CN103417890A - Application of stahlianthus involucratus and extractive of stahlianthus involucratus in preparing drugs for treating and/or preventing cancer - Google Patents
Application of stahlianthus involucratus and extractive of stahlianthus involucratus in preparing drugs for treating and/or preventing cancer Download PDFInfo
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Abstract
The invention relates to the new application of a stahlianthus involucratus, an extractive of the stahlianthus involucratus, a fat-soluble extractive of the stahlianthus involucratus and an essential oil composition of the stahlianthus involucratus, namely the new application of the stahlianthus involucratus, the extractive of the stahlianthus involucratus, the fat-soluble extractive of the stahlianthus involucratus and the essential oil composition of the stahlianthus involucratus in preparing drugs for treating and/or preventing cancer, in particular to the new application of the stahlianthus involucratus, the extractive of the stahlianthus involucratus, the fat-soluble extractive of the stahlianthus involucratus and the essential oil composition of the stahlianthus involucratus in preparing drugs for treating and/or preventing lung cancer.
Description
Technical field
The present invention relates to the new purposes of Stahlianthus hainanensis (Hayata) T. L. Wu, Stahlianthus hainanensis (Hayata) T. L. Wu extract, Stahlianthus hainanensis (Hayata) T. L. Wu liposoluble extract and Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil, treat and/or prevent the new purposes aspect cancer drug in preparation, particularly in preparation, treat and/or prevent the new purposes aspect lung-cancer medicament.
Background technology
Cancer, also claim malignant tumor (Malignant neoplasm), serves as reasons and control the not normal and disease that causes of growth and proliferation of cell mechanism.Cancerous cell except grow out of control, arround also can local invading, normal structure is even transferred to other parts of health via body-internal-circulation system or lymphsystem.
Malignant tumor, because differentiation is immature, growth is very fast, infiltrates the 26S Proteasome Structure and Function that destroys organ, and can shift, thereby serious on the body impact.Malignant tumor, except can causing the local compression and obstructive symptom similar to above-mentioned benign tumor, also can have heating, intractable pain, can occur late period seriously becoming thin, the state of weak, anemia and cachexia.Ectopic endocrine syndrome: some non-endocrine gland tumor energy generation and secreting hormone or hormonal substances, cause the clinical symptoms of endocrine regulation, this tumor is called dystopy incretion tumor, and its caused clinical symptoms is called ectopic endocrine syndrome.This type of tumor mostly is malignant tumor, in the majority with cancer, as gastric cancer, hepatocarcinoma, colon cancer, also is found in sarcoma as fibrosarcoma, leiomyosarcoma etc.The tumor of "amine precursor uptake " decarboxylation" (DNES), also can produce biogenic amine or polypeptide hormone in addition, as carcinoid, pheochromocytoma etc.
Because the product of tumor (comprising that ectopic hormone produces) or abnormal immune reaction (comprising cross immunity, autoimmune and immune complex deposit etc.) cause the system generation pathological changes such as endocrine, nerve, hemopoietic, digestion, osteoarthrosis, kidney, skin, cause corresponding clinical symptoms, be called secondary tumor symptomes complice.
Pulmonary carcinoma is the modal malignant tumor of pulmonary.World Health Organization's investigation report, many countries and regions, the sickness rate of pulmonary carcinoma accounts for the first place of malignant tumor.The male is more than the women, and men and women's ratio is about 4-8:1.City is more than rural area.Age is many more than 40 years old, and a few patients is below 40 years old.Most pulmonary carcinoma originate from the bronchial mucosa epithelium, therefore claim lung bronchogenic carcinoma.Pulmonary carcinoma is that it is called for short or usually calls.The treatment of pulmonary carcinoma be take surgical resection as first-selected.Primary carcinoma is swollen to be confined in broncho-pulmonary, and when metastasis and lymphatic metastasis not yet occur, the latter 5 years survival rates of performing the operation can reach more than 50%.But most patients pathological changes when surgery for the first time is medical has belonged to late period, and most patients have lost the chance of radical excision, therefore, early discovery, early diagnosis and early stage treatment are still current doctor's vital task and responsibility.
Cancer treatment method:
1, surgical operation therapy: excision cancerous issue.
2, X-ray therapy: utilize the high-energy electromagnetic radiation line to act on life entity, biomolecular structure is changed, reach the destruction of cancer cells purpose.It is to the lonizing radiation sensitivity because of cancerous cell that radiation can be treated cancer.The lonizing radiation of application have x-ray treatment and r line to treat two kinds clinically at present.
3, chemotherapy: take the medicine intravascular to whole body, all influential to the health all cells.This therapy is sometimes also referred to as " born of the same parents poison therapy ", because medicine used all is harmful to, or even with toxicity, and cells in vivo, whether no matter malignant cell, all be damaged.
4, Chinese traditional treatment cancer is approved by domestic and international Medical Technologist now widely, the effect of Chinese traditional treatment cancer that a large amount of clinical trials and scientific research result are verified.
Root stock and tuber that Stahlianthus hainanensis (Hayata) T. L. Wu is zingiberaceous plant Folium Zingiberis Radix Notoginseng Stahlianthus involucratus (King ex Bak.) Craib.The tool heat clearing away, dampness removing, pain relieving, removing toxic substances, the effect of hemostasis, for stranguria with turbid discharge, stomachache, aphtha, hemorrhoid, ulcer, traumatic injury, venom, the diseases such as traumatic hemorrhage, be Guangxi Special Traditional Chinese Medicine material.
" China's Zingiberaceae medicinal plants study VI Radix Camptandrae Yunnanensis Analysis of The Essential Oil " (being published in " chromatograph " the 1st the 1st phase of volume in 1984), each composition of Radix Camptandrae Yunnanensis volatile oil shared and identified with gas-liquid chromatograph-mass spectrography by gas-liquid chromatograph, being identified 15 kinds of compositions such as Dihydrostahlianthusone, australene, camphene, nopinene, carene, limonene, cineole, linalool, Camphora, α-capaene, trans-Flos Caryophylli alkene, aromadendrene, γ-muurolene, cadinene, Stahlianthusone.
Application number is CN200510098945.2 " a kind of medicine for external use of Stahlianthus hainanensis (Hayata) T. L. Wu ", a kind of medicine for external use be comprised of Stahlianthus hainanensis (Hayata) T. L. Wu, Sabia schumanniana Diels, Herba asplenii prolongati, Herba Tetrastigmatis yunnanensis, Cortex Aceris Sinensis, Prunus brachypoda Batal. Var.eglandulosa Cheng, Caulis Trachelospermi, ethanol is disclosed, rheumatic arthritis, traumatic injury stasis of blood pain, traumatic injury, traumatic hemorrhage, arthralgia, muscles and bones are sprained to pain, bones and muscles pain, pain caused by ecchymoma, lumbocrural pain, joint aches, wound stasis of blood pain, soft tissue contusion's determined curative effect, the course for the treatment of is shorter, and effect is better.
In addition, Stahlianthus hainanensis (Hayata) T. L. Wu or the therapeutic effect of Rhizoma Stahlianthi Involucrati aspect traumatic injury or rheumatism are all mentioned in the patent application that the patent application that the patent application that publication number is CN1814229 " a kind of damp-repellent pain-relieving medicinal liquor of Radix Vaccinii Fragilis ", publication number are CN1742972 " a kind of Chloranthus multistachys Pei medicine for external use ", publication number are CN1814230 " a kind of Herba Viciae Amoenae activating collaterals to relieve pain medicine ", the patent application " a kind of radix-Gynurae-Bodinieri collateral-flow-activating pain-relieving ointment for treating affection by dampness " that publication number is CN1814228, the patent application " damp-clearing pain-relieving liquor and acupoint application treatment method " that publication number is CN101766729.
In existing open source literature, or mention the volatile oil component of Stahlianthus hainanensis (Hayata) T. L. Wu, or mention the effect of the disease such as Stahlianthus hainanensis (Hayata) T. L. Wu treatment traumatic injury, but have no the effect that open use Stahlianthus hainanensis (Hayata) T. L. Wu, Stahlianthus hainanensis (Hayata) T. L. Wu extract, Stahlianthus hainanensis (Hayata) T. L. Wu liposoluble extract and Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil are used for the treatment of tumor, various cancers, not yet have the people to treat tumor, cancer to Stahlianthus hainanensis (Hayata) T. L. Wu and did comprehensively research.
Summary of the invention
The purpose of this invention is to provide the Stahlianthus hainanensis (Hayata) T. L. Wu plant and treat and/or prevent the application of cancer drug in preparation;
The purpose of this invention is to provide the Stahlianthus hainanensis (Hayata) T. L. Wu extract and treat and/or prevent the application of cancer drug in preparation;
The purpose of this invention is to provide the Stahlianthus hainanensis (Hayata) T. L. Wu extractive of volatile oil and treat and/or prevent the application of cancer drug in preparation;
Another object of the present invention is to provide the preparation method of Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil.
Root stock, tuber that Stahlianthus hainanensis (Hayata) T. L. Wu is zingiberaceous plant Folium Zingiberis Radix Notoginseng Stahlianthus involucratus (King ex Bak.) Craib.Excavate after the time of the year when autumn changes into winter blade is withered and yellow, remove impurity, clean, put rapid iron in boiling water, dry.Record in the 8th page of " Guangxi Chinese crude drug standard " nineteen ninety version.
The another name of Stahlianthus hainanensis (Hayata) T. L. Wu has Radix Notoginseng Rhizoma Zingiberis Recens, Stahlianthus hainanensis (Hayata) T. L. Wu, Rhizoma Stahlianthi Involucrati, Folium Bambusae Radix Notoginseng, Stahlianthus hainanensis (Hayata) T. L. Wu, Cor Gigeriae Galli seven, red husky Rhizoma Zingiberis Recens, sharp Radix Notoginseng, drags top rifle (Zhuang), internal diabetes, DABUSI etc.
According to the inventor, Stahlianthus hainanensis (Hayata) T. L. Wu plant, Stahlianthus hainanensis (Hayata) T. L. Wu extract, Stahlianthus hainanensis (Hayata) T. L. Wu liposoluble extract and Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil are carried out to clinical research and pharmacodynamic study, the tumor cell apoptosis can be bred by inhibition tumor cell, be affected to discovery Stahlianthus hainanensis (Hayata) T. L. Wu plant, Stahlianthus hainanensis (Hayata) T. L. Wu extract, Stahlianthus hainanensis (Hayata) T. L. Wu liposoluble extract and Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil, and the change that affects Cancer Cell cycle, presenting the G1 phase blocks, and reaches the effect of anti-tumor activity, treatment cancer.
On this basis, the inventor is had purpose to extract by the effective ingredient to Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil and after content controls, and it is used for the treatment of and/or the more remarkable effect of prophylaxis of cancer.
The safety of patent clinical practice of the present invention is higher, and at cancer a kind of good medicine of particularly can yet be regarded as in the Drug therapy of pulmonary carcinoma, it has widened the new method of Chinese medicine preparation treatment cancer, especially pulmonary carcinoma, particularly applicable at the basic hospital that can not carry out interventional therapy.
Therefore, the applicant provides Stahlianthus hainanensis (Hayata) T. L. Wu plant, Stahlianthus hainanensis (Hayata) T. L. Wu extract, Stahlianthus hainanensis (Hayata) T. L. Wu liposoluble extract, Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil for the preparation of the new purposes that treats and/or prevents the cancer drug aspect.
The invention discloses the Stahlianthus hainanensis (Hayata) T. L. Wu extract and treat and/or prevent the application of cancer drug in preparation, the preparation of described Stahlianthus hainanensis (Hayata) T. L. Wu extract comprises the following steps: get the chopping of Stahlianthus hainanensis (Hayata) T. L. Wu rhizome, extracting in water decocts twice, the decocting that adds for the first time medical material weight 2-6 doubly to measure boils, and boils 1-3 hour after boiling, and the decocting that adds for the second time medical material weight 1-4 doubly to measure boils 1-3 hour, filter, mix decoction liquor twice, be concentrated into the 0.5-2 of medical material gross weight doubly, obtain.
The invention discloses the Stahlianthus hainanensis (Hayata) T. L. Wu extract and treat and/or prevent the application of cancer drug in preparation, the preparation of described Stahlianthus hainanensis (Hayata) T. L. Wu extract is further comprising the steps of: get the chopping of Stahlianthus hainanensis (Hayata) T. L. Wu rhizome, add alcohol reflux 2-3 time, add for the first time the ethanol that 2-6 times of percent by volume is 40-70%, extract 1-3 hour, add for the second time 1-4 doubly to measure the ethanol extraction 1-3 hour that percent by volume is 40-70%, collecting decoction, filter, and reclaims ethanol, make dry cream, obtain.
The invention also discloses the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract and treat and/or prevent the application of cancer drug in preparation, the preparation of described Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract comprises the following steps: get the chopping of Stahlianthus hainanensis (Hayata) T. L. Wu rhizome, add petroleum ether reflux, extract, 2-3 time, add medical material gross weight 3-5 petroleum ether doubly at every turn, extraction time is 1-3 hour, and merge extractive liquid, filters, reclaim petroleum ether, obtain the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract.
The invention discloses the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract and treat and/or prevent the application of cancer drug in preparation, the preparation of described Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract can also comprise the following steps: get the chopping of Stahlianthus hainanensis (Hayata) T. L. Wu rhizome, adding ethyl acetate backflow extracts 2-3 time, add medical material gross weight 3-5 ethyl acetate doubly at every turn, extraction time is 1-3 hour, and merge extractive liquid, filters, reclaim ethyl acetate, obtain the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract.
The invention discloses the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract and treat and/or prevent the application of cancer drug in preparation, the preparation of described Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract can also comprise the following steps: get the chopping of Stahlianthus hainanensis (Hayata) T. L. Wu rhizome, add water-saturated n-butanol reflux, extract, 2-3 time, add medical material gross weight 3-5 water-saturated n-butanol doubly at every turn, extraction time is 1-3 hour, and merge extractive liquid, filters, reclaim water-saturated n-butanol, obtain the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract.
The invention discloses the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract and treat and/or prevent the application of cancer drug in preparation, the preparation of described Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract can also comprise the following steps: get the chopping of Stahlianthus hainanensis (Hayata) T. L. Wu rhizome, add alcohol reflux 2-3 time, add at every turn medical material gross weight 3-5 doubly, the percent by volume ethanol that is 30-50%, extraction time is 1-3 hour, and merge extractive liquid, filters, reclaim ethanol, obtain concentrated solution.
Described concentrated solution adds petroleum ether and divides 3 extractions, and the 3-5 that each petroleum ether consumption is the concentrated solution volume doubly, collects the petroleum ether phase, and concentrate drying, obtain the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract.
Described concentrated solution adds ethyl acetate and divides 3 extractions, and the 3-5 that each ethyl acetate consumption is the concentrated solution volume doubly, collects the petroleum ether phase, and concentrate drying, obtain the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract.
Described concentrated solution adds water-saturated n-butanol and divides 3 extractions, and the 3-5 that each water-saturated n-butanol consumption is the concentrated solution volume doubly, collects the water-saturated n-butanol phase, and concentrate drying, obtain the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract.
The invention discloses the Stahlianthus hainanensis (Hayata) T. L. Wu volatile-oil composite and treat and/or prevent the application of cancer drug in preparation, contain active ingredient in described Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil, the Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil active ingredient that this active ingredient is extracted by Stahlianthus hainanensis (Hayata) T. L. Wu forms, comprising the monoterpenes compound, the oxide-based compound of monoterpene, sesquiterpenoids, the oxide-based compound of sesquiterpene.
Preferably, described Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil active ingredient comprises: 3,6,7 of 20-30 weight portion, 8-tetrahydro-3,3,6,6-tetramethyl cyclopenta [E] indenes-1 (2H)-one, the camphene of 8-20 weight portion; 1,7 of 6-15 weight portion, 7-trimethyl-bicyclo-[2.2.1] heptan-2-thatch ketone, the aromadendrene of 2-8 weight portion; The caryophyllene oxide of 1-6 weight portion.
More preferably, described Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil active ingredient content comprises: 3,6,7 of 24-28 weight portion, 8-tetrahydro-3,3,6,6-tetramethyl cyclopenta [E] indenes-1 (2H)-one, the camphene of 11-15 weight portion; 1,7 of 8-12 weight portion, 7-trimethyl-bicyclo-[2.2.1] heptan-2-thatch ketone, the aromadendrene of 3-6 weight portion; The caryophyllene oxide of 2-5 weight portion.
The applicant also provides a kind of method for preparing described Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil, and it comprises the steps:
Get Stahlianthus hainanensis (Hayata) T. L. Wu and be ground into coarse powder, add medical material gross weight 3-7 distilled water immersion 0.5-3 hour doubly, by water vapour distillation 3-6 hour, collect the upper strata quintessence oil, obtain Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil.
Preferably, the preparation of Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil comprises the following steps:
Get Stahlianthus hainanensis (Hayata) T. L. Wu and pulverize, add the distilled water immersion 1h of 5 times of medical material gross weights, by water vapour distillation 4h, collect the upper strata quintessence oil and obtain the Stahlianthus hainanensis (Hayata) T. L. Wu volatile-oil composite.
Based on above-mentioned Stahlianthus hainanensis (Hayata) T. L. Wu extract, Stahlianthus hainanensis (Hayata) T. L. Wu liposoluble extract, Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil, a kind of new purposes that the purpose of this invention is to provide above compositions, treat and/or prevent the application of cancer drug in preparation, particularly preparation treats and/or prevents the application of lung-cancer medicament.It can make injection, powder ampoule agent for injection, and freeze-dried powder injection, tablet, pill, powder, granule, mixture, syrup, capsule, drop pill preparation, preferably make injection, freeze-dried powder injection.The medicine of above-mentioned various dosage forms all can be according to the conventional method preparation of pharmaceutical field.
Below the crude drug source of Chinese medicine preparation of the present invention:
Stahlianthus hainanensis (Hayata) T. L. Wu: root stock and the tuber of zingiberaceous plant Folium Zingiberis Radix Notoginseng Stahlianthus involucratus (King ex Bak.) Craib.
The invention discloses the new purposes of Stahlianthus hainanensis (Hayata) T. L. Wu in the treatment cancer, the former plant of Stahlianthus hainanensis (Hayata) T. L. Wu and common process extract have certain therapeutic effect for the treatment cancer, the visible experimental example 1 of result and experimental example 2, and, Stahlianthus hainanensis (Hayata) T. L. Wu toxicity is little, can not produce the strong side effect such as chemotherapy.
The extractive of volatile oil of Stahlianthus hainanensis (Hayata) T. L. Wu has outstanding curative effect, cell experiment as shown in experimental example, its concentration is when 0.5mg/ml, just there is obvious activity, surpassed the control drug group at 24 hours or 36 hours suppression ratio to tumor cell, and its concentration is when 2.0mg/ml is above, its activity of killing cancerous cell can surpass 90% especially.
The accompanying drawing explanation
Fig. 1 is that Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is to human nasopharyngeal epithelioma 1 (CNE-2) proliferation inhibition rate line chart
Fig. 2 is that Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is to lung cancer cell line (A549) proliferation inhibition rate line chart
Fig. 3 is that Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is to hepatoma cell strain (HepG2) proliferation inhibition rate line chart
Fig. 4 is that Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is to breast carcinoma cell strain (MCF-7) proliferation inhibition rate line chart
Fig. 5 is the metamorphosis of Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil to human nasopharyngeal epithelioma 1 CNE-2 apoptosis
Fig. 6 is the metamorphosis of Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil to hepatoma cell strain HepG2 apoptosis
Fig. 7 is the metamorphosis of Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil to lung cancer cell line A549 apoptosis
Fig. 8 is that Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is to human nasopharyngeal epithelioma 1 (CNE-2) early apoptosis Annexin V-PI double-staining fluorescence scatterplot
Fig. 9 Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is to human nasopharyngeal epithelioma 1 (CNE-2) apoptosis rate bar diagram
Figure 10 is that Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is to lung cancer cell line (A549) early apoptosis Annexin V-PI double-staining fluorescence scatterplot
Figure 11 is that Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is to lung cancer cell line (A549) apoptosis rate bar diagram
Figure 12 is that Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is to hepatoma cell strain (HepG2) early apoptosis Annexin V-PI double-staining fluorescence scatterplot
Figure 13 is that Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is to hepatoma cell strain (HepG2) apoptosis rate bar diagram
Figure 14 is that Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is to breast carcinoma cell strain (MCF-7) early apoptosis Annexin V-PI double-staining fluorescence scatterplot
Figure 15 is that Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is to breast carcinoma cell strain (MCF-7) apoptosis rate bar diagram
Figure 16 is the impact of Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil on the CNE-2 cell cycle
Figure 17 is the impact of Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil on the A549 cell cycle
Figure 18 is the impact of Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil on the HepG2 cell cycle
Figure 19 is the impact of Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil on the MCF-7 cell cycle
The specific embodiment
Further illustrate the present invention below by embodiment.It should be understood that embodiments of the invention are for the present invention rather than limitation of the present invention are described.The simple modifications that essence according to the present invention is carried out the present invention all belongs to the scope of protection of present invention.Except as otherwise noted, the percent in the present invention is percetage by weight (ethanol is percent by volume).
Embodiment 1: Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil
Get fresh Stahlianthus hainanensis (Hayata) T. L. Wu 150g and be ground into coarse powder, add the distilled water immersion 0.5h of medical material 450ml, by water vapour distillation 3h, collect the upper strata quintessence oil, obtain Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil 1.7ml.
Gained Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is detected to the component content result: 3,6,7,8-tetrahydro-3,3,6,6-tetramethyl cyclopenta [E] indenes-1 (2H)-one is 24%, camphene is 15%; 1,7,7-trimethyl-bicyclo-[2.2.1] heptan-2-thatch ketone is 12%, and aromadendrene is 3%; Caryophyllene oxide is 2%.
Embodiment 2: Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil
Get fresh Stahlianthus hainanensis (Hayata) T. L. Wu rhizome 150g and pulverize, add the distilled water immersion 1h of 750ml, by water vapour distillation 4h, collect the upper strata quintessence oil, obtain Stahlianthus hainanensis (Hayata) T. L. Wu volatile-oil composite 2.2ml.
Gained Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is detected to the component content result: 3,6,7,8-tetrahydro-3,3,6,6-tetramethyl cyclopenta [E] indenes-1 (2H)-one is 28%, camphene is 11%; 1,7,7-trimethyl-bicyclo-[2.2.1] heptan-2-thatch ketone is 8%, and aromadendrene is 6%; Caryophyllene oxide is 5%.
Embodiment 3: Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil
Get Stahlianthus hainanensis (Hayata) T. L. Wu dry rhizome 150g and pulverize, add 1050ml distilled water immersion 3h, by water vapour distillation 6h, collect the upper strata quintessence oil, obtain Stahlianthus hainanensis (Hayata) T. L. Wu volatile-oil composite 2.0ml.
Gained Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is detected to the component content result: 3,6,7,8-tetrahydro-3,3,6,6-tetramethyl cyclopenta [E] indenes-1 (2H)-one is 30%, camphene is 20%; 1,7,7-trimethyl-bicyclo-[2.2.1] heptan-2-thatch ketone is 15%, and aromadendrene is 2%; Caryophyllene oxide is 1%.
Embodiment 4: the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract
Get Stahlianthus hainanensis (Hayata) T. L. Wu rhizome 5000g chopping, add the petroleum ether reflux, extract, 2 times, add for the first time the 20000ml petroleum ether, extract 2 hours, add for the second time the 15000ml Petroleum ether extraction 2 hours, merge extractive liquid,, filter, reclaim petroleum ether, obtain the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract.
Embodiment 5: the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract
Get Stahlianthus hainanensis (Hayata) T. L. Wu rhizome 5000g chopping, add the petroleum ether reflux, extract, 3 times, add for the first time the 25000ml petroleum ether, extract 3 hours, add for the second time the 20000ml Petroleum ether extraction 2 hours, adding for the third time the 15000ml Petroleum ether extraction time is 1 hour, merge extractive liquid,, filter, reclaim petroleum ether, obtain the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract.
Embodiment 6: the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract
Get Stahlianthus hainanensis (Hayata) T. L. Wu rhizome 5000g chopping, adding ethyl acetate backflow extracts 3 times, add for the first time the 25000ml ethyl acetate, extract 3 hours, add for the second time the 20000ml ethyl acetate to extract 2 hours, add for the third time the 15000ml ethyl acetate, extraction time is 1 hour, and merge extractive liquid, filters, reclaim ethyl acetate, obtain the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract.
Embodiment 7: the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract
Get Stahlianthus hainanensis (Hayata) T. L. Wu rhizome 5000g chopping, add the water-saturated n-butanol reflux, extract, 3 times, add for the first time the 25000ml water-saturated n-butanol, extract 3 hours, add for the second time the 20000ml water-saturated n-butanol to extract 2 hours, adding for the third time the 15000ml water-saturated n-butanol to carry extraction time is 1 hour, merge extractive liquid,, filter, reclaim water-saturated n-butanol, obtain the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract.
Embodiment 8: the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract
Get Stahlianthus hainanensis (Hayata) T. L. Wu dry rhizome 5000g chopping, add alcohol reflux 2 times, add for the first time the ethanol that the 25000ml percent by volume is 40%, extract 3 hours, add for the second time the ethanol extraction 2 hours that the 15000ml percent by volume is 50%, merge extractive liquid,, filter, reclaim ethanol, concentrated solution adds petroleum ether and divides 3 extractions, and each petroleum ether consumption is respectively 5 times, 4 times, 3 times of concentrated solution volume, collects the petroleum ether phase, concentrate drying, obtain the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract.
Embodiment 9: the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract
Get Stahlianthus hainanensis (Hayata) T. L. Wu dry rhizome 5000g chopping, add alcohol reflux 3 times, add for the first time the ethanol that the 20000ml percent by volume is 30%, extract 3 hours, add for the second time the ethanol extraction 2 hours that the 15000ml percent by volume is 40%, add for the third time the ethanol extraction 1 hour that the 10000ml percent by volume is 50%, merge extractive liquid, filter, reclaim ethanol, concentrated solution adds ethyl acetate and divides 3 extractions, each ethyl acetate consumption is respectively 5 times of concentrated solution volume, 4 times, 3 times, collect the ethyl acetate phase, concentrate drying, obtain the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract.
Embodiment 10: the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract
Get Stahlianthus hainanensis (Hayata) T. L. Wu dry rhizome 5000g chopping, add alcohol reflux 3 times, add for the first time the ethanol that the 20000ml percent by volume is 30%, extract 3 hours, add for the second time the ethanol extraction 2 hours that the 15000ml percent by volume is 40%, add for the third time the ethanol extraction 1 hour that the 10000ml percent by volume is 50%, merge extractive liquid, filter, reclaim ethanol, concentrated solution adds water-saturated n-butanol and divides 3 extractions, each water-saturated n-butanol consumption is respectively 5 times of concentrated solution volume, 4 times, 3 times, collect the water-saturated n-butanol phase, concentrate drying, obtain the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract.
Embodiment 11: the Stahlianthus hainanensis (Hayata) T. L. Wu water extract
Get fresh Stahlianthus hainanensis (Hayata) T. L. Wu rhizome 1000g chopping, add for the first time 6000ml and decoct, keep 3h after boiling, add for the second time 4000ml water boiling and extraction 1h.Merge decocting liquid twice, filter, be concentrated into 2000ml, obtain the Stahlianthus hainanensis (Hayata) T. L. Wu water extract.
Embodiment 12: the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract
Get fresh Stahlianthus hainanensis (Hayata) T. L. Wu rhizome 1000g chopping, add alcohol reflux 2 times, add for the first time the ethanol that 6 times of percents by volume are 70%, extract to add for the second time 4 times of amount percents by volume be 40% ethanol extraction 1 hour, collecting decoction 3 hours, filter, reclaim ethanol, make dry cream, obtain the Stahlianthus hainanensis (Hayata) T. L. Wu fat soluble component extract.
Embodiment 13: Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil injection
Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil in embodiment 1-3 and glucose, appropriate cosolvent, appropriate solvent is even, filter, sterilizing, prepare injection.
Embodiment 14: Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil lyophilized injectable powder
Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil in embodiment 1-3 and glucose, appropriate cosolvent, appropriate solvent is even, sterilizing, be sub-packed in the containers such as ampoule or cillin bottle, in sterile closed environment, under low temperature, freeze, then, by reducing ambient pressure, the method for the products temperature that slowly raises makes the solvent distillation in goods, stay the medicine of solid forms, obtain freeze-dried powder of the present invention.
Embodiment 15: Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil
Get Stahlianthus hainanensis (Hayata) T. L. Wu dry rhizome 150g and pulverize, add 600ml distilled water immersion 2h, by water vapour distillation 5h, collect the upper strata quintessence oil, obtain Stahlianthus hainanensis (Hayata) T. L. Wu volatile-oil composite 1.9ml.
Gained Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is detected to the component content result: 3,6,7,8-tetrahydro-3,3,6,6-tetramethyl cyclopenta [E] indenes-1 (2H)-one is 20%, camphene is 8%; 1,7,7-trimethyl-bicyclo-[2.2.1] heptan-2-thatch ketone is 6%, and aromadendrene is 8%; Caryophyllene oxide is 6%.
Effect experiment
Experimental example 1 the present invention is to the Cytostatic to tumor cell effect
Get Stahlianthus hainanensis (Hayata) T. L. Wu volatile-oil composite, ligroin extraction, ethyl acetate extract, water-saturated n-butanol extract, water extract and ethanol extraction in embodiment 2,5,6,7,11,12, with the RPMI1640 culture fluid, the said extracted thing is diluted, 0.22 the degerming of μ m micropore filter sucking filtration, 4 ℃ save backup.
Detect the present invention to the Cytostatic to tumor cell effect with mtt assay.
1 sample and Experimental agents
1.1 tumor cell culture
KB cell (CNE-2), human lung carcinoma cell (A549), human liver cancer cell (HepG2), human breast cancer cell (MCF-7) tumor cell line all, by the RPMI1640 culture medium containing 10% hyclone and 100 μ g/ml penicillin-streptomycins, are put 37 ℃, 5%CO
2, cultivate and go down to posterity in the constant-temperature enclosed incubator under the saturated humidity condition.The 0.02EDTA-0.25% trypsinization, every 2～3d goes down to posterity once.
1.2 Experimental agents
Experimental group: embodiment 2,5,6,7,11,12.
Comparative Examples medicine: cisplatin, Dezhou Deyao Pharmaceutical Co., Ltd of manufacturer; Lot number: the accurate word H37020524 of traditional Chinese medicines; Concentration 3 μ g/ml.The 2MTT method detects the Stahlianthus hainanensis (Hayata) T. L. Wu extract to the Cytostatic to tumor cell effect
Table 1 embodiment of the present invention and the cisplatin suppression ratio to tumor cell proliferation
Experimental result shows: 2 pairs of Cytostatic to tumor cell effects of embodiment obviously are better than other embodiment groups and cisplatin group; Each embodiment group is compared with the cisplatin group, and to Cytostatic to tumor cell, effect has a little less than having by force, but the propagation all embodied tumor cell has inhibitory action.By the above conclusion that obtains:
1, Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil obviously is better than to the inhibited proliferation of tumor cell the extract that Stahlianthus hainanensis (Hayata) T. L. Wu obtains through other extracting method, and obviously is better than cisplatin.。
2, Stahlianthus hainanensis (Hayata) T. L. Wu ligroin extraction, ethyl acetate extract are better than Stahlianthus hainanensis (Hayata) T. L. Wu water-saturated n-butanol extract, Stahlianthus hainanensis (Hayata) T. L. Wu water extract, Stahlianthus hainanensis (Hayata) T. L. Wu ethanol extraction and cisplatin to the inhibited proliferation of tumor cell.
3, Stahlianthus hainanensis (Hayata) T. L. Wu water extract, ethanol extraction and water-saturated n-butanol extract are compared poorly with the cisplatin group to the inhibited proliferation of tumor cell, but can demonstrate tumor cell proliferation is had to inhibitory action equally.
Experimental example 2 Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil anti tumor activity in vitro experiments
1 instrument and reagent
1.1 the instrument volatile oil determination apparatus meets the related standards of an appendix XD determination of volatile oil method of Chinese Pharmacopoeia; Galaxy170S type CO2 cell culture incubator (German Eppendorf company), MLDEL680 type microplate reader (Japanese BIO-RAD company),
Axiovert-40 type inverted phase contrast microscope (Zeiss, Germany company), SW-CJ-2F type superclean bench (Purifying Equipment Co., Ltd., Suzhou).
1.2 reagent hyclone, RPMI1640 culture medium are all purchased from U.S. hyclone company; DMSO, tetramethyl azo azoles blue (MTT) are all purchased from Sigma company; Other reagent such as ether are domestic analytical pure.
1.3 Experimental agents
Experimental group: embodiment 1, Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil.
Comparative Examples medicine: cisplatin, Dezhou Deyao Pharmaceutical Co., Ltd of manufacturer; Lot number: the accurate word H37020524 of traditional Chinese medicines; Concentration 3 μ g/ml.1.4 for the examination tumor cell
KB cell (CNE-2), human lung carcinoma cell (A549), human liver cancer cell (HepG2), human breast cancer cell (MCF-7) provide by Medical Colleges Of Guilin's scientific experiments center.
2 methods and result
2.1 method
2.1.1 medicine preparation
With the RPMI1640 culture fluid, the extractive of volatile oil of embodiment 1 gained (20mg/mL) is diluted in 1:10,1:13,1:20,1:40 ratio, 0.22 μ m micropore filter sucking filtration degerming, 4 ℃ save backup.
2.1.2 experiment grouping:
A: blank group
B:0.5mg/ml Stahlianthus hainanensis (Hayata) T. L. Wu volatilization line of oils
C:1.0mg/ml Stahlianthus hainanensis (Hayata) T. L. Wu volatilization line of oils
D:1.5mg/ml Stahlianthus hainanensis (Hayata) T. L. Wu volatilization line of oils
2.1.3 tumor cell culture
CNE-2, A549, HepG2, MCF-7 tumor cell line all, by the RPMI1640 culture medium containing 10% hyclone and 100 μ g/ml penicillin-streptomycins, are put 37 ℃, 5%CO
2, cultivate and go down to posterity in the constant-temperature enclosed incubator under the saturated humidity condition.The 0.02EDTA-0.25% trypsinization, every 2～3d goes down to posterity once.
2.1.4MTT method detects medicine of the present invention to the Cytostatic to tumor cell effect
The tumor cell of exponential phase is inoculated to 96 well culture plates, 37 ℃, 5%CO by 4000, every hole
2Cultivate 12h in incubator, 24h, 36h, 48h, 60h, after cell attachment, adds respectively cell strain to grow up in 96 porocyte culture plates of monolayer variable concentrations Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil, each concentration repeats 5 multiple holes, set up two matched groups: one is blank group (not adding cell) simultaneously, and one is cell matched group (add RPMI-1640, the concentration of Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is 0).Be placed in 37 ℃, 5%CO
2Cultivate 24h in incubator.Cultivate and finish front 4h, inhale and abandon liquid in culture plate, PBS rinses 3 times, and under the lucifuge condition, every hole adds MTT(5mg/ml) 20 μ l, be placed in 37 ℃, 5%CO
2Continue to cultivate 4h in incubator, remove supernatant after every hole add DMSO150 μ l, horizontal shaking table concussion 10min, elisa reading instrument colorimetric (wavelength 490nm) is surveyed absorbance.Experiment repeats 3 times.Experiment with computing medicine and the control drug inhibitory action to tumor cell as follows.Cell inhibitory rate (%)=(1-drug effect group absorbance/cell matched group absorbance) * 100%.
2.1.5Hoechst33258 dyeing observation of cell form
Get the clean coverslip be soaked in 70% ethanol, be placed in six orifice plates, A group (blank group) is set, B group (0.5mg/ml) C group (1.0mg/ml), D group (1.5mg/ml), with aseptic PBS tri-times, then wash one time with cell culture fluid in aseptic super-clean bench.The different tumor cells (CNE-2, A549, HepG2, MCF-7) of the trophophase of taking the logarithm, every hole inoculation 5x10
4Individual cell culture spends the night.After variable concentrations Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is processed 24h, exhaust culture fluid, PBS washing 3 times, add the 0.5ml fixative, fixes 30 minutes.Remove fixative, wash 2 times with PBS, each 3 minutes, exhaust liquid.Add 0.5ml Hoechst33258 dyeing liquor, dye 10 minutes.Remove dyeing liquor, wash 2 times with PBS, each 3 minutes, exhaust liquid.Drip anti-fluorescent quenching mounting liquid on microscope slide, cover the coverslip that posts cell, allow cells contacting mounting liquid, avoid bubble as far as possible.With about fluorescence exciting wavelength 350nm, emission wavelength 460nm left and right can detect the nucleus that is blue under fluorescence microscope.
2.1.6 flow cytometer detects volatile oil to tumour cell cycle and apoptotic impact
Different tumor cells (HepG2, CNE-2, A549, MCF-7) are divided into four groups: A group (blank group), B group (0.5mg/ml), C group (1.0mg/ml), D group (1.5mg/ml), after different disposal, after collecting all cells (adherent and suspension cell), with PBS, clean once, every group adds respectively 100 μ l buffer, piping and druming evenly, add respectively again 3 μ l PI and 3 μ Annexin-V, lucifuge dyeing 1 hour, add 200 μ l buffer, 200 mesh filter screen filtration cells, be collected in flow cytometer loading pipe.Use 488nm laser as excitation wavelength, 525nm is as detecting wavelength, on flow cytometer to 10,000 cell collection fluorescence intensity data in each sample.
2.1.7 flow cytometer detects the cycle of tumor cell
The different tumor cells of trophophase (CNE-2, A549, HepG2, MCF-7) of taking the logarithm are divided into A group (blank group), B group (0.5mg/ml) C group (1.0mg/ml), D group (1.5mg/ml) is after different disposal, collect all cells (adherent and suspension cell) in the 15ml centrifuge tube after, with PBS, clean once, every group adds respectively 1mL pre-cooling PBS resuspended, and piping and druming evenly.Centrifuge tube is positioned on the vortex oscillator, and concussion limit, limit adds the freezing ethanol of 9ml70% to fix, and places-20 ℃ and spends the night.Before dyeing, with pre-cooling PBS, wash, centrifugal (2000r/min, 4 ℃) 5min, remove fixative, with 500 μ L RNaseA digestion, 37 ℃ of water-bath 30min, add again 25 μ L propidium iodide (propdium iodide, PI) dyeing liquors to mix, room temperature lucifuge dyeing 30min, 200 mesh filter screen filtration cells, be collected in flow cytometer loading pipe.Flow cytometer carries out DNA content and cell cycle analysis, draws the percentage rate in each phase of the cycles of cell.
2.1.8 statistical analysis
All experimental datas are with mean ± standard deviation
Mean, use SPSS17.0 typing and analytical data, between many groups, data relatively adopts one factor analysis of variance (One-Way ANOVA) check, and P<0.05 means that difference has significance.
2.2 result
2.2.1 the present invention is to the growth of tumour cell inhibitory action
Adopt the impact of MTT experimental analysis Experimental agents on tumor cell line (CNE-2, A549, HepG2, MCF-7) propagation, result is as shown in table 2-5 and Fig. 1-4:
Table 2: Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is to human nasopharyngeal carcinoma cell line (CNE-2) proliferation inhibition rate
Table 3: different time human lung carcinoma cell line (A549) proliferation inhibition rate (%, x ± s, n=3)
Table 4: different time human hepatoma cell strain (HepG2) proliferation inhibition rate (%, x ± s, n=3)
Table 5: different time human breast cancer cell strain (MCF-7) proliferation inhibition rate (%, x ± S, n=3)
Annotate:
◇: embodiment group and cisplatin group (positive controls) compare, the P value > 0.05, embodiment group and cisplatin group relatively do not have significant difference, illustrate that the anti-tumor activity of embodiment group is equal to the cisplatin group substantially.
*: embodiment group and cisplatin group (positive controls) are relatively, P value<0.05, embodiment group and cisplatin group relatively have significant difference, from its suppression ratio, are analyzed, the embodiment group is higher than cisplatin group suppression ratio, illustrates that the anti-tumor activity of embodiment group will be higher than the cisplatin group.
△: embodiment group and cisplatin group (positive controls) are relatively, P value<0.01, it is the significant difference that embodiment group and cisplatin group relatively have significance, from its suppression ratio, analyzed, the embodiment group is lower than cisplatin group suppression ratio, illustrates that the anti-tumor activity of embodiment group will or not have anti-tumor activity lower than the cisplatin group.
His-and-hers watches 2-5 experimental data is carried out after statistical analysis known, after effect 24 hours, and the IC of volatile oil of the present invention to tumor cell line (CNE-2, A549, HepG2, MCF-7)
50Be respectively 1.27mg/ml, 1.24mg/ml, 0.84mg/ml, 1.04mg/ml.
Experimental result shows: volatile oil of the present invention all has significant inhibitory action to CNE-2, A549, HepG2, MCF-7 tumor cell line, and its successful is better than control drug cisplatin group.And, with the prolongation of action time and the increase of drug level, its inhibitory action strengthens gradually, show obvious amount-effect and time-effect relationship.
2.2.2Hoechst33258 dyeing observation of cell form
Hoehst33258 staining observation of cell nuclear morphology changes, and result is as shown in Fig. 5-7, and A group karyomorphism is complete, and nuclear membrane is smooth, and chromatin is even, the metamorphosis of the visible division cells core of part; B group, C group, D group be visible nuclear metamorphosis of apoptosis phase all, and nuclear membrane disappears, and the core edge is the burr shape, nuclear staining is deepened, and the part karyopyknosis, is fragmented into the fragment that nuclear membrane is wrapped in, and is graininess, form apoptotic body, obvious apoptosis form (as shown by arrows) occurs.After the inoculation of equivalent amount cell, with the increase of drug dose, the cell number of various dose concentration experimental group also obviously reduces, and side light Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil increases and strengthens gradually with dosage the effect of different tumor cell line apoptosis.
2.2.3 flow cytometer detects the impact of Stahlianthus hainanensis (Hayata) T. L. Wu volatile-oil composite on different tumour cell cycles
Flow cytometry (FCM) result shows as Figure 16-19 table 6-9: after the different tumor cell 24h of different quality concentration Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil effect, with A group (blank group), compare the variation of B, C, D group (dosage group in low) cell quantity:
At CNE-2, MCF-7 cell, in B, C group, G0/G1 phase cell quantity increases, S phase, G2/M phase cell proportion reduce, and D group (1.5mg/ml experimental group) G1 phase cell proportion reduces, may be because Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is arrested in the G1 phase by CNE-2, MCF-7, cause apoptosis of tumor cells, and then cause G1 phase cell proportion to reduce, prompting Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil has obvious G1 phase retardation to CNE-2, MCF-7 cell.
In A549, HepG2 cell, G0/G1 phase cytosis in B, C, D group, S phase and G2/M phase Leukopenia, show that Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil can effectively suppress the normal conversion of cell cycle, cell was piled up in the G1 phase, cell block is in the G1 phase, thereby the mitosis of prevention cell is suppressed cell proliferation.
Table 6: the impact (%) of Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil on the CNE-2 cell cycle
Table 8: the impact (%) of Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil on the HepG2 cell cycle
Table 9: the impact (%) of Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil on the MCF-7 cell cycle
2.2.4 flow cytometer detects the apoptosis of tumor cell
Detect the effect of Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil after 24 hours through flow cytometer, the apoptosis rate of different tumor cells (CNE-2, A549, HepG2, MCF-7) is as shown in Fig. 8-15, can be observed the increase with Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil concentration, the ratio of different tumor cell apoptosis increases gradually.
In the impact on the CNE-2 apoptosis, the B group causes that apoptosis rate is 6%, with blank, compares P=0.085 > 0.05, no significant difference, do not have statistical significance.C group and D organize apoptosis rate and sharply rise, and do not present obvious early apoptosis, directly cause cell in, late period apoptosis or necrocytosis.
In impact on the A549 apoptosis, the B group causes that apoptosis rate is 6.57%, with blank, compares P=0.224 > 0.05, no significant difference, do not have statistical significance.C group and D group relatively have notable difference with blank, and its early apoptosis rate raises with the increase of dosage.
In impact on the HepG2 apoptosis, each group compares P=0.00<0.01. with blank and has obvious statistical significance.The apoptosis rate that B, C are two groups is without obvious difference, and D group apoptosis rate sharply rises, and the cell of most apoptosis is still in early apoptosis.
In the impact on the MCF-7 apoptosis, each group compares P=0.00<0.01 with blank, has obvious statistical significance.Apoptosis rate between tri-groups of B, C, D is without obvious difference.
Conclusion: at flow cytometer, detect in the apoptosis experiment of tumor cell, result shows: in the impact on CNE-2, A549 apoptosis, the B group is compared no significant difference with blank group, not statistically significant, and C group, D group have been compared notable difference with blank group; In impact on HepG2, MCF-7 apoptosis, B, C, D group are compared obvious statistical significance are all arranged with blank group.
The experimental result prompting: (1), to CNE-2, A549 tumor cell, Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil low dosage has no significant effect effect, and middle high dose group has remarkable effect.(2), to HepG2, MCF-7 tumor cell, during Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil is low, high dose group all presents obvious influence.
Experimental result shows, Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil has obvious G1 retardation to CNE-2, MCF-7 cell.To A549, HepG2 cell, can effectively suppress the normal conversion of cell cycle, cell was piled up in the G1 phase, cell block, in the G1 phase, stops the mitosis of cell, cell proliferation is suppressed, thereby reaches the anti-tumor activity effect.
Claims (12)
1. the Stahlianthus hainanensis (Hayata) T. L. Wu plant treats and/or prevents the application of cancer drug in preparation.
2. application as claimed in claim 1 is characterized in that: the Stahlianthus hainanensis (Hayata) T. L. Wu plant treats and/or prevents the application of lung-cancer medicament in preparation.
3. the Stahlianthus hainanensis (Hayata) T. L. Wu extract treats and/or prevents the application of cancer drug in preparation.
4. application according to claim 3 is characterized in that: described Stahlianthus hainanensis (Hayata) T. L. Wu extract is the liposoluble constituent in the Stahlianthus hainanensis (Hayata) T. L. Wu plant.
5. application as described as claim 3 or 4 is characterized in that: described Stahlianthus hainanensis (Hayata) T. L. Wu extract treats and/or prevents the application of lung-cancer medicament in preparation.
6. application as claimed in claim 3 is characterized in that: the Stahlianthus hainanensis (Hayata) T. L. Wu extract is Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil.
7. application as claimed in claim 6, it is characterized in that: in described Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil, contain active ingredient, the Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil active ingredient that this active ingredient is extracted by Stahlianthus hainanensis (Hayata) T. L. Wu forms, comprising the monoterpenes compound, the oxide-based compound of monoterpene, sesquiterpenoids, the oxide-based compound of sesquiterpene.
8. application as claimed in claim 6 is characterized in that: contain active ingredient in described Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil, the Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil active ingredient that this active ingredient is extracted by Stahlianthus hainanensis (Hayata) T. L. Wu forms, comprising the composition of following weight proportion:
3,6,7 of 20-30 weight portion, 8-tetrahydro-3,3,6,6-tetramethyl cyclopenta [E] indenes-1 (2H)-one, the camphene of 8-20 weight portion; 1,7 of 6-15 weight portion, 7-trimethyl-bicyclo-[2.2.1] heptan-2-thatch ketone, the aromadendrene of 2-8 weight portion; The caryophyllene oxide of 1-6 weight portion.
9. application as claimed in claim 6 is characterized in that: contain active ingredient in described Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil, the Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil active ingredient that this active ingredient is extracted by Stahlianthus hainanensis (Hayata) T. L. Wu forms, comprising the composition of following weight proportion:
3,6,7 of 24-28 weight portion, 8-tetrahydro-3,3,6,6-tetramethyl cyclopenta [E] indenes-1 (2H)-one, the camphene of 11-15 weight portion; 1,7 of 8-12 weight portion, 7-trimethyl-bicyclo-[2.2.1] heptan-2-thatch ketone, the aromadendrene of 3-6 weight portion; The caryophyllene oxide of 2-5 weight portion.
10. application as claimed in claim 6, is characterized in that, the preparation of described Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil comprises the following steps:
Get Stahlianthus hainanensis (Hayata) T. L. Wu and be ground into coarse powder, add medical material gross weight 3-7 distilled water immersion 0.5-3 hour doubly, by water vapour distillation 3-6 hour, collect the upper strata quintessence oil, obtain.
11. application as claimed in claim 6, is characterized in that, the preparation of described Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil comprises the following steps:
Get Stahlianthus hainanensis (Hayata) T. L. Wu and be ground into coarse powder, add the distilled water immersion 1h of 5 times of medical material gross weights, by water vapour distillation 4h, collect the upper strata quintessence oil, obtain.
12. as any one described application of claim 6-11, it is characterized in that: Stahlianthus hainanensis (Hayata) T. L. Wu volatile oil treats and/or prevents the application of lung-cancer medicament in preparation.
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| CN201410173805.6A CN104096150B (en) | 2013-07-17 | 2014-04-28 | Stahlianthus hainanensis (Hayata) T. L. Wu and its extract are preparing treatment and/or the application of prevention cancer drug |
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| CN112618676A (en) * | 2020-10-10 | 2021-04-09 | 桂林医学院 | Application of gynura segetum and extract thereof in preparing medicine for treating and/or preventing vascular endothelial injury |
| CN113908240A (en) * | 2020-07-08 | 2022-01-11 | 广东臻宝堂中药技术开发有限公司 | Traditional Chinese medicine conditioning formula for gastric cancer patients and preparation method |
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| CN113908240A (en) * | 2020-07-08 | 2022-01-11 | 广东臻宝堂中药技术开发有限公司 | Traditional Chinese medicine conditioning formula for gastric cancer patients and preparation method |
| CN112618676A (en) * | 2020-10-10 | 2021-04-09 | 桂林医学院 | Application of gynura segetum and extract thereof in preparing medicine for treating and/or preventing vascular endothelial injury |
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