CN103655839B - Preparation method of Yikangbuyuan tablet and application of Yikangbuyuan tablet in medicines for inhibiting mouse lung cancer cell LLC proliferation - Google Patents
Preparation method of Yikangbuyuan tablet and application of Yikangbuyuan tablet in medicines for inhibiting mouse lung cancer cell LLC proliferation Download PDFInfo
- Publication number
- CN103655839B CN103655839B CN201310669846.XA CN201310669846A CN103655839B CN 103655839 B CN103655839 B CN 103655839B CN 201310669846 A CN201310669846 A CN 201310669846A CN 103655839 B CN103655839 B CN 103655839B
- Authority
- CN
- China
- Prior art keywords
- preparation
- extraction
- lung cancer
- application
- cancer cell
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Fee Related
Links
- 238000002360 preparation method Methods 0.000 title claims abstract description 32
- 239000003814 drug Substances 0.000 title claims abstract description 18
- 206010058467 Lung neoplasm malignant Diseases 0.000 title claims abstract description 15
- 229940079593 drug Drugs 0.000 title claims abstract description 15
- 201000005202 lung cancer Diseases 0.000 title claims abstract description 15
- 208000020816 lung neoplasm Diseases 0.000 title claims abstract description 15
- 230000002401 inhibitory effect Effects 0.000 title abstract description 4
- 230000035755 proliferation Effects 0.000 title abstract 2
- 238000000194 supercritical-fluid extraction Methods 0.000 claims abstract description 15
- 238000000874 microwave-assisted extraction Methods 0.000 claims abstract description 11
- 239000009636 Huang Qi Substances 0.000 claims abstract description 7
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 53
- 230000000295 complement effect Effects 0.000 claims description 42
- 238000000605 extraction Methods 0.000 claims description 20
- 239000000284 extract Substances 0.000 claims description 15
- 230000004663 cell proliferation Effects 0.000 claims description 14
- 239000008187 granular material Substances 0.000 claims description 14
- 241001529936 Murinae Species 0.000 claims description 13
- 210000000582 semen Anatomy 0.000 claims description 11
- 239000007788 liquid Substances 0.000 claims description 9
- 241000628997 Flos Species 0.000 claims description 6
- 229940126678 chinese medicines Drugs 0.000 claims description 6
- 239000000835 fiber Substances 0.000 claims description 6
- 229920002472 Starch Polymers 0.000 claims description 5
- 230000000274 adsorptive effect Effects 0.000 claims description 5
- 230000006837 decompression Effects 0.000 claims description 5
- 239000003480 eluent Substances 0.000 claims description 5
- 238000010828 elution Methods 0.000 claims description 5
- 238000002156 mixing Methods 0.000 claims description 5
- 238000004064 recycling Methods 0.000 claims description 5
- 229920005989 resin Polymers 0.000 claims description 5
- 239000011347 resin Substances 0.000 claims description 5
- 235000019698 starch Nutrition 0.000 claims description 5
- 239000008107 starch Substances 0.000 claims description 5
- 230000001629 suppression Effects 0.000 claims description 5
- 229940126680 traditional chinese medicines Drugs 0.000 abstract description 2
- 244000144730 Amygdalus persica Species 0.000 abstract 1
- 244000020518 Carthamus tinctorius Species 0.000 abstract 1
- 235000003255 Carthamus tinctorius Nutrition 0.000 abstract 1
- 241000510667 Conioselinum Species 0.000 abstract 1
- 244000286838 Eclipta prostrata Species 0.000 abstract 1
- 240000002045 Guettarda speciosa Species 0.000 abstract 1
- 235000001287 Guettarda speciosa Nutrition 0.000 abstract 1
- 241000007393 Magnolia hypolampra Species 0.000 abstract 1
- 241000736199 Paeonia Species 0.000 abstract 1
- 235000006753 Platycodon grandiflorum Nutrition 0.000 abstract 1
- 240000003582 Platycodon grandiflorus Species 0.000 abstract 1
- 235000006040 Prunus persica var persica Nutrition 0.000 abstract 1
- 229960004756 ethanol Drugs 0.000 description 20
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 15
- YKRGDOXKVOZESV-WRJNSLSBSA-N Paeoniflorin Chemical compound C([C@]12[C@H]3O[C@]4(O)C[C@](O3)([C@]1(C[C@@H]42)O[C@H]1[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O1)O)C)OC(=O)C1=CC=CC=C1 YKRGDOXKVOZESV-WRJNSLSBSA-N 0.000 description 11
- 238000000034 method Methods 0.000 description 11
- 238000002474 experimental method Methods 0.000 description 8
- 239000000243 solution Substances 0.000 description 8
- 238000012360 testing method Methods 0.000 description 8
- 239000000047 product Substances 0.000 description 7
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 4
- 239000012467 final product Substances 0.000 description 4
- 239000013558 reference substance Substances 0.000 description 4
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 3
- 230000010261 cell growth Effects 0.000 description 3
- 239000003153 chemical reaction reagent Substances 0.000 description 3
- 239000012531 culture fluid Substances 0.000 description 3
- 229960000935 dehydrated alcohol Drugs 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- YKRGDOXKVOZESV-UHFFFAOYSA-N paeoniflorin Natural products O1C(C)(C2(CC34)OC5C(C(O)C(O)C(CO)O5)O)CC3(O)OC1C24COC(=O)C1=CC=CC=C1 YKRGDOXKVOZESV-UHFFFAOYSA-N 0.000 description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 2
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 2
- 239000006285 cell suspension Substances 0.000 description 2
- 230000007812 deficiency Effects 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 239000012528 membrane Substances 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 210000000952 spleen Anatomy 0.000 description 2
- 238000003860 storage Methods 0.000 description 2
- 235000019640 taste Nutrition 0.000 description 2
- DLYUQMMRRRQYAE-UHFFFAOYSA-N tetraphosphorus decaoxide Chemical compound O1P(O2)(=O)OP3(=O)OP1(=O)OP2(=O)O3 DLYUQMMRRRQYAE-UHFFFAOYSA-N 0.000 description 2
- WEEMDRWIKYCTQM-UHFFFAOYSA-N 2,6-dimethoxybenzenecarbothioamide Chemical compound COC1=CC=CC(OC)=C1C(N)=S WEEMDRWIKYCTQM-UHFFFAOYSA-N 0.000 description 1
- 238000002965 ELISA Methods 0.000 description 1
- 241000167880 Hirundinidae Species 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- 208000013738 Sleep Initiation and Maintenance disease Diseases 0.000 description 1
- 229920002334 Spandex Polymers 0.000 description 1
- 102000004142 Trypsin Human genes 0.000 description 1
- 108090000631 Trypsin Proteins 0.000 description 1
- 239000002250 absorbent Substances 0.000 description 1
- 230000002745 absorbent Effects 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 230000036528 appetite Effects 0.000 description 1
- 235000019789 appetite Nutrition 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 230000010165 autogamy Effects 0.000 description 1
- FCPVYOBCFFNJFS-LQDWTQKMSA-M benzylpenicillin sodium Chemical compound [Na+].N([C@H]1[C@H]2SC([C@@H](N2C1=O)C([O-])=O)(C)C)C(=O)CC1=CC=CC=C1 FCPVYOBCFFNJFS-LQDWTQKMSA-M 0.000 description 1
- 235000019658 bitter taste Nutrition 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 230000017531 blood circulation Effects 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 239000000470 constituent Substances 0.000 description 1
- 238000010219 correlation analysis Methods 0.000 description 1
- 239000013078 crystal Substances 0.000 description 1
- 230000029087 digestion Effects 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 239000012738 dissolution medium Substances 0.000 description 1
- 231100000673 dose–response relationship Toxicity 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 239000000945 filler Substances 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 206010022437 insomnia Diseases 0.000 description 1
- 210000003734 kidney Anatomy 0.000 description 1
- 210000003127 knee Anatomy 0.000 description 1
- 238000004900 laundering Methods 0.000 description 1
- 230000031700 light absorption Effects 0.000 description 1
- 238000004811 liquid chromatography Methods 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000002609 medium Substances 0.000 description 1
- 238000000465 moulding Methods 0.000 description 1
- 238000006386 neutralization reaction Methods 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- YTJSFYQNRXLOIC-UHFFFAOYSA-N octadecylsilane Chemical compound CCCCCCCCCCCCCCCCCC[SiH3] YTJSFYQNRXLOIC-UHFFFAOYSA-N 0.000 description 1
- 230000010355 oscillation Effects 0.000 description 1
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 1
- 239000010452 phosphate Substances 0.000 description 1
- 239000000377 silicon dioxide Substances 0.000 description 1
- 239000004759 spandex Substances 0.000 description 1
- 238000010561 standard procedure Methods 0.000 description 1
- 210000002784 stomach Anatomy 0.000 description 1
- 238000005728 strengthening Methods 0.000 description 1
- 229960002385 streptomycin sulfate Drugs 0.000 description 1
- 238000009923 sugaring Methods 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 239000012588 trypsin Substances 0.000 description 1
- 239000000341 volatile oil Substances 0.000 description 1
Landscapes
- Medicines Containing Plant Substances (AREA)
- Medicinal Preparation (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention belongs to the technical field of traditional Chinese medicines, and particularly provides a preparation method and application of a Yikangbuyuan tablet. The Yikangbuyuan tablet is prepared from 280g of radix astragali, 80g of Michelia hedyosperma Lew, 190g of angelica, 220g of yerbadetajo, 190g of rhizoma atractylodis, 150g of safflower, 140g of red paeonia, 140g of peach kernel, 150g of radix achyranthis bidentatae, 120g of hemlock parsley, 70g of fructus aurantii and 100g of balloonflower by supercritical extraction and microwave extraction, so that the penoniflorin content is greatly enhanced. The invention also provides application of the Yikangbuyuan tablet in preparing medicines for inhibiting mouse lung cancer cell LLC proliferation.
Description
Technical field
The invention belongs to technical field of Chinese medicines, be specifically related to a kind of preparation method of Yikang complement sheet and suppressing the application in murine lung cancer cell LLC cell proliferation.
Background technology
Yikang complement particulate level WS-10983(ZD-0983)-2002, be recorded in national standard for traditional Chinese medicines compilation internal medicine taste fascicle.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, there is benefiting QI for activating blood circulation, effect of strengthening spleen, tonifying kidney.For the spiritlessness and weakness that blood stasis due to qi deficiency, deficiency of spleen and stomach cause, short breath, insomnia, soreness of the waist and knees, lack of appetite is forgetful waits disease.
In prior art, Yikang complement granule is not yet had to extract the report adopting supercritical and microwave technology in preparation, and volatile oil is drawn, the method of soak by water, technique is coarse, backward, and impurity is many, cause patient's consumption excessive, be inconvenient to take, had a strong impact on this product and applied clinically.
In prior art, each 1 bag of Yikang complement granule, every bag of 7g, 3 times on the one.Yikang complement granule dosage is large, and granule preparation needs to add a large amount of sugar, and diabetics is not suitable for, and the words taste of not sugaring is not good, and under patient is not easy clothes, compliance is poor.The heavy 0.3g of the every sheet of the Yikang complement sheet adopting the inventive method to be prepared into, only needs 2 at every turn, within 1st, takes 3 times.Dose is greatly reduced under the condition with more active component.This conclusion can be proved by following test.And being prepared into tablet, patient swallows with water, and dosage is little and without bitterness, patient is easy to accept.
The comparison of paeoniflorin content in Yikang complement sheet prepared by test one, distinct methods
L, instrument and reagent Yikang complement of the present invention sheet: by the preparation of embodiment 1 method, use 1510g crude drug, makes 300 through extracting, the heavy 0.3g of every sheet.Former Yikang complement granule, according to WS-10983(ZD-0983)-2002 standard method preparations.Agilent1200 high performance liquid chromatograph; METTLER AE240 electronic analytical balance; Peoniflorin reference substance (Nat'l Pharmaceutical & Biological Products Control Institute).
2, method
Measure according to high performance liquid chromatography (Chinese Pharmacopoeia version in 2010 annex V D).
Chromatographic condition and system suitability octadecylsilane chemically bonded silica are filler; Acetonitrile-0.2% phosphate aqueous solution (15: 85) is mobile phase; Determined wavelength is 230nm.Number of theoretical plate calculates should be not less than 3000 by peoniflorin peak.
The preparation of reference substance solution learns from else's experience the phosphorus pentoxide drying under reduced pressure peoniflorin reference substance of 36 hours in right amount, accurately weighed, adds methanol and makes the solution of every 1ml containing 0.1mg, to obtain final product.
Yikang complement sheet of the present invention is got in the preparation of product need testing solution of the present invention, and porphyrize, gets 2.25g, accurately weighed, put in tool plug conical flask, precision adds the close plug of methanol 25ml., weighed weight, supersound process (power 250w. frequency 33kHz) 1 hour, lets cool, weighed weight again, supply the weight of less loss with methanol, shake up, filter, filter with microporous filter membrane (0.45 μm), to obtain final product.。
The Yikang complement granule of this contrast is got in the preparation of reference product need testing solution, and porphyrize, gets 2.5g, accurately weighed, put in tool plug conical flask, precision adds the close plug of methanol 25ml., weighed weight, supersound process (power 250w. frequency 33kHz) 1 hour, lets cool, weighed weight again, supply the weight of less loss with methanol, shake up, filter, filter with microporous filter membrane (0.45 μm), 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 Yikang complement sheet of the present invention, the content of peoniflorin is 5-8mg/ sheet; And the content of peoniflorin is 4.8mg/ bag in former Yikang complement granule, the paeoniflorin content each serving consumption 2 is 2-4 times of former granule content, and when dose reduces, paeoniflorin content improves a lot.
Above-mentioned research shows, adopt Yikang complement sheet prepared by preparation method of the present invention, active constituent content is far away higher than WS-10983(ZD-0983) the Yikang complement granule prepared of method recorded of-2002 standards.
Summary of the invention
In order to solve above-mentioned technical problem, the invention provides a kind of preparation method of Yikang complement sheet.
Another object of the present invention is to provide a kind of Yikang complement sheet to suppress the application in murine lung cancer cell LLC cell proliferation in preparation.
The object of the invention is by following scheme realize:
A kind of preparation method of Yikang complement sheet, 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, described preparation method is made up of the following step: get numb rare, Radix Angelicae Sinensis, Rhizoma Atractylodis, Semen Persicae, Rhizoma Chuanxiong, 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 300, the heavy 0.3g of every sheet.
In the preparation method of above-mentioned Yikang complement sheet, described CO
2in supercritical extraction, entrainer accounts for the percent by volume of total extractant is 5%.
In the preparation method of above-mentioned Yikang complement sheet, described CO
2in supercritical extraction, extracting pressure is 25MPa.
In the preparation method of above-mentioned Yikang complement sheet, described CO
2in supercritical extraction, extraction temperature is 60 DEG C.
In the preparation method of above-mentioned Yikang complement sheet, described CO
2cO in supercritical extraction
2flow 2ml/g crude drug min.
In the preparation method of above-mentioned Yikang complement sheet, described CO
2in supercritical extraction, extraction time is 200min.
In the preparation method of above-mentioned Yikang complement sheet, described microwave extracting power is 700W.
In the preparation method of above-mentioned Yikang complement sheet, each extraction time of described microwave extracting is 8 minutes.
Above-mentioned Yikang complement sheet suppresses the application in murine lung cancer cell LLC cell proliferation in preparation.
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
Seldom 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, Jiang Mahan, Radix Angelicae Sinensis, Rhizoma Atractylodis, Semen Persicae, Rhizoma Chuanxiong join CO to get Radix Astragali 280g, fiber crops
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 300, the heavy 0.3g of every sheet.
After testing, in finished product, the content of peoniflorin is 7.8mg/ sheet.
Embodiment 2
Seldom 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, Jiang Mahan, Radix Angelicae Sinensis, Rhizoma Atractylodis, Semen Persicae, Rhizoma Chuanxiong join CO to get Radix Astragali 280g, fiber crops
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 300, the heavy 0.3g of every sheet.
After testing, in finished product, the content of peoniflorin is 5.6mg/ sheet.
Embodiment 3
Seldom 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, Jiang Mahan, Radix Angelicae Sinensis, Rhizoma Atractylodis, Semen Persicae, Rhizoma Chuanxiong join CO to get Radix Astragali 280g, fiber crops
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 300, the heavy 0.3g of every sheet.
After testing, in finished product, the content of peoniflorin is 6.2mg/ sheet.
Embodiment 4: Yikang complement sheet suppresses the experimentation data of murine lung cancer cell LLC cell proliferation
1. experiment material
1.1 experiment cell strains
Murine lung cancer cell LLC cell, Shandong University's laboratory cell storehouse, DMEM+10%FBS cellar culture.
1.2 Experimental agents
Drugs: Yikang complement sheet of the present invention: prepare by embodiment 1 method.
Medicinal liquid liquid storage: take 100mg Yikang complement sheet, be dissolved in 5ml dehydrated alcohol, 0.2 μm of frit, 500 μ ldoff pipe subpackages ,-20 DEG C of storages, 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); Penicillin G Sodium Salt(AMRESCO company 1); Streptomycin Sulfate (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) LLC 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 Yikang complement sheet 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, buckle method removes supernatant, pats dry gently with absorbent paper, and every hole, as entered 200 μ l dimethyl sulfoxide, is put 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 Microsoft Excel2007 software and Student t to check, 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 LLC 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 Yikang complement sheet is to LLC cell inhibitory effect influence research (X ± SD)
| Group | Drug level (mg/ml) | Suppression ratio (%) |
| Matched group | 0 | 0 |
| 1 | 5 | 11.69±11.38 |
| 2 | 10 | 25.62±14.10* |
| 3 | 15 | 38.74±16.66** |
| 4 | 20 | 49.68±15.99** |
Note: compare with matched group, * P<O.01; * P<0.001.
5. experiment conclusion
Yikang complement sheet of the present invention can suppress LLC cell proliferation, and reduce the Growth of Cells number of LLC cell, this effect is dose dependent.
Claims (7)
1. the application of Yikang complement sheet in preparation suppression murine lung cancer cell LLC cell proliferation, it is characterized in that, described Yikang complement sheet is 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, described Yikang complement piece preparation method is made up of the following step: get numb rare, Radix Angelicae Sinensis, Rhizoma Atractylodis, Semen Persicae, Rhizoma Chuanxiong, 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 300, the heavy 0.3g of every sheet.
2. the Yikang complement sheet according to claim l suppresses the application in murine lung cancer cell LLC 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 Yikang complement sheet according to claim l suppresses the application in murine lung cancer cell LLC cell proliferation in preparation, it is characterized in that, described CO
2in supercritical extraction, extracting pressure is 25 MPa.
4. the Yikang complement sheet according to claim l suppresses the application in murine lung cancer cell LLC cell proliferation in preparation, it is characterized in that, described CO
2cO in supercritical extraction
2flow 2ml/g crude drug min.
5. the Yikang complement sheet according to claim l suppresses the application in murine lung cancer cell LLC cell proliferation in preparation, it is characterized in that, described CO
2in supercritical extraction, extraction time is 200min.
6. the Yikang complement sheet according to claim l suppresses the application in murine lung cancer cell LLC cell proliferation in preparation, and it is characterized in that, described microwave extracting power is 700W.
7. the Yikang complement sheet according to claim l suppresses the application in murine lung cancer cell LLC cell proliferation in preparation, and it is characterized in that, each extraction time of described microwave extracting is 8 minutes.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310669846.XA CN103655839B (en) | 2013-12-11 | 2013-12-11 | Preparation method of Yikangbuyuan tablet and application of Yikangbuyuan tablet in medicines for inhibiting mouse lung cancer cell LLC proliferation |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310669846.XA CN103655839B (en) | 2013-12-11 | 2013-12-11 | Preparation method of Yikangbuyuan tablet and application of Yikangbuyuan tablet in medicines for inhibiting mouse lung cancer cell LLC proliferation |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN103655839A CN103655839A (en) | 2014-03-26 |
| CN103655839B true CN103655839B (en) | 2015-07-15 |
Family
ID=50295398
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201310669846.XA Expired - Fee Related CN103655839B (en) | 2013-12-11 | 2013-12-11 | Preparation method of Yikangbuyuan tablet and application of Yikangbuyuan tablet in medicines for inhibiting mouse lung cancer cell LLC proliferation |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103655839B (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104288654A (en) * | 2014-10-16 | 2015-01-21 | 济南新起点医药科技有限公司 | Preparation method of fomes officinalis cough and asthma treating tablets and application of fomes officinalis cough and asthma treating tablets in preparation of medicine for inhibiting cell proliferation of mouse lung cancer cell LLC |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1961929A (en) * | 2005-11-09 | 2007-05-16 | 北京奇源益德药物研究所 | 'Yi Kang Bu Yuan' formulation for benefiting qi, moving blood, invigorating spleen and tonifying kidney, its preparation method and quality control method |
| CN102988526A (en) * | 2012-10-08 | 2013-03-27 | 卞毓平 | Method for preparing Bailing tablets and application |
-
2013
- 2013-12-11 CN CN201310669846.XA patent/CN103655839B/en not_active Expired - Fee Related
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1961929A (en) * | 2005-11-09 | 2007-05-16 | 北京奇源益德药物研究所 | 'Yi Kang Bu Yuan' formulation for benefiting qi, moving blood, invigorating spleen and tonifying kidney, its preparation method and quality control method |
| CN102988526A (en) * | 2012-10-08 | 2013-03-27 | 卞毓平 | Method for preparing Bailing tablets and application |
Non-Patent Citations (1)
| Title |
|---|
| 论益气活血法治疗肿瘤;王笑民;《中国中医药信息杂志》;19981231;第5卷(第10期);第11-12页 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN103655839A (en) | 2014-03-26 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN103690646B (en) | Preparation method of Yikangbuyuan tablet and application of Yikangbuyuan tablet in medicine for inhibiting mouse lymphoma cell YAC-1 cell proliferation | |
| CN103655843B (en) | A kind of Yikang complement sheet and its preparation method and application | |
| CN103638450B (en) | A kind of preparation method of Muxiangliqi Tablet and application | |
| CN103656493B (en) | A kind of preparation method of blood stasis dispelling stomach reinforcing sheet and application | |
| CN103690647B (en) | A kind of preparation method of Yikang complement sheet and the application in suppression mouse leukemia cell L6565 cell proliferation thereof | |
| CN103655839B (en) | Preparation method of Yikangbuyuan tablet and application of Yikangbuyuan tablet in medicines for inhibiting mouse lung cancer cell LLC proliferation | |
| CN103655841B (en) | Preparation method of Yikangbuyuan tablet and application of Yikangbuyuan tablet in medicines for inhibiting mouse lymphoid cancer cell P388D1 proliferation | |
| CN103655842B (en) | Preparation method of Yikangbuyuan tablet and application of Yikangbuyuan tablet in medicines for inhibiting mouse lymphomata cell EL4. IL-2 proliferation | |
| CN103655838B (en) | Preparation method of Yikangbuyuan tablet and application of Yikangbuyuan tablet in medicines for inhibiting mouse lymphomata cell L5178Y proliferation | |
| CN103655840B (en) | Preparation method of Yikangbuyuan tablet and application of Yikangbuyuan tablet in medicines for inhibiting mouse leukaemia cell L1210 proliferation | |
| CN103768267A (en) | Preparation method of Qingmei cold tablet and application of Qingmei cold tablet in drugs for inhibiting lung cancer cell H22 from cell proliferation | |
| CN103800498A (en) | Preparation method of Qingmei cold tablet and application of Qingmei cold tablet in drugs for inhibiting mouse prostate cancer cell RM-1 from cell proliferation | |
| CN103751424A (en) | Preparation method of cold tablet containing Artemisia carvifolia and holly root, and application of cold tablet in teratoma cell F9 cell proliferation inhibition medicines. | |
| CN103768211A (en) | Preparation method of Qingmei cold tablet and application of Qingmei cold tablet in drugs for inhibiting lymphomata cell EL4 from cell proliferation | |
| CN103768262A (en) | Preparation method of Qingmei cold tablet and application of Qingmei cold tablet in drugs for inhibiting myeloma cell FO from cell proliferation | |
| CN103768266A (en) | Preparation method of Qingmei cold tablet and application of Qingmei cold tablet in drugs for inhibiting mouse leukaemia cell L6565 from cell proliferation | |
| CN104288658A (en) | Preparation method of fomes officinalis cough and asthma treating tablets and application of fomes officinalis cough and asthma treating tablets in preparation of medicine for inhibiting cell proliferation of mouse myeloma cell SP2/0 | |
| CN103768561B (en) | Only dizzy mind tranquilizing tablet suppresses the application in NonHodgkin's lymhoma hyperproliferation agent in preparation | |
| CN103768207A (en) | Preparation method of Qingmei cold tablet and application of Qingmei cold tablet in drugs for inhibiting lung cancer cell Hepa1-6 from cell proliferation | |
| CN103768265A (en) | Preparation method of Qingmei cold tablet and application of Qingmei cold tablet in drugs for inhibiting lung cancer cell LLC from cell proliferation | |
| CN103800871A (en) | Application of dizziness-relieving and calming tablet in preparing medicine for inhibiting cell proliferation of embryonal carcinoma cell F9 | |
| CN103768264A (en) | Preparation method of Qingmei cold tablet and application of Qingmei cold tablet in drugs for inhibiting teratoma cell P19 from cell proliferation | |
| CN103800569A (en) | Preparation method of Qingmei cold tablet and application of Qingmei cold tablet in drugs for inhibiting lymphoma cell YAC-1 from cell proliferation | |
| CN103768212A (en) | Preparation method of Qingmei cold tablet and application of Qingmei cold tablet in drugs for inhibiting stomach cancer cell MFC from cell proliferation | |
| CN104352929A (en) | Preparation method of fomitopsis officinalis tablet capable of treating cough and asthma and application thereof in preparing drug for inhibiting cell proliferation of mouse liver cancer cell H22 |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| ASS | Succession or assignment of patent right |
Owner name: ZHOU WUYUAN Free format text: FORMER OWNER: SHANDONG ZHONGDA PHARM CO., LTD. Effective date: 20150605 |
|
| C41 | Transfer of patent application or patent right or utility model | ||
| C53 | Correction of patent for invention or patent application | ||
| CB03 | Change of inventor or designer information |
Inventor after: Zhou Wuyuan Inventor after: Cong Ning Inventor after: Zou Benkui Inventor after: Liu Lisheng Inventor before: Wei Shutong Inventor before: Duan Xuewen |
|
| COR | Change of bibliographic data |
Free format text: CORRECT: INVENTOR; FROM: WEI SHUTONG DUAN XUEWEN TO: ZHOU WUYUAN CONG NING ZOU BENKUI LIU LISHENG |
|
| TA01 | Transfer of patent application right |
Effective date of registration: 20150605 Address after: Huaiyin District of Ji'nan City, Shandong province 250117 Yan Ji Road, No. 440 Shandong Province Tumor Hospital Department of Hepatobiliary Surgery Applicant after: Zhou Wuyuan Address before: 250062, No. ten, No. 16369, Lixia District, Ji'nan City, Shandong Province Applicant before: SHANDONG ZHONGDA PHARMACEUTICAL CO., LTD. |
|
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20150715 Termination date: 20191211 |
|
| CF01 | Termination of patent right due to non-payment of annual fee |