CN103156891A - Preparation method of akebia fruit active components and preparation method of preparation of akebia fruit active components and application in preparation of antineoplastic medicines of preparation of foreknowledge sub-active components - Google Patents
Preparation method of akebia fruit active components and preparation method of preparation of akebia fruit active components and application in preparation of antineoplastic medicines of preparation of foreknowledge sub-active components Download PDFInfo
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Abstract
The invention belongs to the technical field of medicines, in particular to a preparation method of akebia fruit active components and a preparation method of the preparation of the akebia fruit active components and the application in the preparation of antineoplastic medicines of the akebia fruit active components. The preparation method comprises steps as below: smashing the akebia fruit medical materials into coarse powders, soaking the coarse powders in alcohol and heating, refluxing and extracting the akebia fruit medical materials for two to three times, filtering the akebia fruit medical materials, and merging filter liquor. Carrying out vacuum concentration, stewing and centrifugation, and therefore sediment I and supernatant fluid I are obtained. Adjusting the potential of hydrogen (PH) of the supernatant fluid I through alkali to 8 to 14, hydrolyzing the supernatant fluid I for 2 to 8 hours under the temperature of 60 Celsius degrees to 100 Celsius degrees, adding acid to adjust the PH of the supernatant fluid to 3 to 7 after the hydrolyzing, carrying out centrifugation, and therefore the sediment II and the supernatant fluid II after the hydrolyzing are obtained. Merging the sediment I and the sediment II, drying the sediment, and therefore the akebia fruit active components are obtained. The active components have significant antineoplastic activity, and can be used for manufacturing medicines of antineoplastic treatment or auxiliary antineoplastic treatment.
Description
Technical field
The invention belongs to medical technical field, be specifically related to a kind of preparation method of Fructus Akebiae active component and preparation method and its application in the preparation antitumor drug of preparation thereof, this active component has significant anti-tumor activity, can be used for preparing antitumor drug or adjuvant therapy medicaments of tumor.
Background technology
Fructus Akebiae is the dry almost ripe fruit of plants of Lardizabalaceae Caulis Akebiae [Akebia quinata (Thunb.) Decne.], threeleaf akebia [Akebia trifoliate (Thunb.) Koidz.] or Caulis Akebiae [Akebia trifoliate (Thunb.) Koidz. var. australis (Diels) Rehd.].Bitter in the mouth, cold.Return liver, gallbladder, stomach, urinary bladder channel.Be used for liver-smoothing, qi-regulating, promoting blood circulation and stopping pain, diuresis, eliminating stagnation.Be used for gastral cavity side of body distending pain, amenorrhea dysmenorrhea, dysuria, sucutaneous nodule mass in the abdomen.Bibliographical information: saponin component is the main component in Fructus Akebiae, have protect the liver, the multiple pharmacologically actives such as relieving convulsion, pain relieving, antifungal, parasite killing, these effects have dependency with the clinical efficacy of Fructus Akebiae.Through research repeatedly, a few saponins in Fructus Akebiae have the antineoplastic activity in addition, through systematic study, prepare the antitumor drug of exploitation take this active component as main component.
Summary of the invention
The object of the present invention is to provide a kind of Fructus Akebiae extract, another object of the present invention is to provide the application in the antitumor field of a kind of preparation method of Fructus Akebiae extract and this extract.
Fructus Akebiae active component of the present invention is achieved by the following technical solution:
Get the Fructus Akebiae medical material, it is ground into coarse powder, with 5-95% soak with ethanol 0.5-2h, 6-12 doubly measures heating and refluxing extraction 1-3 time, and each 1-3h filters merging filtrate; Above-mentioned filtrate is evaporated to 0.2-1.0g raw medicinal herbs/mL in 50-70 ℃, and is static, centrifugal, must precipitate I and supernatant I; The supernatant I is transferred PH to 8-14 with sodium hydroxide, and 60-100 ℃ is hydrolyzed 2-8 hour, adds hydrochloric acid after hydrolysis and transfers PH to 3-7, centrifugal, must precipitate the rear supernatant II of II and hydrolysis; Merge the precipitation I, the precipitation II, drying namely gets the Fructus Akebiae active component.
Extracting purification through this technique can be with saponin component content in the Fructus Akebiae antitumor component greater than 50%; The main saponin of measuring in the precipitation I through the HPLC method is triterpene saponin (3-O-β-D-glucopyranosyl-(1 → 3)-α-L-rhamnopyranosyl-(1 → 2)-α-L-arabopyranose Hederagenin, saponins B, saponins P
DWith saponins A) amount accounted for more than 90% of total saponins amount in the Fructus Akebiae, tool anti-tumor activity, and supernatant I anti-tumor activity is not obvious, but after basic hydrolysis, obtain the precipitation II, measure its main component by the HPLC method identical with the precipitation I, and anti-tumor activity is arranged.
In initial preparation technology, the supernatant II is adsorbed through macroporous adsorptive resins, water, 40% ethanol 70-90% ethanol elution successively after loading are collected the 70-90% ethanol elution, and concentrated, drying gets the extract III.The research discovery, the component of precipitation I, precipitation II and extract III is basic identical (mainly contains 3-O-β-D-glucopyranosyl-(1 → 3)-α-L-rhamnopyranosyl-(1 → 2)-α-L-arabopyranose Hederagenin, saponins B, saponins P
DWith saponins A), and in three part components the content of total saponins all greater than 50%.Yet the yield of precipitation I is 2-8%, and the yield of precipitation II is 2-5%, and the yield of extract III is 0.1-0.3%, and from economizing on resources and economic benefit aspect consideration, this part can be not considered macroporous resin, merges precipitation I and precipitation II in reality; Namely get the Fructus Akebiae active component.Precipitation I and precipitation II in the present invention, use separately or merge the medicine that uses (the preferred merging uses) to may be used to prepare antitumor or be used for tumor aid treatment, this active component can be used as active component and pharmaceutical carrier and is mixed with various preparations on pharmaceutics, the dispersity of Fructus Akebiae antitumor component in preparation can be liquid, can be also solid, the route of administration of said preparation can be oral, injection and topical.
Measure saponin component content with ultraviolet/visible spectrophotometry in the present invention:
1, the preparation of oleanolic acid reference substance solution
Precision takes oleanolic acid reference substance 5mg, is transferred in the 50mL volumetric flask, uses dissolve with methanol, and standardize solution is made into the standard solution of 0.1mg/mL, and stored refrigerated is standby.
2, the oleanolic acid maximum absorption wavelength determines
Accurate reference substance solution and the testing sample solution 1ml of drawing, each 2 parts (portion is done blank, portion is done colour developing), add in the tool plug test tube of the drying of label, volatilize in 60 ℃ of vacuum drying ovens, blank group adds the methanol of 5ml, the colour developing group adds the perchloric acid of 5ml, shake up, colour developing is organized in 70 ℃ of water-baths and is reacted 15min, and jolting constantly, put immediately cooling 15min in ice-water bath after reacting completely, retinue reagent is blank, in Shimadzu UV-2550 ultraviolet spectrophotometer, scans in 200～700nm wave-length coverage.Need testing solution and reference substance solution have absorption maximum at 310nm wavelength place as a result, therefore determine that measuring wavelength is 310nm.
3, the drafting of standard curve
Precision pipettes reference substance solution 0.3,0.6,0.9,1.2,1.5,1.8 mL each 2 parts (portion is done blank, portion is done colour developing), be transferred in the tool plug test tube of 10mL, volatilize in 60 ℃ of vacuum drying ovens, blank group adds the methanol of 5ml, the colour developing group adds the perchloric acid of 5ml, shakes up, and colour developing is organized in 70 ℃ of water-baths and reacted 15min, and jolting constantly, put immediately cooling 15min in ice-water bath after reacting completely, retinue reagent is blank, in Shimadzu UV-2550 ultraviolet spectrophotometer, at 200 ~ 500nm length scanning, the place measures trap at the 310nm wavelength.Take trap (A) as vertical coordinate, oleanolic acid reference substance concentration (C) is abscissa, and the drawing standard curve is made method of least square with trap A to concentration C and returned, and gets the standard curve equation to be: A=0.022C+0.045, R
2=0.9996(n=4), the range of linearity is 7.0～34.3 mg/mL, in the range of linearity, the linear relationship of mass concentration and trap value is good.
4, saponin component content assaying method in sample
Get the Fructus Akebiae medical material appropriate, pulverize, take medical material 34.90g, with 10 times of amount 70% ethanol (being 350ml), extract 2 times, each 1.5 hours, gained solution filters with Medium speed filter paper, is concentrated into below 100ml, is transferred to the volumetric flask of 100ml, be settled to scale, namely get the Fructus Akebiae extract of 0.349g/ml; Pipette Fructus Akebiae extracting solution 1ml, be transferred in the volumetric flask of 250ml, be settled to scale, namely get the Fructus Akebiae need testing solution of 1.396mg/ml.The accurate need testing solution 1ml that draws, 2 parts (portion is done blank, portion is done colour developing), add in the tool plug test tube of drying of label, volatilize in 60 ℃ of vacuum drying ovens, blank group adds the methanol of 5ml, the colour developing group adds the perchloric acid of 5ml, shakes up, and colour developing is organized in 70 ℃ of water-baths and reacted 15min, and jolting constantly, put immediately cooling 15min in ice-water bath after reacting completely, retinue reagent is blank, in Shimadzu UV-2550 ultraviolet spectrophotometer, at 200 ~ 500nm length scanning, measure its trap at the 310nm place.Try to achieve in sample total saponins concentration and calculate content by standard curve.
With the HPLC high-efficient liquid phase technique, the Fructus Akebiae active component is carried out component analysis in the present invention:
1, the preparation of need testing solution: get approximately 50mg of Radix Pulsatillae antitumor component powder, accurately weighed, be placed in the 50mL volumetric flask, dissolve with methanol also is settled to scale, filters, and gets subsequent filtrate 10mL as need testing solution.
2, chromatographic condition: chromatographic column: COSMOSIL(4.6*250mm, 5 μ m) C
18The ODS post; Mobile phase: acetonitrile-water-phosphoric acid=45:55:0.1; Detect wavelength: 203nm; Column temperature: 25 ℃; Flow velocity: 1mL/min.Sample size 20 μ L.
Through component analysis, four main triterpene saponin in the Fructus Akebiae active component are 3-O-β-D-glucopyranosyl-(1 → 3)-α-L-rhamnopyranosyl-(1 → 2)-α-L-arabopyranose Hederagenin, saponins B, saponins P
DSee accompanying drawing for details with saponins A().
Description of drawings
Fig. 1: the HPLC chromatogram of precipitation I
Fig. 2: the HPLC chromatogram of precipitation II
Fig. 3: the HPLC chromatogram of the Fructus Akebiae active component after precipitation I and precipitation II merge
In figure, 1 is that 3-O-β-D-glucopyranosyl-(1 → 3)-α-L-rhamnopyranosyl-(1 → 2)-α-L-arabopyranose Hederagenin, 2 is that saponins B, 3 is saponins P
D, 4 be saponins A.
The specific embodiment:
Relevant Fructus Akebiae antitumor component preparation method specific embodiment:
Embodiment 1: the preparation method of Fructus Akebiae active component, and wherein: the Fructus Akebiae pulverizing medicinal materials is become coarse powder, with 20% soak with ethanol 0.5h, measure heating and refluxing extraction 2 times for 6 times, each 1h filters merging filtrate; Above-mentioned filtrate is evaporated to 0.5g raw medicinal herbs/mL in 50 ℃, must precipitate I and supernatant I, and supernatant I hydro-oxidation sodium is transferred PH to 9,70 ℃ of hydrolysis 7h, and after hydrolysis, hydrochloric acid is transferred pH to 4, and is centrifugal, must precipitate II and hydrolysis and separate rear supernatant II; Drying, precipitation I yield is 5%, precipitation II yield is 3.4%; Merge precipitation I and precipitation II, namely get the Fructus Akebiae active component.Detect through outer/visible spectrophotometry, in the precipitation I, total saponin content is 54%, and in the precipitation II, total saponin content is 74%, and in the Fructus Akebiae active component after merging, the content of total saponins is 60%.
Embodiment 2: the preparation method of Fructus Akebiae active component, and wherein: the Fructus Akebiae pulverizing medicinal materials is become coarse powder, with 80% soak with ethanol 2h, measure heating and refluxing extraction 3 times for 12 times, each 3h filters merging filtrate; Above-mentioned filtrate is evaporated to 1.0g raw medicinal herbs/mL in 70 ℃, centrifugal, must precipitate I and supernatant I, supernatant I hydro-oxidation sodium is transferred PH to 10,80 ℃ of hydrolysis 6h, after hydrolysis, hydrochloric acid is transferred pH to 5, centrifugal, must precipitate the rear supernatant II of II and hydrolysis, drying, precipitation I yield is 5.4%, and precipitation II yield is 3.8%; Merge precipitation I and precipitation II, namely get the Fructus Akebiae active component.Detect through outer/visible spectrophotometry, in the precipitation I, total saponin content is 66%, and in the precipitation II, total saponin content is 85%, and in the Fructus Akebiae active component after merging, the content of total saponins is 74%.All the other are with embodiment 1.
Embodiment 3: the preparation method of Fructus Akebiae active component, and wherein: the Fructus Akebiae pulverizing medicinal materials is become coarse powder, with 50% soak with ethanol 1h, measure heating and refluxing extraction 2 times for 8 times, each 2h filters merging filtrate; Above-mentioned filtrate is evaporated to 0.7g raw medicinal herbs/mL in 60 ℃, centrifugal, must precipitate I and supernatant I, supernatant I hydro-oxidation potassium is transferred PH to 11,90 ℃ of hydrolysis 5h, after hydrolysis, hydrochloric acid is transferred pH to 6, centrifugal, must precipitate the rear supernatant II of II and hydrolysis, drying, precipitation I yield is 4.6%, and precipitation II yield is 3.2%; Merge precipitation I and precipitation II, namely get the Fructus Akebiae active component.Detect through outer/visible spectrophotometry, in the precipitation I, total saponin content is 58%, and in the precipitation II, total saponin content is 84%, and in the Fructus Akebiae active component after merging, the content of total saponins is 68%.All the other are with embodiment 1.
Embodiment 4: the Fructus Akebiae pulverizing medicinal materials is become coarse powder, with 50% soak with ethanol 0.5h, measure heating and refluxing extraction 3 times for 12 times, each 1.5h filters merging filtrate.Above-mentioned filtrate is evaporated to 0.5g raw medicinal herbs/mL in 60 ℃, and is centrifugal, must precipitate I and supernatant I, and supernatant I hydro-oxidation sodium is transferred PH to 12,90 ℃ of hydrolysis 4h, and after hydrolysis, sulphuric acid is transferred pH to 7, and is centrifugal, must precipitate the rear supernatant II of II and hydrolysis; Drying, precipitation I yield is 2.4%, precipitation II yield is 4.8%; Merge precipitation I and precipitation II, namely get the Fructus Akebiae active component.Detect through outer/visible spectrophotometry, in the precipitation I, total saponin content is 62%, and in the precipitation II, total saponin content is 85%, and in the Fructus Akebiae active component after merging, the content of total saponins is 71%.All the other are with embodiment 1.
Embodiment 5: the Fructus Akebiae pulverizing medicinal materials is become coarse powder, with 95% soak with ethanol 2h, measure heating and refluxing extraction 2 times for 6 times, each 3h filters merging filtrate.Above-mentioned filtrate is evaporated to 0.5g raw medicinal herbs/mL in 50 ℃, and is centrifugal, must precipitate I and supernatant I, and supernatant I hydro-oxidation potassium is transferred PH to 13,100 ℃ of hydrolysis 3h, and after hydrolysis, hydrochloric acid is transferred pH to 6, and is centrifugal, must precipitate II and hydrolysis and separate rear supernatant II; Drying, precipitation I yield is 4.4%, precipitation II yield is 3.0%; Merge precipitation I and precipitation II, namely get the Fructus Akebiae active component.Detect through outer/visible spectrophotometry, in the precipitation I, total saponin content is 56%, and in the precipitation II, total saponin content is 78%, and in the Fructus Akebiae active component after merging, the content of total saponins is 63%.All the other are with embodiment 1.
Embodiment 6: the Fructus Akebiae pulverizing medicinal materials is become coarse powder, and 10% soak with ethanol 1.5h measures heating and refluxing extraction 1 time for 8 times, and each 3h filters merging filtrate.Above-mentioned filtrate is evaporated to 0.5g raw medicinal herbs/mL in 50 ℃, and is centrifugal, must precipitate I and supernatant I, and supernatant I hydro-oxidation sodium is transferred PH to 14,60 ℃ of hydrolysis 8h, and after hydrolysis, sulphuric acid is transferred pH to 6, and is centrifugal, must precipitate the rear supernatant II of II and hydrolysis; Drying, precipitation I yield is 5.0%, precipitation II yield is 3.0%; Merge precipitation I and precipitation II, namely get the Fructus Akebiae active component.Detect through outer/visible spectrophotometry, in the precipitation I, total saponin content is 66%, and in the precipitation II, total saponin content is 85%, and in the Fructus Akebiae active component after merging, the content of total saponins is 74%.All the other are with embodiment 1.
Embodiment 7: get the Fructus Akebiae medical material, it is ground into coarse powder, with 20% soak with ethanol 0.5h, measure heating and refluxing extraction 1 time for 6 times, each 1h filters merging filtrate; Above-mentioned filtrate is evaporated to 0.2g raw medicinal herbs/mL in 50 ℃, and is centrifugal, must precipitate I and supernatant I, and supernatant I hydro-oxidation sodium is transferred PH to 12,90 ℃ of hydrolysis 6h, and after hydrolysis, hydrochloric acid is transferred pH to 6, and is centrifugal, must precipitate the rear supernatant II of II and hydrolysis; Drying, precipitation I yield is 3.2%, precipitation II yield is 4.2%; Merge precipitation I and precipitation II, namely get the Fructus Akebiae active component.Detect through outer/visible spectrophotometry, in the precipitation I, total saponin content is 55%, and in the precipitation II, total saponin content is 75%, and in the Fructus Akebiae active component after merging, the content of total saponins is 67%.All the other are with embodiment 1.
Embodiment 8: get the Fructus Akebiae medical material, it is ground into coarse powder, with 80% soak with ethanol 2h, measure heating and refluxing extraction 3 times for 12 times, each 3h filters merging filtrate; Above-mentioned filtrate is evaporated to 1.0g raw medicinal herbs/mL in 70 ℃, and is centrifugal, must precipitate I and supernatant I, and supernatant I hydro-oxidation potassium is transferred PH to 11,100 ℃ of hydrolysis 4h, and after hydrolysis, hydrochloric acid is transferred pH to 5, and is centrifugal, must precipitate the rear supernatant II of II and hydrolysis; Drying, precipitation I yield is 7.4%, precipitation II yield is 2.8%; Merge precipitation I and precipitation II, namely get the Fructus Akebiae active component.Detect through outer/visible spectrophotometry, in the precipitation I, total saponin content is 63%, and in the precipitation II, total saponin content is 87%, and in the Fructus Akebiae active component after merging, the content of total saponins is 77%.All the other are with embodiment 1.
Embodiment 9: get the Fructus Akebiae medical material, it is ground into coarse powder, with 60% soak with ethanol 1h, 6-12 doubly measures heating and refluxing extraction 2 times, and each 2h filters merging filtrate; Above-mentioned filtrate is evaporated to 0.7g raw medicinal herbs/mL in 60 ℃, and is centrifugal, must precipitate I and supernatant I, and supernatant I hydro-oxidation sodium is transferred PH to 13,70 ℃ of hydrolysis 8h, and after hydrolysis, sulphuric acid is transferred pH to 5, and is centrifugal, must precipitate the rear supernatant II of II and hydrolysis; Drying, precipitation I yield is 5.4%, precipitation II yield is 3.8%; Merge precipitation I and precipitation II, namely get the Fructus Akebiae active component.Detect through outer/visible spectrophotometry, in the precipitation I, total saponin content is 60%, and in the precipitation II, total saponin content is 81%, and in the Fructus Akebiae active component after merging, the content of total saponins is 74%.All the other are with embodiment 1.
Embodiment 10: the preparation method of Fructus Akebiae active component, wherein: in the Fructus Akebiae active component, saponin component content is more than 50%.All the other are with embodiment 1.
Embodiment 11: the preparation that described Fructus Akebiae active component is made, wherein: this Fructus Akebiae active component and excipient substance are mixed with the various preparations on pharmaceutics.All the other are with any one in embodiment 1,2,3,4,5,6,7,8,9 and 10.
Embodiment 12: the preparation that described Fructus Akebiae active component is made, wherein: the dispersity of this Fructus Akebiae active component in preparation can be liquid, can be also solid.All the other are with embodiment 11.
Embodiment 13: described Fructus Akebiae active component, wherein: the route of administration of said preparation can be oral, injection and topical.All the other are with embodiment 11.
Embodiment 14, be raw material with the Fructus Akebiae active component of the method for the embodiment of the present invention 1 or embodiment 2 or embodiment 3 or embodiment 4 or embodiment 5 or embodiment 6 or embodiment 7 or embodiment 8 or embodiment 9 or embodiment 10 preparation, carry out lower series preparation preparation:
1, tablet preparation
Prescription (50):
Composition weight (g)
Active component 5.0 of the present invention
Lactose 4.8
Dextrin 9
Microcrystalline Cellulose 6
Magnesium stearate 0.2
According to the operational approach of conventional tablet, above mix homogeneously, wet granulation adds the magnesium stearate mix homogeneously to be pressed into every 500mg at last.
2, capsule preparation
Prescription (90):
Composition weight (g)
Active component 8 of the present invention
Lactose 5
Dextrin 9
Microcrystalline Cellulose 5
According to the operational approach of conventional tablet, above mix homogeneously, wet granulation, fill becomes capsule, every 300mg.
3, cataplasma preparation
Prescription:
Composition weight (g)
Polyacrylate 18
Laurocapram 0.6
Lauric acid 1.8
Propylene glycol 5.4
Ethyl acetate 15
Carry out according to the affected of common cataplasma, stir above-mentioned, coat on the polyester adherent layer, 60 degree cover poly-amino methyl film backing layer after dry 2 minutes, and section gets final product.
4, injectable powder preparation
Prescription:
Composition weight (g)
Active component 0.05 of the present invention
Hydrochloric acid is appropriate
Mannitol 50
Operation according to conventional freeze-dried powder is carried out, and active component of the present invention is added the dissolving of appropriate water for injection, adds mannitol, is settled to 1000ml, regulates pH value between 4.0-5.5, aseptic filtration, lyophilization and get final product.
5, the preparation of aqueous injection
The Fructus Akebiae antitumor component, be dissolved in appropriate water for injection, add the injection water soluble adjuvant of metering, the active carbon that adds 0.1-0.5% is removed pyrogen, regulates PH to 6.0-7.5, after the microporous filter membrane ultrafiltration, the supplementary injection water is to ormal weight, and fill is in ampoule bottle or in infusion bottle, sealing by fusing/roll lid, through after the assay was approved, pack and get final product.
6, drop pill preparation
Prescription:
Composition weight (g)
Active component 0.1g of the present invention
Polyethylene glycol 6000 10g
Macrogol 4000 20g
Polyoxyethylene sorbitan monoleate 0.3g
Ethanol 1ml
Make 1000 balls
Method for making: Fructus Akebiae antitumor component of the present invention is dissolved in ethanol, become active component and disperse ethanol, add in the substrate that the polyethylene glycol 6000, Macrogol 4000, polyoxyethylene sorbitan monoleate of melting form, be stirred well to evenly, take dimethicone as coolant, 15 ℃ of lower drippings become ball, wipe ball, drying, and get final product.
Embodiment 15: the active component with the preparation of the method in the embodiment of the present invention one is raw material, carries out the antitumor component pharmacodynamic study:
1, eliminating evil effect
1.1 the inhibitory action to animal transplanting tumor
Adopt 7402 Liver Cancer Bearing Nude Mices, MFC TCM23 gastric cancer in nude mice and Mice Bearing Lewis lung cancer model, investigate the anti-tumor in vivo effect of Fructus Akebiae antitumor component (YZS).The nude mice of 7402 hepatocarcinoma, MFC TCM23 gastric cancer and A549 pulmonary carcinoma will be inoculated respectively, be divided at random positive controls (cyclophosphamide), solvent control group (blank), high, medium and low three the dosage groups (50 of Fructus Akebiae antitumor component of the present invention (YZS), 100,200mg/kg), gastric infusion is 10 days continuously, and the last administration was put to death animal after 24 hours, separate the oxter tumor and weigh, calculating tumor control rate.Be calculated as follows tumour inhibiting rate: tumour inhibiting rate (%)=[(the average tumor of the average tumor weight-administration of blank group group is heavy)/average tumor of blank group is heavy] * 100%.Experimental result is as follows:
(1) on the impact of 7402 hepatocarcinoma: experimental result shows YZS(50,100, and the tumor that 200mg/kg) can significantly suppress nude mice is heavy, and three dosage suppression ratio are respectively 16.55,44.82 and 57.24%.The results are shown in Table 1.
Table 1 Fructus Akebiae antitumor component (YZS) is right
7402The impact of Liver Cancer Bearing Nude Mice
* P＜0.05, * * P＜0.01 is compared with the blank group
(2) on the impact of MFC TCM23 gastric cancer in nude mice: YZS(50,100,200mg/kg) nude mice tumor weight average there is certain inhibitory action.The results are shown in Table 2.
The impact of table 2 Fructus Akebiae antitumor component (YZS) on MFC TCM23 gastric cancer in nude mice
* P＜0.05, * * P＜0.01 is compared with the blank group
(3) on the impact of A549 pulmonary carcinoma: experimental result shows YZS(50,100,200mg/kg) the heavy suppression ratio of mice A549 lung cancer tumor is respectively 14.21%, 33.73% and 47.34%.The results are shown in Table 3.
The impact of table 3 Fructus Akebiae antitumor component (YZS) on mice A549 pulmonary carcinoma
* P＜0.05 is compared with the blank group
2, centralizing function
2.1 the impact of Fructus Akebiae antitumor component (YZS) on the tumor animal immunologic function
Can obviously the raise killing activity of spleen index, splenocyte conversion ratio and NK cell of tumor-bearing mice of experimental result prompting Fructus Akebiae antitumor component (YZS).
Set up H
22The tumor-bearing mice animal model
,Fructus Akebiae antitumor component (YZS) (50,100,200mg/kg) gastric infusion, continuous 10 days, the last administration was after 24 hours, with sacrifice of animal, get the tumor-bearing mice Thymus and spleen, weigh, calculate respectively thymus index: [heavy (the mg)/body weight (g) of thymus] * 10, spleen index: [heavy (the mg)/body weight (g) of spleen] * 10.
(1) on the impact of mouse thymus exponential sum spleen index
Compare YZS(100,200 mg/kg with the blank group) can obviously improve H
22The spleen index of tumor-bearing mice and thymus index, but cyclophosphamide obviously reduces spleen index and thymus index.Prompting, cyclophosphamide can cause tumor-bearing mice spleen and atrophy of thymus gland.Preliminary explanation YZS may improve the immunity of mice in the performance antitumaous effect, the results are shown in Table 4.
Table 4 Fructus Akebiae antitumor component (YZS) is to H
22The immune impact of tumor-bearing mice (
± s)
| Group | Dosage (mg * kg -1×d) | Spleen index | Thymus index |
| YZS | 200 | 10.56±1.78* | ?3.78±1.89* |
| ? | 100 | 9.14±2.49 | 3.21±1.45 |
| ? | 50 | 8.45±2.89 | 2.79±1.34 |
| Cyclophosphamide | 30 | 7.02±2.10* | ? 1.02±0.49** |
| The blank group | ? | 8.70±2.89 | 2.56±1.23 |
* P＜0.05, * * P＜0.01 is compared with the blank group
(2) impact of human peripheral blood cell
Through analysis of accounts, with the blank group relatively, cyclophosphamide group murine interleukin sum, lymphocyte and erythrocyte significantly reduce (
p＜0.01), YZS respectively organizes murine interleukin sum, lymphocyte, mononuclear cell and neutrophilic granulocyte number and compares with the blank group, do not have significant difference (
p〉0.05).Prompting, cyclophosphamide obviously suppresses the mouse immune cell, and each group of YZS is obviously different from the effect of cyclophosphamide, to mouse peripheral blood cell unrestraint effect.The results are shown in Table 5.
The impact of table 5 Fructus Akebiae antitumor component (YZS) on tumor-bearing mice peripheral blood immunocyte
* P＜0.05, compare with the blank group
3, median lethal dose(LD 50)
Adopt Fructus Akebiae antitumor component powder, be mixed with the solution of desired concn with distilled water, adopt cleaning level mice, provided by the Jiangxi College of Traditional Chinese Medicine Experimental Animal Center.Median lethal dose(LD 50) when the acute toxicity testing result shows the Fructus Akebiae extract oral is 4.90g/kg.
Precipitation I, precipitation II and the Fructus Akebiae active component of the method preparation of embodiment 16, employing embodiment 2 or embodiment 3 or embodiment 4 or embodiment 5 embodiment 6 or embodiment 7 or embodiment 8 or embodiment 9 or embodiment 10 are carried out the research of antitumor drug effect, and effect is similar to Example 15.
The Fructus Akebiae active component of the method preparation of embodiment 17, employing embodiment 2 or embodiment 3 or embodiment 4 or embodiment 5 embodiment 6 or embodiment 7 or embodiment 8 or embodiment 9 or embodiment 10 and excipient substance are mixed with the various preparations on pharmaceutics.
Embodiment 18, the preparation that adopts the described Fructus Akebiae active component of embodiment 17 to make, wherein: the dispersity in preparation can be liquid to the Fructus Akebiae active component as antitumor component, can be also solid.
Embodiment 19, according to described any one Fructus Akebiae active component of method of embodiment 2 or embodiment 3 or embodiment 4 or embodiment 5 embodiment 6 or embodiment 7 or embodiment 8 or embodiment 9 or embodiment 10 as antitumor component, wherein: the Fructus Akebiae active component can be for the preparation of antitumor drug or adjuvant therapy medicaments of tumor as Studies on Anticancer Components.
Claims (7)
1. the application of Fructus Akebiae active component in the preparation antitumor drug, it is characterized in that: the Fructus Akebiae active component can be for the preparation of antitumor drug or adjuvant therapy medicaments of tumor.
2. the preparation method of Fructus Akebiae active component is characterized in that: get the Fructus Akebiae medical material, it is ground into coarse powder, with 5-95% soak with ethanol 0.5-2h, 6-12 doubly measures heating and refluxing extraction 1-3 time, each 1-3h, filtration, merging filtrate; Above-mentioned filtrate is evaporated to 0.2-1.0g raw medicinal herbs/mL in 50-70 ℃, and is centrifugal, must precipitate I and supernatant I; To 8-14,60-100 ℃ is hydrolyzed 2-8 hour to the supernatant I with adjusting PH with base, and after being hydrolyzed, acid adding is transferred pH to 3-7, and is centrifugal, must precipitate II and the rear supernatant II of hydrolysis, merges precipitation I and precipitation II, and drying namely gets the Fructus Akebiae active component.
3. the application of Fructus Akebiae active component in the preparation antitumor drug, it is characterized in that: in the Fructus Akebiae active component, saponin component content is more than 50%; Wherein precipitate in I total saponin content greater than 50%, in the precipitation II, total saponin content greater than 50%, merges in the Fructus Akebiae active component after precipitation I and precipitation II total saponin content greater than 50%.
4. the application of Fructus Akebiae active component in the preparation antitumor drug, it is characterized in that: Fructus Akebiae precipitation I according to claim 2 and precipitation II are used separately or merge to use to may be used to prepare antitumor drug or adjuvant therapy medicaments of tumor because main constituent is basic identical.
5.
According to claim 2The preparation method of described Fructus Akebiae active component is characterized in that: described alkali comprises sodium hydroxide, potassium hydroxide; Described acid comprises hydrochloric acid, sulphuric acid.
6.
Claim 1 or 2 or 3The preparation that described Fructus Akebiae active component is made is characterized in that: this Fructus Akebiae active component and excipient substance are mixed with the various preparations on pharmaceutics.
7.
According to claim 2 or 3Described Fructus Akebiae antitumor component is characterized in that: tablet, capsule, pill, granule, injection, oral liquid, suppository, soft capsule, dispersible tablet, drop pill and subcutaneous administration preparation that this extract can be made clinically or pharmaceutically accept.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN103191161A (en) * | 2013-04-22 | 2013-07-10 | 上海中医药大学 | Application of fructus akebiae extract in preparation of drug for treating primary hepatic carcinoma |
| CN107260777A (en) * | 2017-06-28 | 2017-10-20 | 南京中医药大学 | A kind of fiveleaf akebia fruit with antitumor activity refines total saposins and its application |
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| 《中国药学杂志》 20060831 王家明 《预知子中alpha2常春藤皂苷的HPLC 分析》 1212-1213 1-7 , * |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103191161A (en) * | 2013-04-22 | 2013-07-10 | 上海中医药大学 | Application of fructus akebiae extract in preparation of drug for treating primary hepatic carcinoma |
| CN107260777A (en) * | 2017-06-28 | 2017-10-20 | 南京中医药大学 | A kind of fiveleaf akebia fruit with antitumor activity refines total saposins and its application |
| CN107260777B (en) * | 2017-06-28 | 2020-12-08 | 南京中医药大学 | Refined fiveleaf akebia fruit total saponin with anti-tumor activity and application thereof |
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