CN103694248A - Antitumor compound extracted from guttifer, and preparation method and application thereof - Google Patents
Antitumor compound extracted from guttifer, and preparation method and application thereof Download PDFInfo
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Abstract
The invention relates to an antitumor compound extracted from guttifer, and a preparation method and application thereof. The preparation method of the compound comprises the following steps: (1) taking the medicinal material guttifer, extracting with ethyl acetate, carrying out silica gel column chromatographic separation on the extract, and carrying out petroleum ether-ethyl acetate gradient elution to obtain 11 components A-K; (2) discarding the grease components and garcinia acid components, carrying out HPLC-PDA-MS (high performance liquid chromatography-potato dextrose agar-Murashige-Skoog) repeat analysis to obtain a key component I; (3) carrying out silica gel column chromatographic separation on the component I, carrying out chloroform-methanol gradient elution to obtain a fraction I-2, collecting one fraction for every 30ml, and merging identical components according to the TLC (thin layer chromatography) result to obtain a fraction I-3; (4) carrying out reversed phase RP-18 separation on I-3, carrying out acetonitrile-water gradient elution, collecting one fraction for every 20ml, and merging identical components to obtain a subfraction I-3E; and (5) carrying out purification and acetonitrile elution on I-3E to obtain the compound. The compound can be used for treating tumors.
Description
Technical field
The present invention relates to medical technical field, particularly a kind ofly from Chinese medicinal materials gamboge, extract separated compound, prepare the method for described compound, the medical composition that comprises described compound, and use described compound to prepare the purposes of antitumor drug.
Background technology
Gamboge is that the trunk of Garcinia maingayii gamboge (Garcinia hanburyi Hook f.) is hurt the colloidal resin of rear outflow, has another name called extra large rattan, beautiful Huang, month Huang, cured Huang etc.Originate in South East Asia, there is introducing culture China some areas as Yunnan, Guangxi, Guangdong and other places.
The gamboge beginning is loaded in < < Haiyao Bencao, Oversea Materia Medica > >, it is cold in nature, taste is sour, puckery, pungent, poisonous, there is the effect of broken blood dissipating bind, removing toxic substances, hemostasis, desinsection, the traditional Chinese medical science is used for controlling swollen ulcer drug, stubborn dermatitis is disliked sore, bleeding due to trauma, noma decayed tooth, burn.
At present, reported from gamboge, be divided into from identified more than 20 compounds, be mostly xanthone (xanthone) compounds containing bridged ring, i.e. gambogic acid, Mo Lilin class, morellin class, and gamboge aldehyde, Betulinic acid etc. multiple compounds.
Modern pharmacology research shows, Resina garciniae extract has the anti-microbial activity of wide spectrum, and streptococcus aureus, Pseudomonas aeruginosa etc. is had to stronger bacteriostatic action.And pharmacological evaluation and clinical experience prove, it has antitumor action, can effectively suppress tumor proliferation, and kinds of tumors is had to significant curative effect.
Contriver extracts and obtains having bioactive the compounds of this invention from gamboge, compares have more superior anti-tumor activity with morellic acid.
Summary of the invention
The invention provides a kind of from gamboge, extract separated have bioactive formula (1) compound the purposes that the preparation method of this compound is provided and has prepared medicine for treating tumor thing.
Formula (1) compound has following formula:
The present invention also provides the preparation method of this formula (1) compound, and concrete steps are:
(1) get gamboge medicinal material and extract by ethyl acetate, extract is separated through silica gel column chromatography, and petroleum ether-ethyl acetate gradient elution, obtains 11 component A～K;
(2) discard oil component A, B and morellic acid component D, E, through HPLC-PDA-MS, analyze, obtain emphasis component I;
(3) component I is separated through silica gel column chromatography again, chloroform-methanol gradient elution, and every 30ml collects a flow point, merges the 11st part to the 16th part elutriant, and flow velocity is 2mL/min, obtains flow point I-3;
(4) get I-3 through anti-phase RP-18 chromatographic separation, acetonitrile-water gradient elution, every 20ml collects a flow point, merges the 13rd part to the 18th part elutriant, and flow velocity is 2mL/min, obtains Arius and divides I-3E;
(5) purified, the acetonitrile isocratic elution of I-3E, obtains formula (1) compound.
Preferably, step (1) PetroChina Company Limited. ether-ethyl acetate by volume the ratio of 50～0:1 carry out gradient elution;
Preferred, petroleum ether-ethyl acetate is divided into six gradients and carries out wash-out, is respectively 50:1,10:1,5:1,2:1,1:1,0:1.
Preferably, in step (3) chloroform-methanol by volume the ratio of 50～10:1 carry out gradient elution;
Preferably, in step (4) acetonitrile-water by volume the ratio of 40～90:60～10 carry out gradient elution;
Preferred, acetonitrile-water is divided into five gradients and carries out wash-out, is respectively 40:60,50:50,60:40,80:20,90:10.
Preferably, in step (5), the concentration of acetonitrile is 80%～95%;
Preferred, the concentration of acetonitrile is 80%～85%.
Contriver by physico-chemical property and the Modern spectroscopy section of learning to do (
1h-NMR (Fig. 2),
13c-NMR (Fig. 3), HR-ESI-MS,
1h-
1h COSY and HSQC) the separated compound obtaining has been carried out to Structural Identification, be confirmed that it is structure suc as formula the compound shown in (1).
In one aspect, formula of the present invention (1) compound can be used for the treatment of tumor disease.Contriver finds that the compounds of this invention has the inhibition activity of wide spectrum to tumour cell.Described tumor disease comprises the group that is selected from following composition: neurogliocytoma, adenocarcinoma of lung and ovarian cancer.
Another aspect, the present invention also provides the pharmaceutical composition for the treatment of tumor disease, it comprises formula (1) compound for the treatment of significant quantity, and at least one is selected from following pharmaceutically acceptable non-active ingredient: pharmaceutically acceptable thinner, pharmaceutically acceptable vehicle and pharmaceutically acceptable supporting agent.
Accompanying drawing explanation
Fig. 1 is formula (1) structural formula of compound;
Fig. 2 is formula (1) compound
1h-NMR spectrum;
Fig. 3 is formula (1) compound
13c-NMR spectrum.
Embodiment
compound
Formula (1) compound has following structure:
medical composition/composite
Applicable dispensing path includes, but is not limited to per os, intravenously, rectum, aerosol, non-through intestines, eye, lung, through mucous membrane, through skin, vagina, ear, intranasal, intramuscular injection, subcutaneous injection and topical administration.In addition, only for instance, non-ly through intestines transmission, comprise intramuscular, subcutaneous, intravenously, intramedullary injection, and in sheath, directly in indoor, intraperitoneal, lymph and nasal injection.
In certain embodiments, compound as herein described is with part but not general mode administration.In other embodiments, compound as herein described provides to discharge fast composite form, prolongation release composite form or the middle composite form that discharges.In other embodiments, compound as herein described is local administration.
In certain embodiments, compound as herein described is through being allocated as medical composition.In a particular embodiment, medical composition in the usual way, with one or more physiologically acceptable supporting agents, allocate, described physiologically acceptable supporting agent comprises vehicle and auxiliary agent, and it contributes to active compound to be processed as the preparation that can be used for pharmacy.Suitably composite depends on selected dosing way.
Medical composition refers to the mixture of formula (1) compound and other chemical composition, and described chemical composition is as being supporting agent, stablizer, thinner, dispersion agent, suspension agent, thickening material and/or vehicle.In certain embodiments, medical composition contributes to Mammals administration compound.
In certain embodiments, compound as herein described is through allocating for oral administration.Compound as herein described is allocated with oral dosage form, and only for instance, described oral dosage form comprises tablet, pulvis, pill, sugar-coat ingot, capsule, liquid, gel, syrup, elixir, slurries, suspension and analogue thereof.
In certain embodiments, described medical composition is tablet, capsule, powder ampoule agent for injection, injection, pill and slow releasing tablet.
In one embodiment, formula (1) compound is allocated in the aqueous solution.In a particular embodiment, only for instance, the aqueous solution is selected from physiological compatibility damping fluid, as Han Shi liquid (Hank ' s solution), ringer's solution (Ringer ' s solution) or normal saline buffer solution.
In other embodiments, formula (1) compound is used for offeing medicine through mucous membrane through allotment.In a particular embodiment, through mucous membrane composite, comprise the permeate agent that is suitable for wanting permeability barrier.
At compound as herein described, through allotment, be used for other non-other embodiment through enteral administration, suitably composite comprises water-based or non-aqueous solution.
In certain embodiments; by mixing one or more solid excipients and one or more compounds as herein described, optionally grind gained mixture and add to be applicable to where necessary auxiliary agent post-treatment granular mixture and obtain to obtain tablet or pill the pharmaceutical preparation using for per os.Specifically, suitable vehicle is weighting agent, as sugar, comprises lactose, sucrose, N.F,USP MANNITOL or sorbyl alcohol; Cellulose preparation, as W-Gum, wheat starch, rice starch, yam starch, gelatin, tragacanth, methylcellulose gum, Microcrystalline Cellulose, Vltra tears, Xylo-Mucine; Or other material, as polyvinylpyrrolidone (PVP or polyvidone) or calcium phosphate.In a particular embodiment, optionally add disintegrating agent.Only for instance, disintegrating agent comprises croscarmellose sodium, polyvinylpyrrolidone, agar or Lalgine or its salt, as sodium alginate.
Oral dosage form also comprises the cooperation insertion capsule of being made by gelatin, and the soft seal capsule of being made by gelatin and softening agent (as glycerine or sorbyl alcohol).In a particular embodiment, coordinate and insert the mixture that capsule contains activeconstituents and one or more weighting agents.Only for instance, weighting agent comprises lactose, as tackiness agents such as starch and/or as lubricants such as talcum or Magnesium Stearates, and the optional stablizer using.In other embodiments, soft capsule contains one or more dissolvings or is suspended in the active compound in suitable liquid.Only for instance, suitably liquid comprises one or more fatty oils, liquid paraffin or liquid polyethylene glycol.In addition, optionally add stablizer.
In other embodiments, formula (1) compound is local administration.Composition that can local administration comprises solution, suspension, lotion, gel, paste, swab, balm, newborn frost or ointment.
In other embodiments, formula (1) compound is through allocating for inhalation dosing.The various forms that is suitable for inhalation dosing includes, but is not limited to aerosol, spraying or pulvis.
In order to make those skilled in the art understand better technical scheme of the present invention, below in conjunction with specific embodiment, the present invention is described in further detail.Provide described example only for purpose of explanation, and wish does not limit the scope of claims provided herein.
The preparation method of embodiment 1 formula of the present invention (1) compound
(1) by ethyl acetate, extract gamboge medicinal material (ethyl acetate and gamboge weight ratio are 15:1), extract is separated through silica gel column chromatography, petroleum ether-ethyl acetate (50:1,10:1,5:1,2:1,1:1,0:1) six gradient elutions, flow velocity is 2mL/min, every 30ml collects a flow point, and every 7 parts of elutriants merge, and obtain successively 11 component A～K;
(2) discard after oil component A and B and morellic acid component D and E, then analyze through HPLC-PDA-MS, obtain emphasis component I;
(3) component I is separated through silica gel column chromatography again, chloroform-methanol (50:1～10:1) gradient elution, and every 30ml collects a flow point, merges the 11st part to the 16th part elutriant, and flow velocity is 2mL/min, obtains flow point I-3;
(4) get I-3 through anti-phase RP-18 chromatographic separation, acetonitrile-water (40:60,50:50,60:40,80:20,90:10) gradient elution, every 20ml collects a flow point, merges the 13rd part to the 18th part elutriant, flow velocity is 2mL/min, obtains Arius and divides I-3E;
(5) purified, the 82% acetonitrile isocratic elution of I-3E, obtains formula of the present invention (1) compound.The Structural Identification of embodiment 2 formula of the present invention (1) compound
This compound is yellow jelly, [α] 24D-116.7 (c0.06, MeOH);
1h and
13c NMR data are in Table 1; UV (MeOH) λ
max318,276nm; IR (KBr) v
max3431,2970,2927,1740,1689,1627,1587,1441,1377,1325,1176,1113,955nm; HR-TOF-MS shows quasi-molecular ion peak m/z699.3150 (calcd for C
39h
48o
10na, 699.3145).
The nuclear magnetic data of table 1 formula (1) compound (
1h (500MHz, CDCl
3),
13c (100MHz, CDCl
3) NMR)
The restraining effect of formula (1) compound to people's lung cancer transplanted tumor
1. materials and methods
1.1 tested material
Formula (1) compound: 10mg/ml, purity 97%, lot number 20100121, is prepared by Shanghai Pharmaceutical Inst., Chinese Academy of Sciences.
Vinorelbine tartrate injection liquid (Gai Nuo): Jiangsu Haosen Pharmaceutical Co., Ltd, lot number 100301.
Physiological saline: Shandong Lukang Cisen Pharmaceutical Co., Ltd, lot number 1004234202.
Morellic acid: 10mg/ml, purity 98% (detecting through HPLC), lot number 20100819, is prepared by Kangyuan Pharmaceutical Co., Ltd., Jiangsu Prov.
1.2 transplanted tumor
People's lung cancer NCI-H460 Nude Mice, is inoculated in nude mouse by people's lung cancer NCI-H460 cell strain subcutaneous and set up.Cell inoculum size is 3 * 10
6, inoculation is used after forming and passing for 3 generations again in nude mouse body after transplanted tumor.
1.3 animal
Female BALB/c nude mouse, age in days 35-40 days, body weight 18-22g, is provided by Nanjing Military Command hospital general experimentation on animals section.12 of the negative control groups of every treated animal number, 6 of administration groups.
1.4 test method
The tumor tissue of getting growth animated period cuts into 1.5mm
3left and right, under aseptic condition, is inoculated in nude mouse right side armpit subcutaneous.Vernier caliper measurement transplanted tumor diameter for Nude Mice, treats tumor growth to 100～300mm
3after by animal random packet.Use the method for measuring knurl footpath, dynamically observe the antineoplastic effect of tested medicine.The measurement number of times of diameter of tumor is 3 times weekly, and each measurement also needs to claim mouse heavy simultaneously.Formula (1) compound intravenously administrable, dosage divides and is 12mg/kg, 6mg/kg, 3mg/kg, gives three times weekly.Lid promise intravenously administrable 10mg/kg, gives twice weekly, each 0.2ml, and negative control group is given equivalent physiological saline simultaneously.
1.5 detect index and method of calculation
(1) gross tumor volume (tumor volume, TV), calculation formula is:
(2) relative tumour volume (relative tumor volnme, RTV), calculation formula is:
RTV=TV
t／TV
0
TV wherein
0(d during for minute cage administration
0) gross tumor volume, TV
tgross tumor volume when measuring each time.
(3) relative tumor proliferation rate T/C (%), calculation formula is:
T／C(％)=T
RTV／C
RTV×100
T
rTV: treatment group RTV; C
rTV: negative control group RTV.
Test-results is usingd the evaluation index of relative tumor proliferation rate T/C (%) as anti-tumor activity.
1.6 statistical method
Experimental data represents with mean value and standard deviation, statistical method employing t-check.
2. result
Formula (1) compound the results are shown in Table 2 to the experimental treatment of people's lung cancer NCI-H460.
Formula (1) compound high dose group intravenously administrable has good growth-inhibiting effect to people's lung cancer NCI-H460 mice-transplanted tumor, and preferably T/C (%) is 61.97;
Middle dosage group intravenously administrable 6mg/kg has certain growth-inhibiting effect to people's lung cancer NCI-H460 Nude Mice, and preferably T/C (%) is 73.00;
Low dose group intravenously administrable 3mg/kg has certain growth-inhibiting effect to people's lung cancer NCI-H460 mice-transplanted tumor, and preferably T/C (%) is respectively 75.59;
Lid promise intravenously administrable 10mg/kg has certain growth-inhibiting effect to people's lung cancer NCI-H460 Nude Mice, and preferably T/C (%) is 56.69;
Morellic acid intravenously administrable 12mg/kg has certain growth-inhibiting effect to people's lung cancer NCI-H460 Nude Mice, and preferably T/C (%) is 70.55.
The experimental therapy of table 2 formula (1) compound to people's lung cancer NCI-H460 Nude Mice
D1 divides the RTV of cage administration time P administration group and RTV contrast *: the P<0.05**:P<0.01 of blank group
The extracorporeal anti-tumor function of embodiment 4 the compounds of this invention
1. experiment material
1.1 cell strain
U251: human glioma cell's strain, purchased from Chinese Academy of Sciences's cell bank;
A549: human lung adenocarcinoma cell line, purchased from Chinese Academy of Sciences's cell bank;
SPC-A-1: human lung adenocarcinoma cell line, purchased from Chinese Academy of Sciences's cell bank;
HO-8910: human oophoroma cell line, purchased from Chinese Academy of Sciences's cell bank.
1.2 tested material and reagent
Formula (1) compound: purity 97%, lot number 20100121, is ground by Chinese Academy of Sciences's Shanghai medicine
Study carefully prepared;
0.25% trypsinase-EDTA: U.S. GIBCO company;
MTT solution: purchased from Si Baihui bio tech ltd, Beijing;
PBS damping fluid: Ji Nuo biological medicine technology company limited;
Dimethyl sulfoxide (DMSO) (DMSO): analytical pure.
1.3 experimental technique
1.3.1 cell in vitro is cultivated
The cell Soviet Union of often restoring to norm is placed in incubator at 37 ℃, 5%CO
2and cultivate under saturated humidity condition, until Growth of Cells, during to exponential phase of growth, with 0.25% trypsinase-DTA digestion method, go down to posterity.Absorb old nutrient solution in bottle, with PBS, wash away residual nutrient solution, in bottle, add appropriate Digestive system (0.25% trypsinase-EDTA) again, make Digestive system submergence all cells surface, put and in 37 ℃ of incubators, hatch (time is depending on different cells), put under microscope and observe, after finding that kytoplasm retraction, intercellular substance increase, add immediately the complete culture solution containing serum to stop digestion, centrifugal (1000rpm, 5min), remove supernatant liquor, with counting after nutrient solution re-suspended cell, with cell count 3 * 10
5～5 * 10
5cells/mL is seeded in new culturing bottle, is placed in incubator and cultivates with above-mentioned culture condition, and 2～3d goes down to posterity once.
1.3.2MTT method detects
Growth of Cells is during to exponential phase of growth, with 0.25% trypsinase-DTA digestion, centrifugal (1000rpm, 5min), it is 2 * 10 that cell precipitation is adjusted cell count with perfect medium
4～3 * 10
4cells/mL, 96 holes are cultivated the every hole of version and are inoculated 190 μ L, at 37 ℃, 5%CO
2and cultivate under saturated humidity condition, after 24h, add formula (1) compound, making cumulative volume is 200 μ L, establishes 6 multiple holes, control wells adds PBS solution, continues to cultivate 48h, then adds the MTT that 20 μ L concentration are 5mg/mL, is placed in CO
237 ℃ of incubators are hatched, and discard nutrient solution after 4h, and every hole adds 150 μ LDMSO, and vibration 10min, puts in enzyme micro-plate reader and detect, and measures the OD value in each hole, calculating inhibiting rate.The IC of formula (1) compound to subject cell strain
50with SPSS15.0 software, by probit's weighted regression method, calculate.
Inhibiting rate calculation formula:
2. experimental result
Experimental result shows (as table 3), and formula (1) compound all has restraining effect to each tested tumour cell, and wherein human glioma U251 cell strain is the highest to the susceptibility of medicine.
The IC of table 3 formula (1) compound to each subject cell strain
50(
n=3)
The preparation of embodiment 5 tablets
Working method according to conventional tablet, more than mixes, and wet granulation finally adds Magnesium Stearate to mix and is pressed into tablet, and totally 50, every 500mg.
The preparation of embodiment 6 capsules
Working method according to conventional capsule, more than mixes, wet granulation, filling one-tenth capsule, totally 90, every 300mg.
The preparation of embodiment 7 pills
The compounds of this invention 10.0g
Polyethylene glycol 6000 25.0g
The compounds of this invention was pulverized to 100 mesh sieves, evenly added in the polyethylene glycol 6000 matrix of melting, stirred 30 minutes to even, take dimethyl silicone oil 100 as refrigerant, and 15-4 ℃ of gradient is cooling, and dripping becomes ball, the agent of centrifugal removal surface cool, obtains dripping pill 1000 balls.
The preparation of embodiment 8 powder ampoule agent for injection
The compounds of this invention 1.0g
Hydrochloric acid is appropriate
N.F,USP MANNITOL 50.0g
According to the operation of conventional freeze-dried powder, carry out, the compounds of this invention power 800ml water for injection dissolves, and adds N.F,USP MANNITOL, is settled to 1000ml, regulates pH value between 5.0-6.5, Sterile Filtration, lyophilize and get final product.
The preparation of embodiment 9 injections
The compounds of this invention is dissolved in to dehydrated alcohol, adds 20% Soxylat A 25-7 Viscotrol C to mix, reduction vaporization is removed ethanol, adds appropriate water for injection to be mixed into clear solution, through filtering with microporous membrane, and coating-dividing sealing, flowing steam sterilization and get final product.
The preparation of embodiment 10 slow releasing tablets
The compounds of this invention and polyvidone are dissolved in a small amount of ethanol, reduction vaporization ethanol, gained solid is crossed 100 mesh sieves; Above-mentioned solid and lactose, hypromellose are crossed to 60 mesh sieves and mix, add 3% HPMC (E5) aqueous solution softwood processed in right amount, cross 20 mesh sieves and granulate, forced air drying.Dry particle is crossed the whole grain of 20 mesh sieves, adds the talcum powder of recipe quantity, mixes compressing tablet and get final product.
The above is only the preferred embodiment of the present invention; it should be pointed out that for those skilled in the art, under the premise without departing from the principles of the invention; can also make some improvements and modifications, these improvements and modifications also should be considered as protection scope of the present invention.
Claims (15)
1. a formula (1) compound:
。
2. a medical composition, it comprises the compound according to claim 1 for the treatment of significant quantity, and at least one is selected from following pharmaceutically acceptable non-active ingredient: pharmaceutically acceptable thinner, pharmaceutically acceptable vehicle and pharmaceutically acceptable supporting agent.
3. medical composition according to claim 2, wherein said medical composition is tablet, capsule, powder ampoule agent for injection, injection, pill and slow releasing tablet.
4. according to the purposes of the compound described in claims 1 to 3 or medical composition, it is for the preparation of the medicine that is used for the treatment of tumor disease.
5. purposes according to claim 4, described tumor disease is selected from the group of following composition: neurogliocytoma, adenocarcinoma of lung and ovarian cancer.
6. purposes according to claim 5, described tumor disease is neurogliocytoma.
7. a method of preparing compound according to claim 1, it comprises the following steps:
(1) get gamboge medicinal material and extract by ethyl acetate, extract is separated through silica gel column chromatography, and petroleum ether-ethyl acetate gradient elution, obtains 11 component A～K;
(2) discard oil component and morellic acid component, through HPLC-PDA-MS, analyze, obtain emphasis component I;
(3) component I is separated through silica gel column chromatography again, chloroform-methanol gradient elution, and every 25～35ml collects a flow point, merges the 11st part to the 16th part elutriant, and flow velocity is 2mL/min, obtains flow point I-3;
(4) get I-3 through anti-phase RP-18 chromatographic separation, acetonitrile-water gradient elution, every 15～25ml collects a flow point, merges the 13rd part to the 18th part elutriant, and flow velocity is 2mL/min, obtains Arius and divides I-3E;
(5) purified, the acetonitrile isocratic elution of I-3E, obtains formula (1) compound.
8. method according to claim 7, wherein said step (1) PetroChina Company Limited. ether-ethyl acetate by volume ratio of 50～0:1 is carried out gradient elution.
9. method according to claim 8, wherein said petroleum ether-ethyl acetate is divided into six gradients and carries out wash-out, is respectively 50:1,10:1,5:1,2:1,1:1,0:1.
10. method according to claim 7, in wherein said step (3) chloroform-methanol by volume the ratio of 50～10:1 carry out gradient elution.
11. methods according to claim 7, the middle acetonitrile-water of wherein said step (4) the by volume ratio of 40～90:60～10 carries out gradient elution.
12. methods as claimed in claim 11, wherein acetonitrile-water is divided into five gradients and carries out wash-out, is respectively 40:60,50:50,60:40,80:20,90:10.
13. methods as claimed in claim 7, in wherein said step (5), the concentration of acetonitrile is 80%～95%.
14. methods according to claim 13, the concentration of wherein said acetonitrile is 80%～85%.
15. methods according to claim 14, the concentration of wherein said acetonitrile is 82%.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106039766A (en) * | 2016-05-17 | 2016-10-26 | 辽宁大学 | Method for separating epimers of gambogic acid |
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| US6462041B1 (en) * | 1999-05-21 | 2002-10-08 | Cytovia, Inc. | Gambogic acid, analogs and derivatives as activators of caspases and inducers of apoptosis |
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