature, CO
2Flow 2m1/g crude drug min, extraction time 160min gets supercritical extract, and is standby; Get all the other Chinese medicines, pulverize, add 70% ethanol of 2L, drop in microwave extracting apparatus and carry out microwave extracting, extraction power 500W extracts 2 times, each 6 minutes, combining extraction liquid, concentrated, be added on the D101 macroporous adsorptive resins, 50% ethanol elution is collected 5 times of amount column volume eluents, decompression recycling ethanol, concentrated and dry, get the microwave extraction thing, standby; Above-mentioned supercritical extract and microwave extraction thing are mixed, add starch, 70% ethanol granule processed, drying, tabletting is made 1000, every heavy 0.5g, after testing, in finished product, the content of baicalin is the 2.43mg/ sheet.
Priority Applications (1)
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| CN104997874A (en) * | 2015-08-19 | 2015-10-28 | 陆茹姣 | Traditional Chinese medicine for treating pheochromocytoma |
| CN105497259A (en) * | 2016-01-04 | 2016-04-20 | 四川双鑫生物科技有限公司 | Orally-taken medicine for treating dizziness caused by hyperactivity of liver-yang and preparation method thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1366915A (en) * | 2001-01-22 | 2002-09-04 | 杨孟君 | Nano brain-benefiting blood pressure-reducing preparation medicine and preparation method |
| CN102397394A (en) * | 2011-11-26 | 2012-04-04 | 苏州派腾生物医药科技有限公司 | Preparation method of cephalocathartic hypertension pill |
-
2012
- 2012-10-15 CN CN2012103882692A patent/CN102988577B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1366915A (en) * | 2001-01-22 | 2002-09-04 | 杨孟君 | Nano brain-benefiting blood pressure-reducing preparation medicine and preparation method |
| CN102397394A (en) * | 2011-11-26 | 2012-04-04 | 苏州派腾生物医药科技有限公司 | Preparation method of cephalocathartic hypertension pill |
Non-Patent Citations (4)
| Title |
|---|
| 清脑降压滴丸的制备;郑雅楠等;《现代药物与临床》;20110731;第26卷(第4期);299-301 * |
| 清脑降压片的质量标准研究;邓永中等;《广西中医学院学报》;20081231;第11卷(第2期);52-54 * |
| 邓永中等.清脑降压片的质量标准研究.《广西中医学院学报》.2008,第11卷(第2期),52-54. |
| 郑雅楠等.清脑降压滴丸的制备.《现代药物与临床》.2011,第26卷(第4期),299-301. |
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