Study On The Antitumor Constituents Of Alocasia Macrorrhiza (L.) Schott

Alocasia macrorrhiza（L.）Schott,belongs to the genus of Alocasia of the Araceae family and it is mainly distributed in Southern and Southwestern China.It is commonly used in the traditional Chinese medicine for clearing away heat and detoxifying,promoting Qi and relieving pain,dispersing knot and detumescence.It is mainly used to treat malaria,swelling and pain,and treating various malignant tumors.Modern pharmacological studies have shown that A.macrorrhiza has anti-tumor,anti-inflammatory,antipyretic and analgesic activities.In this study,the fresh rhizomes of A.macrorrhiza were extracted with 95%ethanol.Then extract with CH₂Cl₂,Et OAc,and n-Bu OH.Then the dichloromethane fraction was isolated and purified using various chromatographic methods,such as normal phase chromatography,reverse phase chromatography,gel filtration chromatography and Semi-HPLC.A total of 46 compounds were identified by chemical and spectroscopic techniques.The identified compounds were:（1）Alkaloids:（2S,3R,5S,6R）-2-methyl-3,5-dihydroxy-6-（9-phenylnonyl）piperidine（1*）,（2R,3R,4R,6S）-N,2-dimethyl-3,4-dihydroxy-6-（9-phenylnonyl）piperidine（2*）,（2S,3S,4S,6R）-N,2-dimethyl-3,4-dihydroxy-6-（9-phenylnonyl）piperidine（3*）,（2S,3S,4S,6R）-2-methyl-3,4-dihydroxy-6-（9-phenylnonyl）piperidine（4*）,（2R,3R,6S）-2-hydroxy methyl-3-hydroxy-6-（9-phenylnonyl）piperidine（5*）,（2S,3R,6R）-2-hydroxymethyl-3-hydroxy-6-（9-phenylnonyl）piperidine（6*）,（2R,3S,6S）-2-methyl-3-hydroxy-6-（7-phenylnonyl）piperidine（7*）,（2S,3R,6R）-2-methyl-6-（9-phenylnonyl）piperidin-3-ol（8）,（2S,3S,6S）-2-methyl-6-（9-phenylnonyl）piperidin-3-ol（9）,（-）-（2R,3S,6S）-N,2-dimethyl-3-hydroxy-6-（9-phenylnonyl）piperidine（10）,（2R,3R,4S,6S）-2-methyl-6-（9-phenylnonyl）piperidine-3,4-diol（11）,（2S,3R,6R）-2-methyl-6-（1-phenylnonan-5-one-9-yl）piperidin-3-ol（12）,1H-indole-3-carbaldehyde（13）,5-hydroxy-1H-indole-3-glyoxylate methyl ester（14）,5-hydroxy-1H-indole-3-carboxylic acid ethyl ester（15）,（S）-（-）-N-benzoylphenylalaninol（16）,（2）Terpenoids:2,19α-dihydroxy-3-oxo-24-noroleana-1,4,12-trien-28-oic acid（17*）,2-hydroxy-3-oxo-24-noroleana-1,4,12-trien-28-oic acid（18）,2-hydroxy-3-oxo-24-norursa-1,4,12-trien-28-oic acid（19）,4-oxo-19α-hydroxy-3,24-dinor-2,4-secoolean-12-en-2,28-dioic acid（20）,oleanolic acid（21）,ursolic acid（22）,2α,3β,23-trihydroxyurs-12,18-dien-28-oic acidβ-D-glucopyranoside ester（23）,quadranosideⅧ（24）,3β-hydroxy-cycloart-24-en-26-carboxylic acid（25）,（6S,9R）-9-hydroxy-4-megastigmen-3-one（26）,（3）pinoresinol（27）,epipinoresinol（28）,syringaresinol（29）,lirioresinol-A（30）,4-hydroxymellein（31）,8-methoxymellein（32）,（4）Sterols:5,8-epidioxy-（3β,5α,8α,22E）-ergosta-6,9（11）,22-trien-3-ol（33）,（3β,5α,8α,22E）-5,8-diepoxy-ergosta-6,22-dien-3-ol（34）,（22E,24R-5α,6α）-epoxyergosta-8,22-diene-3β,7α-diol（35）,（5）Others:simplicildone C（36*）,simplicildone B（37）,preussomerin BG1（38）,preussomerin C（39）,preussomerin SA1（40）,preussomerin A（41）,preussomerin B（42）,virescenoside B（43）,p-hydroxybenzaldehyde（44）,syringylethanone（45）,4,8-dihydroxy-1-tetralone（46）,1-linoleoylglycerol（47）,linoleic acid（48）.The identified48 compounds included 16 alkaloids（1-16）,10 terpenoids（17-26）,6phenylpropanoids（27-32）,3 sterols（33-35）,and 13 other compounds（36-48）.Among them,1-7,17,and 36 were 9 novel compounds.The antitumor activities of the isolated compounds were evaluated in this study.Compounds 1-6,and 8-11 were evaluated for their anti-proliferative activities against Hep G2,AGS,MCF-7 and CNE-1 tumor cell lines by MTT method.The results showed that all the ten compounds were indicated obvious inhibitory activity comparable to the positive control（cisplatin）.Among them,compounds 8,9,and 11 exhibited pronounced antiproliferative against AGS(IC₅₀=9.54±0.43,8.77±0.45,8.14±0.59μM)than cisplatin(IC₅₀=11.39±1.22μM).In addition,all 10 compounds were tested cytotoxic to Hacat cells and the toxicity was comparable to cisplatin.Based on the activity assay results,the colony formation assay,the cell morphology observation and Cell apoptosis analysis were performed in this study and suggested that compounds 3-5 all significantly inhibit the proliferation of AGS cells,obviously change the cell morphology of AGS cells,and induce AGS cell apoptosis in a dose-dependent manner.Taken together,we suggest that piperidine alkaloids may exert antiproliferative effects by inducing tumor cell apoptosis.This study provided the chemical basis for further development of A.macrorrhiza.