Chemical constituents and bioactivities of the fruits and leaves of phaleria macrocarpa (scheff.) boerl

Abstract

Phytochemicals and bioactivities of the fruits and the leaves of Phaleria macrocarpa have been studied. Successive extraction of the dried fruits using cold extraction followed by fractionation and purification afforded two benzophenones identified as 2,6,4'-trihydroxy-4-methoxybenzophenone and 6,4'-dihydroxy-4-methoxybenzophenone-2-O-ß-D-glucopyranoside and two triterpenoids identified as 24-methylenecycloartan-3-one and 24-methyl-9,19-cyclolanost-25-en-3-ol. Meanwhile, cold extraction of the dried leaves followed by fractionation and purification yielded two sterols known as stigmasterol and ß-sitosterol. Fractionation and purification process were done using vacuum liquid chromatography and column chromatography, respectively. The structures of isolated compounds were elucidated on the basis of their spectral data including 1D and 2D Nuclear Magnetic Resonance (1H, 13C, DEPT, COSY, HMQC, HMBC), Infrared, Ultraviolet spectroscopies and mass spectrometry. The antibacterial activities of all crude extracts and isolated compounds were performed using disc diffusion method followed by minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) against two Gram-positive bacterial strains, Bacillus subtilis and Staphylococcus aureus together with two Gram-negative bacterial strains including Escherichia coli and Pseudomonas putida. The results have demonstrated that all extracts and isolated compounds exhibited weak activity against all tested bacteria with the MBC values exceeded 1000 µg/mL. Evaluation of antioxidant activity using 2,2-diphenyl-1-picrylhydydrazyl (DPPH) assay has revealed strong free radical scavenging properties of ethyl acetate and ethanol extracts from the fruits with SC50 of 14.00 and 19.97 µg/mL, respectively. The chloroform extract from the fruits and all extracts from the leaves exhibited moderate antioxidant activities with SC50 ranging between 58.07 – 94.10 µg/mL. Among the isolated compounds, 2,6,4'-trihydroxy-4-methoxy- benzophenone exhibited high free radical scavenging activity with SC50 of 29.73 µg/mL whereas the other isolated compounds were inactive against DPPH. MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay was carried out to evaluate the cytotoxic activity of crude extracts and isolated compounds. MTT assay of CHO, MCF-7, MDA-MB-231 and MDA-MB-468 cells were treated with all extracts and isolated compounds whereas the cell viability of 3T3, HeLa and HT-29 were treated by extracts of the fruits and two isolated benzophenones. The result has demonstrated good activity of dichloromethane extract from the leaves against MDA-MB-231 cell line (IC50 70 µg/mL) and moderate cytotoxic effect of ethanol extract from the fruits towards MDA-MB-468 and HT-29 cell lines with the same IC50 of 100 µg/mL. Chloroform extract from the fruits was found to have moderate cytotoxic properties against HT-29 cell line (IC50 100 µg/mL). All isolated compounds were found to have weak cytotoxic activity towards all tested cell lines.