Studies On Chemical Constituents From The Leaves Of Cajanus Cajan And Anticancer Activity

Pigeon pea[Cajanus cajan(Linn.)Millsp.]is a plant of leguminosae.The leaves of C ajanus cajan is a traditional Chinese herbal medicine.The main chemical components of C.cajan leaf are found as flavonoids and stilbenes by literatures.Various pharmacological eff ects including antipox virus,anti-inflammatory,alleviating chickenpox and swelling,anti-t umor,anti-oxidation,anti-osteoporosis,hypolipidemic,anti-microbial,and neuroprotection of C.cajan leaf or its chemical components have been revealed.Cajanonic acid A,cajanola ctone A.and cajastilbene H,which are new compounds with novel stilbenoid skeletone and isolated from the C cajan leaves by our research group showed interesting effects such as anti-osteoporosis,aiti-hyperlipidemic,anti-hyperglycemic.Further studies on chemical con stituents with novel bioactivity and structure from the leaves of C.cajan are meaningful for drug discovery and quality control of C.cajan leaves.ObjectiveSince flavonoids and stilbene compounds found from Cajanus cajan leaves have showed multiple pharmacological activities.It is valuable to continue to explore the chemical constituents with bioactivity and novel structure from the leaves of C.cajan.For the isolated new compound,molecular docking will be carried out to predict their probable anti-tumor activity.Such work will provide proof for demonstrating the efficacy and and mechanism of effect of this herbal medicine.Methods(1)A sample of dried and crushedCajanus cajan leaves is extracted with boiling 95%ethanol for 3 times by reflux,to give an ethanol extract.The extract is dissolved with ethanol and mixed throughly with silica gel(100-200 mesh),which has a weighed quantity,and performed a column chromatography on a silica gel column,eluting with mixed petroleum ether and acetone(95:5-70:30.v/v)and ethanol respectively,to afford a lipophilic fraction and a great polarity fraction.The above fractions are separated and purified by colum chrommatographies including normal-phase and reversed-phase silica gel,hydroxypropyl dextran gel(Sephadex LH-20),as well as preparative high performance liquid chromatography(HPLC),and the chemical structures were identified by comprehensive analyses on the data of 1H-NMR,13C-NMR,2D-NMR,MS,IR and UV.(2)The interaction of new chemical components isolated from Cajanus cajanleaves with drug-target in cancer cells is analyzed by AutoDock molecular docking technique,to predict probable anticaner effect and mechanism.(3)SRB assay is used to learn the inhibitory effect of the new compounds on the proliferation of human cancer cell lines which overexpress the drug-target molecules which are predicted by molecular docking analyse.Results(1)In this study,a total of sixteen compounds were isolated from Cajanus cajan Leaves,They are 3-hydroxy-5-methoxy-4-(2-hydroxy-3-methylbut-3-en-1-yl)stilbene(1),ethy heptadecanoate(2),cajanolatone A(3),pinostrobin(4),1-heneicosanol(5)longistylin C(6),β-Sitosterol(7),lupeol(8),3-hydroxy-5-methoxy-4-(3-methyl-2butenyl)bibenzyl(9),5-hydroxy-7,4’dimethoxyflavanone(10),Stigmasterol(11),3-methoxy5-hydroxystilbene(12),longistylinA(13),Cajanotone(14),Betulinic acid(15),4’,5,7Trihydroxyflavanone(16).Among them 3-hydroxy-5-methoxy-4-(2-hydroxy-3-methylbut3-en-1-yl)-stilbene(1)is a new compound,and 1-heneicosanol(5)are isolated and identified fromCajanus cajanfor the first time.(2)Through the anti-tumor targeted molecular docking of the compound 1 isolated from Cajanus cajan,the targets with the absolute value of the binding energy score greater than 8 were JAK2,PIK3CA,MTOR In which JAK2 and MTOR,which are overexpressed in human cancers such as gastric cancer,breast cancer,lung cancer,etc,possessed absolute value of binding energy score great than 9,suggested that the compound 1 may have strong binding with these two tumor target proteins and produce antitumor activity.(3)In this study,the inhibitory effect of the compoundl on human gastric cancer(SGC790)cells was tested.The experimental results showed that the compound 1 had an inhibitory effect on SGC790 cells with an IC50 value as 49.34±1.79 μg/mL.Conclusions(1)In this study,16 compounds were isolated from Cajanus cajan leaves and their chemical structures were identified by means of spectroscopic analyses,among them 3hydroxy-5-methoxy-4-(2-hydroxy-3-methylbut-3-en-1-yl)-stilbene(1)was a new compound,1-heneicosanol(5)were isolated and identified from this plant for the first time.Such findings provide new scientific basis for studies of chemical constituents and pharmacodynamic substance for plant Cajanus cajan.(2)The interaction of the compound 1 on anti-tumor-related targets analyzed by Autodock molecular docking technique showed that the absolute binding energy scores of the target proteins JAK2 and mTOR with compound 1 were 9.2 and 9.0,speculating that 1 may have inhibitory effect on the growth ofgastric cancer.breast cancer,and lung cancer.Our in vitro experiments proofed that compound 1 inhibit the proliferation of human gastric cancer(SGC790)cells,with IC50 value of 49.34± 1.79 μg/mL.