Studies On Chemical Constituents From Ganoderma Lucidum And Their Cytotoxic Activities

BACKGROUND AND OBJECTIVE At present chemotherapy,surgery and radiotherapy are the main ways to treat the malignant tumors,and chemotherapy is also very important.But the toxic reactions of medicine and the emergence of resistance are the main causes of chemotherapy failure.So how to improve the therapeutic index and break through these two barriers of chemotherapy have been hot in research for the moment.Now finding anticancer chemical compositions,with low toxicity and high efficiency from Chinese herbal medicines,has become one of the development strategies of anticancer drug.Ganoderma lucidum is Basidiomycotina,Hymenomycetes,Homobasidiomycetes,Aphyllophorales,Ganodermataceae,Ganoderma fruiting bodies,which as a traditional Chinese medicine in China has 2,000 years of application history.Habitually alleged Ganoderma lucidum is Ganoderma lucidum(Leyss.ex Fr.)Karst fruiting body part,which contains a variety of chemical constituents,including polysaccharides,triterpenes,sterols,alkaloids,furan derivatives,amino peptide,inorganic elements,fatty acids etc.Triterpenoids which has a lot of pharmacological activities,such as anti-tumor effect,the protection of liver function,resisting bacteria and eliminating inflammation,antihypertensive effect etc.Among which,the anti-tumor effect of Ganoderma triterpenoids has caused people great attention,but the study of the isolation and purification and their mechanism of action of anti-tumor active ingredients of Ganoderma triterpenoids have been a hot issue.STUDY METHODS Reflux extraction,liquid-liquid extraction,silica gel thin-layer chromatography,gel permeation chromatographyand semi-preparative HPLC and other means were used to carry out the extraction and purification of Ganoderma systems;structures were identified by spectral analysis and chemical methods and other means;MTT assay was used to detect test drugs' anti-tumor active,tolerance and reversal activity;cell apoptosis were detected by flow cytometry.RESULTS1.We had get 3 new compounds from Ganoderma lucidum,which were named Compd.7,Compd.14,Compd.17.And 14 known chemical compounds were:ergosterol(1),ganoderol B(2),ganodermanontriol(3),ganoderic acid S1(4),ergosterol peroxide(5),ergosta-7,22-dien-3?-ol(6),7-oxo-ganoderic acid Z(8),ganodermatriol(9),ganoderiol F(10),ganodermanondiol(11),ganoderone A(12),lucialdehyde C(13),ganoderic acid DM(15),5?-lanosta-7,9(11),24-triene-15?,26-dihydroxy-3-one(16).There was one compound without structural identification,because of its quality very little,which was named Compound 18.2.Compd.11 had the growth inhibition on K562 tumor cells for 48 h,and its IC50 values was 2.56?g/ml;Compd.5,Compd.9,Compd.10,Compd.11,Compd.16 and Compd.18 could obviously inhibit the growth of HL-60 tumor cells,and their IC50 values were 2.8?g/ml,4.32?g/ml,2.93?g/ml,3.55?g/ml,1.92?g/ml,2.15?g/ml respectively;MDA-MB-453,BT474 and KBv200 tumor cells were not sensitive to almost compounds,but only the effect of Compd.18 was obvious,and their IC50 values were 2.71?g/ml,10.13?g/ml,6.26?g/ml respectively;SW-620 cells was very sensitive to Compd.17 and Compd.18,and their IC50 values were 3.71?g/ml,2.75?g/ml respectively;Compd.11 and Compd.18 had the growth inhibition on HCT-116 tumor cells,and their IC50 values were 4.96?g/ml,11.03?g/ml respectively;Compd.9,Compd.16 and Compd.17 had the growth inhibition on OE-19 tumor cells,and their IC50 values were 6.40?g/ml,8.15?g/ml,6.25?g/ml respectively;EC-9706,KB and K562/A02 were not sensitive to all of compounds.3.Apoptosis AnnexinV-FITC Joint PI staining showed that in the 2.5?g/ml and10?g/ml concentrations,the apoptotic rate of Compd.13 on K562 cell for 48 h increased with increase of concentration.But Compd.11,Compd.15 and Compd.8,these three compounds had little effect on cell apoptosis.4.Under the concentration of IC10 value,we had tested the reversal activity of every compound with ADR.On resistant cell line KBv200 for 48 h,the reversal times of compd.2,compd.8,compd.10,compd.11,compd.12 and ganoderiol B in the10?g/ml concentration were 6.58 times,4.91 times,7.05 times,4.01 times,4.71 times and 3.42 times respectively;in the 10?g/ml and 5?g/ml concentrations,the reversal times of compd.17 and compd.7 were 11.12,5.25 and 13.27,5.96 respectively;in the5?g/ml,the reversal times of compd.3 was 3.23;in the 2.5?g/ml concentration,the reversal times of compd.9 was 3.97.On resistant cell line K562/A02 for 48 h,the reversal times of ganoderiol B in the 10?g/ml concentration was 5.53;in the 10?g/ml and 5?g/ml concentrations,the reversal times of compd.17 were 15.75 and 7.36respectively;the reversal times of Compd.16 was 16.37 in the 2.5?g/ml concentration.CONCLUSION Ganoderma component chemicals had some anti-tumor activity,which could more obviously inhibit the growth of human leukemia cells(K562,HL-60)and Human colon cancer cells(SW-620,HCT-116)and Human esophageal carcinoma cells(OE-19)than the human breast cancer,and hardly affected the growth of the multidrug resistance tumor cells(K562/A02,KBv200);and compd.18 can obviously inhibit the growth of most tumor cells,the same to the resistance tumor cells,which showed that it can be used as a potential anti-cancer drugs.Detected by flow cytometry,the result of Cell apoptosis showed that the main active ingredient inducing apoptosis were the neutral component of Ganoderma triterpenoids.They also had a certain role in reversing multidrug resistance of tumor cells K562/A02 and KBv200 on adriamycin,especially the two new compounds—compound 7 and compound 17.