Studies On The Chemical Constituents From Clausena Hainanensis Huang Ex Xing And Their Antitumor Activities

There are about 30 species of the genus Clausena(Rutaceae)in worldwide,which distributed in the tropical and subtropical regions of the eastern hemisphere.There are about 10 species and 2 variety in China,growing the south of the Yangtze River,most species distributed in Hainan,Yunnan,Guangxi and Guangdong provinces.The fruit of the Clausena is not only a common tropical fruit,it have been used as folk medicinal plants.It has the functions of analgesia and detoxification.Studies on the chemical constituents of the Clausena have shown that there are many types of compounds in the Clausena,including alkaloids,coumarins,limonoids and so on.The results of modern pharmacological studies on the chemical constituents isolated from the Clausena show that most of this have a wide range of biological activities,such as anti-tumor,anti-inflammatory,antibacterial,anti-viral,and hepatoprotective.C.hainanensis is an endemic plant in Hainan.Until now,no research about its chemical composition and its pharmacological activity has been reported.Based on the work we did to evaluate the antitumor activity of the 90%ethanol extract from the branches and leaves of C.hainanensis in the previous study,and the results showed that it had significant anti-tumor activity.In order to develop and utilize this plant resource and display its medicinal value,we conducted a systematic study about its chemical composition.We take C.hainanensis as the research object,systematically studied the chemical composition of its branches and leaves.Then we conducted preliminary anti-tumor activity experiments on the compounds which have good activity and can make the theoretical and material basis for the development of effective and low-toxic anti-tumor drugs.The dried branches and leaves of C.hainanensis were crushed,then extracted by 90%ethanol,we recycle ethanol under reduced pressure.The extractum has extracted by prtroleum ether and ethyl acetate,respectively.Then the extracts are isolated and purified with the method of reversed-phase Cis silica gel column chromatography and HPLC,silica gel,Sephadex LH-20.We used MS,NMR,UV,IR spectroscopy technology and their physical and chemical properties,as well as the data compared to the literature to identify the structures.Sixty-six coumpounds were isolated from the extract of C.hainanensis,which were identified as clausehainanine A(1),clausehainanine B(2),clausehainanine D(3),clausehainanine E(4),clausehainanine C(5),indizoline(6),clauemarazole E(7),clauemarazole C(8),claulansine B(9),claulansine A(10),mafaicheenamine A(11),claulansine F(12),7-methoxymurrayacine(13),mafaicheenamine C(14),clausenaline B(15),claulansine G(16),clausenaline D(17),3-formylcarbazole(18),3-formyl-6-methoxy-carbazole(19),murrayanine(20),3-formyl-2-hydroxy-6-methoxy carbazole(21),methyl carbazole-3-carboxylate(22),mukonine(23),clausine X(24),8-methoxypsoralen(25),6,8-dimethoxy-4,5-dimethyl-3-methyleneisochromanl-one(26),isogosferol(27),imperatorin(28),pabularinone(29),heraclenol(30),lansiumarin A(31),melicotriphyllin C(32),8-[(2E)-6-oxo-3,7-dimethyloct-2-enyloxylpsoralen(33),clausenalansimin A(34),(E,E)-8-(7-hydroxy-3,7-dimethylocta-2,5-dienyloxy)psoralen(35),melicotriphyllin A(36),claucoumarin D(37),claucoumarin A(38),clauhainanin A(39),indicolactone(40),9-[3-methyl-4-(4-methyl-5-oxo-tetrapydro-furan-2-yl)-but-2-enyloxy]-fur-o[3,2-g]chromen-7-one(41),N-phenethylbenzamide(42),N-benzoyltyramine(43),N-benzoyltyrarnine methyl ether(44),tembamid(45),(E)-N-methyl cinnamon amide(46),(E)-N-(4-methoxyphenethyl)-2-methylbut-2-enamide(47),N-2-phenylethylcinnamamide(48),N-(2-hydroxy-2-phenylethyl)cinnamide(49),clausenalansamide B(50),benzenepropanamide(51),SB-204900(52),aurantiamide(53),aurantiamide acetate(54),4-methoxy-2(1H)-quinolinone(55),4-methoxy-N-methyl-2-quinol-one(56),isohematinic acid(57),lansamide 4(58),(-)-homoclausenamide(59),lansamide 3(60),6-O-methyl-epi-cisneoclausenamide(61),zeta-clausenamide(62),N-methyl-2-pyrolidinone(63),coumaric acid(64),isololiolide(65),pinoresinol(66).Among them,compound 1-24 were carbazole alkaloid,25-41 were coumarins,42-63 were amide alkaloids,64 was a phenolic,65 was a terpenoid,66 was a lignin.Compounds 1-5 and 39 were new compounds.The compounds 64-66 were isolated from the plant of the Clausena for the first time.Finally,we used the modified MTT to evaluate the antitumor activities of compounds 1-8,10,15-16,33-34,36-37 and 39 which were isolated from C.hainanensis.The results showed that the IC50 value of the coumarin compounds was 1.36-18.96 ?M,and the IC50 value of the carbazole alkaloids was 0.12-15.86 ?M.The tested compounds all have significant antitumor activity,compounds 1,2 and 39 were even stronger than that of the positive control.This provides some basis for the study of the antitumor activity of C.hainanensis,at the same time,it provides a new resource to expore efficient,low-toxic antitumor drugs.