CN104940426A - Preparation method and application of osteoproliferation removal tablets - Google Patents
Preparation method and application of osteoproliferation removal tablets Download PDFInfo
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- CN104940426A CN104940426A CN201510424205.7A CN201510424205A CN104940426A CN 104940426 A CN104940426 A CN 104940426A CN 201510424205 A CN201510424205 A CN 201510424205A CN 104940426 A CN104940426 A CN 104940426A
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Abstract
The invention belongs to the field of traditional Chinese medicine preparations, and particularly provides a preparation method of osteoproliferation removal tablets. The osteoproliferation removal tablets comprise the active pharmaceutical ingredients of 208 g of prepared rehmannia roots, 140 g of caulis spatholobi, 140 g of herba epimedii, 140 g of rhizoma drynariae, 140 g of rhizoma cibotii, 70 g of semen raphani, 70 g of wolfberry fruits, 140 g of cynomorium songaricum and 70 g of himalayan teasel roots. The osteoproliferation removal tablets are prepared from cationic resin in a supercritical extraction mode, the content is greatly increased, and the dosage is decreased. The invention further provides the application of the osteoproliferation removal tablets in preparing medicine for restraining proliferation of human tissue cell lymphoma U937 cells.
Description
Technical field
The present invention relates to technical field of traditional Chinese medicine preparation, be specifically related to preparation method and application that a kind of bone increases the sheet that disappears.
Background technology
Bone increases the sheet that disappears and is recorded in Ministry of Public Health standard, be made up as crude drug of Radix Rehmanniae Preparata 208g, Caulis Spatholobi 140g, Herba Epimedii 140g, Rhizoma Drynariae 140g, Rhizoma Cibotii 140g, Semen Raphani 70g, Fructus Lycii 70g, Herba Cynomorii 140g, interrupted 70g, above nine tastes, Rhizoma Cibotii, Herba Epimedii are ground into fine powder, seven tastes such as all the other Radix Rehmanniae Preparata, decoct with water secondary, each 1.5 hours, gradation filters, merging filtrate is also evaporated to the thick paste that relative density is 1.3-1.35, mixes with above-mentioned fine powder, dry, pulverize, make granule, dry, tabletting, sugar coating or film-coat, to obtain final product.The liver and the kidney tonifying, invigorates blood circulation.For bone matrix hyperplasia such as the waist kneecap joint aches caused by Liver and kidney two void.
In prior art, not yet there is bone to increase and disappear sheet at the report extracting employing cationic resin and supercritical technology in preparation, and adopt the method for pulverizing and 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.
In prior art, green bone increases the every sheet 0.5g of sheet that disappears, each 2,3 times on the one, and the bone adopting the present invention to be prepared into increases the every sheet 0.3g of sheet that disappears, but its active component contained increases greatly.
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 bone to increase the preparation method of the sheet that disappears.
The application that another object of the present invention is to provide a kind of bone to increase to disappear sheet to suppress in human tissue cell lymphoma U937 cell proliferation in preparation.
Technical scheme: the object of the invention is by following scheme realize:
A kind of bone increases the preparation method of the sheet that disappears, by Radix Rehmanniae Preparata 208g, Caulis Spatholobi 140g, Herba Epimedii 140g, Rhizoma Drynariae 140g, Rhizoma Cibotii 140g, Semen Raphani 70g, Fructus Lycii 70g, Herba Cynomorii 140g, interrupted 70g makes as crude drug, described method is made up of the following step: get Radix Rehmanniae Preparata 208g, Caulis Spatholobi 140g, Herba Epimedii 140g, Rhizoma Drynariae 140g, Rhizoma Cibotii 140g, Semen Raphani 70g, Fructus Lycii 70g, Herba Cynomorii 140g, interrupted 70g, pulverize, add 20 respectively, 16, the water boiling and extraction of 14 times amount three times, each 1 hour, merge three extracting solution, filter, remove slag and get liquid, relative density 1.10 when being concentrated into 65 DEG C, adding ethanol makes the percent by volume of alcohol content reach 50%, precipitation, leave standstill, filter, obtain alcohol deposit fluid, alcohol deposit fluid is adsorbed through 10kg cation exchange resin 001 × 7, loading flow velocity is 2ml/min, effluent is for subsequent use, the ammonia of cation exchange resin 10L 0.5mol/L carries out eluting, collect eluent, merge the effluent after cationic exchange resin adsorption and ammonia eluent, reclaim ethanol, 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 1-3m1/g crude drug min, extraction time 150-180min, obtain supercritical extract, add starch, 70% ethanol granule, dry, tabletting, make 1000, the heavy 0.3g of every sheet.
The percent by volume that described CO2 supercritical extraction entrainer accounts for total extractant is 5%.
The extracting pressure 20MPa of described CO2 supercritical extraction, temperature 40 DEG C, CO2 flow 2ml/g crude drug min, extraction time 160min.
Described bone increases the application of sheet in preparation suppression human tissue cell lymphoma U937 cell proliferation that disappear, described method is made up of the following step: get Radix Rehmanniae Preparata 208g, Caulis Spatholobi 140g, Herba Epimedii 140g, Rhizoma Drynariae 140g, Rhizoma Cibotii 140g, Semen Raphani 70g, Fructus Lycii 70g, Herba Cynomorii 140g, interrupted 70g, pulverize, add 20 respectively, 16, the water boiling and extraction of 14 times amount three times, each 1 hour, merge three extracting solution, filter, remove slag and get liquid, relative density 1.10 when being concentrated into 65 DEG C, adding ethanol makes the percent by volume of alcohol content reach 50%, precipitation, leave standstill, filter, obtain alcohol deposit fluid, alcohol deposit fluid is adsorbed through 10kg cation exchange resin 001 × 7, loading flow velocity is 2ml/min, effluent is for subsequent use, the ammonia of cation exchange resin 10L 0.5mol/L carries out eluting, collect eluent, merge the effluent after cationic exchange resin adsorption and ammonia eluent, reclaim ethanol, 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 1-3m1/g crude drug min, extraction time 150-180min, obtain supercritical extract, add starch, 70% ethanol granule, dry, tabletting, make 1000, the heavy 0.3g of every sheet.
Described bone increases the sheet that disappears suppresses in human tissue cell lymphoma U937 cell proliferation application in preparation, and described in the preparation method that bone increases the sheet that disappears, CO2 supercritical extraction entrainer accounts for the percent by volume of total extractant is 5%.
Described bone increases the application of sheet in preparation suppression human tissue cell lymphoma U937 cell proliferation that disappear, the extracting pressure 20MPa of CO2 supercritical extraction described in the preparation method that bone increases the sheet that disappears, temperature 40 DEG C, CO2 flow 2ml/g crude drug min, extraction time 160min.
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 Radix Rehmanniae Preparata 208g, Caulis Spatholobi 140g, Herba Epimedii 140g, Rhizoma Drynariae 140g, Rhizoma Cibotii 140g, Semen Raphani 70g, Fructus Lycii 70g, Herba Cynomorii 140g, interrupted 70g, pulverize, adds the water boiling and extraction three times of 20,16,14 times amount respectively, each 1 hour, merge three extracting solution, filter, remove slag and get liquid, relative density 1.10 when being concentrated into 65 DEG C, adding ethanol makes the percent by volume of alcohol content reach 50%, precipitation, leaves standstill, filter, obtain alcohol deposit fluid, alcohol deposit fluid is adsorbed through 10kg cation exchange resin 001 × 7, loading flow velocity is 2ml/min, effluent is for subsequent use, the ammonia of cation exchange resin 10L 0.5mol/L carries out eluting, collect eluent, merge the effluent after cationic exchange resin adsorption and ammonia eluent, reclaim ethanol, 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 50 C, CO2 flow 1m1/g crude drug min, extraction time 180min, obtain supercritical extract, add starch, 70% ethanol granule, dry, tabletting, make 1000, the heavy 0.3g of every sheet.
Embodiment 2: get Radix Rehmanniae Preparata 208g, Caulis Spatholobi 140g, Herba Epimedii 140g, Rhizoma Drynariae 140g, Rhizoma Cibotii 140g, Semen Raphani 70g, Fructus Lycii 70g, Herba Cynomorii 140g, interrupted 70g, pulverize, adds the water boiling and extraction three times of 20,16,14 times amount respectively, each 1 hour, merge three extracting solution, filter, remove slag and get liquid, relative density 1.10 when being concentrated into 65 DEG C, adding ethanol makes the percent by volume of alcohol content reach 50%, precipitation, leaves standstill, filter, obtain alcohol deposit fluid, alcohol deposit fluid is adsorbed through 10kg cation exchange resin 001 × 7, loading flow velocity is 2ml/min, effluent is for subsequent use, the ammonia of cation exchange resin 10L 0.5mol/L carries out eluting, collect eluent, merge the effluent after cationic exchange resin adsorption and ammonia eluent, reclaim ethanol, join in CO2 supercritical extraction device, ethanol is as entrainer, the percent by volume that entrainer accounts for total extractant is 6%, extracting pressure 30MPa, temperature 30 DEG C, CO2 flow 3m1/g crude drug min, extraction time 150min, obtain supercritical extract, add starch, 70% ethanol granule, dry, tabletting, make 1000, the heavy 0.3g of every sheet.
Embodiment 3: get Radix Rehmanniae Preparata 208g, Caulis Spatholobi 140g, Herba Epimedii 140g, Rhizoma Drynariae 140g, Rhizoma Cibotii 140g, Semen Raphani 70g, Fructus Lycii 70g, Herba Cynomorii 140g, interrupted 70g, pulverize, adds the water boiling and extraction three times of 20,16,14 times amount respectively, each 1 hour, merge three extracting solution, filter, remove slag and get liquid, relative density 1.10 when being concentrated into 65 DEG C, adding ethanol makes the percent by volume of alcohol content reach 50%, precipitation, leaves standstill, filter, obtain alcohol deposit fluid, alcohol deposit fluid is adsorbed through 10kg cation exchange resin 001 × 7, loading flow velocity is 2ml/min, effluent is for subsequent use, the ammonia of cation exchange resin 10L 0.5mol/L carries out eluting, collect eluent, merge the effluent after cationic exchange resin adsorption and ammonia eluent, reclaim ethanol, join in CO2 supercritical extraction device, ethanol is as entrainer, the percent by volume that entrainer accounts for total extractant is 5%, extracting pressure 20MPa, temperature 40 DEG C, CO2 flow 2m1/g crude drug min, extraction time 160min, obtain supercritical extract, add starch, 70% ethanol granule, dry, tabletting, make 1000, the heavy 0.3g of every sheet.
Embodiment 4: bone increases the experimentation data that the sheet that disappears suppresses human tissue cell lymphoma U937 cell proliferation
1 experiment material
1.1 experiment cell strains: human tissue cell lymphoma U937 cell, Jilin Province just with Pharmaceutical Group Plc laboratory cell storehouse, DMEM+10%FBS cellar culture.
1.2 Experimental agents: drugs: bone of the present invention increases the sheet that disappears: prepare by embodiment 3 method.Medicinal liquid liquid storage: take 100mg bone and increase and to disappear sheet content, be dissolved in 5ml dehydrated alcohol, 0.2 μm of frit, 500 μ l doff pipe subpackages ,-20 DEG C of storage, 0.2 μm of frit dehydrated alcohol prepares against the use of matched group simultaneously.
1.3 experiment reagents: 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.11810-033 Lot.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); PBS (laboratory autogamy);
1.4 experiment equipments: Lycra inverted microscope (German Leica model: DM1L); Visible-ultraviolet light microwell plate detector (MD company of U.S. model: SPECTRA MAX 190); 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/N 020579); 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) human tissue cell lymphoma U937 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, discard culture fluid, PBS fine laundering 3 times, add 3ml 0.25% 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 and increase the sheet solution that disappears to the marrow, 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, 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 experimental results: statistical result showed after mtt assay experiment, compare with matched group, when dosage reaches 5mg/ml, to human tissue cell's lymphoma U937 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.
The increasing of table 1 bone disappears sheet to human tissue cell lymphoma U937 cell inhibitory effect influence research (X ± SD)
Note: compare with matched group, * P<0.01; * P<0.001
4 experiment conclusion: bone increases the sheet that disappears can suppress human tissue cell lymphoma U937 cell proliferation, and reduce the Growth of Cells number of human tissue cell lymphoma U937 cell, this effect is dose dependent.
Claims (6)
1. a bone increases the preparation method of the sheet that disappears, by Radix Rehmanniae Preparata 208g, Caulis Spatholobi 140g, Herba Epimedii 140g, Rhizoma Drynariae 140g, Rhizoma Cibotii 140g, Semen Raphani 70g, Fructus Lycii 70g, Herba Cynomorii 140g, interrupted 70g makes as crude drug, it is characterized in that described method is made up of the following step: get Radix Rehmanniae Preparata 208g, Caulis Spatholobi 140g, Herba Epimedii 140g, Rhizoma Drynariae 140g, Rhizoma Cibotii 140g, Semen Raphani 70g, Fructus Lycii 70g, Herba Cynomorii 140g, interrupted 70g, pulverize, add 20 respectively, 16, the water boiling and extraction of 14 times amount three times, each 1 hour, merge three extracting solution, filter, remove slag and get liquid, relative density 1.10 when being concentrated into 65 DEG C, adding ethanol makes the percent by volume of alcohol content reach 50%, precipitation, leave standstill, filter, obtain alcohol deposit fluid, alcohol deposit fluid is adsorbed through 10kg cation exchange resin 001 × 7, loading flow velocity is 2ml/min, effluent is for subsequent use, the ammonia of cation exchange resin 10L0.5mol/L carries out eluting, collect eluent, merge the effluent after cationic exchange resin adsorption and ammonia eluent, reclaim ethanol, 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, adds starch, 70% ethanol granule, and dry, tabletting, makes 1000, the heavy 0.3g of every sheet.
2. bone increases the preparation method of the sheet that disappears according to claim 1, it is characterized in that described CO
2the percent by volume that supercritical extraction entrainer accounts for total extractant is 5%.
3. bone increases the preparation method of the sheet that disappears according to claim 1, it is characterized in that described CO
2the extracting pressure 20MPa of supercritical extraction, temperature 40 DEG C, CO
2flow 2ml/g crude drug min, extraction time 160min.
4. bone increases the application of sheet in preparation suppression human tissue cell lymphoma U937 cell proliferation that disappear according to claim 1, it is characterized in that described method is made up of the following step: get Radix Rehmanniae Preparata 208g, Caulis Spatholobi 140g, Herba Epimedii 140g, Rhizoma Drynariae 140g, Rhizoma Cibotii 140g, Semen Raphani 70g, Fructus Lycii 70g, Herba Cynomorii 140g, interrupted 70g, pulverize, add 20 respectively, 16, the water boiling and extraction of 14 times amount three times, each 1 hour, merge three extracting solution, filter, remove slag and get liquid, relative density 1.10 when being concentrated into 65 DEG C, adding ethanol makes the percent by volume of alcohol content reach 50%, precipitation, leave standstill, filter, obtain alcohol deposit fluid, alcohol deposit fluid is adsorbed through 10kg cation exchange resin 001 × 7, loading flow velocity is 2ml/min, effluent is for subsequent use, the ammonia of cation exchange resin 10L0.5mol/L carries out eluting, collect eluent, merge the effluent after cationic exchange resin adsorption and ammonia eluent, reclaim ethanol, 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, adds starch, 70% ethanol granule, and dry, tabletting, makes 1000, the heavy 0.3g of every sheet.
5. bone increases the sheet that disappears suppresses in human tissue cell lymphoma U937 cell proliferation application in preparation according to claim 4, it is characterized in that CO described in the preparation method that bone increases the sheet that disappears
2the percent by volume that supercritical extraction entrainer accounts for total extractant is 5%.
6. bone increases the sheet that disappears suppresses in human tissue cell lymphoma U937 cell proliferation application in preparation according to claim 4, it is characterized in that CO described in the preparation method that bone increases the sheet that disappears
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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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN105687257A (en) * | 2016-03-04 | 2016-06-22 | 南京多宝生物科技有限公司 | Poria cocos formula granule preparation method and application |
| CN107468821A (en) * | 2017-10-11 | 2017-12-15 | 南京正宽医药科技有限公司 | A kind of bone increases the preparation method and application for the piece that disappears |
| CN112516218A (en) * | 2019-09-18 | 2021-03-19 | 中国药科大学 | Melanoidin in prepared rehmannia root and application thereof |
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| CN101104035A (en) * | 2007-02-13 | 2008-01-16 | 通化正和药业有限公司 | Medicine composition for treating hyperosteogeny disease and preparation method |
| CN102228513A (en) * | 2011-06-29 | 2011-11-02 | 江苏大学 | Medicinal composition for treating diabetes or diabetic complications and preparation method thereof |
| CN103494902A (en) * | 2013-10-08 | 2014-01-08 | 南京正亮医药科技有限公司 | Preparation method and application of pseudo-ginseng injury tablets |
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- 2015-07-19 CN CN201510424205.7A patent/CN104940426A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101104035A (en) * | 2007-02-13 | 2008-01-16 | 通化正和药业有限公司 | Medicine composition for treating hyperosteogeny disease and preparation method |
| CN102228513A (en) * | 2011-06-29 | 2011-11-02 | 江苏大学 | Medicinal composition for treating diabetes or diabetic complications and preparation method thereof |
| CN103494902A (en) * | 2013-10-08 | 2014-01-08 | 南京正亮医药科技有限公司 | Preparation method and application of pseudo-ginseng injury tablets |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105687257A (en) * | 2016-03-04 | 2016-06-22 | 南京多宝生物科技有限公司 | Poria cocos formula granule preparation method and application |
| CN107468821A (en) * | 2017-10-11 | 2017-12-15 | 南京正宽医药科技有限公司 | A kind of bone increases the preparation method and application for the piece that disappears |
| CN112516218A (en) * | 2019-09-18 | 2021-03-19 | 中国药科大学 | Melanoidin in prepared rehmannia root and application thereof |
| CN112516218B (en) * | 2019-09-18 | 2022-02-01 | 中国药科大学 | Melanoidin in prepared rehmannia root and application thereof |
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