CN101057872A - Anti tumor active part of American cockroach prepared with reverse phase material and medical application - Google Patents
Anti tumor active part of American cockroach prepared with reverse phase material and medical application Download PDFInfo
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Abstract
The invention relates to a Periplaneta americana extract of effective part, the preparation method and medical application. The extract is anti-tumor. The product is got from fresh or dry Periplaneta americana through processes of extraction with alcohol and water, reversed material column chromatography, alcohol washing and refining. The product can be used to prepare drug or health-care product of treating nasopharyngeal carcinoma and human oral epidermoid carcinoma.
Description
Technical field
The present invention relates to medical technical field, particularly, the present invention relates to a kind of American-cockroach-extract effective site of expecting to have cell toxicant and anti-tumor function and preparation method thereof and medical usage.
Technical background
Periplaneta americana (Periplaneta americana) is an Insecta Pterigota Blattaria Blattidae Periplaneta insecticide, is commonly called as " Blatta seu periplaneta ", and its beginning of being used as medicine is stated from Shennong's Herbal, wherein it is classified as middle product, meaning " flavor: salty, cold; Control: the hard cold and heat of Blood stasis, removing mass is poly-, laryngopharynx closes, the endogenous cold loss of fecundity " [Sun Xingyan etc., Shennong's Herbal, the Commercial Press, 1955:90]." Blatta seu periplaneta " sees supplementary Amplifications of the Compendium of Materia Medica for its popular name head, also is called stone Rhizoma Zingiberis Recens, sliding worm; There is its different title method the each place, as: steal oily mother-in-law, tea woman, kitchen range ant etc.Blattidae (Blattidae) is a huge insecticide family, has survived on earth 3.2 hundred million years, belongs to the biology that the same epoch occur with dinosaur.It is the strongest, the most ancient, one of the most successful insecticide monoid of procreation so far of vitality in the world undoubtedly.Nineteen sixty Princis reported once that cockroach worldwide had 3500 kinds approximately, totally 253 kinds [Feng Pingzhang, Guo Yuyuan, Wu Fuzhen, Chinese Blatta seu periplaneta kind and control, Beijing: China Science Tech Publishing House, in April, 1997] of the existing record of China.Mainly be distributed in the indoor or open-air of the torrid zone, subtropical zone.And periplaneta americana is one of a few household cockroach kind wherein.It mainly inhabits kitchen, dining room, warehouse etc. as indoor pest and locates to damage food, clothes, books etc.Get food and pollution people's food, propagate multiple diseases such as dysentery, typhoid fever, ascarid, pinworm.Detect 25 kinds in pathogenic bacterium in 226 periplaneta americana body surfaces collecting from kitchen and warehouse etc. and the intestinal, infection rate is up to 99.9%[Pai H H etc., Isolation of bacteria withantibiotic resistance from household cockroaches (Periplaneta americana andBlattella germanica), Acta Tropica, 2005,93 (3): 259].Periplaneta americana is one of urban health epidemic prevention department and customs quarantine control department keypoint control object as insect.Yet, in today that science and technology is maked rapid progress, in the face of the thousands of kinds of various agent for killing at it, this insecticide colony is still very active, this shows to have some special material and its distinctive mechanism of action in its body, has caused more domestic and international scholars' concern.
In recent years along with the attention of China to traditional Chinese medicine The Study on Resources exploitation, some scholar has carried out the research of application and development aspect to this insect that can't thoroughly do away with for thousands of years, and obtained gratifying achievements, excavated out the positive rank value of this traditional insect.The product made from periplaneta americana ethanol extract purification " rehabilitation is new " is in the good efficacy of traumatic surfaces such as burn, scald." the Xinmailong injection " that purification refine forms from the periplaneta americana ethanol extract, tool strengthening heart and boosting pressure, microcirculation improvement, the excited breathing, diuresis, the blood flow of the increase heart, brain, lung, kidney." Xinmailong injection " cures mainly: right heart failure, but also treating acute and chronic heart failure, and hemorrhagic shock, hypotension, ventricular premature contraction, sick hole syndrome, ischemic cardiovascular and cerebral vascular disease is country's two kind new medicines (Chinese medicine) of examining by SFDA in 2004.
Obtain the antibacterial peptide composition with extraction separation in the periplaneta americana and test its bacteriostasis by escherichia coli, the colibacillary quantity of result reduces [Lan Jianglin gradually in certain testing time, Zhou Xianzhi, Zhuo Kan, Deng, periplaneta americana (Periplaneta americana L.) antibacterial peptide bactericidal action preliminary observation, University Of Agriculture and Forestry In Fujian's journal (natural science edition), 2004,33 (2): 166.].Show that by electron microscopic observation the periplaneta americana antibacterial peptide at first makes skin and the cytoplasma membrane damage of antibacterial, forms opening, causes content to leak and dead, last thalline disintegrate fragmentates.Think that the periplaneta americana antibacterial peptide is not general bacteriostasis to escherichia coli, but direct killing action.Screening in the body that the oily alcohol extract of Blattidae insecticide Australian cockroach (Periplaneta australasie Fabricus) Scorpio (go wing and foot) is carried out, proof Australian cockroach extract all has significant antitumaous effect [Huang Houpin to S-180 and the xenotransplantation of human esophagus cancer mice, Cheng Caifen, Li Shufang, the antitumaous effect of Blatta seu periplaneta oil, Chinese herbal medicine, 1981,12 (1): 35].Cheng Caifen etc. get oily liquid to the heteroplastic antitumaous effect of human esophagus cancer mice with Australian cockroach with 95% soak with ethanol, the result shows: every day 1 time, each subcutaneous injection is in 2 gram crude drug/kilograms, and successive administration 5 days can make the tumor piece significantly dwindle [Cheng Caifen, Lin Wenqin, yellow thick engaging, Blatta seu periplaneta oil is to the heteroplastic antitumaous effect of human esophagus cancer mice, Chinese herbal medicine, 986,17 (6): 38].Chen Liming [Chen Liming, clinical and the experimentation of Blatta seu periplaneta extract AT2 antitumaous effect, combination of Chinese and Western medicine magazine, 1986, (11): 648] Blatta seu periplaneta extract AT2 antitumor action is studied, pharmacological evaluation shows: AT2 has anti-tumor activity, can strengthen the Turnover of Mouse Peritoneal Macrophages phagocytic function, and spleen weight is increased, at the external conversion reaction that increases the T lymphocyte to Con A.
AT2 treatment primary hepatocarcinoma 49 examples, clinical observation shows that this product has relief of symptoms, and alpha-fetoprotein is descended, and prolongs patient's effect of life cycle.The husky quiet pretty [Sha Jingshu such as grade of information institute of Shanghai Institute of Pharmaceutical Industry, Mao Hongkui, eliminating mass the liver benefiting sheet, pharmacy circular, 1987,22 (9): 569] experimental studies results to eliminating mass the liver benefiting sheet (I) shows: eliminating mass the liver benefiting sheet (I) has effects such as removing blood stasis with potent drugs, eliminate indigestion, detumescence and detoxifcation.Some mouse tumor there is inhibitory action,, the macrophage function of mice is had tangible potentiation animal immune function unrestraint effect.Eliminating mass the liver benefiting sheet (I) is by the Blatta seu periplaneta oral tablet that adult makes through extraction of living, in being applicable to, the treatment of late primary liver cancer., and compare with other western medicines in treatment group 46 examples in 49 routine patients through clinic trial, the result is: (I) the annual rate of depositing in group treatment back is 14.89%, and none example survival of matched group is more than 1 year.Clinical showing, patient's subjective symptom has improvement after the medication, and hepatalgia alleviates or disappearance, appetite increase, weight increase, abdominal distention alleviate.The untoward reaction such as scratch where it itches of slight xerostomia, constipation, whole body is arranged after part patient's medication.Aspect clinical practice, Blatta seu periplaneta worm extract for treating alive primary lung cancer has good result.Liao Jianying obtains good result [Liao Jianying, malignant fibrohistiocytoma 1 example, Dali College journal, 2003,2 (3): 92 are newly treated in rehabilitation] with " rehabilitation is new " treatment malignant fibrohistiocytoma.There is relevant report to show to have the effect of certain raise immunity simultaneously at antineoplastic by Blatta seu periplaneta extract that [Luo Zhihong accounts for Chinese chapter, Li Yanhong etc., the Chinese medicine rehabilitation newly effect in the head-neck malignant tumor treatment is observed, China's combination of Chinese and Western medicine hals,Nasen und Ohrenheilkunde is assorted, and 1998,6 (3): 132; Yan Fengxiang etc., the Chinese medicine rehabilitation new in body immunopharmacology feature, DaLi Medicine Academy journal, 1991,3 (1): 21].
Be clear and definite periplaneta americana medical material anti-tumor effective component, instruct and further understand fully the structure of matter basis of wherein playing main pharmacological, illustrate the multiple bioactive mechanism of action of periplaneta americana from the molecular pharmacology basis, the inventor has launched detailed pharmacy technical study to it.The present invention uses polyamide to be separating tool, and the ethanol extract of this raw medicinal herbs has been carried out segmentation; And adopt two strain human body tumour cell strains: the growth inhibition test of human oral cavity epithelial JEG-3 (KB) and KB cell (CNE) is followed the tracks of, search out the anti-tumor effective component of tool cytotoxic activity, and carried out checking in the antitumous effect animal body with suppressing mice S180 sarcoma growth test in this effective site body.The final discovery: some position activity that the periplaneta americana ethanol extract obtains after the reversed material segmentation obviously is better than other position, and some active site cell toxicant and anti-tumor activity be better than the corresponding activity of crude extract, thereby constitutes the present invention.
Summary of the invention
The effective part extract that the purpose of this invention is to provide a kind of periplaneta americana tool cell toxicant and anti-tumor activity;
Another object of the present invention has provided a kind of method for preparing the effective part extract of periplaneta americana tool cell toxicant and anti-tumor activity;
Another object of the present invention has provided the purposes that American-cockroach-extract effective site is used to prepare tumour medicines such as anti-treatment of esophagus cancer, gastric cancer, ovarian cancer, colon cancer, nasopharyngeal carcinoma, leukemia, hepatocarcinoma, pulmonary carcinoma and oral epithelium cancer;
Another object of the present invention has provided a kind of pharmaceutical composition that American-cockroach-extract effective site is used to prepare anti-curing oncoma that contains.
The early stage research of periplaneta americana focuses mostly in the solvent crude extract of polypide, has set forth a kind of optimization extracting mode [Yan Shaohong etc., national inventing patent CN 1255117C] of periplaneta americana crude extract as Yan Shaohong etc.; Liu Yaobei uses the acetic acid extraction Blatta seu periplaneta extract, and hepatoma carcinoma cell BEL7404 and BEL7721 are carried out the cancer cell multiplication inhibition test, finds that crude extract can have the effect [Liu Yaobei etc., national inventing patent CN1199991C] of obvious anticancer growth; The Japanese plum nanmu removes degrease from the ethanol extract of periplaneta americana, with removing pigment and allergic component after the carbon dioxide absorption, be used for the treatment of cardiovascular disease [Japanese plum nanmu etc., national inventing patent CN 1067243C]; Liao Ji etc. have reported the health food [Liao Ji etc., national inventing patent CN 1230183C] for preparing blood fat reducing, cholesterol regulating, blood sugar regulation, anti-curing oncoma and angiocardiopathy preventing, anti rheumatism action with No. 6 extracting solution from the cockroach polypide; Xu Yan and disclose a kind of similar crude extract preparation method [Xu Yanhe, national inventing patent CN 1548058A]; Yan Shaohong etc. disclose the comprehensive utilization of a kind of periplaneta americana crude extract extracting mode [Yan Shaohong etc., national inventing patent CN 1593467A]; But, do not report with the further investigation of effective site in the above patent about concrete periplaneta americana antineoplastic agent.Yet according to the more deep research of inventor's later stage, find that its anti-tumor active ingredient can obtain enrichment by process optimization, the periplaneta americana ethanol extract that the inventor adopts water or water-alcohol solvent extraction to obtain is aided with reversed material chromatography means and can obtains several effective sites, the activity that suppresses tumor growth in its anticancer propagation and the body is higher than crude extract, creative and the novelty of effective site preparation method provided by the present invention is described, finishes the present invention in view of the above.
Particularly, the preparation method of the effective part extract of Central America of the present invention big Lian tool cell toxicant and anti-tumor activity is as follows: reclaim solvent, reversed-phase bonded silica RP-8 and/or RP-18 class material column chromatography, washing, alcohol-water solution gradient elution method purification with solvent extraction through the periplaneta americana polypide of pulverizing or do not pulverize, concentration extraction; The interior antitumor activity effective site of increment of the anticancer that obtains and body is through being evaporated to no pure flavor, concentrating under reduced pressure or lyophilization, pulverize product.Specifically comprise the following steps:
Animal drug among the present invention can be the arbitrary position of periplaneta americana, promptly can be periplaneta americana (Periplaneta americana) head, body, wing, also can be their mixture, preferred Scorpio.Can be dry Scorpio, also can be the Scorpio or the live body Scorpio of undried.
The inventor finds, the periplaneta americana ethanol extract that the present invention obtains has important biological through each extract of polyamide, and external cytotoxic activity test to human oral cavity epithelial JEG-3 (KB) and KB cell (CNE) shows: this type of periplaneta americana ethanol extract is inhibited to growth of tumour cell through each extract of reversed-phase bonded silica RP-8 and/or the segmentation of RP-18 class material.In addition, the periplaneta americana ethanol extract that obtains of the present invention through the extract of polyamide to ICR kind mice body in the growth of S180 sarcoma have the obvious suppression effect.Illustrate all that below the periplaneta americana ethanol extract might develop into new control tumour medicine through reversed-phase bonded silica RP-8 and/or RP-18 class material each extract of column chromatography or its mixture.
The periplaneta americana ethanol extract can combine with spoke material or carrier pharmaceutically commonly used through reversed-phase bonded silica RP-8 and/or RP-18 class material each extract of column chromatography or its mixture, has growth of tumour cell and suppresses the active pharmaceutical composition that can be used for anti-curing oncoma thereby prepare.Above-mentioned various kinds of drug compositions can adopt drug forms such as injection, tablet, capsule, aerosol, suppository, membrane, drop pill, externally-applied liniment, ointment, can also adopt the known controlled release of modern pharmaceutical circle or slow release formulation or nanometer formulation.
Periplaneta americana ethanol extract of the present invention through reversed-phase bonded silica RP-8 and/or RP-18 class material each extract of column chromatography or its mixture can with antitumor drug that has now gone on the market such as platinum medicine cisplatin (DDP), camptothecine irinotecan (Irinatecan, CPT-11), the vinca alkaloids medicine loses carbon vincaleucoblastine (Vinorebine, the NVB nvelbine), deoxidation born of the same parents former times class medicine gemcitabine (Gemcitabine, Gemzar, strong selecting), etoposide (Etoposide), paclitaxel (Paclitaxel) etc. is united use, prepare and have tumor growth and suppress active cytotoxicity compositions, can be used for treating tumor disease.Such pharmaceutical composition can adopt drug forms such as injection, tablet, capsule, aerosol, suppository, membrane, drop pill, externally-applied liniment, ointment, can also adopt the known controlled release of modern pharmaceutical circle or slow release formulation or nanometer formulation.These pharmaceutical compositions can be used for the treatment of tumor disease, delay purposes such as tumor worsens, prolongation tumour patient life span.
Specific embodiments
In order to understand essence of the present invention better, below respectively with such periplaneta americana ethanol extract through reversed-phase bonded silica RP-8 and/or RP-18 class material each extract of column chromatography or its mixture to the inhibitory action of two kinds of tumor cell lines growths and the The pharmacological results of S180 sarcoma growth inhibited effect test in to ICR kind mice body, its new purposes in the antitumor drug research field is described.The present invention has also carried out the oral administration gavage acute toxicity test of ICR mice with this activity extract, the toxicity of this effective site to animal is described, the safety when being reflected in the human body use thus.Pharmacology embodiment has provided the part activity data of representative compounds.
Mandatory declaration be: pharmacology embodiment of the present invention is used to illustrate the present invention rather than limitation of the present invention.Essence according to the present invention all belongs to the scope of protection of present invention to the simple modifications that the present invention carries out.
Embodiment 1: periplaneta americana ethanol extract and through the preparation of each extract of reversed-phase bonded silica RP-18 column chromatography gained
1.1 3 kilograms of the preparation periplaneta americana medical materials of periplaneta americana ethanol extract add 10000 milliliters of 55% ethanol, soaking at room temperature is extracted, and extracts each 24 hours 3 times.Merge extractive liquid, is evaporated to driedly, and vacuum drying promptly gets the periplaneta americana ethanol extract.
1.2 preparation periplaneta americana ethanol extract 20 grams of the different extracts of reversed-phase bonded silica RP-18 column chromatography segmentation, mix sample with 30 gram RP-18 (10~30 order) absorption, upper prop, elder generation's water is eluted to colourless, water lotion is got 10 grams and is evaporated to dried, vacuum drying promptly gets extract (I) [periplaneta americana ethanol extract water lotion]; Continuous be eluted to colourlessly with 10% ethanol-water solution, get 10 milliliters of eluents and be evaporated to driedly, vacuum drying promptly gets extract (II) [periplaneta americana ethanol extract 10% pure washing liquid]; Continuous be eluted to colourlessly with 20% ethanol-water solution, get 10 milliliters of eluents and be evaporated to driedly, vacuum drying promptly gets extract (III) [periplaneta americana ethanol extract 20% pure washing liquid]; Continuous be eluted to colourlessly with 30% ethanol-water solution, get 10 milliliters of eluents and be evaporated to driedly, vacuum drying promptly gets extract (IV) [periplaneta americana ethanol extract 30% pure washing liquid]; Continuous be eluted to colourlessly with 40% ethanol-water solution, get 10 milliliters of eluents and be evaporated to driedly, vacuum drying promptly gets extract (V) [periplaneta americana ethanol extract 40% pure washing liquid]; Continuous be eluted to colourlessly with 50% ethanol-water solution, get 10 milliliters of eluents and be evaporated to driedly, vacuum drying promptly gets extract (VI) [periplaneta americana ethanol extract 50% pure washing liquid]; Continuous be eluted to colourlessly with 60% ethanol-water solution, get 10 milliliters of eluents and be evaporated to driedly, vacuum drying promptly gets extract (VII) [periplaneta americana ethanol extract 60% pure washing liquid]; Continuous be eluted to colourlessly with 70% ethanol-water solution, get 10 milliliters of eluents and be evaporated to driedly, vacuum drying promptly gets extract (VIII) [periplaneta americana ethanol extract 70% pure washing liquid]; Continuous be eluted to colourlessly with 80% ethanol-water solution, get 10 milliliters of eluents and be evaporated to driedly, vacuum drying promptly gets extract (IX) [periplaneta americana ethanol extract 80% pure washing liquid]; Continuous be eluted to colourlessly with 95% ethanol-water solution, get 10 milliliters of eluents and be evaporated to driedly, vacuum drying promptly gets extract (X) [periplaneta americana ethanol extract 95% pure washing liquid].
Embodiment 2: the periplaneta americana active site is to the cytotoxicity test of oral squamous carcinoma cell (KB)
1.1 specimen: the periplaneta americana ethanol extract for preparing according to embodiment 1 and each extract part (being extract (I)~extract (X)) that obtains through the RP-18 column chromatography.
1.2 cell toxicity test KB (oral epithelium cancer) cell contains 10% hyclone, the streptomycin of 100U/ ml penicillin and 100U/ milliliter with RPMI 1640 culture medium culturings in the culture medium.Cell is with every hole 5 * 10
3Concentration join in 96 orifice plates, contain 5%CO at 37 ℃
2Cultivated 24 hours in the incubator of humid air.The mensuration of cell survival rate is with improveing mtt assay.Cell is through after 24 hours hatch, the dimethyl sulfoxide solution of the different extracts (I) of the periplaneta americana that will newly join respectively~extract (X) joins in each hole with Concentraton gradient, make different extract (extract (I)~extract (the X)) ultimate densities of the big Lian in Central America, hole be respectively 200,66.7,22.2 and 7.4 mcg/ml.After 72 hours, the phosphate buffer that adds 10 microlitre MTT (5 mg/ml), continue 37 ℃ of cultivations after 4 hours again, removed unconverted MTT in centrifugal 5 minutes, add 200 microlitre dimethyl sulfoxines in every hole, with the MTT crystal Jia Za (formazan) of dissolving and reducing, formed formazan microplate reader colorimetric under the 570nm wavelength, cell survival rate is by the ratio calculation of sample with respect to reference substance.Wherein the different extracts of periplaneta americana are to KB cell 503nhibiting concentration (IC
50) obtain by dose effect curve.
1.3 result:
The different extracts of table 1 reverse phase silica gel RP-18 column chromatography are to the cytotoxicity test of oral squamous carcinoma cell (KB)
Sample | IC 50(mcg/ml) |
American cockroach alcohol extract extract (I) [American cockroach alcohol extract water lotion extract (II) [American cockroach alcohol extract 10% pure washing lotion] extract (III) [American cockroach alcohol extract 20% pure washing lotion] extract (IV) [American cockroach alcohol extract 30% pure washing lotion] extract (V) [American cockroach alcohol extract 40% pure washing lotion] extract (VI) [American cockroach alcohol extract 50% pure washing lotion] extract (VII) [American cockroach alcohol extract 60% pure washing lotion] extract (VIII) [American cockroach alcohol extract 70% pure washing lotion] extract (IX) [American cockroach alcohol extract 80% pure washing lotion] extract (X) [American cockroach alcohol extract 95% pure washing lotion] positive control cis-platinum | 105.4 >200 >200 >200 >200 16.35 7.3 2.1 9.3 84.7 41.4 1.61 |
The positive control amycin | 0.17 |
The result shows, periplaneta americana ethanol extract extract (V) [periplaneta americana ethanol extract 40% pure washing liquid], extract (VI) [periplaneta americana ethanol extract 50% pure washing liquid], extract (VII) [periplaneta americana ethanol extract 60% pure washing liquid], extract (VIII) [periplaneta americana ethanol extract 70% pure washing liquid], extract (IX) [periplaneta americana ethanol extract 80% pure washing liquid] and extract (X) [periplaneta americana ethanol extract 95% pure washing liquid] have strong cytotoxic activity to oral squamous carcinoma cell (KB), and it suppresses oral squamous carcinoma cell increment activity all above the periplaneta americana ethanol extract.
1.5 experiment conclusion: the KB cell is Cytotoxic effective tool and the evaluation index of test compounds to tumor cell.This experiment shows that periplaneta americana ethanol extract some extract behind reverse phase silica gel RP-18 column sectional has stronger cytotoxicity to the KB cell, might develop into the new medicine with antitumor action.
Embodiment 3: the periplaneta americana ethanol extract is through through the cytotoxic activity of each extract of reverse phase silica gel RP-18 post to nasopharyngeal carcinoma CBE cell
1.1CNE (nasopharyngeal carcinoma) cell contains 10% hyclone, the streptomycin of 100U/ ml penicillin and 100U/ milliliter with RPMI 1640 culture medium culturings in the culture medium.With every hole 5 * 10
3The dense clothing of cell joins in 96 orifice plates, contains 5%CO at 37 ℃
2Cultivated 24 hours in the incubator of humid air.
1.2 the mensuration of cell survival rate is with improveing mtt assay, concrete grammar such as embodiment 2.
Table 2 is through the cytotoxicity test of each extract of reverse phase silica gel RP-18 post to nasopharyngeal carcinoma cell (CNE)
Sample | IC 50(mcg/ml) |
American cockroach alcohol extract extract (I) [American cockroach alcohol extract water lotion extract (II) [American cockroach alcohol extract 10% pure washing lotion] extract (III) [American cockroach alcohol extract 20% pure washing lotion] extract (IV) [American cockroach alcohol extract 30% pure washing lotion] extract (V) [American cockroach alcohol extract 40% pure washing lotion] extract (VI) [American cockroach alcohol extract 50% pure washing lotion] extract (VII) [American cockroach alcohol extract 60% pure washing lotion] extract (VIII) [American cockroach alcohol extract 70% pure washing lotion] extract (IX) [American cockroach alcohol extract 80% pure washing lotion] extract (X) [American cockroach alcohol extract 95% pure washing lotion] positive control cis-platinum positive control adriamycin | 151.8 >200 >200 >200 >200 23.46 7.04 2.4 16.59 74.61 111.12 3.45 0.18 |
1.3 the result shows, periplaneta americana ethanol extract extract (V) [periplaneta americana ethanol extract 40% pure washing liquid], extract (VI) [periplaneta americana ethanol extract 50% pure washing liquid], extract (VII) [periplaneta americana ethanol extract 60% pure washing liquid], extract (VIII) [periplaneta americana ethanol extract 70% pure washing liquid], extract (IX) [periplaneta americana ethanol extract 80% pure washing liquid] and extract (X) [periplaneta americana ethanol extract 95% pure washing liquid] have strong cytotoxic activity to nasopharyngeal carcinoma cell (CNE), and it suppresses nasopharyngeal carcinoma cell increment activity all above the periplaneta americana ethanol extract.
1.4 experiment conclusion: nasopharyngeal carcinoma CNE cell is Cytotoxic effective tool and the evaluation index of test compounds to tumor cell.This experiment shows that the periplaneta americana ethanol extract has stronger cytotoxicity through some extract of reverse phase silica gel RP-18 post to nasopharyngeal carcinoma CNE cell, might develop into the new medicine with the effect of treatment nasopharyngeal carcinoma.
Embodiment 4: periplaneta americana ethanol extract extract (VII) [periplaneta americana ethanol extract 60% pure washing liquid] is to sarcoma S180
Experiment
1.1 method and step:
1.1.1 animal grouping, number and weigh: get 20 of the white mice of body weight 18-22 gram, 10 of picked at random are done the administration group, and all the other 10 is matched group.
1.1.2 press 1500 milligrams/kilogram of experimental design concentration, with sterile saline preparation periplaneta americana ethanol extract extract (VII) [periplaneta americana ethanol extract 60% pure washing liquid].
1.1.3 respectively organize mice by corresponding dosage gastric infusion 14 days, 1 time/day.Put to death animal on the 15th day, dissect, take out the tumor body and weigh.Calculate tumor control rate according to following formula.
Suppression ratio=(the average tumor of the average tumor weight-test group of matched group is heavy)/average tumor of matched group heavy * 100%
1.2 result: this test serves as effective with tumor control rate greater than 30%.The suppression ratio of periplaneta americana ethanol extract extract (VII) test group is 51%.This experimental result shows that periplaneta americana ethanol extract extract (VII) [periplaneta americana ethanol extract 60% pure washing liquid] has significant curative effect to S180 solid type tumor.
1.3 experiment conclusion: the S180 solid tumor models is the effective tool and the evaluation index of the tested thing antitumor of test, inhibition tumor growth.This experiment shows that periplaneta americana ethanol extract extract (VII) [periplaneta americana ethanol extract 60% pure washing liquid] has stronger antitumor activity to the S180 solid tumor, very likely develops into the new medicine with treatment function of tumor.
Embodiment 5: periplaneta americana ethanol extract extract (VII) [periplaneta americana ethanol extract 60% pure washing liquid] acute toxicity (LD
50) test
1.1 experiment material:
Treat test agent: extract (VII) [periplaneta americana ethanol extract 60% pure washing liquid];
Solvent: 0.5%CMC-Na;
Animal: the ICR mice, the male and female dual-purpose, 19.0~20.9g is available from Shanghai Slac Experimental Animal Co., Ltd..
1.2 experimental technique:
1.2.1 preparation sample suspension: take by weighing periplaneta americana ethanol extract extract (VII) sample and make the suspension that concentration is 50 mg/ml.
1.2.2 administration:, irritate stomach and give periplaneta americana ethanol extract extract (VII) suspension with 12 mices (male and female half and half); The administration volume is 40 milliliters/kilogram, and dosage is 2000 milligrams/kilogram.
1.2.3 observed and recorded: the response situation of mice after observation and the record administration.
1.3 experimental result: observe after the administration 10 days animal feed and active situation continuously, do not see that mice is any abnormal response, do not have the phenomena of mortality to take place.
1.4 experiment conclusion: in general, in the acute primality test of medicine for treating tumor thing, half lethal dose>2.0 gram/kilogram persons half to be considered as toxicity less, perhaps nontoxic substantially.Periplaneta americana ethanol extract extract (VII) satisfies this requirement, so belong to low toxicity or nontoxic substantially effective site.Expect that clinical use has higher safety coefficient.
Embodiment 6: the preparation method of periplaneta americana ethanol extract extract (VII) capsule that is prepared by embodiment 1
5,000 milligrams of the periplaneta americana ethanol extract extracts (VII) for preparing with embodiment 1, with raw material: (starch+microcrystalline Cellulose) be (1: 1) in proportion, wherein starch: be inserted in after microcrystalline Cellulose (1: 1) is granulated in No. 0 Capsules.Adorn 100 altogether, each capsule weighs 100 milligrams.
Claims (8)
1. the active site of a periplaneta americana is characterized by this active site and utilizes reversed material to prepare.
2. according to the preparation method of the periplaneta americana active site of claim 1, it is characterized by: with the periplaneta americana polypide of solvent extraction through pulverizing or not pulverizing, concentration extraction reclaims solvent; Through the reversed material column chromatography, the alcoholic solvent eluting is used in washing then with concentrated solution; Eluent is evaporated to does not have the alcohol flavor, drying, pulverize product.
3. according to the preparation method of claim 2, it is characterized by: solvent is pure water mixed solvent, and wherein alcohol is methanol, ethanol, ethylene glycol or their mixture; Extract 1-5 time, doubly measure with 1-10 at every turn, extracted 0.5-24 hour at every turn; Reversed material comprises reversed-phase bonded silica RP-8 and/or RP-18 class material, and MCI class reversed material; Alcoholic solvent is meant alcohol solvent.
4. according to the preparation method of claim 2, it is characterized by the column chromatography used medium is reversed-phase bonded silica RP-8 and/or RP-18 class material; Alcoholic solvent is the ethanol of 10-98%.
5. according to the preparation method of claim 2, wherein the temperature during concentrating under reduced pressure is no more than 100 ℃, and drying is meant vacuum decompression drying and lyophilization.
6. the American-cockroach-extract effective site of one of claim 1-5 is used to prepare the purposes of preventing and treating tumour medicine, it is characterized in that nasopharyngeal carcinoma and oral epithelium cancer are had tangible cytotoxic activity and in-vivo tumour inhibition activity.
7. pharmaceutical composition with cytotoxicity and anti-tumor function according to claim 6, it contains American-cockroach-extract effective site and pharmaceutically suitable carrier of one of claim 1-5 for the treatment of effective dose.
8. according to the pharmaceutical composition of claim 6, it can be tablet, granule, capsule, oral liquid, injection, drop pill, transdermal patch, externally-applied liniment or aerosol.
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CN102319268A (en) * | 2011-08-12 | 2012-01-18 | 云南省腾冲制药厂 | A kind of injection that chronic heart failure levies and preparation method thereof of treating |
CN103222988A (en) * | 2013-01-15 | 2013-07-31 | 四川好医生攀西药业有限责任公司 | Periplaneta americana extract and its preparation method and use |
CN105878290A (en) * | 2015-07-06 | 2016-08-24 | 四川好医生攀西药业有限责任公司 | Periplaneta americana preparation and applications thereof in preparing medicines for treating colon cancer |
CN105878289A (en) * | 2015-07-06 | 2016-08-24 | 四川好医生攀西药业有限责任公司 | Periplaneta americana preparation and applications thereof in preparing medicines for treating gastric cancer |
CN107693546A (en) * | 2016-12-16 | 2018-02-16 | 四川好医生攀西药业有限责任公司 | A kind of medicine containing American cockroach and Sorafenib and its application |
CN107693545A (en) * | 2016-12-07 | 2018-02-16 | 四川好医生攀西药业有限责任公司 | A kind of combination medicine for treating non-small cell lung cancer |
CN107714731A (en) * | 2016-12-01 | 2018-02-23 | 四川好医生攀西药业有限责任公司 | A kind of medicine containing American cockroach and taxol and its application |
CN107714732A (en) * | 2016-12-16 | 2018-02-23 | 四川好医生攀西药业有限责任公司 | A kind of medicine containing American cockroach and capecitabine and its application |
CN109172611A (en) * | 2018-11-09 | 2019-01-11 | 大理大学 | Extraction and the purification process of pulmonary fibrosis active site are treated in a kind of American cockroach |
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CN102319268B (en) * | 2011-08-12 | 2013-01-23 | 云南省腾冲制药厂 | Preparation method of injection used for treating chronic congestive cardiac failure |
CN102319268A (en) * | 2011-08-12 | 2012-01-18 | 云南省腾冲制药厂 | A kind of injection that chronic heart failure levies and preparation method thereof of treating |
CN103222988A (en) * | 2013-01-15 | 2013-07-31 | 四川好医生攀西药业有限责任公司 | Periplaneta americana extract and its preparation method and use |
CN103222988B (en) * | 2013-01-15 | 2015-10-28 | 四川好医生攀西药业有限责任公司 | A kind of American-cockroach-extract and its preparation method and application |
CN105878290B (en) * | 2015-07-06 | 2019-12-06 | 四川好医生攀西药业有限责任公司 | periplaneta americana preparation and application thereof in preparation of medicine for treating colon cancer |
CN105878290A (en) * | 2015-07-06 | 2016-08-24 | 四川好医生攀西药业有限责任公司 | Periplaneta americana preparation and applications thereof in preparing medicines for treating colon cancer |
CN105878289A (en) * | 2015-07-06 | 2016-08-24 | 四川好医生攀西药业有限责任公司 | Periplaneta americana preparation and applications thereof in preparing medicines for treating gastric cancer |
CN105878289B (en) * | 2015-07-06 | 2019-12-17 | 四川好医生攀西药业有限责任公司 | Periplaneta americana preparation and application thereof in preparation of medicine for treating gastric cancer |
CN107714731A (en) * | 2016-12-01 | 2018-02-23 | 四川好医生攀西药业有限责任公司 | A kind of medicine containing American cockroach and taxol and its application |
CN107693545A (en) * | 2016-12-07 | 2018-02-16 | 四川好医生攀西药业有限责任公司 | A kind of combination medicine for treating non-small cell lung cancer |
CN107693545B (en) * | 2016-12-07 | 2020-08-25 | 四川好医生攀西药业有限责任公司 | Combined medicine for treating non-small cell lung cancer |
CN107693546A (en) * | 2016-12-16 | 2018-02-16 | 四川好医生攀西药业有限责任公司 | A kind of medicine containing American cockroach and Sorafenib and its application |
CN107714732A (en) * | 2016-12-16 | 2018-02-23 | 四川好医生攀西药业有限责任公司 | A kind of medicine containing American cockroach and capecitabine and its application |
CN109172611A (en) * | 2018-11-09 | 2019-01-11 | 大理大学 | Extraction and the purification process of pulmonary fibrosis active site are treated in a kind of American cockroach |
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